Results of liver transplantation in patients with acute liver failure due to Amanita phalloides and paracetamol (acetaminophen) intoxication

PubMed Central

Grąt, Michał; Hołówko, Wacław; Masior, Łukasz; Wronka, Karolina M.; Grąt, Karolina; Stypułkowski, Jan; Patkowski, Waldemar; Krawczyk, Marek

2015-01-01

Introduction Amanita phalloides and paracetamol intoxications are responsible for the majority of acute liver failures. Aim To assess survival outcomes and to analyse risk factors affecting survival in the studied group. Material and methods Of 1369 liver transplantations performed in the Department of General, Transplant, and Liver Surgery, Medical University of Warsaw before December 2013, 20 (1.46%) patients with Amanita phalloides (n = 13, 0.95%) and paracetamol (n = 7, 0.51%) intoxication were selected for this retrospective study. Overall and graft survival at 5 years were set as primary outcome measures. Results Five-year overall survival after liver transplantation in the studied group was 53.57% and 53.85% in patients with paracetamol and Amanita phalloides poisoning, respectively (p = 0.816). Five-year graft survival was 26.79% for patients with paracetamol and 38.46% with Amanita phalloides intoxication (p = 0.737). Risk factors affecting patient survival were: pre-transplant bilirubin concentration (p = 0.023) and higher number of red blood cells (p = 0.013) and fresh frozen plasma (p = 0.004) transfused intraoperatively. Likewise, higher number of red blood cells (p = 0.012) and fresh frozen plasma (p = 0.007) transfused were risk factors affecting 5-year graft survival. Surprisingly, donor and recipient blood type incompatibility was neither the risk factor for 5-year overall survival (p = 0.939) nor the risk factor for 5-year graft survival (p = 0.189). Conclusions In selected intoxicated patients urgent liver transplantation is the only successful modality of treatment. Risk factors affecting survival are in correspondence with the patient's pre-transplant status (bilirubin level in serum) and intraoperative status (number of red blood cells and fresh frozen plasma transfused). PMID:27350835

Approach to intraoperative consultation for donor liver biopsies.

PubMed

Melin, Claire; Miick, Ronald; Young, Nancy A; Ortiz, Jorge; Balasubramanian, Manjula

2013-02-01

As demand for organs to treat end-stage liver disease increases, donor livers once deemed only marginally suitable for donation are being considered for transplantation. Pathologists are increasingly being asked to evaluate these livers for acceptability. This article provides guidelines for frozen section evaluation of livers for transplantation. This article concentrates on the histopathologic features of transplant suitability with appropriate clinicopathologic correlation for the practicing pathologist. Recommendations for proper handling and sampling of tissue are discussed. Relative and absolute contraindications as well as artifacts and benign conditions are emphasized. Sources include a compilation of the authors' experiences in academic and community liver transplantation centers. In addition, relevant medical literature was reviewed, as well as Web sites specializing in organ transplantation, such as Transplant Pathology Internet Services and the Organ Procurement and Transplantation Network. Malignancy and extensive necrosis in the liver are absolute contraindications to transplantation. Evaluation of macrosteatosis, fibrosis, hepatitis, and necrosis depends on the severity of disease and correlation with the clinical situation. Donor age of greater than 60 years does not preclude transplantation. Artifacts and benign conditions need to be understood to prevent wastage of precious organs and to ensure that an appropriate organ is provided for the recipient.

9 CFR 319.182 - Braunschweiger and liver sausage or liverwurst.

Code of Federal Regulations, 2013 CFR

2013-01-01

... liverwurst. 319.182 Section 319.182 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE... made from fresh, cured, and/or frozen pork, beef, and/or veal and at least 30 percent pork, beef, and/or veal livers computed on the weight of the fresh livers. It may also contain pork and/or beef fat...

9 CFR 319.182 - Braunschweiger and liver sausage or liverwurst.

Code of Federal Regulations, 2012 CFR

2012-01-01

... liverwurst. 319.182 Section 319.182 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE... made from fresh, cured, and/or frozen pork, beef, and/or veal and at least 30 percent pork, beef, and/or veal livers computed on the weight of the fresh livers. It may also contain pork and/or beef fat...

9 CFR 319.182 - Braunschweiger and liver sausage or liverwurst.

Code of Federal Regulations, 2014 CFR

2014-01-01

... liverwurst. 319.182 Section 319.182 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE... made from fresh, cured, and/or frozen pork, beef, and/or veal and at least 30 percent pork, beef, and/or veal livers computed on the weight of the fresh livers. It may also contain pork and/or beef fat...

9 CFR 319.182 - Braunschweiger and liver sausage or liverwurst.

Code of Federal Regulations, 2010 CFR

2010-01-01

... liverwurst. 319.182 Section 319.182 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE... made from fresh, cured, and/or frozen pork, beef, and/or veal and at least 30 percent pork, beef, and/or veal livers computed on the weight of the fresh livers. It may also contain pork and/or beef fat...

9 CFR 319.182 - Braunschweiger and liver sausage or liverwurst.

Code of Federal Regulations, 2011 CFR

2011-01-01

... liverwurst. 319.182 Section 319.182 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE... made from fresh, cured, and/or frozen pork, beef, and/or veal and at least 30 percent pork, beef, and/or veal livers computed on the weight of the fresh livers. It may also contain pork and/or beef fat...

Contribution of Transjugular Liver Biopsy in Patients with the Clinical Presentation of Acute Liver Failure

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miraglia, Roberto, E-mail: rmiraglia@ismett.edu; Luca, Angelo; Gruttadauria, Salvatore

2006-12-15

Purpose. Acute liver failure (ALF) treated with conservative therapy has a poor prognosis, although individual survival varies greatly. In these patients, the eligibility for liver transplantation must be quickly decided. The aim of this study was to assess the role of transjugular liver biopsy (TJLB) in the management of patients with the clinical presentation of ALF. Methods. Seventeen patients with the clinical presentation of ALF were referred to our institution during a 52 month period. A TJLB was performed using the Cook Quick-Core needle biopsy. Clinical data, procedural complications, and histologic findings were evaluated. Results. Causes of ALF were virusmore » hepatitis B infection in 7 patients, drug toxicity in 4, mushroom in 1, Wilson's disease in 1, and unknown origin in 4. TJLB was technically successful in all patients without procedure-related complications. Tissue specimens were satisfactory for diagnosis in all cases. In 14 of 17 patients the initial clinical diagnosis was confirmed by TJLB; in 3 patients the initial diagnosis was altered by the presence of unknown cirrhosis. Seven patients with necrosis <60% were successfully treated with medical therapy; 6 patients with submassive or massive necrosis ({>=}85%) were treated with liver transplantation. Four patients died, 3 had cirrhosis, and 1 had submassive necrosis. There was a strict statistical correlation (r = 0.972, p < 0.0001) between the amount of necrosis at the frozen section examination and the necrosis found at routine histologic examination. The average time for TJLB and frozen section examination was 80 min. Conclusion. In patients with the clinical presentation of ALF, submassive or massive liver necrosis and cirrhosis are predictors of poor prognosis. TLJB using an automated device and frozen section examination can be a quick and effective tool in clinical decision-making, especially in deciding patient selection and the best timing for liver transplantation.« less

Robust transcriptional tumor signatures applicable to both formalin-fixed paraffin-embedded and fresh-frozen samples

PubMed Central

Cheng, Jun; He, Jun; Liu, Huaping; Cai, Hao; Hong, Guini; Zhang, Jiahui; Li, Na; Ao, Lu; Guo, Zheng

2017-01-01

Formalin-fixed paraffin-embedded (FFPE) samples represent a valuable resource for clinical researches. However, FFPE samples are usually considered an unreliable source for gene expression analysis due to the partial RNA degradation. In this study, through comparing gene expression profiles between FFPE samples and paired fresh-frozen (FF) samples for three cancer types, we firstly showed that expression measurements of thousands of genes had at least two-fold change in FFPE samples compared with paired FF samples. Therefore, for a transcriptional signature based on risk scores summarized from the expression levels of the signature genes, the risk score thresholds trained from FFPE (or FF) samples could not be applied to FF (or FFPE) samples. On the other hand, we found that more than 90% of the relative expression orderings (REOs) of gene pairs in the FF samples were maintained in their paired FFPE samples and largely unaffected by the storage time. The result suggested that the REOs of gene pairs were highly robust against partial RNA degradation in FFPE samples. Finally, as a case study, we developed a REOs-based signature to distinguish liver cirrhosis from hepatocellular carcinoma (HCC) using FFPE samples. The signature was validated in four datasets of FFPE samples and eight datasets of FF samples. In conclusion, the valuable FFPE samples can be fully exploited to identify REOs-based diagnostic and prognostic signatures which could be robustly applicable to both FF samples and FFPE samples with degraded RNA. PMID:28036264

FROZEN THIN SECTIONS OF FRESH TISSUE FOR ELECTRON MICROSCOPY, WITH A DESCRIPTION OF PANCREAS AND LIVER

PubMed Central

Christensen, A. Kent

1971-01-01

A simple method has been developed that allows frozen thin sections of fresh-frozen tissue to be cut on a virtually unmodified ultramicrotome kept at room temperature. A bowl-shaped Dewar flask with a knifeholder in its depths replaces the stage of the microtome; a bar extends down into the bowl from the microtome's cutting arm and bears the frozen tissue near its lower end. When the microtome is operated, the tissue passes a glass or diamond knife in the depths of the bowl as in normal cutting. The cutting temperature is maintained by flushing the bowl with cold nitrogen gas, and can be set anywhere from about -160°C up to about -30°C. The microtome is set for a cutting thickness of 540–1000 A. Sections are picked up from the dry knife edge, and are placed on membrane-coated grids, flattened with the polished end of a copper rod, and either dried in nitrogen gas or freeze-dried. Throughout the entire process the tissue is kept cold and does not come in contact with any solvent. The morphology seen in frozen thin sections of rat pancreas and liver generally resembles that in conventional preparations, although freezing damage and low contrast limit the detail that can be discerned. Among unusual findings is a frequent abundance of mitochondrial granules in material prepared by this method. PMID:4942776

Campylobacters and their bacteriophages from chicken liver: The prospect for phage biocontrol.

PubMed

Firlieyanti, Antung S; Connerton, Phillippa L; Connerton, Ian F

2016-11-21

Consumption of foods containing chicken liver has been associated with Campylobacter enteritis. Campylobacters can contaminate the surface of livers post-mortem but can also arise through systemic infection of colonising bacteria in live birds. The use of bacteriophage to reduce levels of Campylobacter entering the food chain is a promising intervention approach but most phages have been isolated from chicken excreta. This study examined the incidence and contamination levels of Campylobacter and their bacteriophage in UK retail chicken liver. Using enrichment procedures, 87% of 109 chicken livers were surface contaminated with Campylobacter and 83% contaminated within internal tissues. Direct plating on selective agar allowed enumeration of viable bacteria from 43% of liver samples with counts ranging from 1.8->3.8log 10 CFU/cm 2 for surface samples, and 3.0->3.8log 10 CFU/g for internal tissue samples. Three C. jejuni isolates recovered from internal liver tissues were assessed for their ability to colonise the intestines and extra-intestinal organs of broiler chickens following oral infection. All isolates efficiently colonised the chicken intestines but were variable in their abilities to colonise extra-intestinal organs. One isolate, CLB104, could be recovered by enrichment from the livers and kidneys of three of seven chickens. Campylobacter isolates remained viable within fresh livers stored at 4°C over 72h and frozen livers stored at -20°C over 7days in atmospheric oxygen, and therefore constitute a risk to human health. Only three Campylobacter-specific bacteriophages were isolated, and these exhibited a limited host range against the Camplylobacter chicken liver isolates. All were identified as group III virulent bacteriophage based on their genome size of 140kb. The application of broad host range group II virulent phages (8log 10 PFU/g) to liver homogenates containing C. jejuni strains of diverse origin at 4°C resulted in modest but significant reductions in the viable counts ranging from 0.2 to 0.7log 10 CFU/g. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Helicobacter marmotae sp. nov. isolated from livers of woodchucks and intestines of cats.

PubMed

Fox, James G; Shen, Zeli; Xu, Shilu; Feng, Yan; Dangler, Charles A; Dewhirst, Floyd E; Paster, Bruce J; Cullen, John M

2002-07-01

Woodchucks (Marmota monax) have a high incidence of hepatocellular carcinoma (HCC) associated with chronic infection with woodchuck hepatitis virus (WHV) and serve as a model of hepatitis B virus-associated HCC in humans. Helicobacter hepaticus, an enterohepatic helicobacter in mice, is known to cause hepatocellular adenomas and carcinomas in susceptible mouse strains. In long-term chemical bioassays conducted with B6C3F(1) mice, H. hepaticus has been regarded as a confounding factor because of its tumor-promoting activity. In order to determine if woodchucks harbor a Helicobacter sp. that might play a role in potentiating hepatic inflammation or neoplasia, a study was undertaken to determine whether woodchucks' livers were infected with a Helicobacter sp. Frozen liver samples from 20 (17 WHV-infected and 3 noninfected) woodchucks, 10 with WHV-associated hepatic tumors and 10 without tumors, were cultured by microaerobic techniques and analyzed by using genus- and species-specific helicobacter PCR primers. A 1,200-bp Helicobacter sp.-specific sequence was amplified from 14 liver samples. Southern hybridization confirmed the specific identity of the PCR products. Nine of the 10 livers with tumors had positive Helicobacter sp. identified by PCR, whereas 5 of the 10 livers without tumors were positive. By use of 16S rRNA species-specific primers for H. marmotae, two additional liver samples from the nontumor group had positive PCR amplicons confirmed by Southern hybridization. A urease-, catalase-, and oxidase-positive bacterium was isolated from one liver sample from a liver tumor-positive woodchuck. By 16S rRNA analysis and biochemical and phenotypic characteristics, the organism was classified as a novel Helicobacter sp. Subsequently, four additional bacterial strains isolated from feces of cats and characterized by biochemical, phenotypic, and 16S rRNA analysis were determined to be identical to the woodchuck isolate. We propose the name Helicobacter marmotae sp. nov. for these organisms. Further studies are required to ascertain if this novel Helicobacter sp. plays a tumor promotion role in hepadnavirus-associated tumors in woodchucks or causes enterohepatic disease in cats.

Helicobacter marmotae sp. nov. Isolated from Livers of Woodchucks and Intestines of Cats

PubMed Central

Fox, James G.; Shen, Zeli; Xu, Shilu; Feng, Yan; Dangler, Charles A.; Dewhirst, Floyd E.; Paster, Bruce J.; Cullen, John M.

2002-01-01

Woodchucks (Marmota monax) have a high incidence of hepatocellular carcinoma (HCC) associated with chronic infection with woodchuck hepatitis virus (WHV) and serve as a model of hepatitis B virus-associated HCC in humans. Helicobacter hepaticus, an enterohepatic helicobacter in mice, is known to cause hepatocellular adenomas and carcinomas in susceptible mouse strains. In long-term chemical bioassays conducted with B6C3F1 mice, H. hepaticus has been regarded as a confounding factor because of its tumor-promoting activity. In order to determine if woodchucks harbor a Helicobacter sp. that might play a role in potentiating hepatic inflammation or neoplasia, a study was undertaken to determine whether woodchucks' livers were infected with a Helicobacter sp. Frozen liver samples from 20 (17 WHV-infected and 3 noninfected) woodchucks, 10 with WHV-associated hepatic tumors and 10 without tumors, were cultured by microaerobic techniques and analyzed by using genus- and species-specific helicobacter PCR primers. A 1,200-bp Helicobacter sp.-specific sequence was amplified from 14 liver samples. Southern hybridization confirmed the specific identity of the PCR products. Nine of the 10 livers with tumors had positive Helicobacter sp. identified by PCR, whereas 5 of the 10 livers without tumors were positive. By use of 16S rRNA species-specific primers for H. marmotae, two additional liver samples from the nontumor group had positive PCR amplicons confirmed by Southern hybridization. A urease-, catalase-, and oxidase-positive bacterium was isolated from one liver sample from a liver tumor-positive woodchuck. By 16S rRNA analysis and biochemical and phenotypic characteristics, the organism was classified as a novel Helicobacter sp. Subsequently, four additional bacterial strains isolated from feces of cats and characterized by biochemical, phenotypic, and 16S rRNA analysis were determined to be identical to the woodchuck isolate. We propose the name Helicobacter marmotae sp. nov. for these organisms. Further studies are required to ascertain if this novel Helicobacter sp. plays a tumor promotion role in hepadnavirus-associated tumors in woodchucks or causes enterohepatic disease in cats. PMID:12089272

NASA's Rodent Research Project: Validation of Flight Hardware, Operations and Science Capabilities for Conducting Long Duration Experiments in Space

NASA Technical Reports Server (NTRS)

Choi, S. Y.; Beegle, J. E.; Wigley, C. L.; Pletcher, D.; Globus, R. K.

2015-01-01

Research using rodents is an essential tool for advancing biomedical research on Earth and in space. Rodent Research (RR)-1 was conducted to validate flight hardware, operations, and science capabilities that were developed at the NASA Ames Research Center. Twenty C57BL/6J adult female mice were launched on Sept 21, 2014 in a Dragon Capsule (SpaceX-4), then transferred to the ISS for a total time of 21-22 days (10 commercial mice) or 37 (10 validation mice). Tissues collected on-orbit were either rapidly frozen or preserved in RNA later at less than or equal to -80 C (n=2/group) until their return to Earth. Remaining carcasses were rapidly frozen for dissection post-flight. The three controls groups at Kennedy Space Center consisted of: Basal mice euthanized at the time of launch, Vivarium controls, housed in standard cages, and Ground Controls (GC), housed in flight hardware within an environmental chamber. FLT mice appeared more physically active on-orbit than GC, and behavior analysis are in progress. Upon return to Earth, there were no differences in body weights between FLT and GC at the end of the 37 days in space. RNA was of high quality (RIN greater than 8.5). Liver enzyme activity levels of FLT mice and all control mice were similar in magnitude to those of the samples that were optimally processed in the laboratory. Liver samples collected from the intact frozen FLT carcasses had RNA RIN of 7.27 +/- 0.52, which was lower than that of the samples processed on-orbit, but similar to those obtained from the control group intact carcasses. Nonetheless, the RNA samples from the intact carcasses were acceptable for the most demanding transcriptomic analyses. Adrenal glands, thymus and spleen (organs associated with stress response) showed no significant difference in weights between FLT and GC. Enzymatic activity was also not significantly different. Over 3,000 tissues collected from the four groups of mice have become available for the Biospecimen Sharing Program. Together, these validation flight findings demonstrate the capability to support long-duration RR on the ISS to achieve both basic science and biomedical objectives.

Effect of freezing on the rheological, chemical and colour properties of Serpa cheese.

PubMed

Alvarenga, Nuno; Canada, João; Sousa, Isabel

2011-02-01

The effect of freezing on the properties of a raw ewes'-milk semi-soft cheese (Serpa cheese) was studied using small amplitude oscillatory (SAOS) and texture measurements, colour and chemical parameters. The freezing was introduced at three different stages of the ripening process (28, 35 and 42 days), and the cheeses were maintained frozen for 12 months. Cheeses were submitted to a slow or fast freezing method, and to different storage temperatures: -10 and -20°C (three replicates for each set conditions). Chemical data showed that only the proteolysis indicators exhibited differences between frozen and non-frozen samples; frozen samples showed higher values of NPN than the non-frozen samples, indicating that the freezing process did not prevent the secondary proteolysis of cheese. Frozen samples showed a significantly (P<0·05) stronger structure than the non-frozen, as indicated by hardness. However, the differences between the frozen and non-frozen samples were not significantly for storage modulus (G' 1Hz) and loss tangent (tan δ 1Hz) (P>0·05). Freezing affected mainly colour parameters: frozen samples were more luminous, and more yellow-green. The results allowed us to conclude that the damages caused by freezing to cheese properties could be minimized if this type of storage is introduced at the end of ripening (42 d) using a freezing temperature of -20°C.

A fresh liver agar substrate for rearing small numbers of forensically important blow flies (Diptera: Calliphoridae).

PubMed

Gruner, Susan V; Slone, Daniel H

2014-05-01

Forensically important calliphorids can be reared on a mixture of beef liver and agar. Small pieces of meat, especially fresh or frozen beef liver, will desiccate in 2-6 h, but this simple-to-make feeding substrate remains moist for at least 12 h at 25 and 30 degrees C without desiccation, even in small (5 g) amounts. We determined the survivorship of small numbers of Chrysomya megacephala (F.) (first-instar larvae to adult eclosion) raised on 5 g of liver agar and fresh beef liver. We found that all larvae raised on 5 g of liver died due to desiccation, but survivorship on 5 g of liver agar was equivalent to that on larger (50 g) pieces of either liver agar or beef liver.

Spatial considerations during cryopreservation of a large volume sample.

PubMed

Kilbride, Peter; Lamb, Stephen; Milne, Stuart; Gibbons, Stephanie; Erro, Eloy; Bundy, James; Selden, Clare; Fuller, Barry; Morris, John

2016-08-01

There have been relatively few studies on the implications of the physical conditions experienced by cells during large volume (litres) cryopreservation - most studies have focused on the problem of cryopreservation of smaller volumes, typically up to 2 ml. This study explores the effects of ice growth by progressive solidification, generally seen during larger scale cryopreservation, on encapsulated liver hepatocyte spheroids, and it develops a method to reliably sample different regions across the frozen cores of samples experiencing progressive solidification. These issues are examined in the context of a Bioartificial Liver Device which requires cryopreservation of a 2 L volume in a strict cylindrical geometry for optimal clinical delivery. Progressive solidification cannot be avoided in this arrangement. In such a system optimal cryoprotectant concentrations and cooling rates are known. However, applying these parameters to a large volume is challenging due to the thermal mass and subsequent thermal lag. The specific impact of this to the cryopreservation outcome is required. Under conditions of progressive solidification, the spatial location of Encapsulated Liver Spheroids had a strong impact on post-thaw recovery. Cells in areas first and last to solidify demonstrated significantly impaired post-thaw function, whereas areas solidifying through the majority of the process exhibited higher post-thaw outcome. It was also found that samples where the ice thawed more rapidly had greater post-thaw viability 24 h post-thaw (75.7 ± 3.9% and 62.0 ± 7.2% respectively). These findings have implications for the cryopreservation of large volumes with a rigid shape and for the cryopreservation of a Bioartificial Liver Device. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Efficient Recovery of Fluoroquinolone-Susceptible and Fluoroquinolone-Resistant Escherichia coli Strains From Frozen Samples

PubMed Central

Lautenbach, Ebbing; Santana, Evelyn; Lee, Abby; Tolomeo, Pam; Black, Nicole; Babson, Andrew; Perencevich, Eli N.; Harris, Anthony D.; Smith, Catherine A.; Maslow, Joel

2010-01-01

We assessed the rate of recovery of fluoroquinolone-resistant and fluoroquinolone-susceptible Escherichia coli isolates from culture of frozen perirectal swab samples compared with the results for culture of the same specimen before freezing. Recovery rates for these 2 classes of E. coli were 91% and 83%, respectively. The majority of distinct strains recovered from the initial sample were also recovered from the frozen sample. The strains that were not recovered were typically present only in low numbers in the initial sample. These findings emphasize the utility of frozen surveillance samples. PMID:18279070

Efficient recovery of fluoroquinolone-susceptible and fluoroquinolone-resistant Escherichia coli strains from frozen samples.

PubMed

Lautenbach, Ebbing; Santana, Evelyn; Lee, Abby; Tolomeo, Pam; Black, Nicole; Babson, Andrew; Perencevich, Eli N; Harris, Anthony D; Smith, Catherine A; Maslow, Joel

2008-04-01

We assessed the rate of recovery of fluoroquinolone-resistant and fluoroquinolone-susceptible Escherichia coli isolates from culture of frozen perirectal swab samples compared with the results for culture of the same specimen before freezing. Recovery rates for these 2 classes of E. coli were 91% and 83%, respectively. The majority of distinct strains recovered from the initial sample were also recovered from the frozen sample. The strains that were not recovered were typically present only in low numbers in the initial sample. These findings emphasize the utility of frozen surveillance samples.

A fresh liver agar substrate for rearing small numbers of forensically important blow flies (Diptera: Calliphoridae)

USGS Publications Warehouse

Gruner, Susan V.; Slone, Daniel H.

2014-01-01

Forensically important calliphorids can be reared on a mixture of beef liver and agar. Small pieces of meat, especially fresh or frozen beef liver, will desiccate in 2–6 h, but this simple-to-make feeding substrate remains moist for at least 12 h at 25 and 30°C without desiccation, even in small (5 g) amounts. We determined the survivorship of small numbers of Chrysomya megacephala (F.) (first-instar larvae to adult eclosion) raised on 5 g of liver agar and fresh beef liver. We found that all larvae raised on 5 g of liver died due to desiccation, but survivorship on 5 g of liver agar was equivalent to that on larger (50 g) pieces of either liver agar or beef liver.

Molecular constituents of colorectal cancer metastatic to the liver by imaging infrared spectroscopy

NASA Astrophysics Data System (ADS)

Coe, James V.; Chen, Zhaomin; Li, Ran; Nystrom, Steven V.; Butke, Ryan; Miller, Barrie; Hitchcock, Charles L.; Allen, Heather C.; Povoski, Stephen P.; Martin, Edward W.

2015-03-01

Infrared (IR) imaging spectroscopy of human liver tissue slices has been used to identify and characterize liver metastasis of colorectal origin which was surgically removed from a consenting patient and frozen without formalin fixation or dehydration procedures, so that lipids and water remain in the tissues. First, a k-means clustering analysis, using metrics from the IR spectra, identified groups within the image. The groups were identified as tumor or nontumor regions by comparing to an H and E stain of the same sample after IR imaging. Then, calibrant IR spectra of protein, several fats, glycogen, and polyvinyl alcohol were isolated by differencing spectra from different regions or groups in the image space. Finally, inner products (or scores) of the IR spectra at each pixel in the image with each of the various calibrants were calculated showing how the calibrant molecules vary in tumor and nontumor regions. In this particular case, glycogen and protein changes enable separation of tumor and nontumor regions as shown with a contour plot of the glycogen scores versus the protein scores.

A Limited Survey of Heavy Metal Concentrations in Fresh and Frozen Cuttlefish Ink and Mantle Used As Food.

PubMed

Conficoni, Daniele; Alberghini, Leonardo; Bissacco, Elisa; Contiero, Barbara; Giaccone, Valerio

2018-02-01

Cuttlefish ink is consumed as a delicacy worldwide. The current study is the first assessment of heavy metal concentrations in cuttlefish ink versus mantle under different storage methods. A total of 212 samples (64 of fresh mantle, 42 of frozen mantle, 64 of fresh ink, and 42 of frozen ink) were analyzed for the detection of the following heavy metals: arsenic (As), chromium (Cr), iron (Fe), lead (Pb), mercury (Hg), and cadmium (Cd). The median As concentrations were 12.9 mg/kg for fresh mantle, 8.63 mg/kg for frozen mantle, 10.8 mg/kg for frozen ink, and 0.41 mg/kg for fresh ink. The median Cr concentrations were 0.06 mg/kg for fresh mantle and frozen ink, 0.03 mg/kg for frozen mantle, and below the limit of quantification (LOQ) for fresh ink. The median Fe concentrations were 4.08 mg/kg for frozen ink, 1.51 mg/kg for fresh mantle, 0.73 mg/kg for frozen mantle, and below the LOQ for fresh ink. The median Pb concentrations of almost all samples were below the LOQ; only two frozen ink, one fresh ink, one frozen mantle, and one fresh mantle sample exceeded the limit stipulated by the European Union. The Hg concentrations were statistically similar among the four categories of samples; the median Hg concentrations were below the LOQ, and the maximum concentrations were found in frozen ink, at 1.62 mg/kg. The median Cd concentrations were 0.69 mg/kg for frozen ink and 0.11 mg/kg for frozen mantle, fresh mantle and fresh ink concentrations were below the LOQ, and in 11.3% of the tested samples, Cd concentrations were higher than the European Union limit. The probability of samples having a Cd concentration above the legal limit was 35.75 times higher in frozen than in fresh products. Fresh ink had significantly lower concentrations of As, Cr, Fe, and Cd, but the concentrations of Hg and Pb were not significantly different from those of other products. Frozen ink had significantly higher concentrations of Cd, Cr, and Fe, but concentrations of As were lower than those in fresh mantle, pointing out a possible role for the freezing process and for different fishing zones as risk factors for heavy metal contamination.

Effect of different cooking methods on color, phytochemical concentration, and antioxidant capacity of raw and frozen brassica vegetables.

PubMed

Pellegrini, Nicoletta; Chiavaro, Emma; Gardana, Claudio; Mazzeo, Teresa; Contino, Daniele; Gallo, Monica; Riso, Patrizia; Fogliano, Vincenzo; Porrini, Marisa

2010-04-14

This study evaluated the effect of common cooking practices (i.e., boiling, microwaving, and basket and oven steaming) on the phytochemical content (carotenoids, chlorophylls, glucosinolates, polyphenols, and ascorbic acid), total antioxidant capacity (TAC), and color changes of three generally consumed Brassica vegetables analyzed fresh and frozen. Among cooking procedures, boiling determined an increase of fresh broccoli carotenoids and fresh Brussels sprout polyphenols, whereas a decrease of almost all other phytochemicals in fresh and frozen samples was observed. Steaming procedures determined a release of polyphenols in both fresh and frozen samples. Microwaving was the best cooking method for maintaining the color of both fresh and frozen vegetables and obtaining a good retention of glucosinolates. During all cooking procedures, ascorbic acid was lost in great amount from all vegetables. Chlorophylls were more stable in frozen samples than in fresh ones, even though steaming methods were able to better preserve these compounds in fresh samples than others cooking methods applied. The overall results of this study demonstrate that fresh Brassica vegetables retain phytochemicals and TAC better than frozen samples.

Genomewide association study of liver abscess in beef cattle.

PubMed

Keele, J W; Kuehn, L A; McDaneld, T G; Tait, R G; Jones, S A; Keel, B N; Snelling, W M

2016-02-01

Fourteen percent of U.S. cattle slaughtered in 2011 had liver abscesses, resulting in reduced carcass weight, quality, and value. Liver abscesses can result from a common bacterial cause, , which inhabits rumen lesions caused by acidosis and subsequently escapes into the blood stream, is filtered by the liver, and causes abscesses in the liver. Our aim was to identify SNP associated with liver abscesses in beef cattle. We used lung samples as a DNA source because they have low economic value, they have abundant DNA, and we had unrestricted access to sample them. We collected 2,304 lung samples from a beef processing plant: 1,152 from animals with liver abscess and 1,152 from animals without liver abscess. Lung tissue from pairs of animals, 1 with abscesses and another without, were collected from near one another on the viscera table to ensure that pairs of phenotypically extreme animals came from the same lot. Within each phenotype (abscess or no abscess), cattle were pooled by slaughter sequence into 12 pools of 96 cattle for each phenotype for a total of 24 pools. The pools were constructed by equal volume of frozen lung tissue from each animal. The DNA needed to allelotype each pool was then extracted from pooled lung tissue and the BovineHD Bead Array (777,962 SNP) was run on all 24 pools. Total intensity (TI), an indicator of copy number variants, was the sum of intensities from red and green dyes. Pooling allele frequency (PAF) was red dye intensity divided TI. Total intensity and PAF were weighted by the inverse of their respective genomic covariance matrices computed over all SNP across the genome. A false discovery rate ≤ 5% was achieved for 15 SNP for PAF and 20 SNP for TI. Genes within 50 kbp from significant SNP were in diverse pathways including maintenance of pH homeostasis in the gastrointestinal tract, maintain immune defenses in the liver, migration of leukocytes from the blood into infected tissues, transport of glutamine into the kidney in response to acidosis to facilitate production of bicarbonate to increase pH, aggregate platelets to liver injury to facilitate liver repair, and facilitate axon guidance. Evidence from the 35 detected SNP associations combined with evidence of polygenic variation indicate that there is adequate genetic variation in incidence rate of liver abscesses, which could be exploited to select sires for reduced susceptibility to subacute acidosis and associated liver abscess.

Detection and Characterization of Circulating Tumour Cells from Frozen Peripheral Blood Mononuclear Cells

PubMed Central

Lu, David; Graf, Ryon P.; Harvey, Melissa; Madan, Ravi A.; Heery, Christopher; Marte, Jennifer; Beasley, Sharon; Tsang, Kwong Y.; Krupa, Rachel; Louw, Jessica; Wahl, Justin; Bales, Natalee; Landers, Mark; Marrinucci, Dena; Schlom, Jeffrey; Gulley, James L.; Dittamore, Ryan

2015-01-01

Retrospective analysis of patient tumour samples is a cornerstone of clinical research. CTC biomarker characterization offers a non-invasive method to analyse patient samples. However, current CTC technologies require prospective blood collection, thereby reducing the ability to utilize archived clinical cohorts with long-term outcome data. We sought to investigate CTC recovery from frozen, archived patient PBMC pellets. Matched samples from both mCRPC patients and mock samples, which were prepared by spiking healthy donor blood with cultured prostate cancer cell line cells, were processed “fresh” via Epic CTC Platform or from “frozen” PBMC pellets. Samples were analysed for CTC enumeration and biomarker characterization via immunofluorescent (IF) biomarkers, fluorescence in-situ hybridization (FISH) and CTC morphology. In the frozen patient PMBC samples, the median CTC recovery was 18%, compared to the freshly processed blood. However, abundance and localization of cytokeratin (CK) and androgen receptor (AR) protein, as measured by IF, were largely concordant between the fresh and frozen CTCs. Furthermore, a FISH analysis of PTEN loss showed high concordance in fresh vs. frozen. The observed data indicate that CTC biomarker characterization from frozen archival samples is feasible and representative of prospectively collected samples. PMID:28936240

Fresh vs Frozen Samples and Ambient Temperature Have Little Effect on Detection of Colorectal Cancer or Adenomas by a Fecal Immunochemical Test in a Colorectal Cancer Screening Cohort in Germany.

PubMed

Chen, Hongda; Werner, Simone; Brenner, Hermann

2017-10-01

Fecal immunochemical tests (FITs) are used in colorectal cancer (CRC) screening. We compared detection of CRCs and colorectal neoplasms by FITs using fresh samples (collected into buffer-filled tubes) vs frozen samples, and we assessed the effects of seasonal variations in ambient temperature on test performance. We performed a prospective study of 3466 individuals (50% male; mean age, 62 years) undergoing screening colonoscopies at 20 gastroenterology practices in southern Germany from November 2008 through September 2014. Frozen stool samples (collected and frozen by patients through February 2012, n = 1644) and fresh stool samples (collected by patients into buffer-filled tubes after February 2012, n = 1822) were obtained; hemoglobin (Hgb) concentrations were measured by using a commercial, quantitative FIT (cutoff value for positive result, 17 μg Hgb/g feces). Colonoscopy results were used as the gold standard, with results categorized as CRC, advanced adenoma, non-advanced adenoma, or no colorectal neoplasm. Differences in detection of colorectal neoplasms with fresh vs frozen samples were compared by using Wilcoxon rank sum test (continuous variables) and Fisher exact test (categorical variables). We also compared test performance when samples were collected during different seasons (based on outdoor temperature less than 8°, 8°-15°, or more than 15°). Of the samples analyzed by FIT, 12.8% of frozen stool samples (95% confidence interval [CI], 11.3%-14.5%) and 8.7% of fresh stool samples (95% CI, 7.5%-10.1%) had positive results (P value for difference < .001). When adjusting the Hgb cutoff value to produce the same percentage of positive results for fresh and frozen samples (10% and 5%), FIT with frozen vs fresh samples detected colorectal neoplasms with similar levels of sensitivity and specificity. For example, at cutoff values that produced 5% positive results for each sample type, FIT detected advanced neoplasms with 27.8% sensitivity when frozen samples were used (95% CI, 21.4%-35.1%) and 25.6% sensitivity when fresh samples were used (95% CI, 19.8%-32.1%). Specificity values were 97.7% when frozen samples were used (95% CI, 96.8%-98.4%) and 97.6% when fresh samples were used (95% CI, 96.7%-98.3%). We did not observe any differences in detection of neoplasms during different seasons that were based on outdoor temperature. In a prospective study of 3466 individuals who underwent screening colonoscopies and received FITs, we found that use of fresh vs frozen samples slightly affected positivity rates and the proportions of CRCs or adenomas detected at the recommended Hgb cutoff value. However, after we adjusted Hgb cutoff values to produce equal proportions of positive results for fresh vs frozen samples, the performance of the FIT was similar with each sample type. Season of sample collection (based on outdoor temperature) did not affect detection of CRC using either sample type in this study from Middle Europe. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

The use of frozen plasma samples in thromboelastometry.

PubMed

Schoergenhofer, Christian; Buchtele, Nina; Schwameis, Michael; Bartko, Johann; Jilma, Bernd; Jilma-Stohlawetz, Petra

2017-11-01

Thromboelastometry is increasingly used in the clinical and scientific setting. The use of frozen plasma samples may be useful in overcoming certain limitations such as local and timely availability. Whole blood (WB) samples of 20 healthy volunteers were obtained, and plasma was generated. NATEM (n = 20), EXTEM (n = 20) and INTEM (n = 8) analyses were performed in WB, fresh plasma and frozen and thawed plasma. Dabigatran (500, 1000 ng/ml), rivaroxaban (100, 200 ng/ml) or alteplase (333 ng/ml) were added ex vivo to WB, and thromboelastometry was performed in WB and in frozen and thawed plasma samples. Clot formation time, mean clot firmness and the area under the curve were significantly altered in plasma compared to WB. In INTEM and EXTEM analysis, clotting time (CT) was comparable between WB (100%) and fresh (INTEM 114% and EXTEM 93%, ratio of the means) and frozen plasma samples (85 and 99%), whereas in NATEM analysis, the CT increased in fresh (193%) and frozen plasma samples (130%). Dabigatran dose-dependently increased the CT approximately 5- and 9-fold in WB and even more pronounced 10- and 26-fold in plasma. Accordingly, rivaroxaban dose-dependently increased the CT 2- and 2.7-fold in WB, and 3.5- and 4-fold in plasma samples. Hyperfibrinolysis was achieved by addition of alteplase in all WB samples and was reproducible in plasma samples. In conclusion, thromboelastometry, especially INTEM and EXTEM analyses, is possible using frozen and stored plasma samples with comparable results to the corresponding whole blood samples.

Effect of gamma-irradiation on frozen shrimps for decontamination of pathogenic bacteria

NASA Astrophysics Data System (ADS)

Ito, Hitoshi; Rashid, Harun Or; Sangthong, Naruemon; Adulyatham, Pitaya; Rattagool, Pongpen; Ishigaki, Isao

1993-07-01

Twelve samples of imported frozen shrimps were used in this study. The total aerobic bacteria were at 2 × 10 4 to 6 × 10 6 per gram. A few of Vibrio parahaemolyticus, V. mimicus, V. alginolyticus, V. vulnificus, V. fluvialis and Listeria monocytogenes were isolated from many samples. However, Salmonella was not detected in any of the samples. After exposure to 4-5 kGy of gamma-rays, the total aerobic bacteria in frozen shrimps were reduced by approximately 2-3 log cycles. The dose necessary to reduce the vibrio isolates and Aeromonas hydrophila at a level of below 10 -4 per gram was about 3 kGy in frozen shrimps, whereas about 3.5 kGy was required for L. monocytogenes and Salmonella typhimurium. In this study, unpleasant off-odor was clearly detected in the non-frozen shrimps irradiated at 2.5 kGy. On the other hand, off-odor was negligible in the frozen product below 5 kGy irradiation. No remarkable changes of peroxide values were also obtained up to 9 kGy of irradiation in the frozen shrimps. However peroxide values of non-frozen shrimps were clearly increased even irradiated at 4 kGy. Trimethylamine content was not changed at doses below 10 kGy in both of frozen and non-frozen shrimps. Shelf-life of defrosted shrimps were extended ca. 2 times under non-frozen market conditions.

The effects of soy on freezable bread dough: a magnetic resonance study.

PubMed

Simmons, Amber L; Vodovotz, Yael

2012-11-15

Hygroscopic soy ingredients were hypothesised to slow the rate of water migration in unleavened bread dough during frozen storage. Thawed soy (18% dry weight) and wheat dough samples were assessed using non-destructive nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI) for up to 8 wks frozen storage time. MRI suggested a spatially homogeneous, net increase in proton mobility with frozen storage and, with solution state proton NMR, distinct "free" and "bound" states were discerned. T(2) relaxation times of the majority proton population suggested increased mobility with frozen storage time, and statistical difference from the fresh sample was seen later for the soy samples than the wheat samples. As seen by (13)C-solid state NMR, the crystallinity of the starch was not affected by either soy addition or frozen storage. In conclusion, addition of soy to bakery products led to slightly enhanced preservation of "fresh" characteristics of the dough during frozen storage. Copyright © 2012 Elsevier Ltd. All rights reserved.

Effect of sugar and acid on the acceptability of frozen yogurt to a student population.

PubMed

Guinard, J X; Little, C; Marty, C; Palchak, T R

1994-05-01

One hundred and forty-one college students tasted and rated on a nine-point hedonic scale their degree of liking for nine samples of vanilla frozen yogurt varying in sugar and lactic acid. Subjects were also asked to complete a questionnaire about consumption of frozen yogurt and other dairy products. Degree of liking differed significantly among samples, and the samples best liked were those with the lowest acidity, .23 to .29%, independent of sugar concentration. Degree of liking of frozen yogurt failed to correlate with dairy product consumption or hunger at the time of testing. No significant difference existed between male and female students for overall degree of liking of frozen yogurt or overall dairy product intake, yet the questionnaire revealed a significantly higher consumption of frozen yogurt among female students. The results of this study suggest that, for the student population tested, frozen yogurt should combine the sensory properties of ice cream (low acidity) with the nutritional properties of yogurt (low fat, active enzyme culture).

Frozen section analysis of margins for head and neck tumor resections: reduction of sampling errors with a third histologic level.

PubMed

Olson, Stephen M; Hussaini, Mohammad; Lewis, James S

2011-05-01

Frozen section analysis is an essential tool for assessing margins intra-operatively to assure complete resection. Many institutions evaluate surgical defect edge tissue provided by the surgeon after the main lesion has been removed. With the increasing use of transoral laser microsurgery, this method is becoming even more prevalent. We sought to evaluate error rates at our large academic institution and to see if sampling errors could be reduced by the simple method change of taking an additional third section on these specimens. All head and neck tumor resection cases from January 2005 through August 2008 with margins evaluated by frozen section were identified by database search. These cases were analyzed by cutting two levels during frozen section and a third permanent section later. All resection cases from August 2008 through July 2009 were identified as well. These were analyzed by cutting three levels during frozen section (the third a 'much deeper' level) and a fourth permanent section later. Error rates for both of these periods were determined. Errors were separated into sampling and interpretation types. There were 4976 total frozen section specimens from 848 patients. The overall error rate was 2.4% for all frozen sections where just two levels were evaluated and was 2.5% when three levels were evaluated (P=0.67). The sampling error rate was 1.6% for two-level sectioning and 1.2% for three-level sectioning (P=0.42). However, when considering only the frozen section cases where tumor was ultimately identified (either at the time of frozen section or on permanent sections) the sampling error rate for two-level sectioning was 15.3 versus 7.4% for three-level sectioning. This difference was statistically significant (P=0.006). Cutting a single additional 'deeper' level at the time of frozen section identifies more tumor-bearing specimens and may reduce the number of sampling errors.

Scanning electron microscopy of high-pressure-frozen sea urchin embryos.

PubMed

Walther, P; Chen, Y; Malecki, M; Zoran, S L; Schatten, G P; Pawley, J B

1993-12-01

High-pressure-freezing permits direct cryo-fixation of sea urchin embryos having a defined developmental state without the formation of large ice crystals. We have investigated preparation protocols for observing high-pressure-frozen and freeze-fractured samples in the scanning electron microscope. High-pressure-freezing was superior to other freezing protocols, because the whole bulk sample was reasonably well frozen and the overall three-dimensional shape of the embryos was well preserved. The samples were either dehydrated by freeze-substitution and critical-point-drying, or imaged in the partially hydrated state, using a cold stage in the SEM. During freeze-substitution the samples were stabilized by fixatives. The disadvantage of this method was that shrinking and extraction effects, caused by the removal of the water, could not be avoided. These disadvantages were avoided when the sample was imaged in the frozen-hydrated state using a cold-stage in the SEM. This would be the method of choice for morphometric studies. Frozen-hydrated samples, however, were very beam sensitive and many structures remained covered by the ice and were not visible. Frozen-hydrated samples were partially freeze-dried to make visible additional structures that had been covered by ice. However, this method also caused drying artifacts when too much water was removed.

Bile duct hamartomas (von Mayenburg complexes) mimicking liver metastases from bile duct cancer: MRC findings

PubMed Central

Nagano, Yasuhiko; Matsuo, Kenichi; Gorai, Katsuya; Sugimori, Kazuya; Kunisaki, Chikara; Ike, Hideyuki; Tanaka, Katsuaki; Imada, Toshio; Shimada, Hiroshi

2006-01-01

We present a case of a 72-year-old man with a common bile duct cancer, who was initially believed to have multiple liver metastases based on computed tomography findings, and in whom magnetic resonance cholangiography (MRC) revealed a diagnosis of bile duct hamartomas. At exploration for pancreaticoduodenectomy, liver palpation revealed disseminated nodules at the surface of the liver. These nodules showed gray-white nodular lesions of about 0.5 cm in diameter scattered on the surface of both liver lobes, which were looked like multiple liver metastases from bile duct cancer. Frozen section of the liver biopsy disclosed multiple bile ducts with slightly dilated lumens embedded in the collagenous stroma characteristics of multiple bile duct hamartomas (BDHs). Only two reports have described the MRC features of bile duct hamartomas. Of all imaging procedures, MRC provides the most relevant features for the imaging diagnosis of bile duct hamartomas. PMID:16534895

EROD activity and biliary fluorescence in Schroederichthys chilensis (Guichenot 1848): biomarkers of PAH exposure in coastal environments of the South Pacific Ocean.

PubMed

Fuentes-Rios, Daniel; Orrego, Rodrigo; Rudolph, Anny; Mendoza, Gonzalo; Gavilán, Juan F; Barra, Ricardo

2005-10-01

Schroederichthys chilensis is a common shark that lives in Chilean coastal environments. In this work, the relationship between liver 7-ethoxyresorufin-O-deethylase dealkylation (EROD) activity and Fluorescent Aromatic Compounds (FAC) in bile of S. chilensis sampled in three bays with different degrees of pollution were performed including a reference area. Sixty individuals were collected, 20 for each site; (10 males and 10 females per site) livers and bile samples were obtained and immediately frozen. EROD activity and FAC were measured according to three standard methods. EROD activity and FAC were higher in polluted areas than in the reference area. Synchronous Fluorescence Spectra of the bile from the fish collected at the most polluted area showed a peak at 347nm representing a metabolite corresponding to 1-hydroxypyrene. The low EROD activity in the reference area is likely related to the low level of PAH in sediments. We propose that this species is a good indicator of exposure to FACs, since it presents a series of characteristics that make it suitable for monitoring PAH exposure in coastal zones.

Effect of blanching treatments on antioxidant activity of frozen green capsicum (Capsicum annuum L. var bell pepper) using radical scavenging activity (DPPH) assay

NASA Astrophysics Data System (ADS)

Azizzuddin, Norafida; Abdullah, Aminah

2016-11-01

Blanching treatments are needed to deactivate enzymes in frozen vegetables. Antioxidant activity using DPPH radical scavenging activity assay were evaluated in steaming, boiling water, and microwave blanching at different temperature, time and microwave power level on frozen green capsicum. Green capsicum was chosen for frozen treatment compared to other capsicum with different maturity index because of the firm texture. The objective of this study was to compare the antioxidant activity of frozen green capsicum between conventional and Oxi Count Kit® assay for DPPH radical scavenging activity. Results showed frozen green capsicum blanched using microwave at high level/90 seconds (sample J) contained higher level of DPPH in both conventional method and Oxi Count Kit® compared to other treatments. However, there were no significant differences between sample J and fresh sample (sample A). Overall, the sequences from highest to lowest in blanching treatments for both DPPH conventional method, and DPPH Oxi Count Kit® were J (microwave high level/90 seconds) > A (Fresh) > H (Microwave Medium Level/120 seconds) > D (Boiling Water 80°C/150 seconds) > K (Microwave High Level/120 seconds) > I (Microwave Medium Level/150 seconds) > F (Microwave Low Level/150 seconds)> B (Steam 100°C/150 seconds) > E (Boiling Water 100°C /120 seconds) > G (Microwave Low Level /180 seconds)> C (Steam 100°C/180 seconds). Almost all frozen green capsicum samples showed no significant differences for comparison between test using DPPH conventional method and Oxi Count Kit®. Frozen storage for 0, and 3rd months showed no significant differences which indicate no changes on antioxidant activity during frozen storage at -18°C.

Variable X-chromosome inactivation and enlargement of pericentral glutamine synthetase zones in the liver of heterozygous females with OTC deficiency.

PubMed

Musalkova, Dita; Sticova, Eva; Reboun, Martin; Sokolova, Jitka; Krijt, Jakub; Honzikova, Jitka; Gurka, Jiri; Neroldova, Magdalena; Honzik, Tomas; Zeman, Jiri; Jirsa, Milan; Dvorakova, Lenka; Hrebicek, Martin

2018-06-01

Ornithine transcarbamylase (OTC) deficiency is an X-linked disorder that causes recurrent and life-threatening episodes of hyperammonemia. The clinical picture in heterozygous females is highly diverse and derives from the genotype and the degree of inactivation of the mutated X chromosome in hepatocytes. Here, we describe molecular genetic, biochemical, and histopathological findings in the livers explanted from two female patients with late-onset OTC deficiency. Analysis of X-inactivation ratios by DNA methylation-based assays showed remarkable intra-organ variation ranging from 46:54 to 82:18 (average 70:30, n = 37), in favor of the active X chromosome carrying the mutation c.583G>C (p.G195R), in the first patient and from 75:25 to 90:10 (average 82:18, n = 20) in favor of the active X chromosome carrying the splicing mutation c.663+1G>A in the second patient. The X-inactivation ratios in liver samples correlated highly with the proportions of OTC-positive hepatocytes calculated from high-resolution image analyses of the immunohistochemically detected OTC in frozen sections that was performed on total area > 5 cm 2 . X-inactivation ratios in blood in both female patients corresponded to the lower limit of the liver values. Our data indicate that the proportion of about 20-30% of hepatocytes expressing the functional OTC protein is not sufficient to maintain metabolic stability. X-inactivation ratios assessed in liver biopsies taken from heterozygous females with X-linked disorders should not be considered representative of the whole liver.

Survey of Salmonella and Campylobacter contamination of whole, raw poultry on retail sale in Wales in 2003.

PubMed

Meldrum, R J; Tucker, I D; Smith, R M M; Edwards, C

2005-07-01

A survey of the Salmonella and Campylobacter contamination of raw, whole chickens available to consumers in Wales was performed between March and December 2003. In total, 736 samples were taken, and overall contamination rates of 73.1% for Campylobacter and 5.7% for Salmonella were found. This survey follows a survey performed during 2001 to 2002 by Welsh local authorities and the National Public Health Service for Wales that established updated baseline rates for both pathogens in raw, whole chicken available to consumers in Wales. This survey indicated no difference in Campylobacter rates between fresh and frozen samples or between samples taken from retailers and local butchers, but significant differences existed in Salmonella rates between fresh and frozen samples and between those sampled from retailers and butchers, with frozen chickens and samples taken from retailers having significantly higher rates. However, the difference in Salmonella isolation rate between retailers and butchers was found to be due to the differences in the proportions of fresh and frozen chickens sampled from these locations, with a significantly higher number of frozen chickens (with a higher Salmonella rate) being sampled from retailers.

Post-sampling release of free fatty acids - effects of heat stabilization and methods of euthanasia.

PubMed

Jernerén, Fredrik; Söderquist, Marcus; Karlsson, Oskar

2015-01-01

The field of lipid research has made progress and it is now possible to study the lipidome of cells and organelles. A basic requirement of a successful lipid study is adequate pre-analytical sample handling, as some lipids can be unstable and postmortem changes can cause substantial accumulation of free fatty acids (FFAs). The aim of the present study was to investigate the effects of conductive heat stabilization and euthanasia methods on FFA levels in the rat brain and liver using liquid chromatography tandem mass spectrometry. The analysis of brain homogenates clearly demonstrated phospholipase activity and time-dependent post-sampling changes in the lipid pool of snap frozen non-stabilized tissue. There was a significant increase in FFAs already at 2min, which continued over time. Heat stabilization was shown to be an efficient method to reduce phospholipase activity and ex vivo lipolysis. Post-sampling effects due to tissue thawing and sample preparation induced a massive release of FFAs (up to 3700%) from non-stabilized liver and brain tissues compared to heat stabilized tissue. Furthermore, the choice of euthanasia method significantly influenced the levels of FFAs in the brain. The FFAs were decreased by 15-44% in the group of animals euthanized by pentobarbital injection compared with CO2 inhalation or decapitation. Our results highlight the importance of considering euthanasia methods and pre-analytical treatment in lipid analysis, factors which may otherwise interfere with the outcome of the experiments. Copyright © 2014 Elsevier Inc. All rights reserved.

Rheological properties and bread quality of frozen yeast-dough with added wheat fiber.

PubMed

Adams, Vivian; Ragaee, Sanaa M; Abdel-Aal, El-Sayed M

2017-01-01

The rheological characteristics of frozen dough are of great importance in bread-making quality. The effect of addition of commercial wheat aleurone and bran on rheological properties and final bread quality of frozen dough was studied. Wheat aleurone (A) and bran (B) containing 240 g kg -1 and 200 g kg -1 arabinoxylan (AX), respectively, were incorporated into refined wheat flour at 150 g kg -1 substitution level (composite A and B, respectively). Dough samples of composite A and B in addition to two reference dough samples, refined flour (ref A) and whole wheat flour (ref B) were stored at -18°C for 9 weeks. Frozen stored composite dough samples contained higher amounts of bound water, less freezable water and exhibited fewer modifications in gluten network during frozen storage based on data from differential scanning calorimetry and nuclear magnetic resonance spectroscopy. Bread made from composite frozen dough had higher loaf volume compared to ref A or ref B throughout the storage period. The incorporation of wheat fiber into refined wheat flour produced dough with minimum alterations in its rheological properties during 9 weeks of frozen storage compared to refined and 100% wheat flour dough samples. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

High quality copy number and genotype data from FFPE samples using Molecular Inversion Probe (MIP) microarrays

PubMed Central

Wang, Yuker; Carlton, Victoria EH; Karlin-Neumann, George; Sapolsky, Ronald; Zhang, Li; Moorhead, Martin; Wang, Zhigang C; Richardson, Andrea L; Warren, Robert; Walther, Axel; Bondy, Melissa; Sahin, Aysegul; Krahe, Ralf; Tuna, Musaffe; Thompson, Patricia A; Spellman, Paul T; Gray, Joe W; Mills, Gordon B; Faham, Malek

2009-01-01

Background A major challenge facing DNA copy number (CN) studies of tumors is that most banked samples with extensive clinical follow-up information are Formalin-Fixed Paraffin Embedded (FFPE). DNA from FFPE samples generally underperforms or suffers high failure rates compared to fresh frozen samples because of DNA degradation and cross-linking during FFPE fixation and processing. As FFPE protocols may vary widely between labs and samples may be stored for decades at room temperature, an ideal FFPE CN technology should work on diverse sample sets. Molecular Inversion Probe (MIP) technology has been applied successfully to obtain high quality CN and genotype data from cell line and frozen tumor DNA. Since the MIP probes require only a small (~40 bp) target binding site, we reasoned they may be well suited to assess degraded FFPE DNA. We assessed CN with a MIP panel of 50,000 markers in 93 FFPE tumor samples from 7 diverse collections. For 38 FFPE samples from three collections we were also able to asses CN in matched fresh frozen tumor tissue. Results Using an input of 37 ng genomic DNA, we generated high quality CN data with MIP technology in 88% of FFPE samples from seven diverse collections. When matched fresh frozen tissue was available, the performance of FFPE DNA was comparable to that of DNA obtained from matched frozen tumor (genotype concordance averaged 99.9%), with only a modest loss in performance in FFPE. Conclusion MIP technology can be used to generate high quality CN and genotype data in FFPE as well as fresh frozen samples. PMID:19228381

Frozen Section Evaluation of Margin Status in Primary Squamous Cell Carcinomas of the Head and Neck: A Correlation Study of Frozen Section and Final Diagnoses.

PubMed

Layfield, Eleanor M; Schmidt, Robert L; Esebua, Magda; Layfield, Lester J

2018-06-01

Frozen section is routinely used for intraoperative margin evaluation in carcinomas of the head and neck. We studied a series of frozen sections performed for margin status of head and neck tumors to determine diagnostic accuracy. All frozen sections for margin control of squamous carcinomas of the head and neck were studied from a 66 month period. Frozen and permanent section diagnoses were classified as negative or malignant. Correlation of diagnoses was performed to determine accuracy. One thousand seven hundred and ninety-six pairs of frozen section and corresponding permanent section diagnoses were obtained. Discordances were found in 55 (3.1%) pairs. In 35 pairs (1.9%), frozen section was reported as benign, but permanent sections disclosed carcinoma. In 21 cases, the discrepancy was due to sampling and in the remaining cases it was an interpretive error. In 20 cases (1.1%), frozen section was malignant, but the permanent section was interpreted as negative. Frozen section is an accurate method for evaluation of operative margins for head and neck carcinomas with concordance between frozen and permanent results of 97%. Most errors are false negative results with the majority of these being due to sampling issues.

Maximizing Science Return from Future Rodent Experiments on the International Space Station (ISS): Tissue Preservation

NASA Technical Reports Server (NTRS)

Choi, S. Y.; Lai, S.; Klotz, R.; Popova, Y.; Chakravarty, K.; Beegle, J. E.; Wigley, C. L.; Globus, R. K.

2014-01-01

To better understand how mammals adapt to long duration habitation in space, a system for performing rodent experiments on the ISS is under development. Rodent Research-1 is the first flight and will include validation of both on-orbit animal support and tissue preservation. To evaluate plans for on-orbit sample dissection and preservation, we simulated conditions for euthanasia, tissue dissection, and prolonged sample storage on the ISS, and we also developed methods for post-flight dissection and recovery of high quality RNA from multiple tissues following prolonged storage in situ for future science return. Livers and spleens from mice were harvested under conditions that simulated nominal, on-orbit euthanasia and dissection procedures including storage at minus 80 degrees Centigrade for 4 months. The RNA recovered was of high quality (RNA Integrity Number, RNA Integrity Number (RIN) greater than 8) and quantity, and the liver enzyme contents and activities (catalase, glutathione reductase, GAPDH) were similar to positive controls, which were collected under standard laboratory conditions. We also assessed the impact of possible delayed on-orbit dissection scenarios (off-nominal) by dissecting and preserving the spleen (RNA, later) and liver (fast-freezing) at various time points post-euthanasia (from 5 minutes up to 105 minutes). The RNA recovered was of high quality (spleen, RIN greater than 8; liver, RIN greater than 6) and liver enzyme activities were similar to positive controls at all time points, although an apparent decline in select enzyme activities was evident at 105 minutes. Additionally, various tissues were harvested from either intact or partially dissected, frozen carcasses after storage for approximately 2 months; most of the tissues (brain, heart, kidney, eye, adrenal glands and muscle) were of acceptable RNA quality for science return, whereas some tissues (small intestine, bone marrow and bones) were not. These data demonstrate: 1) The protocols developed for future flight experiments will support science return despite delayed preservation post-euthanasia or prolonged storage, and 2) High-quality RNA samples from many different tissues can be recovered by dissection following prolonged storage of the tissue in situ at minus 80 degrees Centigrade. These findings have relevance both to high-value, ground-based experiments when sample collection capability is severely constrained, and to future spaceflight experiments that entail on-orbit sample recovery by the ISS crew.

Hepatic gene expression of Caucasian and African-American patients with obesity-related non-alcoholic fatty liver disease.

PubMed

Stepanova, Maria; Hossain, Noreen; Afendy, Arian; Perry, Kellie; Goodman, Zachary D; Baranova, Ancha; Younossi, Zobair

2010-05-01

There is increasing data suggesting that African Americans with NAFLD tend to have less progressive liver disease. The aim of this study is to assess differences in the hepatic gene expression of African-American and Caucasian patients with NAFLD who had undergone bariatric surgery. A total of 94 patients (81 NAFLD and 13 weight-matched controls with normal liver biopsy) were included. Of the entire cohort, 73 were Caucasians and 21 were African Americans. All patients were undergoing bariatric surgery. Two liver biopsies were obtained at the time of surgery. One biopsy was snap-frozen for gene expression and the other biopsy was stained for pathologic assessment. Liver biopsy confirmed that 24 patients from our cohort had NASH while 57 had only simple steatosis. Snap-frozen liver biopsy specimens of these patients were then used for the RNA extraction. cDNA probes were hybridized with customized microarray gene chips containing 5,220 relevant genes. Gene expression profiles were compared between groups using significance analysis of microarrays algorithm. In comparison to all Caucasian patients, African-American patients had over-expression of EPB41L1, IGF2, FAH, ACSL4, FUT4, CYP3A (q values < 10(-4)). In comparison to Caucasian NAFLD patients, African-American NAFLD patients showed over-expression of EPB41L1 and ACSL4 genes. Finally, in comparison to Caucasian NASH patients, African-American NASH patients showed over-expression of GSTM 2, GSTM4 and GSTM5 as well as FH and ASCL4 genes. Some genes highlighted by this analysis, particularly cytochrome CYP3A and glutathione transferases GSTM2, 4, 5, were previously implicated in the pathogenesis of NASH. African-American patients with biopsy-proven obesity-related NAFLD and NASH have a specific hepatic gene expression pattern that may explain their differences from Caucasian patients with NAFLD in developing progressive liver disease.

Biobanking of fresh frozen tissue from clinical surgical specimens: transport logistics, sample selection, and histologic characterization.

PubMed

Botling, Johan; Micke, Patrick

2011-01-01

Access to high-quality fresh frozen tissue is critical for translational cancer research and molecular -diagnostics. Here we describe a workflow for the collection of frozen solid tissue samples derived from fresh human patient specimens after surgery. The routines have been in operation at Uppsala University Hospital since 2001. We have integrated cryosection and histopathologic examination of each biobank sample into the biobank manual. In this way, even small, macroscopically ill-defined lesions can be -procured without a diagnostic hazard due to the removal of uncharacterized tissue from a clinical -specimen. Also, knowledge of the histomorphology of the frozen tissue sample - tumor cell content, stromal components, and presence of necrosis - is pivotal before entering a biobank case into costly molecular profiling studies.

Radiation Exposure Alters Expression of Metabolic Enzyme Genes In Mice

NASA Technical Reports Server (NTRS)

Wotring, Virginia E.; Mangala, L. S.; Zhang, Y.; Wu, H.

2010-01-01

Most pharmaceuticals are metabolized by the liver. The health of the liver, especially the rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. Because of the importance of the liver in drug metabolism it is important to understand the effects of spaceflight on the enzymes of the liver. Exposure to cosmic radiation is one aspect of spaceflight that can be modeled in ground experiments. This study is an effort to examine the effects of adaptive mechanisms that may be triggered by early exposure to low radiation doses. Using procedures approved by the JSC Animal Care & Use Committee, C57 male mice were exposed to Cs-137 in groups: controls, low dose (50 mGy), high dose (6Gy) and a fourth group that received both radiation doses separated by 24 hours. Animals were anesthetized and sacrificed 4 hours after their last radiation exposure. Livers were removed immediately and flash-frozen in liquid nitrogen. Tissue was homogenized, RNA extracted and purified (Absolutely RNA, Agilent). Quality of RNA samples was evaluated (Agilent Bioanalyzer 2100). Complementary DNA was prepared from high-quality RNA samples, and used to run RT-qPCR screening arrays for DNA Repair and Drug Metabolism (SuperArray, SABiosciences/Qiagen; BioRad Cfx96 qPCR System). Of 91 drug metabolism genes examined, expression of 7 was altered by at least one treatment condition. Genes that had elevated expression include those that metabolize promethazine and steroids (4-8-fold), many that reduce oxidation products, and one that reduces heavy metal exposure (greater than 200-fold). Of the 91 DNA repair and general metabolism genes examined, expression of 14 was altered by at least one treatment condition. These gene expression changes are likely homeostatic and could lead to development of new radioprotective countermeasures.

Sample storage-induced changes in the quantity and quality of soil labile organic carbon

PubMed Central

Sun, Shou-Qin; Cai, Hui-Ying; Chang, Scott X.; Bhatti, Jagtar S.

2015-01-01

Effects of sample storage methods on the quantity and quality of labile soil organic carbon are not fully understood even though their effects on basic soil properties have been extensively studied. We studied the effects of air-drying and frozen storage on cold and hot water soluble organic carbon (WSOC). Cold- and hot-WSOC in air-dried and frozen-stored soils were linearly correlated with those in fresh soils, indicating that storage proportionally altered the extractability of soil organic carbon. Air-drying but not frozen storage increased the concentrations of cold-WSOC and carbohydrate in cold-WSOC, while both increased polyphenol concentrations. In contrast, only polyphenol concentration in hot-WSOC was increased by air-drying and frozen storage, suggesting that hot-WSOC was less affected by sample storage. The biodegradability of cold- but not hot-WSOC was increased by air-drying, while both air-drying and frozen storage increased humification index and changed specific UV absorbance of both cold- and hot-WSOC, indicating shifts in the quality of soil WSOC. Our results suggest that storage methods affect the quantity and quality of WSOC but not comparisons between samples, frozen storage is better than air-drying if samples have to be stored, and storage should be avoided whenever possible when studying the quantity and quality of both cold- and hot-WSOC. PMID:26617054

Methods of human body odor sampling: the effect of freezing.

PubMed

Lenochova, Pavlina; Roberts, S Craig; Havlicek, Jan

2009-02-01

Body odor sampling is an essential tool in human chemical ecology research. However, methodologies of individual studies vary widely in terms of sampling material, length of sampling, and sample processing. Although these differences might have a critical impact on results obtained, almost no studies test validity of current methods. Here, we focused on the effect of freezing samples between collection and use in experiments involving body odor perception. In 2 experiments, we tested whether axillary odors were perceived differently by raters when presented fresh or having been frozen and whether several freeze-thaw cycles affected sample quality. In the first experiment, samples were frozen for 2 weeks, 1 month, or 4 months. We found no differences in ratings of pleasantness, attractiveness, or masculinity between fresh and frozen samples. Similarly, almost no differences between repeatedly thawed and fresh samples were found. We found some variations in intensity; however, this was unrelated to length of storage. The second experiment tested differences between fresh samples and those frozen for 6 months. Again no differences in subjective ratings were observed. These results suggest that freezing has no significant effect on perceived odor hedonicity and that samples can be reliably used after storage for relatively long periods.

Humidity-controlled preparation of frozen-hydrated biological samples for cryogenic coherent x-ray diffraction microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takayama, Yuki; Nakasako, Masayoshi; RIKEN Harima Institute/SPring-8, 1-1-1 Kouto, Mikaduki, Sayo, Hyogo 679-5148

2012-05-15

Coherent x-ray diffraction microscopy (CXDM) has the potential to visualize the structures of micro- to sub-micrometer-sized biological particles, such as cells and organelles, at high resolution. Toward advancing structural studies on the functional states of such particles, here, we developed a system for the preparation of frozen-hydrated biological samples for cryogenic CXDM experiments. The system, which comprised a moist air generator, microscope, micro-injector mounted on a micromanipulator, custom-made sample preparation chamber, and flash-cooling device, allowed for the manipulation of sample particles in the relative humidity range of 20%-94%rh at 293 K to maintain their hydrated and functional states. Here, wemore » report the details of the system and the operation procedure, including its application to the preparation of a frozen-hydrated chloroplast sample. Sample quality was evaluated through a cryogenic CXDM experiment conducted at BL29XUL of SPring-8. Taking the performance of the system and the quality of the sample, the system was suitable to prepare frozen-hydrated biological samples for cryogenic CXDM experiments.« less

Expression of CD163 in the liver of patients with viral hepatitis.

PubMed

Hiraoka, Atsushi; Horiike, Norio; Akbar, Sk Md Fazle; Michitaka, Kojiro; Matsuyama, Takami; Onji, Morikazu

2005-01-01

CD163 is a marker of activated macrophages, and increased levels of soluble CD163 have been detected in sera obtained from patients with hepatitis. The aim of this study was to detect the expression of CD163 in the liver from patients with viral hepatitis. Frozen sections of liver specimens were obtained from 5 patients with acute viral hepatitis (AH) and from 23 patients with chronic viral hepatitis (CH). The expression of CD163 in the liver was determined immunohistochemically using monoclonal antibody to human CD163. Double immunostaining was done to assess those cell types that express CD163 in the liver. The frequencies of CD163-positive cells were significantly higher both in the portal areas and in the hepatic lobules in the liver of patients with AH compared to those with CH (p < 0.05). Double immunostaining revealed that most of the CD163-positive cells were macrophages and Kupffer cells, because they expressed CD68. The expression of CD163 was very low in endothelial cells and liver stellate cells. This study shows that macrophages are activated in hepatitis liver.

Long-term frozen storage of urine samples: a trouble to get PCR results in Schistosoma spp. DNA detection?

PubMed

Fernández-Soto, Pedro; Velasco Tirado, Virginia; Carranza Rodríguez, Cristina; Pérez-Arellano, José Luis; Muro, Antonio

2013-01-01

Human schistosomiasis remains a serious worldwide public health problem. At present, a sensitive and specific assay for routine diagnosis of schistosome infection is not yet available. The potential for detecting schistosome-derived DNA by PCR-based methods in human clinical samples is currently being investigated as a diagnostic tool with potential application in routine schistosomiasis diagnosis. Collection of diagnostic samples such as stool or blood is usually difficult in some populations. However, urine is a biological sample that can be collected in a non-invasive method, easy to get from people of all ages and easy in management, but as a sample for PCR diagnosis is still not widely used. This could be due to the high variability in the reported efficiency of detection as a result of the high variation in urine samples' storage or conditions for handling and DNA preservation and extraction methods. We evaluate different commercial DNA extraction methods from a series of long-term frozen storage human urine samples from patients with parasitological confirmed schistosomiasis in order to assess the PCR effectiveness for Schistosoma spp. detection. Patients urine samples were frozen for 18 months up to 7 years until use. Results were compared with those obtained in PCR assays using fresh healthy human urine artificially contaminated with Schistosoma mansoni DNA and urine samples from mice experimentally infected with S. mansoni cercariae stored frozen for at least 12 months before use. PCR results in fresh human artificial urine samples using different DNA based extraction methods were much more effective than those obtained when long-term frozen human urine samples were used as the source of DNA template. Long-term frozen human urine samples are probably not a good source for DNA extraction for use as a template in PCR detection of Schistosoma spp., regardless of the DNA method of extraction used.

Curation of Frozen Samples

NASA Technical Reports Server (NTRS)

Fletcher, L. A.; Allen, C. C.; Bastien, R.

2008-01-01

NASA's Johnson Space Center (JSC) and the Astromaterials Curator are charged by NPD 7100.10D with the curation of all of NASA s extraterrestrial samples, including those from future missions. This responsibility includes the development of new sample handling and preparation techniques; therefore, the Astromaterials Curator must begin developing procedures to preserve, prepare and ship samples at sub-freezing temperatures in order to enable future sample return missions. Such missions might include the return of future frozen samples from permanently-shadowed lunar craters, the nuclei of comets, the surface of Mars, etc. We are demonstrating the ability to curate samples under cold conditions by designing, installing and testing a cold curation glovebox. This glovebox will allow us to store, document, manipulate and subdivide frozen samples while quantifying and minimizing contamination throughout the curation process.

Effect of Freezing Conditions on Distances and Their Distributions Derived from Double Electron Electron Resonance (DEER): A Study of Doubly-Spin-Labeled T4 Lysozyme

PubMed Central

Georgieva, Elka R.; Roy, Aritro S.; Grigoryants, Vladimir M.; Borbat, Petr P.; Earle, Keith A.; Scholes, Charles P.; Freed, Jack H.

2012-01-01

Pulsed dipolar ESR spectroscopy, DEER and DQC, require frozen samples. An important issue in the biological application of this technique is how the freezing rate and concentration of cryoprotectant could possibly affect the conformation of biomacromolecule and/or spin-label. We studied in detail the effect of these experimental variables on the distance distributions obtained by DEER from a series of doubly spin-labeled T4 lysozyme mutants. We found that the rate of sample freezing affects mainly the ensemble of spin-label rotamers, but the distance maxima remain essentially unchanged. This suggests that proteins frozen in a regular manner in liquid nitrogen faithfully maintain the distance-dependent structural properties in solution. We compared the results from rapidly freeze-quenched (≤100 μs) samples to those from commonly shock-frozen (slow freeze, 1s or longer) samples. For all the mutants studied we obtained inter-spin distance distributions, which were broader for rapidly frozen samples than for slowly frozen ones. We infer that rapid freezing trapped a larger ensemble of spin label rotamers; whereas, on the time-scale of slower freezing the protein and spin-label achieve a population showing fewer low-energy conformers. We used glycerol as a cryoprotectant in concentrations of 10% and 30% by weight. With 10% glycerol and slow freezing, we observed an increased slope of background signals, which in DEER is related to increased local spin concentration, in this case due to insufficient solvent vitrification, and therefore protein aggregation. This effect was considerably suppressed in slowly frozen samples containing 30% glycerol and rapidly frozen samples containing 10% glycerol. The assignment of bimodal distributions to tether rotamers as opposed to protein conformations is aided by comparing results using MTSL and 4-Bromo MTSL spin-labels. The latter usually produce narrower distance distributions. PMID:22341208

Maximizing Science Return from Future Rodent Experiments on the International Space Station (ISS): Tissue Preservation

NASA Technical Reports Server (NTRS)

Choi, S. Y.; Lai, S.; Klotz, R.; Popova, Y.; Chakravarty, K.; Beegle, J. E.; Wigley, C. L.; Globus, R. K.

2014-01-01

To better understand how mammals adapt to long duration habitation in space, a system for performing rodent experiments on the ISS is under development; Rodent Research-1 is the first flight and will include validation of both on-orbit animal support and tissue preservation. To evaluate plans for on-orbit sample dissection and preservation, we simulated conditions for euthanasia, tissue dissection, and prolonged sample storage on the ISS, and we also developed methods for post-flight dissection and recovery of high quality RNA from multiple tissues following prolonged storage in situ for future science. Mouse livers and spleens were harvested under conditions that simulated nominal, on-orbit euthanasia and dissection operations including storage at -80 C for 4 months. The RNA recovered was of high quality (RNA Integrity Number, RIN(is) greater than 8) and quantity, and the liver enzyme contents and activities (catalase, glutathione reductase, GAPDH) were similar to positive controls, which were collected under standard laboratory conditions. We also assessed the impact of possible delayed on-orbit dissection scenarios (off-nominal) by dissecting and preserving the spleen (RNAlater) and liver (fast-freezing) at various time points post-euthanasia (from 5 min up to 105 min). The RNA recovered was of high quality (spleen, RIN (is) greater than 8; liver, RIN (is) greater than 6) and liver enzyme activities were similar to positive controls at all time points, although an apparent decline in select enzyme activities was evident at the latest time (105 min). Additionally, various tissues were harvested from either intact or partially dissected, frozen carcasses after storage for approximately 2 months; most of the tissues (brain, heart, kidney, eye, adrenal glands and muscle) were of acceptable RNA quality for science return, whereas some tissues (small intestine, bone marrow and bones) were not. These data demonstrate: 1) The protocols developed for future flight experiments will support science return despite delayed preservation post-euthanasia or prolonged storage, and 2) Many additional tissues for gene expression analysis can be obtained by dissection following prolonged storage of the tissue in situ at -80 C. These findings have relevance both to high value, ground-based experiments when sample collection capability is severely constrained, and to all future spaceflight experiments that entail on-orbit sample recovery by the ISS crew.

Long-Term Frozen Storage of Urine Samples: A Trouble to Get PCR Results in Schistosoma spp. DNA Detection?

PubMed Central

Fernández-Soto, Pedro; Velasco Tirado, Virginia; Carranza Rodríguez, Cristina; Pérez-Arellano, José Luis; Muro, Antonio

2013-01-01

Background Human schistosomiasis remains a serious worldwide public health problem. At present, a sensitive and specific assay for routine diagnosis of schistosome infection is not yet available. The potential for detecting schistosome-derived DNA by PCR-based methods in human clinical samples is currently being investigated as a diagnostic tool with potential application in routine schistosomiasis diagnosis. Collection of diagnostic samples such as stool or blood is usually difficult in some populations. However, urine is a biological sample that can be collected in a non-invasive method, easy to get from people of all ages and easy in management, but as a sample for PCR diagnosis is still not widely used. This could be due to the high variability in the reported efficiency of detection as a result of the high variation in urine samples’ storage or conditions for handling and DNA preservation and extraction methods. Methodology/Principal Findings We evaluate different commercial DNA extraction methods from a series of long-term frozen storage human urine samples from patients with parasitological confirmed schistosomiasis in order to assess the PCR effectiveness for Schistosoma spp. detection. Patientś urine samples were frozen for 18 months up to 7 years until use. Results were compared with those obtained in PCR assays using fresh healthy human urine artificially contaminated with Schistosoma mansoni DNA and urine samples from mice experimentally infected with S. mansoni cercariae stored frozen for at least 12 months before use. PCR results in fresh human artificial urine samples using different DNA based extraction methods were much more effective than those obtained when long-term frozen human urine samples were used as the source of DNA template. Conclusions/Significance Long-term frozen human urine samples are probably not a good source for DNA extraction for use as a template in PCR detection of Schistosoma spp., regardless of the DNA method of extraction used. PMID:23613907

An Array-Based Analysis of MicroRNA Expression Comparing Matched Frozen and Formalin-Fixed Paraffin-Embedded Human Tissue Samples

PubMed Central

Zhang, Xiao; Chen, Jiamin; Radcliffe, Tom; LeBrun, Dave P.; Tron, Victor A.; Feilotter, Harriet

2008-01-01

MicroRNAs (miRNAs) are small, noncoding RNAs that suppress gene expression at the posttranscriptional level via an antisense RNA-RNA interaction. miRNAs used for array-based profiling are generally purified from either snap-frozen or fresh samples. Because tissues found in most pathology departments are available only in formalin-fixed and paraffin-embedded (FFPE) states, we sought to evaluate miRNA derived from FFPE samples for microarray analysis. In this study, miRNAs extracted from matched snap-frozen and FFPE samples were profiled using the Agilent miRNA array platform (Agilent, Santa Clara, CA). Each miRNA sample was hybridized to arrays containing probes interrogating 470 human miRNAs. Seven cases were compared in either duplicate or triplicate. Intrachip and interchip analyses demonstrated that the processes of miRNA extraction, labeling, and hybridization from both frozen and FFPE samples are highly reproducible and add little variation to the results; technical replicates showed high correlations with one another (Kendall tau, 0.722 to 0.853; Spearman rank correlation coefficient, 0.891 to 0.954). Our results showed consistent high correlations between matched frozen and FFPE samples (Kendall tau, 0.669 to 0.815; Spearman rank correlation coefficient, 0.847 to 0.948), supporting the use of FFPE-derived miRNAs for array-based, gene expression profiling. PMID:18832457

Flow cytometric sorting of fresh and frozen-thawed spermatozoa in the western lowland gorilla (Gorilla gorilla gorilla).

PubMed

O'Brien, J K; Stojanov, T; Crichton, E G; Evans, K M; Leigh, D; Maxwell, W M C; Evans, G; Loskutoff, N M

2005-08-01

We adapted flow cytometry technology for high-purity sorting of X chromosome-bearing spermatozoa in the western lowland gorilla (Gorilla gorilla gorilla). Our objectives were to develop methodologies for liquid storage of semen prior to sorting, sorting of liquid-stored and frozen-thawed spermatozoa, and assessment of sorting accuracy. In study 1, the in vitro sperm characteristics of gorilla ejaculates from one male were unchanged (P > 0.05) after 8 hr of liquid storage at 15 degrees C in a non-egg yolk diluent (HEPES-buffered modified Tyrode's medium). In study 2, we examined the efficacy of sorting fresh and frozen-thawed spermatozoa using human spermatozoa as a model for gorilla spermatozoa. Ejaculates from one male were split into fresh and frozen aliquots. X-enriched samples derived from both fresh and frozen-thawed human semen were of high purity, as determined by fluorescence in situ hybridization (FISH; 90.7%+/-2.3%, overall), and contained a high proportion of morphologically normal spermatozoa (86.0%+/-1.0%, overall). In study 3, we processed liquid-stored semen from two gorillas for sorting using a modification of methods for human spermatozoa. The sort rate for enrichment of X-bearing spermatozoa was 7.3+/-2.5 spermatozoa per second. The X-enriched samples were of high purity (single-sperm PCR: 83.7%) and normal morphology (79.0%+/-3.9%). In study 4 we examined frozen-thawed gorilla semen, and the sort rate (8.3+/-2.9 X-bearing sperm/sec), purity (89.7%), and normal morphology (81.4%+/-3.4%) were comparable to those of liquid-stored semen. Depending on the male and the type of sample used (fresh or frozen-thawed), 0.8-2.2% of gorilla spermatozoa in the processed ejaculate were present in the X-enriched sample. These results demonstrate that fresh or frozen-thawed gorilla spermatozoa can be flow cytometrically sorted into samples enriched for X-bearing spermatozoa. Copyright 2005 Wiley-Liss, Inc.

Impact of intraoperative factor concentrates on blood product transfusions during orthotopic liver transplantation.

PubMed

Colavecchia, A Carmine; Cohen, David A; Harris, Jesse E; Thomas, Jeena M; Lindberg, Scott; Leveque, Christopher; Salazar, Eric

2017-12-01

Major bleeding in orthotopic liver transplantation is associated with significant morbidity and mortality. Limited literature exists regarding comparative effectiveness of prothrombin complex concentrate and fibrinogen concentrate during orthotopic liver transplantation on blood product utilization. This retrospective, single-institution study evaluated the impact of prothrombin complex concentrate and fibrinogen concentrate on blood product utilization during orthotopic liver transplantation from December 2013 to April 2016. This study included patients age 18 years or older and excluded patients who received simultaneous heart or lung transplantation or did not meet documentation criteria. A propensity score matching technique was used to match patients who were exposed to prothrombin complex concentrate with unexposed patients, at a 2 to 1 ratio, to control for selection bias. During this study, 212 patients received orthotopic liver transplantation with 39 prothrombin complex concentrate exposures. The matched study population included 39 patients who were exposed to prothrombin complex concentrate and 78 unexposed patients. Overall, 84.6% of patients who were exposed to prothrombin complex concentrate also received concomitant fibrinogen concentrate, whereas only 2% of patients in the control group received fibrinogen concentrate. After propensity score matching, no other factors that were included in the model differed significantly or had a standardized mean difference of 0.11 or greater. There was no statistical difference in the utilization of red blood cells or fresh frozen plasma for the exposed group versus the unexposed group after matching (mean ± standard deviation: red blood cell units, 12.4 ± 8.0 units vs. 9.7 ± 5.6 units [p = 0.058]; fresh-frozen plasma units, 10.0 ± 6.3 vs. 12.7 ± 9.7 units [p = 0.119], respectively). The intraoperative use of prothrombin complex concentrate and fibrinogen concentrate during orthotopic liver transplantation did not reduce intraoperative blood product requirements at a single institution. © 2017 AABB.

Cryopreservation of human spermatozoa. I. Effects of holding procedure and seeding on motility, fertilizability, and acrosome reaction.

PubMed

Critser, J K; Huse-Benda, A R; Aaker, D V; Arneson, B W; Ball, G D

1987-04-01

Three experiments were conducted to evaluate effects of holding semen at +5.0 degrees C for 30 minutes or -5.0 degrees C for 10 minutes and ice crystal induction (seeding) on frozen-thawed human spermatozoa. In experiment 1, spermatozoa were frozen, and postthaw motility was evaluated immediately (0 hour) and 24 hours later. At both 0 and 24 hours, nonfrozen control samples had higher motility than all other treatment groups. At 0 hour postthaw, motility was higher in samples held at -5.0 degrees C for 10 minutes with no significant effect of seeding. At 24 hours, samples held at -5.0 degrees C for 10 minutes and seeded, but not samples held at -5.0 degrees C and not seeded, had higher motility than samples held at +5.0 degrees C. In experiment 2, semen samples were frozen, and fertilizability was evaluated in a zona-free hamster egg penetration assay. Seeded samples had a higher frequency of sperm penetration than either nonfrozen or nonseeded samples. In experiment 3, nonfrozen controls and frozen treatment groups were evaluated for the frequency of survival and acrosomal integrity. Seeded samples had higher frequencies of survival and loss of acrosomal integrity than nonseeded samples. All frozen-thawed samples had a lower frequency of survival and a higher frequency of loss of acrosomal integrity than nonfrozen controls. Although altered patterns of fertilizability and acrosomal integrity are induced, collectively these data suggest that incorporating a holding temperature of -5.0 degrees C for 10 minutes and seeding may result in a superior protocol for freezing human spermatozoa.

Image sampling in static telepathology for frozen section diagnosis.

PubMed

Della Mea, V; Cataldi, P; Boi, S; Finato, N; Dalla Palma, P; Beltrami, C A

1999-10-01

A frozen section diagnostic service is often not directly available in small rural or mountain hospitals. In these cases, it could be possible to provide frozen section diagnosis through telepathology systems. Telepathology is based on two main methods: static and dynamic. The former is less expensive, but involves the crucial problem of image sampling. To characterise the differences in image sampling for static telepathology when undertaken by pathologists with different experience. As a test field, a previously studied telepathology method based on multimedia email was adopted. Using this method, three pathologists with different levels of experience sampled images from 155 routine frozen sections and sent them to a distant pathology institute, where diagnoses were made on digital images. After the telepathology diagnoses, the glass slides of both the frozen sections and the definitive sections were sent to the remote pathologists for review. Four of 155 transmissions were considered inadequate by the remote pathologist. In the remaining 151 cases, the telepathology diagnosis agreed with the gold standard in 146 (96.7%). There was no significant divergence between the three pathologists in their sampling of the images. Each case comprised five images on average, acquired in four minutes. The overall time for transmission was about 19 minutes. The results suggest that in routine frozen section diagnosis an inexperienced pathologist can sample images sufficiently well to permit remote diagnosis. However, as expected, the internet is too unreliable for such a time dependent task. An improvement in the system would involve integrated real time features, so that there could be interaction between the two pathologists.

Artificial insemination of cranes with frozen semen

USGS Publications Warehouse

Gee, G.F.; Sexton, T.J.; Lewis, J.C.

1979-01-01

For the first time (1978) artificial insemination (AI) with frozen greater sandhill crane (Grus canadensis tabida) semen resulted in fertile eggs and chicks. During the 2 year (1977-78) study, 6 of 27 eggs produced were fertile. Three chicks hatched. Semen samples used for insemination were frozen and stored in liquid nitrogen for two months or less. Recent improvements in the laboratory indicated that a more effective sample can be prepared and greater fertility rates should be expected.

The Coagulation Profile of End-Stage Liver Disease and Considerations for Intraoperative Management.

PubMed

Forkin, Katherine T; Colquhoun, Douglas A; Nemergut, Edward C; Huffmyer, Julie L

2018-01-01

The coagulopathy of end-stage liver disease results from a complex derangement in both anticoagulant and procoagulant processes. With even minor insults, cirrhotic patients experience either inappropriate bleeding or clotting, or even both simultaneously. The various phases of liver transplantation along with fluid and blood product administration may contribute to additional disturbances in coagulation. Thus, anesthetic management of patients undergoing liver transplantation to improve hemostasis and avoid inappropriate thrombosis in the perioperative environment can be challenging. To add to this challenge, traditional laboratory tests of coagulation are difficult to interpret in patients with end-stage liver disease. Viscoelastic coagulation tests such as thromboelastography (Haemonetics Corporation, Braintree, MA) and rotational thromboelastometry (TEM International, Munich, Germany) have helped to reduce transfusion of allogeneic blood products, especially fresh frozen plasma, but have also lead to the increased use of fibrinogen-containing products. In general, advancements in surgical techniques and anesthetic management have led to significant reduction in blood transfusion requirements during liver transplantation. Targeted transfusion protocols and pharmacologic prevention of fibrinolysis may further aid in the management of the complex coagulopathy of end-stage liver disease.

Investigation of the usefulness of fluorescein sodium fluorescence in stereotactic brain biopsy.

PubMed

Thien, Ady; Han, Julian Xinguang; Kumar, Krishan; Ng, Yew Poh; Rao, Jai Prashanth; Ng, Wai Hoe; King, Nicolas Kon Kam

2018-02-01

Intraoperative frozen section assessment, to confirm acquisition of pathological tissues, is used in stereotactic brain biopsy to minimise sampling errors. Limitations include the dependence on dedicated neuro-oncology pathologists and an increase in operative duration. We investigated the use of intraoperative fluorescein sodium, and compared it to frozen section assessment, for confirming pathological tissue samples in the stereotactic biopsy of gadolinium-contrast-enhancing brain lesions. This prospective observational study consisted of 18 consecutive patients (12 men; median age, 63 years) who underwent stereotactic biopsy of gadolinium-contrast-enhancing brain lesions with intravenous fluorescein sodium administration. Twenty-three specimens were obtained and examined for the presence of fluorescence using a microscope with fluorescence visualisation capability. Positive and negative predictive values were calculated based on the fluorescence status of the biopsy samples with its corresponding intraoperative frozen section and definitive histopathological diagnosis. Nineteen specimens (83%) were fluorescent and four (17%) were non-fluorescent. All 19 fluorescent specimens were confirmed to be lesional on intraoperative frozen section assessment and were suitable for histopathological diagnosis. Three of the non-fluorescent specimens were confirmed to be lesional on intraoperative frozen section assessment. One non-fluorescent specimen was non-diagnostic on frozen section and histological assessments. The positive predictive value was 100% and the negative predictive value was 25%. Fluorescein sodium fluorescence is as accurate as frozen section assessment in confirming sampling of pathological tissue in the stereotactic biopsy of gadolinium-contrast-enhancing brain lesions. Fluorescein sodium fluorescence-guided stereotactic biopsy is a useful addition to the neurosurgical armamentarium.

Freezing of homogenized sputum samples for intermittent storage.

PubMed

Holz, O; Mücke, M; Zarza, P; Loppow, D; Jörres, R A; Magnussen, H

2001-08-01

Among the reasons that restrict the application of sputum induction in outpatient settings is the need for processing of samples within 2 h after induction. The aim of our study was to assess whether freezing is suitable for intermediate storage of sputum samples before processing. We compared differential cell counts between two sputum aliquots derived from the same sample. One aliquot was processed within 2 h after production and one, after it had been frozen under addition of dimethyl-sulfoxid (DMSO) and stored up to 10 days at -20 degrees C. Thirty-five samples were frozen immediately prior to preparation of cytospins, and 10 samples were frozen at an even earlier stage, directly after homogenization. In both sets of experiments we observed a significant relationship between frozen and native samples regarding macrophages, neutrophils and eosinophils, as indicated by respective intraclass correlation coefficients of 0.96, 0.96, and 0.93 in the first, and of 0.92, 0.96 and 0.77 in the second experiments. Our results indicate that the freezing of sputum samples at different stages of processing does not alter sputum morphology to an extent that affects the results of differential cell counts.

Sensory descriptive quantitative analysis of unpasteurized and pasteurized juçara pulp (Euterpe edulis) during long-term storage

PubMed Central

da Silva, Paula Porrelli Moreira; Casemiro, Renata Cristina; Zillo, Rafaela Rebessi; de Camargo, Adriano Costa; Prospero, Evanilda Teresinha Perissinotto; Spoto, Marta Helena Fillet

2014-01-01

This study evaluated the effect of pasteurization followed by storage under different conditions on the sensory attributes of frozen juçara pulp using quantitative descriptive analysis (QDA). Pasteurization of packed frozen pulp was performed by its immersion in stainless steel tank containing water (80°C) for 5 min, followed by storage under refrigerated and frozen conditions. A trained sensory panel evaluated the samples (6°C) on day 1, 15, 30, 45, 60, 75, and 90. Sensory attributes were separated as follows: appearance (foamy, heterogeneous, purple, brown, oily, and creamy), aroma (sweet and fermented), taste (astringent, bitter, and sweet), and texture (oily and consistent), and compared to a reference material. In general, unpasteurized frozen pulp showed the highest score for foamy appearance, and pasteurized samples showed highest scores to creamy appearance. Pasteurized samples remained stable regarding brown color development while unpasteurized counterparts presented increase. Color is an important attribute related to the product identity. All attributes related to taste and texture remained constant during storage for all samples. Pasteurization followed by storage under frozen conditions has shown to be the best conservation method as samples submitted to such process received the best sensory evaluation, described as foamy, slightly heterogeneous, slightly bitter, and slightly astringent. PMID:25473489

Sensory descriptive quantitative analysis of unpasteurized and pasteurized juçara pulp (Euterpe edulis) during long-term storage.

PubMed

da Silva, Paula Porrelli Moreira; Casemiro, Renata Cristina; Zillo, Rafaela Rebessi; de Camargo, Adriano Costa; Prospero, Evanilda Teresinha Perissinotto; Spoto, Marta Helena Fillet

2014-07-01

This study evaluated the effect of pasteurization followed by storage under different conditions on the sensory attributes of frozen juçara pulp using quantitative descriptive analysis (QDA). Pasteurization of packed frozen pulp was performed by its immersion in stainless steel tank containing water (80°C) for 5 min, followed by storage under refrigerated and frozen conditions. A trained sensory panel evaluated the samples (6°C) on day 1, 15, 30, 45, 60, 75, and 90. Sensory attributes were separated as follows: appearance (foamy, heterogeneous, purple, brown, oily, and creamy), aroma (sweet and fermented), taste (astringent, bitter, and sweet), and texture (oily and consistent), and compared to a reference material. In general, unpasteurized frozen pulp showed the highest score for foamy appearance, and pasteurized samples showed highest scores to creamy appearance. Pasteurized samples remained stable regarding brown color development while unpasteurized counterparts presented increase. Color is an important attribute related to the product identity. All attributes related to taste and texture remained constant during storage for all samples. Pasteurization followed by storage under frozen conditions has shown to be the best conservation method as samples submitted to such process received the best sensory evaluation, described as foamy, slightly heterogeneous, slightly bitter, and slightly astringent.

Effect of freezing on electrical properties and quality of thawed chicken breast meat

PubMed Central

Wei, Ran; Wang, Peng; Han, Minyi; Chen, Tianhao; Xu, Xinglian; Zhou, Guanghong

2017-01-01

Objective The objective of this research was to study the electrical properties and quality of frozen-thawed chicken breast meat and to investigate the relationship between these parameters at different times of frozen storage. Methods Thawed samples of chicken breast muscles were evaluated after being kept in frozen storage at −18°C for different periods of time (1, 2, 3, 4, 5, 6, 7, and 8 months). Results The results showed that water-holding capacity (WHC) and protein solubility decreased while thiobarbituric acid-reactive substances content increased with increasing storage time. The impedance module of samples decreased during 8-month frozen storage. Pearson correlation coefficients showed that the impedance change ratio (Q value) was significantly (p<0.05) related to pH, color, WHC, lipid oxidation and protein solubility, indicating a good relationship between the electrical properties and qualities of frozen-thawed chicken breast meat. Conclusion Impedance measurement has a potential to assess the quality of frozen chicken meat combining with quality indices. PMID:27554358

Blood oxygen saturation of frozen tissue determined by hyper spectral imaging

NASA Astrophysics Data System (ADS)

Braaf, Boy; Nadort, Annemarie; Faber, Dirk; ter Wee, Rene; van Leeuwen, Ton; Aalders, Maurice

2008-02-01

A method is proposed for determining blood oxygen saturation in frozen tissue. The method is based on a spectral camera system equipped with an Acoustic-Optical-Tuneable-Filter. The HSI-setup is validated by measuring series of unfrozen and frozen samples of a hemoglobin-solution, a hemoglobin-intralipid mixture and whole blood with varying oxygen saturation. The theoretically predicted linear relation between oxygen saturation and absorbance was observed in both the frozen sample series and the unfrozen series. In a final proof of principal, frozen myocardial tissue was measured. Higher saturation values were recorded for ventricle and atria tissue compared to the septum and connective tissue. These results are not validated by measurements with another method. The formation of methemoglobin during freezing and the presence of myoglobin in the tissue turned out to be possible sources of error.

The case study of drillbit and borehole frozen water of the subglacial Lake Vostok, East Antarctica for microbial content

NASA Astrophysics Data System (ADS)

Bulat, Sergey; Doronin, Maxim; Dominique, Marie; Lipenkov, Vladimir; Lukin, Valery; Karlov, Denis; Demchenko, Leonid; Khilchenko, Margarita

The objective was to estimate microbial content and diversity in the subglacial Lake Vostok (buried beneath 4-km thick East Antarctic ice sheet) by studying the uppermost water layer which entered the borehole upon lake entry (February 5, 2012) and then shortly frozen within. The samples of so-called drillbit water frozen on a drill bit upon lake enter (RAE57) along with re-drilled so-called borehole-frozen water (RAE58) were provided for the study with the ultimate goal to discover the life in this extreme icy environment. The comprehensive analyses (constrained by Ancient DNA research criteria) of the first lake water samples - drillbit- (one sample) and borehole-frozen (3 different depths 5G-2N-3425, 3429 et 3450m), are nearly got finished. If the drillbit water sample was heavily polluted with drill fluid (at ratio 1:1), re-drilled borehole-frozen samples were proved to be rather clean but still strongly smelling kerosene and containing numerous micro-droplets of drill fluid making the ice non-transparent. The cell concentrations measured by flow cytofluorometry showed 167 cells per ml in the drillbit water sample while in borehole-frozen samples ranged from 5.5 (full-cylinder 3429m deep frozen water ice core) to 38 cells per ml (freeze-centre of 3450m deep moon-shape ice core). DNA analyses came up with total 44 bacterial phylotypes discovered by sequencing of different regions (v3-v5, v4-v8, v4-v6 et full-gene) of 16S rRNA genes. Amongst them all but two were considered to be contaminants (were present in our contaminant library, including drill fluid findings). The 1st remaining phylotype successfully passing all contamination criteria proved to be hitherto-unknown type of bacterium (group of clones, 3 allelic variants) showing less than 86% similarity with known taxa. Its phylogenetic assignment to bacterial divisions or lineages was also unsuccessful despite of the RDP has classified it belonging to OD1 uncultured Candidate Division. The 2nd phylotype was less remarkable and still dubious in terms of contamination. It was presented by just one clone and showed 93% similarity with Janthinobacterium sp of Oxalobacteraceae (Beta-Proteobacteria) - well-known ‘water-loving’ bacteria. No archaea were detected in lake water frozen samples. Thus, the unidentified and unclassified bacterial w123-10 phylotype for the first time discovered in the uppermost water layer in subglacial Lake Vostok might represent ingenious cell populations in the lake, making the life in the lake less elusive. The proof may come (as well as novel phylotype discoveries) with farther analyses (e.g., sample screening with w123-10-specific primers, 16S rRNA v4 region amplicon sequencing) of existing and newly requested moon-shape samples of borehole-frozen water which are on a way to laboratories. We are deeply grateful to Jean Robert Petit and Jean Martins, UJF-CNRS, Grenoble (France) for assistance in conducting some analyses.

Microbiological quality of five potato products obtained at retail markets.

PubMed Central

Duran, A P; Swartzentruber, A; Lanier, J M; Wentz, B A; Schwab, A H; Barnard, R J; Read, R B

1982-01-01

The microbiological quality of frozen hash brown potatoes, dried hash brown potatoes with onions, frozen french fried potatoes, dried instant mashed potatoes, and potato salad was determined by a national sampling at the retail level. A wide range of results was obtained, with most sampling units of each products having excellent microbiological quality. Geometric mean aerobic plate counts were as follows: dried hash brown potatoes, 270/g; frozen hash brown potatoes with onions, 580/g; frozen french fried potatoes 78/g; dried instant mashed potatoes, 1.1 x 10(3)/g; and potato salad, 3.6 x 10(3)/g. Mean values of coliforms, Escherichia coli, and Staphylococcus aureus were less than 10/g. PMID:6758695

Influence of the freezing method on the changes that occur in grape samples after frozen storage.

PubMed

Santesteban, Luis G; Miranda, Carlos; Royo, José B

2013-09-01

Sample freezing is frequently used in oenological laboratories as a compromise solution to increase the number of samples that can be analysed, despite the fact that some grape characteristics are known to change after frozen storage. However, freezing is usually performed using standard freezers, which provide a slow freezing. The aim of this work was to evaluate whether blast freezing would decrease the impact of standard freezing on grape composition. Grape quality parameters were assessed in fresh and in frozen stored samples that had been frozen using three different procedures: standard freezing and blast freezing using either a blast freezer or an ultra-freezer. The implications of frozen storage in grape samples reported in earlier research were observed for the three freezing methods evaluated. Although blast freezing improved repeatability for the most problematic parameters (tartaric acidity, TarA; total phenolics, TP), the improvement was not important from a practical point of view. However, TarA and TP were relatively repeatable among the three freezing procedures, which suggests that freezing had an effect on these parameters independently of the method used . According to our results, the salification potential of the must is probably implied in the changes observed for TarA, whereas for TP the precipitation of protoanthocyanins after association with cell wall material is hypothesized to cause the lack of repeatability between fresh and frozen grapes. Blast freezing would not imply a great improvement if implemented in oenological laboratories, at least for the parameters included in this study. © 2013 Society of Chemical Industry.

Urine phenobarbital drug screening: potential use for compliance assessment in neonates.

PubMed

Guillet, Ronnie; Kwon, Jennifer M; Chen, Sixaio; McDermott, Michael P

2012-02-01

This study was done to determine if urine phenobarbital measurements provide a reliable indicator of presence of the drug in neonates. Urine was collected from neonates treated with phenobarbital for clinical indications within 4 to 6 hours of clinically indicated collection of serum phenobarbital levels. Urine samples were also collected from control neonates not treated with phenobarbital. One aliquot was assayed fresh, another frozen at -30°C and assayed 1 to 3 months later. Phenobarbital was assayed using the ONLINE TDM Roche/Hitachi automated clinical chemistry analyzer. Serum and urine concentrations were compared as were fresh and frozen urine measurements. Serum phenobarbital ranged from 5.6 to 52.7 μg/mL. Matched urine samples were 56.6 ± 12.5% of the serum level. Frozen samples were 98.3 ± 8.0% of the fresh samples. Urine phenobarbital concentrations, either fresh or frozen, can be used in neonates as a noninvasive estimate of drug levels.

Frozen tissue preparation for high-resolution multiplex histological analyses of human brain specimens.

PubMed

Shao, Fangjie; Jiang, Wenhong; Gao, Qingqing; Li, Baizhou; Sun, Chongran; Wang, Qiyuan; Chen, Qin; Sun, Bing; Shen, Hong; Zhu, Keqing; Zhang, Jianmin; Liu, Chong

2017-10-01

The availability of a comprehensive tissue library is essential for elucidating the function and pathology of human brains. Considering the irreplaceable status of the formalin-fixation-paraffin-embedding (FFPE) preparation in routine pathology and the advantage of ultra-low temperature to preserve nucleic acids and proteins for multi-omics studies, these methods have become major modalities for the construction of brain tissue libraries. Nevertheless, the use of FFPE and snap-frozen samples is limited in high-resolution histological analyses because the preparation destroys tissue integrity and/or many important cellular markers. To overcome these limitations, we detailed a protocol to prepare and analyze frozen human brain samples that is particularly suitable for high-resolution multiplex immunohistological studies. As an alternative, we offered an optimized procedure to rescue snap-frozen tissues for the same purpose. Importantly, we provided a guideline to construct libraries of frozen tissue with minimal effort, cost and space. Taking advantage of this new tissue preparation modality to nicely preserve the cellular information that was otherwise damaged using conventional methods and to effectively remove tissue autofluorescence, we described the high-resolution landscape of the cellular composition in both lower-grade gliomas and glioblastoma multiforme samples. Our work showcases the great value of fixed frozen tissue in understanding the cellular mechanisms of CNS functions and abnormalities.

Bacteria-killing ability of fresh blood plasma compared to frozen blood plasma

DOE PAGES

Jacobs, Anne C.; Fair, Jeanne Marie

2015-10-09

In recent years, the bacteria-killing assay (BKA) has become a popular technique among ecoimmunologists. New variations of that assay allow researchers to use smaller volumes of blood, an important consideration for those working on small-bodied animals. However, this version of the assay requires access to a lab with a nanodrop spectrophotometer, something that may not be available in the field. One possible solution is to freeze plasma for transport; however, this assumes that frozen plasma samples will give comparable results to fresh ones. Here, we tested this assumption using plasma samples from three species of birds: chickens (Gallus gallus), ash-throatedmore » flycatchers (Myiarchus cinerascens), and western bluebirds (Sialia mexicana). Chicken plasma samples lost most or all of their bacterial killing ability after freezing. This did not happen in flycatchers and bluebirds; however, frozen plasma did not produce results comparable to those obtained using fresh plasma. Finally, we caution researchers using the BKA to use fresh samples whenever possible, and to validate the use of frozen samples on a species-by-species basis.« less

Bacteria-killing ability of fresh blood plasma compared to frozen blood plasma.

PubMed

Jacobs, Anne C; Fair, Jeanne M

2016-01-01

In recent years, the bacteria-killing assay (BKA) has become a popular technique among ecoimmunologists. New variations of that assay allow researchers to use smaller volumes of blood, an important consideration for those working on small-bodied animals. However, this version of the assay requires access to a lab with a nanodrop spectrophotometer, something that may not be available in the field. One possible solution is to freeze plasma for transport; however, this assumes that frozen plasma samples will give comparable results to fresh ones. We tested this assumption using plasma samples from three species of birds: chickens (Gallus gallus), ash-throated flycatchers (Myiarchus cinerascens), and western bluebirds (Sialia mexicana). Chicken plasma samples lost most or all of their bacterial killing ability after freezing. This did not happen in flycatchers and bluebirds; however, frozen plasma did not produce results comparable to those obtained using fresh plasma. We caution researchers using the BKA to use fresh samples whenever possible, and to validate the use of frozen samples on a species-by-species basis. Copyright © 2015 Elsevier Inc. All rights reserved.

Bacteria-killing ability of fresh blood plasma compared to frozen blood plasma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacobs, Anne C.; Fair, Jeanne Marie

In recent years, the bacteria-killing assay (BKA) has become a popular technique among ecoimmunologists. New variations of that assay allow researchers to use smaller volumes of blood, an important consideration for those working on small-bodied animals. However, this version of the assay requires access to a lab with a nanodrop spectrophotometer, something that may not be available in the field. One possible solution is to freeze plasma for transport; however, this assumes that frozen plasma samples will give comparable results to fresh ones. Here, we tested this assumption using plasma samples from three species of birds: chickens (Gallus gallus), ash-throatedmore » flycatchers (Myiarchus cinerascens), and western bluebirds (Sialia mexicana). Chicken plasma samples lost most or all of their bacterial killing ability after freezing. This did not happen in flycatchers and bluebirds; however, frozen plasma did not produce results comparable to those obtained using fresh plasma. Finally, we caution researchers using the BKA to use fresh samples whenever possible, and to validate the use of frozen samples on a species-by-species basis.« less

Freezing-thawing and sub-sampling influence the marination performance of chicken breast meat.

PubMed

Bowker, B; Zhuang, H

2017-09-01

Vacuum-tumbling marination is often used to improve the yield and quality of whole or portioned broiler breast fillets. The relationship between the marination performance of whole Pectoralis major muscles and breast fillet sub-samples is not well understood. The objective was to determine the effects of sub-sampling and freezing-thawing on the marination performance and cook loss of broiler breast meat. Paired right and left breast fillets were marinated as whole fillets or sub-samples (cranial and mid-caudal portions). Samples were marinated at 48 h postmortem (fresh) or stored at -20°C and then thawed prior to marination (frozen-thawed). Samples were vacuum-tumbled in 20% wt/wt brine (5% NaCl, 3% STP) and weighed pre-marination, during marination (15, 30, and 45 min), and 24 h post-marination. Samples were then cooked to 75°C for determination of cook loss. Marinade uptake was greater in caudal sub-samples than intact fillets and cranial sub-samples after 15 min of marination (P < 0.0001). After 30 min, marinade uptake was greater in caudal sub-samples and intact fillets than cranial sub-samples (P < 0.05). After 45 min, marinade uptake for fresh samples was greatest in intact fillets and lowest in cranial sub-samples. For frozen-thawed samples, marinade uptake at 45 min was greater in caudal sub-samples and intact fillets than cranial sub-samples (P < 0.0001). Marinade uptake in sub-samples at 30 min was greater in frozen-thawed versus fresh fillets (P < 0.05). Differences in marinade retention were not observed. Cook loss was similar between fresh and frozen-thawed samples but was greater in sub-samples compared to intact fillets (P < 0.0001). Correlations between marinade uptake in intact fillets and cranial sub-samples were greater in fresh (r = 0.64 to 0.78) than frozen-thawed samples (r = 0.39 to 0.59). Correlations between marinade uptake in intact fillets and caudal sub-samples were greater in frozen-thawed (r = 0.79 to 0.82) than fresh samples (r = 0.46 to 0.63). Data suggest that the relationships between marination performance of whole breast fillets and fillet sub-samples are dependent upon prior sample handling and intra-fillet sampling location. Published by Oxford University Press on behalf of Poultry Science Association 2017.

Frozen yogurt with added inulin and isomalt.

PubMed

Isik, U; Boyacioglu, D; Capanoglu, E; Erdil, D Nilufer

2011-04-01

The objective of this study was to produce a frozen yogurt containing low fat and no added sugar. Samples containing 5% polydextrose, 0.065% aspartame and acesulfame-K mixture, and different levels of inulin and isomalt (5.0, 6.5, and 8.0%) were produced at pilot scale and analyzed for their physical and chemical properties including proximate composition, viscosity, acidity, overrun, melting rate, heat shock stability, as well as sensory characteristics, and viability of lactic acid bacteria. With the addition of inulin and isomalt, viscosity increased by 19 to 52% compared with that of sample B (reduced-fat control). The average calorie values of samples substituted with sweeteners were about 43% lower than that of original sample. Low-calorie frozen yogurt samples melted about 33 to 48% slower than the reduced-fat control sample at 45 min. Based on quantitative descriptive profile test results, statistically significant differences among products were observed for hardness, iciness, foamy melting, whey separation, and sweetness characteristics. The results of principal component analysis showed that the sensory properties of the sample containing 6.5% inulin and 6.5% isomalt were similar to those of control. Lactic acid bacteria counts of frozen yogurt were found to be between 8.12 and 8.49 log values, 3 mo after the production. The overall results showed that it is possible to produce an attractive frozen yogurt product with the incorporation of inulin and isomalt with no added sugar and reduced fat. Copyright © 2011 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Volatile generation in bell peppers during frozen storage and thawing using selected ion flow tube mass spectrometry (SIFT-MS).

PubMed

Wampler, Brendan; Barringer, Sheryl A

2012-06-01

To determine volatile formation during storage and thawing, whole, pureed, blanched, and raw green and red bell peppers (Capsicum annuum) were frozen quickly or slowly then stored at -18 °C for up to 7 mo, with and without SnCl(2) addition during thawing. Headspace analysis was performed by a Selected Ion Flow Tube Mass Spectrometer (SIFT-MS). After blanching, (Z)-3-hexenal had a large significant decrease in concentration since it is a heat labile compound while most other volatiles did not change in concentration. The freezing process increased volatile levels in the puree only. Slow freeze peppers had higher levels of some LOX generated volatiles during storage than quick freeze. During frozen storage of blanched samples (E)-2-hexenal, (Z and E)-hexen-1-ol, and (E)-2-pentenal increased likely because of nonenzymatic autoxidation of fatty acids while other volatiles remained constant. In Raw Whole peppers, (Z)-3-hexenal, hexanal, and 2-pentylfuran were generated during storage likely because the LOX enzyme is still active during frozen storage. However, blanched samples had higher concentrations of (E)-2-hexenal, (Z and E)-hexen-1-ol, 1-penten-3-one, and (E)-2-heptenal because of enzymatic destruction of these volatiles in the raw samples. The levels of many of the volatiles in the raw samples, including (Z)-3-hexenal, (E)-2-hexenal, (Z and E)-hexen-1-ol, hexanal, (E)-2-pentenal, and 2-pentylfuran, appeared to peak around 34 d after freezing. Pureed samples had significantly higher levels of volatiles than the whole samples, and volatiles peaked earlier. Green bell pepper volatile levels were always higher than red bell pepper. Significantly higher volatile formation occurred during thawing than it did during frozen storage. Studying and monitoring the headspace volatiles with a SIFT-MS can give information that will help manufacturers better understand how the volatiles in bell peppers change during frozen storage. This will give valuable information to processors on how to minimize volatile changes during storage of frozen peppers. © 2012 Institute of Food Technologists®

Assessment of frozen storage duration effect on quality characteristics of various horse muscles.

PubMed

Seong, Pil Nam; Seo, Hyun Woo; Kim, Jin-Hyoung; Kang, Geun Ho; Cho, Soo-Hyun; Chae, Hyun Seok; Park, Beom Young; Van Ba, Hoa

2017-12-01

The study aimed at assessing the effects of frozen storage duration on quality characteristics, lipid oxidation and sensory quality of various horse muscles. Five representative muscles: longissimus dorsi (LD), gluteus medius (GM), semimembranosus (SM), biceps femoris (BF), and triceps brachii (TB) at 24 h post-mortem obtained from 28-mo-old Jeju female breed horses (n = 8) were used in the present investigation. The muscles were vacuum-packaged and frozen at -20°C for 120, 240, and 360 days. All the samples were analyzed for thawing and cooking losses, pH, Warner-Bratzler shear forces (WBSF), color traits, total volatile basic nitrogen (TVBN), thiobarbituric acid reactive substances (TBARS) and sensory traits. The muscle samples analyzed on day 0 of frozen storage (fresh, non-frozen) were used for comparison. Results revealed that thawing and cooking losses significantly (p<0.05) increased in all the muscles after 120 days and then remained unchanged up to 360 days of frozen storage. The TBARS and TVBN contents significantly increased as increasing frozen storage time up to 360 days (p<0.05). While, significant decreases in WBSF values were observed for all the muscles with increased frozen storage time (p<0.05). Frozen storage variously affected the color traits of the muscles for instance; the redness of LD, GM, and BF muscles showed a decreasing tendency during frozen storage while it was not changed in TB and SM muscles. Furthermore, the frozen storage did not produce detrimental effects on sensory quality as it did not cause flavor and juiciness defects whereas it partially improved the tenderness of all the muscles studied. Based on the results obtained from our work, it is concluded that frozen storage could be applied to increase the long-term shelf life of horsemeat while still retaining its sensory quality.

Quantitative Method for Analysis of Hydrocodone, Hydromorphone and Norhydrocodone in Human Plasma by Liquid Chromatography-tandem Mass Spectrometry

DTIC Science & Technology

2013-03-01

ratio ranges obtained for the six standards. Twelve samples were analyzed to demonstrate the efficiency of the extraction procedure. Drug and internal...frozen (−70 ◦C). Refrigerated samples were tested after 2 months of storage ; frozen samples were tested for up to 1 year from stor- age date. The...freeze–thaw stability was evaluated by analyzing three subject samples with known drug concentrations and two quality control samples at concentrations

Effect of moderate dietary restriction on visceral organ weight, hepatic oxygen consumption, and metabolic proteins associated with energy balance in mature pregnant beef cows.

PubMed

Wood, K M; Awda, B J; Fitzsimmons, C; Miller, S P; McBride, B W; Swanson, K C

2013-09-01

Twenty-two nonlactating multiparous pregnant beef cows (639 ± 68 kg) were used to investigate the effect of dietary restriction on the abundance of selected proteins regulating cellular energy metabolism. Cows were fed at either 85% (n = 11; LOW) or 140% (n = 11; HIGH) of total NE requirements. The diet consisted of a haylage-based total mixed ration containing 20% wheat straw. Cows were slaughtered by block (predicted date of parturition), beginning 83 d after the initiation of dietary treatments and every week thereafter for 6 wk, such that each block was slaughtered at approximately 250 d of gestation. Tissue samples from liver, kidney, sternomandibularis muscle, ruminal papilli (ventral sac), pancreas, and small intestinal muscosa were collected at slaughter and snap frozen in liquid N2. Western blots were conducted to quantify abundance of proliferating cell nuclear antigen (PCNA), ATP synthase, ubiquitin, and Na/K+ ATPase for all tissues; PPARγ, PPARγ coactivator 1 α (PGC-1α), and 5´-adenosine monophosphate-activated protein kinase (AMPK) and the activated form phosphorylated-AMPK (pAMPK) for liver, muscle, and rumen; phosphoenolpyruvate carboxykinase (PEPCK) for liver and kidney; and uncoupling protein 2 (UCP2) for liver. Statistical analysis was conducted using Proc Mixed in SAS and included the fixed effects of dietary treatment, cow age, block, and the random effect of pen. Dietary treatments resulted in cows fed HIGH having greater (P ≤ 0.04) ADG and final BW than cows fed LOW. Abundance of ubiquitin in muscle was greater (P = 0.009) in cows fed LOW, and PCG-1 α in liver was greater (P = 0.03) in cows fed HIGH. Hepatic O2 consumption was greater in HIGH (P ≤ 0.04). Feed intake can influence the abundance of important metabolic proteins and suggest that protein degradation may increase in muscle from moderately nutrient restricted cows and that energy metabolism in liver increases in cows fed above NE requirements.

Oxidative status of human milk and its variations during cold storage.

PubMed

Miranda, María; Muriach, María; Almansa, Inmaculada; Jareño, Enrique; Bosch-Morell, Francisco; Romero, Francisco J; Silvestre, Dolores

2004-01-01

Breastfeeding and human milk are widely accepted as optimal for human infants' nutrition. Nowadays lifestyle often makes it difficult to maintain or even initiate human lactation. This situation is mostly related to the workload of women away from home. New approaches are needed to enable maternal lactation under these circumstances. Human breastmilk storage for differed use is one possibility. The aim of this study was to assess changes in glutathione peroxidase (GPx) activity and in the concentration of the lipid peroxidation marker, malondialdehyde (MDA), when human milk was kept refrigerated or frozen. Thirty-two human milk samples were assayed for GPx activity and MDA concentration. Samples were divided in three aliquot portions, the first to be immediately analysed, the second to be refrigerated at 4 degrees C and analysed 24 h thereafter, and the third to be frozen at -20 degrees C and assayed after 10 days. GPx activity was significantly decreased in refrigerated and in frozen milk, when compared to their control samples. MDA was increased only in refrigerated milk but not in frozen samples. Thus, freezing seems better than refrigeration in order to prevent lipid peroxidation in stored human milk samples.

Oxidative stability of red palm oils blended chicken nuggets during frozen storage

NASA Astrophysics Data System (ADS)

Kamaruzaman, Nurkhuzaiah; Babji, Abdul Salam

2014-09-01

The effects of red palm oils known as Naturally Vitamin Rich Oil (NVRO) on the lipid stability of chicken nuggets were determined. Lipid oxidation was analyzed during frozen storage (-18 °C) for up to 4 months. Thiobarbituric acid (TBA) values and peroxide value (PV) for all samples chicken nuggets increased throughout 3 months of frozen storage and then start to decrease thereafter. Chicken nuggets formulated with NVRO, NVRO-100 and NVRO-50 showed lower TBA values and PV compared to the samples prepared with chicken fat. However, among NVRO, there were not significantly different for most of the months. It was concluded that the utilization of red palm oils in chicken nuggets significantly reduced the oxidation of lipid, which was indicated by the PV throughout 4 months of frozen storage.

System for maintaining materials at freezer temperatures for shipping

DOEpatents

Schabron, John F [Laramie, WY; Sorini-Wong, Susan S [Laramie, WY

2007-08-28

At least one embodiment of the inventive technology relates to a frozen environmental sample temperature control system that comprises a frozen formulation having water in an amount from substantially 87% to 78% by weight of the formulation, and salt in an amount from substantially 13% to 22% by weight of the formulation, the system further including at least one container containing the frozen formulation; and a cooler having insulating material disposed between an outer wall and an inner surface that defines an inner chamber into which the at least one container and the at least one frozen environmental sample may be placed for storage and/or transport. Various embodiments may incorporate specific types of insulating material and/or adaptations to an inner surface of the cooler to enhance the insulation effected thereby.

Indication and method of frozen section in vaginal radical trachelectomy.

PubMed

Chênevert, Jacinthe; Têtu, Bernard; Plante, Marie; Roy, Michel; Renaud, Marie-Claude; Grégoire, Jean; Grondin, Katherine; Dubé, Valérie

2009-09-01

Vaginal radical trachelectomy (VRT) is a fertility-sparing surgical technique used as an alternative to radical hysterectomy in early stage cervical carcinoma. With the advent of VRT, preoperative evaluation of the surgical margin has become imperative, because if the tumor is found within 5 mm of the endocervical margin, additional surgical resection is required. In a study published earlier from our center, we came to the conclusion that a frozen section should be conducted only when a cancerous lesion is grossly visible, and that it could be omitted in normal-looking specimens or VRT with nonspecific lesions. Since then, 53 VRT have been performed in our center, and frozen sections were conducted according to these recommendations. Fifteen VRT were grossly normal, 24 had a nonspecific lesion and 14 showed a grossly visible lesion. Final margins were satisfactory on all 15 grossly normal specimens. Of the 24 VRT with nonspecific lesions, 2 cases for which no frozen section was performed had unsatisfactory final margins (<5 mm). Of the 14 VRT with grossly visible lesions, 3 cases were inadequately evaluated by frozen section due to sampling errors, which led to unsatisfactory final margin assessment. These results confirm that a frozen section can be omitted on normal looking VRT specimens, but contrary to results published earlier, we recommend that a frozen section be performed on all VRT with nonspecific lesions. As for VRT with a grossly visible lesion, frozen section evaluation is still warranted, and we recommend increasing the sampling to improve the adequacy of frozen sections.

Comparative performance of isolation methods using Preston broth, Bolton broth and their modifications for the detection of Campylobacter spp. from naturally contaminated fresh and frozen raw poultry meat.

PubMed

Seliwiorstow, T; De Zutter, L; Houf, K; Botteldoorn, N; Baré, J; Van Damme, I

2016-10-03

The performance of different isolation methods was evaluated for the detection of Campylobacter from naturally contaminated raw poultry meat. Therefore, fresh and frozen poultry meat samples were analysed using the standard procedure (ISO 10272-1:2006), enrichment in Preston broth, and enrichment in modified Bolton broth (supplemented with (i) potassium clavulanate (C-BB), (ii) triclosan (T-BB), (iii) polymyxin B (P-BB)). The enrichment cultures were streaked onto both modified charcoal cefoperazone deoxycholate agar (mCCDA) and RAPID'Campylobacter agar (RCA). Moreover, direct plating on mCCDA and RCA was performed to quantify Campylobacter. In total, 33 out of 59 fresh retail meat samples (55.9%) were Campylobacter positive. For both fresh and frozen poultry meat samples, enrichment in Bolton broth (ISO 10272-1:2006) resulted in a higher number of positive samples than enrichment in Preston broth. Supplementation of Bolton broth with potassium clavulanate (C-BB) and triclosan (T-BB) enhanced the Campylobacter recovery from fresh poultry meat compared to non-supplemented Bolton broth, although the use of C-BB was less applicable than T-BB for Campylobacter recovery from frozen samples. Additionally, the use of RCA resulted in a higher isolation rate compared to mCCDA. The present study demonstrates the impact of culture medium on the recovery of Campylobacter from fresh and frozen naturally contaminated poultry meat samples and can support laboratories in choosing the most appropriate culturing method to detect Campylobacter. Copyright © 2016 Elsevier B.V. All rights reserved.

Colloid centrifugation of fresh stallion semen before cryopreservation decreased microorganism load of frozen-thawed semen without affecting seminal kinetics.

PubMed

Guimarães, T; Lopes, G; Pinto, M; Silva, E; Miranda, C; Correia, M J; Damásio, L; Thompson, G; Rocha, A

2015-01-15

Freezability of equine semen may be influenced by microorganism population of semen. The objective of this study was to verify the effect of single-layer density gradient centrifugation (SLC) of fresh semen before cryopreservation on semen's microbial load (ML) and sperm cells kinetics after freezing-thawing. For that, one ejaculate was collected from 20 healthy stallions and split into control (C) samples (cryopreserved without previous SLC) and SLC samples (subjected to SLC). Semen cryopreservation was performed according to the same protocol in both groups. Microbial load of each microorganism species and total microbial load (TML) expressed in colony-forming units (CFU/mL) as well as frozen-thawed sperm kinetics were assessed in both groups. Additional analysis of the TML was performed, subdividing the frozen-thawed samples in "suitable" (total motility ≥ 30%) and "unsuitable" (total motility < 30%) semen for freezing programs, and comparing the C and SLC groups within these subpopulations. After thawing, SLC samples had less (P < 0.05) TML (88.65 × 10(2) ± 83.8 × 10(2) CFU/mL) than C samples (155.69 × 10(2) ± 48.85 × 10(2) CFU/mL), mainly due to a reduction of Enterococcus spp. and Bacillus spp. A relationship between post-thaw motility and SLC effect on ML was noted, as only in samples with more than 30% total motility was ML reduced (P < 0.05) by SLC (from 51.33 × 10(2) ± 33.26 × 10(2) CFU/mL to 26.68 × 10(2) ± 12.39 × 10(2) CFU/mL in "suitable" frozen-thawed semen vs. 240.90 × 10(2) ± 498.20 × 10(2) to 139.30 × 10(2) ± 290.30 × 10(2) CFU/mL in "unsuitable" frozen-thawed semen). The effect of SLC on kinetics of frozen-thawed sperm cells was negligible. Copyright © 2015 Elsevier Inc. All rights reserved.

Challenges of biological sample preparation for SIMS imaging of elements and molecules at subcellular resolution

NASA Astrophysics Data System (ADS)

Chandra, Subhash

2008-12-01

Secondary ion mass spectrometry (SIMS) based imaging techniques capable of subcellular resolution characterization of elements and molecules are becoming valuable tools in many areas of biology and medicine. Due to high vacuum requirements of SIMS, the live cells cannot be analyzed directly in the instrument. The sample preparation, therefore, plays a critical role in preserving the native chemical composition for SIMS analysis. This work focuses on the evaluation of frozen-hydrated and frozen freeze-dried sample preparations for SIMS studies of cultured cells with a CAMECA IMS-3f dynamic SIMS ion microscope instrument capable of producing SIMS images with a spatial resolution of 500 nm. The sandwich freeze-fracture method was used for fracturing the cells. The complimentary fracture planes in the plasma membrane were characterized by field-emission secondary electron microscopy (FESEM) in the frozen-hydrated state. The cells fractured at the dorsal surface were used for SIMS analysis. The frozen-hydrated SIMS analysis of individual cells under dynamic primary ion beam (O 2+) revealed local secondary ion signal enhancements correlated with the water image signals of 19(H 3O) +. A preferential removal of water from the frozen cell matrix in the Z-axis was also observed. These complications render the frozen-hydrated sample type less desirable for subcellular dynamic SIMS studies. The freeze-drying of frozen-hydrated cells, either inside the instrument or externally in a freeze-drier, allowed SIMS imaging of subcellular chemical composition. Morphological evaluations of fractured freeze-dried cells with SEM and confocal laser scanning microscopy (CLSM) revealed well-preserved mitochondria, Golgi apparatus, and stress fibers. SIMS analysis of fractured freeze-dried cells revealed well-preserved chemical composition of even the most highly diffusible ions like K + and Na + in physiologically relevant concentrations. The high K-low Na signature in individual cells provided a rule-of-thumb criterion for the validation of sample preparation. The fractured freeze-dried cells allowed 3-D SIMS imaging and localization of 13C 15N labeled molecules and therapeutic drugs containing an elemental tag. Examples are shown to demonstrate that both diffusible elements and molecules are prone to artifact-induced relocation at subcellular scale if the sample preparation is compromised. The sample preparation is problem dependent and may vary widely between the diverse sample types of biological systems and the type of instrument used for SIMS analysis. The sample preparation, however, must be validated so that SIMS can be applied with confidence in biology and medicine.

TL and ESR based identification of gamma-irradiated frozen fish using different hydrolysis techniques

NASA Astrophysics Data System (ADS)

Ahn, Jae-Jun; Akram, Kashif; Shahbaz, Hafiz Muhammad; Kwon, Joong-Ho

2014-12-01

Frozen fish fillets (walleye Pollack and Japanese Spanish mackerel) were selected as samples for irradiation (0-10 kGy) detection trials using different hydrolysis methods. Photostimulated luminescence (PSL)-based screening analysis for gamma-irradiated frozen fillets showed low sensitivity due to limited silicate mineral contents on the samples. Same limitations were found in the thermoluminescence (TL) analysis on mineral samples isolated by density separation method. However, acid (HCl) and alkali (KOH) hydrolysis methods were effective in getting enough minerals to carry out TL analysis, which was reconfirmed through the normalization step by calculating the TL ratios (TL1/TL2). For improved electron spin resonance (ESR) analysis, alkali and enzyme (alcalase) hydrolysis methods were compared in separating minute-bone fractions. The enzymatic method provided more clear radiation-specific hydroxyapatite radicals than that of the alkaline method. Different hydrolysis methods could extend the application of TL and ESR techniques in identifying the irradiation history of frozen fish fillets.

Comparison of techniques for stabilizing hemoglobins of rainbow trout (Salmo gairdneri) during frozen storage

USGS Publications Warehouse

Reinitz, G.L.

1976-01-01

1. The stability of hemoglobin of rainbow trout under frozen conditions in oxyform, carboxyform, and cyanometform was examined.2. Carboxyhemoglobin retained its original electrophoretic banding pattern after 14 days of frozen storage, whereas oxyform and cyanometform hemoglobins did not.3. Banding patterns changed in some samples in all treatment groups after 21 days of storage.

Prevalence of Listeria spp and Listeria monocytogenes in meat and fermented fish in Malaysia.

PubMed

Hassan, Z; Purwati, E; Radu, S; Rahim, R A; Rusul, G

2001-06-01

Fermented fish and meat samples were purchased from supermarket and wet market for microbiological analysis of Listeria species and Listeria monocytogenes contamination. Listeria species were isolated from 17 (73.9%) of 23 samples of imported frozen beef, 10 (43.5%) of the 23 samples of local beef and 14 (56%) of the 25 samples of fermented fish from wet market. Listeria monocytogenes occurred in 15 (75%) of the frozen beef samples, 6 (30.4%) of the 23 samples of local meat and 3 (12%) of the 25 samples from fermented fish. Listeria species was not isolated from any of the 23 samples of imported frozen beef from supermarket and from the 5 samples of buffalo meat examined. This highlights the possibility of Listeria spp or L. monocytogenes to persist in meat and fermented fish in wet market and raises the problem of illness due to the handling and consumption of Listeria-contaminated meat or fermented fish are likely as evidence by the high contamination rates of samples sold at the wet market.

Effect of ELISA kit manufacturing process and incubation time on progesterone concentration measured in dog serum for ovulation diagnosis - Short communication.

PubMed

Thuróczy, Julianna; Reiczigel, Jenő; Balogh, Lajos

2016-09-01

Twenty-two serum samples of healthy bitches were tested with the frozen and lyophilised version of the same ELISA kit (Quanticheck, Faculty of Veterinary Science, Budapest, Hungary). Samples were chosen on the basis of their progesterone (P4) concentrations, which were between 1.00 and 20.00 ng/mL. As it is well known, this range has the highest clinical relevance in ovulation diagnosis. Both types of microplates were read at 15-min intervals from the 15th until the 90th minute (min) of incubation, and the results were compared with those of frozen plates at 60 min of incubation as 100 percent. Lyophilised microplates gave on average 18 percent higher results than the frozen version at equal incubation times. The highest difference between lyophilised and frozen samples was observed at 45 and 60 min of incubation. Ninety-four percent of the reaction in the frozen microplate occurred in the first 15 min, and during the subsequent 30 min the reaction seemingly stopped. After the 45th min of incubation, this 94 percent increased to 108 percent in the subsequent 30 min, which remained the final approximate result at the end of the 90 min of incubation. In contrast to the frozen microplate, the measured concentration increased continuously in the lyophilised version and reached the highest level at the 60th min. The results of the lyophilised microplate reached the same level at 30 min of incubation as those of the frozen version at 60 min. In conclusion, a mechanical increase or decrease of the incubation time does not generate a linear change in the test results. This study demonstrated that the results of a series of samples collected from the same bitch cannot be compared if they are measured with different laboratory methods or different ELISA kits.

[Undifferentiated (embryonal) sarcoma of the liver. Report of a case].

PubMed

Orozco, H; Mercado, M A; Takahashi, T; Chan, C; Quintanilla, L; Jiménez, M; Sosa, R; Esquivel, E

1991-01-01

A 15-year-old woman who was studied because an abdominal mass at the Instituto Nacional de la Nutricion Salvador Zubiran (INNSZ) is reported. The history revealed only malaise and mild abdominal pain. At physical exploration, an abdominal mass in the upper right quadrant was found. Liver function tests were normal. Abdominal ultrasound and computerized tomography revealed a large cystic mass of the right hepatic lobe. She underwent exploratory laparotomy. Intraoperative frozen sections of the biopsies demonstrated undifferentiated sarcoma of the liver, and an extended right trisegmentectomy was performed. Postoperative outcome was uneventful. Adjuvant treatment with doxorubicin and dacarbazine was given, and at six months of follow-up, the patient is alive without any evidence of recurrence. Clinical and histopathologic features of this rare malignant tumor are discussed, as well as the therapeutic choices.

Freeze chromatography method and apparatus

DOEpatents

Scott, C.D.

1987-04-16

A freeze chromatography method and apparatus are provided which enable separation of the solutes contained in a sample. The apparatus includes an annular column construction comprising cylindrical inner and outer surfaces defining an annular passage therebetween. One of the surfaces is heated and the other cooled while passing an eluent through the annular passageway so that the eluent in contact with the cooled surface freezes and forms a frozen eluent layer thereon. A mixture of solutes dissolved in eluent is passed through the annular passageway in contact with the frozen layer so that the sample solutes in the mixture will tend to migrate either toward or away the frozen layer. The rate at which the mixture flows through the annular passageway is controlled so that the distribution of the sample solutes approaches that at equilibrium and thus a separation between the sample solutes occurs. 3 figs.

[Adaptability of sweet corn ears to a frozen process].

PubMed

Ramírez Matheus, Alejandra O; Martínez, Norelkys Maribel; de Bertorelli, Ligia O; De Venanzi, Frank

2004-12-01

The effects of frozen condition on the quality of three sweet corn ears (2038, 2010, 2004) and the pattern (Bonanza), were evaluated. Biometrics characteristics like ear size, ear diameter, row and kernel deep were measured as well as chemical and physical measurement in fresh and frozen states. The corn ears were frozen at -95 degrees C by 7 minutes. The yield and stability of the frozen ears were evaluated at 45 and 90 days of frozen storage (-18 degrees C). The average commercial yield as frozen corn ear for all the hybrids was 54.2%. The industry has a similar value range of 48% to 54%. The ear size average was 21.57 cm, row number was 15, ear diameter 45.54 mm and the kernel corn deep was 8.57 mm. All these measurements were found not different from commercial values found for the industry. All corn samples evaluated showed good stability despites the frozen processing and storage. Hybrid 2038 ranked higher in quality.

The BUME method: a new rapid and simple chloroform-free method for total lipid extraction of animal tissue

NASA Astrophysics Data System (ADS)

Löfgren, Lars; Forsberg, Gun-Britt; Ståhlman, Marcus

2016-06-01

In this study we present a simple and rapid method for tissue lipid extraction. Snap-frozen tissue (15-150 mg) is collected in 2 ml homogenization tubes. 500 μl BUME mixture (butanol:methanol [3:1]) is added and automated homogenization of up to 24 frozen samples at a time in less than 60 seconds is performed, followed by a 5-minute single-phase extraction. After the addition of 500 μl heptane:ethyl acetate (3:1) and 500 μl 1% acetic acid a 5-minute two-phase extraction is performed. Lipids are recovered from the upper phase by automated liquid handling using a standard 96-tip robot. A second two-phase extraction is performed using 500 μl heptane:ethyl acetate (3:1). Validation of the method showed that the extraction recoveries for the investigated lipids, which included sterols, glycerolipids, glycerophospholipids and sphingolipids were similar or better than for the Folch method. We also applied the method for lipid extraction of liver and heart and compared the lipid species profiles with profiles generated after Folch and MTBE extraction. We conclude that the BUME method is superior to the Folch method in terms of simplicity, through-put, automation, solvent consumption, economy, health and environment yet delivering lipid recoveries fully comparable to or better than the Folch method.

Disease Modeling and Gene Therapy of Copper Storage Disease in Canine Hepatic Organoids.

PubMed

Nantasanti, Sathidpak; Spee, Bart; Kruitwagen, Hedwig S; Chen, Chen; Geijsen, Niels; Oosterhoff, Loes A; van Wolferen, Monique E; Pelaez, Nicolas; Fieten, Hille; Wubbolts, Richard W; Grinwis, Guy C; Chan, Jefferson; Huch, Meritxell; Vries, Robert R G; Clevers, Hans; de Bruin, Alain; Rothuizen, Jan; Penning, Louis C; Schotanus, Baukje A

2015-11-10

The recent development of 3D-liver stem cell cultures (hepatic organoids) opens up new avenues for gene and/or stem cell therapy to treat liver disease. To test safety and efficacy, a relevant large animal model is essential but not yet established. Because of its shared pathologies and disease pathways, the dog is considered the best model for human liver disease. Here we report the establishment of a long-term canine hepatic organoid culture allowing undifferentiated expansion of progenitor cells that can be differentiated toward functional hepatocytes. We show that cultures can be initiated from fresh and frozen liver tissues using Tru-Cut or fine-needle biopsies. The use of Wnt agonists proved important for canine organoid proliferation and inhibition of differentiation. Finally, we demonstrate that successful gene supplementation in hepatic organoids of COMMD1-deficient dogs restores function and can be an effective means to cure copper storage disease. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Acquisition of thin coronal sectional dataset of cadaveric liver.

PubMed

Lou, Li; Liu, Shu Wei; Zhao, Zhen Mei; Tang, Yu Chun; Lin, Xiang Tao

2014-04-01

To obtain the thin coronal sectional anatomic dataset of the liver by using digital freezing milling technique. The upper abdomen of one Chinese adult cadaver was selected as the specimen. After CT and MRI examinations verification of absent liver lesions, the specimen was embedded with gelatin in stand erect position and frozen under profound hypothermia, and the specimen was then serially sectioned from anterior to posterior layer by layer with digital milling machine in the freezing chamber. The sequential images were captured by means of a digital camera and the dataset was imported to imaging workstation. The thin serial section of the liver added up to 699 layers with each layer being 0.2 mm in thickness. The shape, location, structure, intrahepatic vessels and adjacent structures of the liver was displayed clearly on each layer of the coronal sectional slice. CT and MR images through the body were obtained at 1.0 and 3.0 mm intervals, respectively. The methodology reported here is an adaptation of the milling methods previously described, which is a new data acquisition method for sectional anatomy. The thin coronal sectional anatomic dataset of the liver obtained by this technique is of high precision and good quality.

Frozen and fresh ovarian tissue require different culture media to promote in vitro development of bovine preantral follicles.

PubMed

Castro, Simone Vieira; Carvalho, Adeline Andrade; Silva, Cleidson Manoel Gomes; Santos, Francielli Weber; Campello, Cláudio Cabral; de Figueiredo, José Ricardo; Rodrigues, Ana Paula Ribeiro

2014-10-01

The aim of this study was to evaluate the efficiency of different media in the in vitro culture of bovine preantral follicles that were used either fresh or following slow freezing treatment. Frozen and fresh noncultured or cultured ovarian fragments were processed for histological, viability, and cell proliferation analyses. For cryopreservation, a solution containing 1.5 M ethylene glycol was frozen in a programmable biological freezer. After thawing, a portion of the samples was destined for frozen controls. The remainder were cultured in vitro for 5 days in three media: α-MEM, McCoy, or M199. Samples from these culture media were collected on days 1 and 5 for quantification of reactive oxygen species (ROS) and for hormonal assays. In fresh-cultured tissues, the percentage of morphologically normal follicles was significantly higher when cultured in M199 compared to that in the other media. In frozen-cultured tissues, McCoy medium was significantly superior to the other media, and was the only treatment that helped in maintaining the viability similar to fresh and frozen controls. Upon quantification of the nucleolus organizer region, we observed greater proliferation of granulosa cells in the frozen-cultured tissues with McCoy medium, and lesser proliferation in fresh-cultured tissues only with α-MEM. In frozen-cultured tissues, ROS levels were highest at day 1 and progressively reduced during culture, independent of the media used. In conclusion, under the conditions used in this study, the M199 and McCoy media are recommended for the culture of follicles derived from fresh and frozen ovarian tissues, respectively.

Effect of freezing, irradiation, and frozen storage on survival of Salmonella in concentrated orange juice.

PubMed

Niemira, Brendan A; Sommers, Christopher H; Boyd, Glenn

2003-10-01

Six strains of Salmonella (Anatum F4317, Dublin 15480, Enteritidis 13076, Enteritidis WY15159, Stanley H0588, and Typhimurium 14028) were individually inoculated into orange juice concentrate (OJC) and frozen to -20 degrees C. The frozen samples were treated with 0 (nonirradiated), 0.5, 1.0, or 2.0 kGy of gamma radiation and held frozen for 1 h, and the surviving bacterial population was assessed. The strains showed significant variability in their response to freezing and to freezing in combination with irradiation. The response was dose dependent. Relative to the nonfrozen, nonirradiated control, the reduction following the highest dose (2.0 kGy) ranged from 1.29 log CFU/ml (Salmonella Typhimurium) to 2.17 log CFU/ml (Salmonella Stanley). Samples of OJC inoculated with Salmonella Enteritidis WY15159 and irradiated were stored at -20 degrees C for 1, 2, 7, or 14 days, and the surviving population was determined. Relative to the nonfrozen, nonirradiated control, after 14 days, the population was reduced by 1.2 log CFU/ml in the nonirradiated samples and by 3.3 log CFU/ml following treatment with 2.0 kGy. The combination of frozen storage plus irradiation resulted in greater overall reductions than either process alone.

Photochemical Production of Singlet Oxygen from Dissolved Organic Matter in Ice.

PubMed

Fede, Alexis; Grannas, Amanda M

2015-11-03

Dissolved natural organic matter (DOM) is a ubiquitous component of natural waters and an important photosensitizer. A variety of reactive oxygen species (ROS) are known to be produced from DOM photochemistry, including singlet oxygen, 1O2. Recently, it has been determined that humic-like substances and unknown organic chromophores are significant contributors to sunlight absorption in snowpack; however, DOM photochemistry in snow/ice has received little attention in the literature. We recently showed that DOM plays an important role in indirect photolysis processes in ice, producing ROS and leading to the efficient photodegradation of a probe hydrophobic organic pollutant, aldrin.1 ROS scavenger experiments indicated that 1O2 played a significant role in the indirect photodegradation of aldrin. Here we quantitatively examine 1O2 photochemically produced from DOM in frozen and liquid aqueous solutions. Steady-state 1O2 production is enhanced up to nearly 1000 times in frozen DOM samples compared to liquid samples. 1O2 production is dependent on the concentration of DOM, but the nature of the DOM source (terrestrial vs microbial) does not have a significant effect on 1O2 production in liquid or frozen samples, with different source types producing similar steady-state concentrations of 1O2. The temperature of frozen samples also has a significant effect on steady-state 1O2 production in the range of 228-262 K, with colder samples producing more steady-state 1O2. The large enhancement in 1O2 in frozen samples suggests that it may play a significant role in the photochemical processes that occur in snow and ice, and DOM could be a significant, but to date poorly understood, oxidant source in snow and ice.

Comparison of human papillomavirus detection between freshly frozen tissue and paraffin embedded tissue of invasive cervical cancer

PubMed Central

2010-01-01

Background Human Papillomavirus (HPV) detection results comparing paraffin embedded cervical tissue and other cervical specimens have been done with varying degrees of agreement. However, studies comparing freshly frozen specimens and paraffin embedded specimens of invasive cervical carcinomas are lacking. The aim of the study was to compare HPV detection using SPF10 broad-spectrum primers PCR followed by DEIA and genotyping by LiPA25 (version 1) between freshly frozen cervical tissue samples and paraffin embedded blocks of cervical tissue from the same patient. There were 171 pairs of paraffin embedded and freshly frozen samples analyzed from cervical carcinoma cases from Kampala, Uganda. Results 88.9% (95% CI: 83.2%-93.2%) of paraffin embedded samples were HPV positive compared with 90.1% (95% CI: 84.6%-94.1%) of freshly frozen samples, giving an overall agreement in HPV detection between fresh tissue and paraffin embedded tissue at 86.0% (95% CI: 79.8%-90.8%). Although the proportion of HPV positive cases in freshly frozen tissue was higher than those in paraffin blocks, the difference was not statistically significant (p > 0.05). In both types of tissues, single HPV infections were predominant, with HPV16 accounting for 47% of positive cases. Comparison in the overall agreement, taking into accounts not only positivity in general, but also HPV types, showed a 65% agreement (complete agreement of 59.7%, partial agreement of 5.3%) and complete disagreement of 35.0%. HPV detection in squamous cell carcinomas (SCC) and adenocarcinomas (ADC) was similar in fresh tissue or paraffin blocks (p ≥ 0.05). p16 immunostaining in samples that had at least one HPV negative results showed that 24 out of 25 cases had an over-expressed pattern. Conclusions HPV DNA detection was lower among ADC as compared to SCC. However, such differences were minimized when additional p16 testing was added, suggesting that the technical issues may largely explain the HPV negative cases. PMID:20846370

Relative IGF-1 and IGF-2 gene expression in maternal and fetal tissues from diabetic swine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wolverton, C.K.; Leaman, D.W.; White, M.E.

1990-02-26

Fourteen pregnant, crossbred gilts were utilized in this study. Seven gilts were injected with alloxan (50 mg/kg) at day 75 of gestation to induce diabetes. Gilts underwent caesarean section on day 105 of gestation. Samples were collected from maternal skeletal muscle, adipose tissue, uterus and endometrium; and from fetal skeletal muscle, adipose tissue, placenta, liver, lung, kidney, heart, brain and spleen. Tissues were frozen in liquid nitrogen for later analysis of IGF-1 and IGF-2 gene expression. Samples were pooled and total RNA was isolated using the guanidine isothiocynate method. Total mRNA was analyzed by dot blot hybridization. Blots were probedmore » with {sup 32}P-cDNA for porcine IGF-1 and rat IGF-2. IGF-1 gene expression in maternal tissues was unaffected by diabetes. Maternal diabetes increased IGF-2 mRNA in maternal adipose tissue but exhibited no effect in muscle or uterus. Expression of IGF-2 by maternal endometrium was decreased by diabetes. Maternal diabetes induced an increase in IGF-1 gene expression in muscle and placenta while causing an increase in IGF-2 expression in fetal liver and placenta. IGF-2 mRNA was lower in lung from fetuses of diabetic mothers than in controls. These results suggest that maternal diabetes alters IGF-1 and IGF-2 gene expression in specific tissues and differential regulation of these genes appears to exist in the mother and developing fetus.« less

Normothermic machine perfusion of donor livers without the need for human blood products

PubMed Central

Matton, Alix P. M.; Burlage, Laura C.; van Rijn, Rianne; de Vries, Yvonne; Karangwa, Shanice A.; Nijsten, Maarten W.; Gouw, Annette S. H.; Wiersema‐Buist, Janneke; Adelmeijer, Jelle; Westerkamp, Andrie C.; Lisman, Ton

2018-01-01

Normothermic machine perfusion (NMP) enables viability assessment of donor livers prior to transplantation. NMP is frequently performed by using human blood products including red blood cells (RBCs) and fresh frozen plasma (FFP). Our aim was to examine the efficacy of a novel machine perfusion solution based on polymerized bovine hemoglobin‐based oxygen carrier (HBOC)‐201. Twenty‐four livers declined for transplantation were transported by using static cold storage. Upon arrival, livers underwent NMP for 6 hours using pressure‐controlled portal and arterial perfusion. A total of 12 livers were perfused using a solution based on RBCs and FFPs (historical cohort), 6 livers with HBOC‐201 and FFPs, and another 6 livers with HBOC‐201 and gelofusine, a gelatin‐based colloid solution. Compared with RBC + FFP perfused livers, livers perfused with HBOC‐201 had significantly higher hepatic adenosine triphosphate content, cumulative bile production, and portal and arterial flows. Biliary secretion of bicarbonate, bilirubin, bile salts, and phospholipids was similar in all 3 groups. The alanine aminotransferase concentration in perfusate was lower in the HBOC‐201–perfused groups. In conclusion, NMP of human donor livers can be performed effectively using HBOC‐201 and gelofusine, eliminating the need for human blood products. Perfusing livers with HBOC‐201 is at least similar to perfusion with RBCs and FFP. Some of the biomarkers of liver function and injury even suggest a possible superiority of an HBOC‐201–based perfusion solution and opens a perspective for further optimization of machine perfusion techniques. Liver Transplantation 24 528–538 2018 AASLD. PMID:29281862

Effect of inulin on the physicochemical properties, flow behavior and probiotic survival of frozen yogurt.

PubMed

Rezaei, Rahil; Khomeiri, Morteza; Aalami, Mehran; Kashaninejad, Mahdi

2014-10-01

This study investigated the effect of inulin (0, 1 and 2 %), on some physicochemical properties of frozen yogurt, as well as its effect on flow behavior and probiotic survival. The results showed that the addition of inulin improved overrun, viscosity and melting properties significantly (p < 0.05); when added at 2 % level, it also had significant effect on pH. Total acceptability of samples revealed that frozen yogurt with 2 % inulin had the most appealing sensory characteristics. The flow behavior of all samples showed their pseudoplastic nature; power law was the best model to predict their flow behavior. In terms of probiotic survival, the sample with 2 % inulin significantly improved the viability of Lactobacillus acidophilus and Bifidobacterium lactis.

Logistics and quality control for DNA sampling in large multicenter studies.

PubMed

Nederhand, R J; Droog, S; Kluft, C; Simoons, M L; de Maat, M P M

2003-05-01

To study associations between genetic variation and disease, large bio-banks need to be created in multicenter studies. Therefore, we studied the effects of storage time and temperature on DNA quality and quantity in a simulation experiment with storage up to 28 days frozen, at 4 degrees C and at room temperature. In the simulation experiment, the conditions did not influence the amount or quality of DNA to an unsatisfactory level. However, the amount of extracted DNA was decreased in frozen samples and in samples that were stored for > 7 days at room temperature. In a sample of patients from 24 countries of the EUROPA trial obtained by mail with transport times up to 1 month DNA yield and quality were adequate. From these results we conclude that transport of non-frozen blood by ordinary mail is usable and practical for DNA isolation for polymerase chain reaction in clinical and epidemiological studies.

DNA methylation profiling of genomic DNA isolated from urine in diabetic chronic kidney disease: A pilot study

PubMed Central

Sexton-Oates, Alexandra; Carmody, Jake; Ekinci, Elif I.; Dwyer, Karen M.; Saffery, Richard

2018-01-01

Aim To characterise the genomic DNA (gDNA) yield from urine and quality of derived methylation data generated from the widely used Illuminia Infinium MethylationEPIC (HM850K) platform and compare this with buffy coat samples. Background DNA methylation is the most widely studied epigenetic mark and variations in DNA methylation profile have been implicated in diabetes which affects approximately 415 million people worldwide. Methods QIAamp Viral RNA Mini Kit and QIAamp DNA micro kit were used to extract DNA from frozen and fresh urine samples as well as increasing volumes of fresh urine. Matched buffy coats to the frozen urine were also obtained and DNA was extracted from the buffy coats using the QIAamp DNA Mini Kit. Genomic DNA of greater concentration than 20μg/ml were used for methylation analysis using the HM850K array. Results Irrespective of extraction technique or the use of fresh versus frozen urine samples, limited genomic DNA was obtained using a starting sample volume of 5ml (0–0.86μg/mL). In order to optimize the yield, we increased starting volumes to 50ml fresh urine, which yielded only 0–9.66μg/mL A different kit, QIAamp DNA Micro Kit, was trialled in six fresh urine samples and ten frozen urine samples with inadequate DNA yields from 0–17.7μg/mL and 0–1.6μg/mL respectively. Sufficient genomic DNA was obtained from only 4 of the initial 41 frozen urine samples (10%) for DNA methylation profiling. In comparison, all four buffy coat samples (100%) provided sufficient genomic DNA. Conclusion High quality data can be obtained provided a sufficient yield of genomic DNA is isolated. Despite optimizing various extraction methodologies, the modest amount of genomic DNA derived from urine, may limit the generalisability of this approach for the identification of DNA methylation biomarkers of chronic diabetic kidney disease. PMID:29462136

DNA methylation profiling of genomic DNA isolated from urine in diabetic chronic kidney disease: A pilot study.

PubMed

Lecamwasam, Ashani; Sexton-Oates, Alexandra; Carmody, Jake; Ekinci, Elif I; Dwyer, Karen M; Saffery, Richard

2018-01-01

To characterise the genomic DNA (gDNA) yield from urine and quality of derived methylation data generated from the widely used Illuminia Infinium MethylationEPIC (HM850K) platform and compare this with buffy coat samples. DNA methylation is the most widely studied epigenetic mark and variations in DNA methylation profile have been implicated in diabetes which affects approximately 415 million people worldwide. QIAamp Viral RNA Mini Kit and QIAamp DNA micro kit were used to extract DNA from frozen and fresh urine samples as well as increasing volumes of fresh urine. Matched buffy coats to the frozen urine were also obtained and DNA was extracted from the buffy coats using the QIAamp DNA Mini Kit. Genomic DNA of greater concentration than 20μg/ml were used for methylation analysis using the HM850K array. Irrespective of extraction technique or the use of fresh versus frozen urine samples, limited genomic DNA was obtained using a starting sample volume of 5ml (0-0.86μg/mL). In order to optimize the yield, we increased starting volumes to 50ml fresh urine, which yielded only 0-9.66μg/mL A different kit, QIAamp DNA Micro Kit, was trialled in six fresh urine samples and ten frozen urine samples with inadequate DNA yields from 0-17.7μg/mL and 0-1.6μg/mL respectively. Sufficient genomic DNA was obtained from only 4 of the initial 41 frozen urine samples (10%) for DNA methylation profiling. In comparison, all four buffy coat samples (100%) provided sufficient genomic DNA. High quality data can be obtained provided a sufficient yield of genomic DNA is isolated. Despite optimizing various extraction methodologies, the modest amount of genomic DNA derived from urine, may limit the generalisability of this approach for the identification of DNA methylation biomarkers of chronic diabetic kidney disease.

Impact of donor health on corneal biochemistry--an unexpected caveat from a pilot study.

PubMed

Kryczka, Tomasz; Chrapusta, Stanisław J; Szaflik, Jacek P; Szaflik, Jerzy; Midelfart, Anna

2014-03-12

To test the possibility that some chronic systemic maladies not directly related to the function of the eye may significantly and permanently disturb corneal metabolism. Contents of selected low molecular weight metabolites were compared among corneas collected from donors who died suddenly of an accident or non-poisoning suicide, or met a sudden non-accidental death from unidentified causes, or died of a chronic cardiovascular disease or of idiopathic liver cirrhosis (N=4 for each group). Corneal buttons were halved; one half was snap-frozen and stored at -80°C, and the other half was stored at +4°C in Eusol-C for 8 days and then was snap-frozen and stored at -80°C until analyzed. Metabolite contents were assessed using high-resolution magic angle spinning proton NMR spectroscopy. Significant between-group differences in corneal biochemical profiles were identified. Most of them were reduced or nullified by the Eusol-C storage, suggesting their link to differences in in vivo corneal environment. The corneas from donors with liver cirrhosis or cardiovascular diseases differed considerably from the remaining ones, both before and after the Eusol-C storage. Various chronic systemic diseases that are not directly related to the function of the eye markedly affect corneal biochemistry. Some of the alterations are likely related to a permanent aberration in corneal metabolism. A study is warranted in larger donor groups on the effect of idiopathic liver cirrhosis and cardiovascular diseases on corneal metabolism and/or a retrospective analysis of the long-term outcome of keratoplasty and other grafting procedures employing materials from these donor groups.

Effect of Frozen Human Epidermis Storage Duration and Cryoprotectant on Barrier Function using Two Model Compounds

PubMed Central

Barbero, Ana M.; Frasch, H. Frederick

2015-01-01

Skin is commonly stored frozen and then thawed prior to use for in-vitro permeation experiments. Does frozen storage of skin alter its barrier property? Numerous studies have found contradictory answers to this question. In this study, the steady state flux and lag time of diethyl phthalate (DEP) were measured for fresh human skin and skin frozen at −85 °C for 1, 2, 3, 6, 9, 12, and 18 months, with 10% glycerol as cryoprotective agent. No significant differences in steady state flux were found between fresh and previously frozen samples (P = 0.6). For lag time, a significant (P = 0.002) difference was found among all groups but comparisons with fresh skin were not significant. Does glycerol have a cryoprotective effect? The steady state flux and lag time of DEP and caffeine were measured through human skin stored at −85 °C for up to 12 months with and without 10 % glycerol. No significant differences in steady state flux or lag time were found between samples stored with or without glycerol for either DEP or caffeine (P ≥ 0.17). These findings support the use of frozen skin to measure the passive permeation of chemicals in studies unconcerned with viability and metabolism. PMID:26606593

Using immunoglobulin Y as an alternative antibody for the detection of hepatitis A virus in frozen liver sections.

PubMed

Bentes, Gentil Arthur; Lanzarini, Natália Maria; Lima, Lyana Rodrigues Pinto; Manso, Pedro Paulo de Abreu; da Silva, Alexandre Dos Santos; Mouta Junior, Sergio da Silva E; Guimarães, Juliana Rodrigues; de Moraes, Marcia Terezinha Baroni; Pelajo-Machado, Marcelo; Pinto, Marcelo Alves

2015-06-01

An increasing amount of research has been conducted on immunoglobulin Y (IgY) because the use of IgY offers several advantages with respect to diagnostic testing, including its easy accessibility, low cost and translatability to large-scale production, in addition to the fact that it can be ethically produced. In a previous work, immunoglobulin was produced and purified from egg yolks (IgY) reactive to hepatitis A virus (HAV) antigens. In the present work, this anti-HAV-specific IgY was used in an indirect immunofluorescence assay to detect viral antigens in liver biopsies that were obtained from experimentally infected cynomolgus monkeys. Fields that were positive for HAV antigen were detected in liver sections using confocal microscopy. In conclusion, egg yolks from immunised hens may be a reliable source for antibody production, which can be employed for immunological studies.

Differentiation of fresh and frozen-thawed fish samples using Raman spectroscopy coupled with chemometric analysis.

PubMed

Velioğlu, Hasan Murat; Temiz, Havva Tümay; Boyaci, Ismail Hakki

2015-04-01

The potential of Raman spectroscopy was investigated in terms of its capability to discriminate the species of the fish samples and determine their freshness according to the number of freezing/thawing cycles they exposed. Species discrimination analysis was carried out on sixty-four fish samples from six different species, namely horse mackerel (Trachurus trachurus), European anchovy (Engraulis encrasicolus), red mullet (Mullus surmuletus), Bluefish (Pomatamus saltatrix), Atlantic salmon (Salmo salar) and flying gurnard (Trigla lucerna). Afterwards, fish samples were exposed to different numbers of freezing/thawing cycles and separated into three batches, namely (i) fresh, (ii) once frozen-thawed (OF) and (iii) twice frozen-thawed (TF) samples, in order to perform the freshness analysis. Raman data collected were used as inputs for chemometric analysis, which enabled us to develop two main PCA models to successfully terminate the studies for both species discrimination and freshness determination analysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Hepatitis C virus in sickle cell disease.

PubMed Central

Hassan, Mohamed; Hasan, Syed; Giday, Samuel; Alamgir, Laila; Banks, Alpha; Frederick, Winston; Smoot, Duane; Castro, Oswaldo

2003-01-01

PURPOSE: To determine the prevalence of hepatitis C virus antibodies (anti-HCV) in patients with sickle cell disease. PATIENTS AND METHODS: Between 1983 and 2001, 150 patients from the Howard University Hospital Center for Sickle Cell Disease were screened for HCV antibody (52% women, 48% men, mean age 34 years). Frozen serum samples from 56 adult sickle cell patients who had participated in previous surveys (1983-92) of HIV and HTLV-1 serology and who were tested in 1992 for anti-HCV antibody--when commercial ELISA test (Ortho) became available--were included in this paper. Of the 150 patients in the study, 132 had sickle cell anemia genotype (SS), 15 had sickle cell hemoglobin-C disease (SC) and three had sickle beta thalassemia. Clinical charts were reviewed for history of blood transfusion, IV drug abuse, homosexuality, tattooing, iron overload, and alcohol abuse. RESULTS: Antibodies to HCV were detected in 53 patients (35.3%). Of the 55 patients who had frozen serum samples tested in 1992, 32 (58%) were reactive for anti-HCV, while only 21 of the 95 patients (22%) tested after 1992 were positive for HCV antibodies (P<0.001). Thirty-nine of 77 patients (51%) who received more than 10 units of packed red blood cells were positive for HCV antibody, and only 14 of 61 patients (23%) who received less than 10 units of packed red blood cells transfusion were positive for HCV antibodies (P<0.001). None of the 12 patients who never received transfusion were positive for HCV antibody. In the 53 anti-HCV positive patients, the mean alanine amino-transferase (ALT) value was 98- and 81 U/L, respectively, for males and females. These values were normal for the HCV-antibody negative patients. The aspartate amino-transferase (AST) and the total bilirubin were also higher in the anti-HCV positive patients compared to patients in the anti-HCV negative group. Forty-four patients (57.1%) who were transfused more than 10 units developed iron overload defined by a serum ferritin level higher than 1,000 ng/ml. A total of 20 of the patients with iron overload underwent liver biopsies. Seven of these 20 patients (35%) were HCV positive. These patients often had more severe liver disease and higher degree of iron deposition. CONCLUSION: The prevalence of HCV antibody and iron overload is directly related to the number of blood transfusions in patients with sickle cell disease. The prevalence of HCV infection has decreased significantly, since blood donor screening for HCV became available. Chronic HCV infection and iron overload place sickle cell patients at risk for significant liver disease. PMID:14620705

Histology as a Valid Tool To Differentiate Fresh from Frozen-Thawed Marinated Fish.

PubMed

Meistro, Serena; Pezzolato, Marzia; Muscolino, Daniele; Giarratana, Filippo; Baioni, Elisa; Panebianco, Antonio; Bozzetta, Elena

2016-08-01

European Commission Regulation (EU) 1276/2011 requires that fishery products intended for raw consumption be frozen at -20°C for not less than 24 h or at -35°C for at least 15 h in order to kill viable parasites other than trematodes. But because marinating processes are not always effective in destroying nematode larvae, raw marinated fish preparations should be frozen before consumption. This study evaluated the performance of a standardized histological method to distinguish between fresh and frozen-thawed raw marinated fish. Sixty anchovy (Engraulis encrasicolus) fillets were sampled: 30 were marinated at +4°C for 24 h, and 30 were frozen at -20°C for 24 h before being marinated for 24 h. All 60 samples were fixed in formalin, processed for paraffin embedding, cut, and stained with hematoxylin and eosin. The slide preparations were examined microscopically by three independent histopathologists and classified as frozen-thawed or negative according to standard operating procedure criteria in use at our laboratory. Performance evaluation of the method showed 100% sensitivity (95% confidence interval [CI], 88.4 to 100%) and 100% specificity (95% CI, 88.4 to 100%), and the interrater agreement (Cohen's kappa) was 1 (95% CI, 0.85 to 1). Histology proved a valid and reliable tool to distinguish fresh from frozen-thawed marinated fish. It can be applied to deliver safe raw fishery products to consumers in order to minimize the risk of anisakidosis.

Study of a scanning HIFU therapy protocol, Part II: Experiment and results

NASA Astrophysics Data System (ADS)

Andrew, Marilee A.; Kaczkowski, Peter; Cunitz, Bryan W.; Brayman, Andrew A.; Kargl, Steven G.

2003-04-01

Instrumentation and protocols for creating scanned HIFU lesions in freshly excised bovine liver were developed in order to study the in vitro HIFU dose response and validate models. Computer-control of the HIFU transducer and 3-axis positioning system provided precise spatial placement of the thermal lesions. Scan speeds were selected in the range of 1 to 8 mm/s, and the applied electrical power was varied from 20 to 60 W. These parameters were chosen to hold the thermal dose constant. A total of six valid scans of 15 mm length were created in each sample; a 3.5 MHz single-element, spherically focused transducer was used. Treated samples were frozen, then sliced in 1.27 mm increments. Digital photographs of slices were downloaded to computer for image processing and analysis. Lesion characteristics, including the depth within the tissue, axial length, and radial width, were computed. Results were compared with those generated from modified KZK and BHTE models, and include a comparison of the statistical variation in the across-scan lesion radial width. [Work supported by USAMRMC.

Filter paper-assisted cell transfer (FaCT) technique: A novel cell-sampling technique for intraoperative diagnosis of central nervous system tumors.

PubMed

Kawamura, Jumpei; Kamoshida, Shingo; Shimakata, Takaaki; Hayashi, Yurie; Sakamaki, Kuniko; Denda, Tamami; Kawai, Kenji; Kuwao, Sadahito

2017-04-01

Intraoperative diagnosis of central nervous system (CNS) tumors provides critical guidance to surgeons in the determination of surgical resection margins and treatment. The techniques and preparations used for the intraoperative diagnosis of CNS tumors include frozen sectioning and cytologic methods (squash smear and touch imprint). Cytologic specimens, which do not have freezing artifacts, are important as an adjuvant tool to frozen sections. However, if the amount of submitted tissue samples is limited, then it is difficult to prepare both frozen sections and squash smears or touch imprint specimens from a single sample at the same time. Therefore, the objective of this study was to derive cells directly from filter paper on which tumor samples are placed. The authors established the filter paper-assisted cell transfer (FaCT) smear technique, in which tumor cells are transferred onto a glass slide directly from the filter paper sample spot after the biopsy is removed. Cell yields and diagnostic accuracy of the FaCT smears were assessed in 40 CNS tumors. FaCT smears had ample cell numbers and well preserved cell morphology sufficient for cytologic diagnosis, even if the submitted tissues were minimal. The overall diagnostic concordance rates between frozen sections and FaCT smears were 90% and 87.5%, respectively (no significant differences). When combining FaCT smears with frozen sections, the diagnostic concordance rate rose to 92.5%. The current results suggest that the FaCT smear technique is a simple and effective processing method that has significant value for intraoperative diagnosis of CNS tumors. Cancer Cytopathol 2017;125:277-282. © 2016 American Cancer Society. © 2017 American Cancer Society.

Cryopreserved and frozen hyaline cartilage imaged by environmental scanning electron microscope. An experimental and prospective study.

PubMed

Sastre, Sergi; Suso, Santiago; Segur, Josep-Maria; Bori, Guillem; Carbonell, José-Antonio; Agustí, Elba; Nuñez, Montse

2008-08-01

To obtain images of the articular surface of osteochondral grafts (fresh, frozen, and cryopreserved in RPMI) using an environmental scanning electron microscope (ESEM). To evaluate and compare the main morphological aspects of the chondral surface of the fresh, frozen, and cryopreserved grafts as visualized via ESEM. The study was based on osteochondral fragments from the internal condyle of the knee joint of New Zealand rabbits, corresponding to the chondral surface from fresh, frozen, and cryopreserved samples. One hundred ESEM images were obtained from each group and then classified according to a validated system. The kappa index and the corresponding concordance index were calculated, and the groups were compared by Pearson's chi-squared test (p < 0.05). The articular surface of cryopreserved osteochondral grafts had fewer even surfaces and filled lacunae and a higher number of empty lacunae as compared to fresh samples; these differences correspond to images of cell membrane lesions that lead to destruction of the chondrocyte. Frozen grafts showed more hillocky and knobby surfaces than did fresh grafts; they also had a greater number of empty chondrocyte lacunae. ESEM is useful for obtaining images of the surface of osteochondral grafts. When compared to fresh samples, cryopreservation in RPMI medium produces changes in the surface of hyaline cartilage, but to a lesser extent than those produced by freezing.

Blind Biobanking of the Prostatectomy Specimen: Critical Evaluation of the Existing Techniques and Development of the New 4-Level Tissue Extraction Model With High Sampling Efficacy.

PubMed

Tolkach, Yuri; Eminaga, Okyaz; Wötzel, Fabian; Huss, Sebastian; Bettendorf, Olaf; Eltze, Elke; Abbas, Mahmoud; Imkamp, Florian; Semjonow, Axel

2017-03-01

Fresh tissue is mandatory to perform high-quality translation studies. Several models for tissue extraction from prostatectomy specimens without guidance by frozen sections are already introduced. However, little is known about the sampling efficacy of these models, which should provide representative tissue in adequate volumes, account for multifocality and heterogeneity of tumor, not violate the routine final pathological examination, and perform quickly without frozen section-based histological control. The aim of the study was to evaluate the sampling efficacy of the existing tissue extraction models without guidance by frozen sections ("blind") and to develop an optimized model for tissue extraction. Five hundred thirty-three electronic maps of the tumor distribution in prostates from a single-center cohort of the patients subjected to radical prostatectomy were used for analysis. Six available models were evaluated in silico for their sampling efficacy. Additionally, a novel model achieving the best sampling efficacy was developed. The available models showed high efficacies for sampling "any part" from the tumor (up to 100%), but were uniformly low in efficacy to sample all tumor foci from the specimens (with the best technique sampling only 51.6% of the all tumor foci). The novel 4-level extraction model achieved a sampling efficacy of 93.1% for all tumor foci. The existing "blind" tissue extraction models from prostatectomy specimens without frozen sections control are suitable to target tumor tissues but these tissues do not represent the whole tumor. The novel 4-level model provides the highest sampling efficacy and a promising potential for integration into routine. Prostate 77: 396-405, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Disparities in Intratumoral Steroidogenesis

DTIC Science & Technology

2013-07-01

Tuesday shipment only) by overnight express for next day delivery on dry ice. Frozen specimens will be shipped on dry ice to the following address...Samples will be labeled with the study subject number and date of surgery. Frozen samples will be batch shipped (Monday and Tuesday shipment only) by... Morris MJ, de Bono JS, Ryan CJ, Denmeade SR, Smith MR, et al. Phase II multicenter study of abiraterone acetate plus prednisone therapy in patients

A rapid and efficient DNA extraction protocol from fresh and frozen human blood samples.

PubMed

Guha, Pokhraj; Das, Avishek; Dutta, Somit; Chaudhuri, Tapas Kumar

2018-01-01

Different methods available for extraction of human genomic DNA suffer from one or more drawbacks including low yield, compromised quality, cost, time consumption, use of toxic organic solvents, and many more. Herein, we aimed to develop a method to extract DNA from 500 μL of fresh or frozen human blood. Five hundred microliters of fresh and frozen human blood samples were used for standardization of the extraction procedure. Absorbance at 260 and 280 nm, respectively, (A 260 /A 280 ) were estimated to check the quality and quantity of the extracted DNA sample. Qualitative assessment of the extracted DNA was checked by Polymerase Chain reaction and double digestion of the DNA sample. Our protocol resulted in average yield of 22±2.97 μg and 20.5±3.97 μg from 500 μL of fresh and frozen blood, respectively, which were comparable to many reference protocols and kits. Besides yielding bulk amount of DNA, our protocol is rapid, economical, and avoids toxic organic solvents such as Phenol. Due to unaffected quality, the DNA is suitable for downstream applications. The protocol may also be useful for pursuing basic molecular researches in laboratories having limited funds. © 2017 Wiley Periodicals, Inc.

The effect of cryopreservation on goat semen characteristics related to sperm freezability.

PubMed

Dorado, J; Muñoz-Serrano, A; Hidalgo, M

2010-08-01

Seminal quality parameters were used to evaluate the effect of freeze-thawing procedure on goat sperm characteristics, and to relate possible changes in sperm parameters to cryopreservation success. Semen samples (n=110) were frozen with TRIS and milk-based extenders and thawed. Sperm quality parameters (motility, morphology and acrosome) were compared between fresh and frozen-thawed samples. Sperm freezability was judged by classifying the semen samples as "suitable" or "not suitable" according to the sperm quality parameters assessed before and after thawing. Fertility data was obtained after cervical insemination with frozen semen doses. The ejaculates were grouped into two categories according to their fertility results. In experiment 1, significant differences were found between semen extenders (P<0.001), bucks (P<0.05) and ejaculates within the same male (P<0.05) in terms of sperm quality. There was no seasonal effect (P>0.05) on the majority of the sperm parameters assessed after thawing. Moreover, significant differences (P<0.001) in semen parameters assessed in fresh semen and frozen-thawed samples were found between groups. The effect of the freeze-thawing procedure on sperm quality parameters was also different (P<0.05) between extenders within the same group. The number of sperm quality parameters that had changed after cryopreservation was lower in "suitable" semen samples before and after thawing. In experiment 2, no differences (P>0.05) in semen parameters assessed in fresh semen and frozen-thawed samples were found between groups. The effect of freezing and thawing on sperm quality parameters were different (P<0.05) between extenders within the same group. Only mean beat cross frequency (BCF) values were significantly higher (P<0.05) in TRIS diluted samples that led to successful pregnancies after artificial insemination. In conclusion, CASA-derived motility parameters, together with traditional semen assessment methods, give valuable information on sperm quality before and after freezing. Therefore, the identification of ejaculates as "good" or "bad" based on fresh and post-thaw semen parameters studied in the present experiment were good indicators of goat semen freezability, although the fertilizing capacity of frozen-thawed goat spermatozoa are not revealed by this quality study. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Feasibility of using the National Marine Mammal Tissue Bank for retrospective exploratory studies of perfluorinated alkyl acids.

PubMed

Lynch, Jennifer M; Ragland, Jared M; Reagen, William K; Wolf, Susan T; Malinsky, Michelle D; Ellisor, Michael B; Moors, Amanda J; Pugh, Rebecca S; Reiner, Jessica L

2018-05-15

Perfluorinated alkyl acids (PFAAs) have been used for 50+ years in materials such as stain-resistant treatments for paper and clothing, lubricants, and foam fire extinguishers. PFAAs are characterized by a fully fluorinated alkyl chain with a terminal acid group. Their long half-lives and ubiquitous environmental distribution create considerable concern for wildlife and human exposure. There is interest in examining temporal trends of PFAAs using the National Marine Mammal Tissue Bank (NMMTB), but NMMTB tissues are frozen and cryohomogenized in polytetrafluoroethylene (PTFE)-based materials. Because PTFE supplies may leach PFAAs into samples, this study mimicked collection, processing and storage steps of NMMTB samples and measured PFAA leaching to determine the feasibility of using this sample archive for PFAA temporal trends. We also explored concentrations in Atlantic white-sided dolphin (Lagenorhynchus acutus, WSDs) and rough-toothed dolphin (Steno bredanensis, RTDs) blubber (n=3 and 0) and liver (n=48 and 12, respectively). The materials used in NMMTB protocols may add up to 0.968ng/g perfluorooctanoic acid (PFOA), 0.090ng/g perfluorononanoic acid (PNFA), and 0.221ng/g perfluorooctane sulfonate (PFOS) to each archived sample. Leaching of PFNA and PFOS from supplies compared to dolphin levels was negligible, but PFOA contributions were substantially higher than levels found in most dolphin liver samples. Therefore, monitoring PFOA temporal trends from the NMMTB would require careful consideration. RTDs had significantly higher levels of PFOS and PFNA than WSDs. Both species have similar life history, trophic status, and foraging behaviors in deep pelagic waters, so differences could be from latitudinal variation in contamination. RTDs stranded in Florida; WSDs stranded farther north mostly in Massachusetts. Juveniles had significantly higher levels of PFOS and PFNA than adults in both species, suggesting growth dilution as they approach maturity. PFOS significantly decreased after 2001 in both species as expected based on changes in production. Published by Elsevier B.V.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sundaramurthi, Prakash; Patapoff, Thomas W.; Suryanarayanan, Raj

To study the crystallization of trehalose in frozen solutions and to understand the phase transitions during the entire freeze-drying cycle. Aqueous trehalose solution was cooled to -40 C in a custom-designed sample holder. The frozen solution was warmed to -18 C and annealed, and then dried in the sample chamber of the diffractometer. XRD patterns were continuously collected during cooling, annealing and drying. After cooling, hexagonal ice was the only crystalline phase observed. However, upon annealing, crystallization of trehalose dihydrate was evident. Seeding the frozen solution accelerated the solute crystallization. Thus, phase separation of the lyoprotectant was observed in frozenmore » solutions. During drying, dehydration of trehalose dihydrate yielded a substantially amorphous anhydrous trehalose. Crystallization of trehalose, as trehalose dihydrate, was observed in frozen solutions. The dehydration of the crystalline trehalose dihydrate to substantially amorphous anhydrate occurred during drying. Therefore, analyzing the final lyophile will not reveal crystallization of the lyoprotectant during freeze-drying. The lyoprotectant crystallization can only become evident by continuous monitoring of the system during the entire freeze-drying cycle. In light of the phase separation of trehalose in frozen solutions, its ability to serve as a lyoprotectant warrants further investigation.« less

Subchronic treatment with acai frozen pulp prevents the brain oxidative damage in rats with acute liver failure.

PubMed

de Souza Machado, Fernanda; Kuo, Jonnsin; Wohlenberg, Mariane Farias; da Rocha Frusciante, Marina; Freitas, Márcia; Oliveira, Alice S; Andrade, Rodrigo B; Wannmacher, Clovis M D; Dani, Caroline; Funchal, Claudia

2016-12-01

Acai has been used by the population due to its high nutritional value and its benefits to health, such as its antioxidant properties. The aim of this study was to evaluate the protective effect of acai frozen pulp on oxidative stress parameters in cerebral cortex, hippocampus and cerebellum of Wistar rats treated with carbon tetrachloride (CCl 4 ). Thirty male Wistar rats (90-day-old) were orally treated with water or acai frozen pulp for 14 days (7 μL/g). On the 15th day, half of the animals received treatment with mineral oil and the other half with CCl 4 (3.0 mL/kg). The cerebral cortex, hippocampus and cerebellum were dissected and used for analysis of creatine kinase activity (CK), thiobarbituric acid reactive substances (TBARS), carbonyl, sulfhydryl, and the activity of antioxidant enzymes catalase (CAT) and superoxide dismutase (SOD). Statistical analysis was performed by ANOVA followed by Tukey's post-test. CCl 4 was able to inhibit CK activity in all tissues tested and to provoke lipid damage in cerebral cortex and cerebellum, and protein damage in the three tissues tested. CCl 4 enhanced CAT activity in the cerebral cortex, and inhibited CAT activity in the hippocampus and cerebellum and reduced SOD activity in all tissues studied. Acai frozen pulp prevented the inhibition of CK, TBARS, carbonyl and CAT activity in all brain structures and only in hippocampus for SOD activity. Therefore, acai frozen pulp has antioxidant properties and maybe could be useful in the treatment of some diseases that affect the central nervous system that are associated with oxidative damage.

The effect of Arabic gum on frozen dough properties and the sensory assessments of the bread produced.

PubMed

Tavakoli, Hamid Reza; Jonaidi Jafari, Nematollah; Hamedi, Hassan

2017-04-01

The use of hydrocolloids in frozen dough has become frequent as bread improvers due to their anti-staling effect. Nevertheless, the impact of both different frozen storage and Arabic gum level in non-prefermented flat dough with following thawing procedure have not been studied. This work intended to study the effect of three different ratio of Arabic gum on rheological properties of 1, 7, and 30 days of frozen storage and the quality of the bread made from. In order to gain the least detrimental effects on gluten network, we used rapid rate freezing and microwave heating in thawing stage. Rheological results showed that the unfrozen samples to which Arabic gum had been added rendered the highest resistance to extension. The resistance of gum fortified samples were less than fresh dough, however the decline was not significant in 3.0% Arabic gum dough kept in a month storage (p > .05). The similar findings were obtained for extensibility and adhesiveness; in which the maximum incorporation of Arabic gum lessen the destructive impact of long freezing storage. Addition of 3% gum could be able to retard staling through an increment in hydrophilic bonds between water molecules and amylose during thawing (p < .05). The overall rating of Arabic gum enriched samples was similar with bread made from non-frozen dough, even after 30 days of storage as indicated by the sensory evaluation of breads. Producing a chapatti-like fermented bread without long fermentation period. Formulation a frozen dough without using chemical additives. Introducing a proper use of a new defrosting method with the aim of achieving a better texture. Improvement in retarding staling by the use of Gum Arabic after 7 days. © 2016 Wiley Periodicals, Inc.

Effect of different concentrations of egg yolk and virgin coconut oil in Tris-based extenders on chilled and frozen-thawed bull semen.

PubMed

Tarig, A A; Wahid, H; Rosnina, Y; Yimer, N; Goh, Y M; Baiee, F H; Khumran, A M; Salman, H; Ebrahimi, M

2017-07-01

The aim of this study was to evaluate the effects of 8% virgin coconut oil (VCO) combined with different percentages of egg yolk in Tris extender on the quality of chilled and frozen-thawed bull semen. A total of 24 ejaculates from four bulls were collected using an electroejaculator. Semen samples were diluted with 8% VCO in Tris extender which contained different concentrations 0% (control), 4%, 8%, 12%, 16% and 20% egg yolk. The diluted semen samples were divided into two fractions: one was chilled and stored at 4°C until evaluation after 24, 72, and 144h; the second fraction was processed by chilling for 3h at 4°C to equilibrate, then packaged in 0.25ml straws and frozen and stored in liquid nitrogen at -196°C until evaluation after 7 and 14 days. Both chilled and frozen semen samples were then thawed at 37°C and assessed for general motility using computer-assisted semen analysis (CASA), viability, acrosome integrity, and morphology (eosin-nigrosin), membrane integrity (hypo-osmotic swelling test) and lipid peroxidation (thiobarbituric acid-reactive substances (TBARS)). The results indicate treatments with 8%, 12%, 16% and 20% egg yolk with 8% VCO had greater sperm quality (P<0.05) as compared with the control. The treatment with 20% egg yolk had the greatest sperm quality (P<0.05) among the treated groups for both chilled and frozen-thawed semen. In conclusion, the use of 8% VCO combined with 20% egg yolk in a Tris-based extender enhanced the values for chilled and frozen-thawed quality variables of bull sperm. Copyright © 2017 Elsevier B.V. All rights reserved.

Immunoreactive LH in long-term frozen human urine samples.

PubMed

Singh, Gurmeet Kaur Surindar; Jimenez, Mark; Newman, Ron; Handelsman, David J

2014-04-01

Urine provides a convenient non-invasive alternative to blood sampling for measurement of certain hormones. Urinary luteinizing hormone (LH) measurements have been used for endocrinology research and anti-doping testing. However, the commercially available LH immunoassays are developed and validated for human blood samples but not urine so that LH assays intended for use with urine samples need thorough validation. Therefore, the present study evaluated the measurement of urinary LH immunoreactivity using previously validated immunofluorometric (IF) and immunochemiluminometric (ICL) LH assays after prolonged frozen storage. LH was measured in serial urine samples following administration of a single injection of one of two doses of recombinant human chorionic hormone (rhCG) with assays run at the end of study (2008) and again after four years of frozen (-20 °C) storage where samples were stored without adding preservatives. The ICL assay showed quantitatively reproducible LH measurements after prolonged -20 °C storage. However, the IF immunoassay gave consistently lower LH levels relative to ICL (2008) with a further proportionate reduction after four years of sample storage (2012). Yet, both the assays displayed similar patterns of the time-course of urine LH measurement both before and after four years of frozen storage. In conclusion, we found that both immunoassays are suitable for urinary LH measurements with ICL assay being more robust for quantitative urinary LH measurement such as for anti-doping purposes, whereas the IF could be applicable for research studies where urine LH levels are compared within-study but not in absolute terms. Copyright © 2013 John Wiley & Sons, Ltd.

Blood use in liver transplantation

PubMed Central

Lewis, J. H.; Bontempo, F. A.; Cornell, F.; Ki̋ss, J. E.; Larson, P.; Ragni, M. V.; Rice, E. O.; Spero, J. A.; Starzl, T. E.

2010-01-01

During the first 5 years (1981–1985) of the liver transplantation program in Pittsburgh, a total (preoperative, intraoperative, and postoperative) of 18,668 packed red cell units, 23,627 fresh-frozen plasma units, 20,590 platelet units, and 4241 cryoprecipitate units was transfused for the procedures. This represents 3 to 9 percent of the total of blood products supplied by the Central Blood Bank to its 32 member hospitals. Six hundred thirty-six (636) transplants were performed on 485 patients in two hospitals: the Presbyterian University Hospital (564 beds) and Children’s Hospital of Pittsburgh (236 beds). All of the blood components used in the operations were procured and released by the Central Blood Bank. This report describes some of these findings. PMID:3296340

Is sperm cryopreservation at -150 degree C a feasible alternative?

PubMed

Medrano, A; Cabrera, F; González, F; Batista, M; Gracia, A

2002-01-01

A series of experiments was carried out to validate a -150 degree C ultra-low temperature freezer for its possible use to properly freeze and store semen. In the first part, crude sample handling was simulated to see whether temperature of stored samples was maintained within a safe range; also, the freezing point and latent heat of fusion plateau of a semen extender were monitored. In the second part, buck semen was (i) frozen in liquid nitrogen and stored in the ultra-low freezer, (ii) frozen and stored in the ultra-low freezer, and (iii) frozen and stored in liquid nitrogen, to compare sperm cryosurvival between freezing methods. Both, frequent removal of samples and long opening of the freezer door did not negatively affect stored sample temperature; latent heat of fusion plateau was 5 minutes long. Semen stored either at -150 degree C or at -196 degree C cryosurvived similarly after 2 days and after 2 months of cryopreservation.

Ufmylation and FATylation Pathways are Down Regulated in Human Alcoholic and Non Alcoholic Steatohepatitis, and Mice Fed DDC, where Mallory-Denk Bodies (MDBs) Form

PubMed Central

Liu, H; Li, J; Tillman, B; French, BA; French, SW

2014-01-01

We previously reported the mechanisms involved in the formation of Mallory-Denk bodies (MDBs) in mice fed DDC. To further provide clinical evidence as to how ubiquitin-like protein (Ubls) modification, gene transcript expression in Ufmylation and FATylation were investigated in human archived formalin-fixed, paraffin-embedded (FFPE) liver biopsies and frozen liver sections from DDC re-fed mice were used. Real-time PCR analysis showed that all Ufmylation molecules (Ufm1, Uba5, Ufc1, Ufl1 and UfSPs) were significantly down regulated, both in DDC re-fed mice livers and patients’ livers where MDBs had formed, indicating that gene transcript changes were limited to MDB-forming livers where the protein quality control system was down regulated. FAT10 and subunits of the immunoproteasome (LMP2 and LMP7) were both up regulated as previously shown. An approximate 176- and 5-fold up regulation (respectively) of FAT10 were observed in the DDC re-fed mice liver and in the livers of human alcoholic hepatitis with MDBs present, implying that there was an important role played by this gene. The FAT10-specific E1 and E2 enzymes Uba6 and USE1, however, were found to be down regulated both in patients’ livers and in the liver of DDC re-fed mice. Interestedly, the down regulation of mRNA levels was proportionate to MDB abundance in the liver tissues. Our results show the first systematic demonstration of transcript regulation of Ufmylation and FATylation in the liver of patients who form MDBs, where protein quality control is down regulated. This was also shown in livers of DDC re-fed mice where MDBs had formed. PMID:24893112

Ufmylation and FATylation pathways are downregulated in human alcoholic and nonalcoholic steatohepatitis, and mice fed DDC, where Mallory-Denk bodies (MDBs) form.

PubMed

Liu, H; Li, J; Tillman, B; French, B A; French, S W

2014-08-01

We previously reported the mechanisms involved in the formation of Mallory-Denk bodies (MDBs) in mice fed DDC. To further provide clinical evidence as to how ubiquitin-like protein (Ubls) modification, gene transcript expression in Ufmylation and FATylation were investigated in human archived formalin-fixed, paraffin-embedded (FFPE) liver biopsies and frozen liver sections from DDC re-fed mice were used. Real-time PCR analysis showed that all Ufmylation molecules (Ufm1, Uba5, Ufc1, Ufl1 and UfSPs) were significantly downregulated, both in DDC re-fed mice livers and patients' livers where MDBs had formed, indicating that gene transcript changes were limited to MDB-forming livers where the protein quality control system was downregulated. FAT10 and subunits of the immunoproteasome (LMP2 and LMP7) were both upregulated as previously shown. An approximate 176- and 5-fold upregulation (respectively) of FAT10 was observed in the DDC re-fed mice liver and in the livers of human alcoholic hepatitis with MDBs present, implying that there was an important role played by this gene. The FAT10-specific E1 and E2 enzymes Uba6 and USE1, however, were found to be downregulated both in patients' livers and in the liver of DDC re-fed mice. Interestedly, the downregulation of mRNA levels was proportionate to MDB abundance in the liver tissues. Our results show the first systematic demonstration of transcript regulation of Ufmylation and FATylation in the liver of patients who form MDBs, where protein quality control is downregulated. This was also shown in the livers of DDC re-fed mice where MDBs had formed. Copyright © 2014 Elsevier Inc. All rights reserved.

A Molecular Framework for Understanding DCIS

DTIC Science & Technology

2016-10-01

frozen patient biopsies, these have been annotated by our pathologist and prepared to be taken on for sequencing. The tissue includes DCIS, IDC...stroma adjacent to DCIS/IDC and normal tissue . We have initiated the RNA sequencing from these samples and also the DNA sequencing 15. SUBJECT TERMS DCIS...before they reach 55. Utilizing a unique bank of frozen mammary biopsies, containing samples with DCIS alone, and a combination of DCIS and IDC, we aim

Scanning electron microscopy of hepatic ultrastructure: secondary, backscattered, and transmitted electron imaging.

PubMed

Miyai, K; Abraham, J L; Linthicum, D S; Wagner, R M

1976-10-01

Several methods of tissue preparation and different modes of operation of the scanning electron microscope were used to study the ultrastructure of rat liver. Rat livers were perfusion fixed with buffered 2 per cent paraformaldehyde or a mixture of 1.5 per cent paraformaldehyde and 1 per cent glutaraldehyde and processed as follows. Tissue blocks were postfixed in buffered 2 per cent osmium tetroxide followed sequentially by the ligand-mediated osmium binding technique, dehydration and cryofracture in ethanol, and critical point drying. They were then examined without metal coating in the scanning electron microscope operating in the secondary electron and backscattered electron modes. Fifty-micrometer sections were cut with a tissue sectioner, stained with lead citrate, postfixed with osmium, dehydrated, critical point dried, and examined in the secondary electron and back-scattered electron modes. Frozen sections (0.25 to 0.75 mum. thick) were cut by the method of Tokuyasu (Toluyasu KT: J Cell Biol 57:551, 1973) and their scanning transmission electron microscope images were examined either with a scanning transmission electron microscope detector or with a conversion stub using the secondary electron detector. Secondary electron images of the liver prepared by ligand-mediated osmium binding and subsequent cryofracture revealed such intracellular structures as cisternae of the endoplasmic reticulum, lysosomes, mitochondria, lipid droplets, nucleolus and nuclear chromatin, as well as the usual surface morphology, Lipocytes in the perisinusoidal space were readily identified. Backscattered electron images. Unembedded frozen sections had little drying artifact and were virtually free of freezing damage. The scanning transmission electron microscope image revealed those organelles visualized by the secondary electron mode in the ligand-mediated osmium binding-treated tissue.

Effect of osmotic dehydration pretreatment and glassy state storage on the quality attributes of frozen mangoes under long-term storage.

PubMed

Zhao, Jin-Hong; Xiao, Hong-Wei; Ding, Yang; Nie, Ying; Zhang, Yu; Zhu, Zhen; Tang, Xuan-Ming

2017-05-01

Changes in the quality of frozen mango cuboids were investigated during long-term glassy state storage with and without osmotic dehydration pretreatment. The mango cuboids were dehydrated in mixed solutions (sucrose: glucose: fructose in a ratio of 3.6:1:3) of different concentrations (30, 40, and 50% (wt/wt)) prior to freezing and then stored at -55 °C (in the glassy state) for 6 months. The results revealed that compared with the untreated samples, osmotic pretreatment decreased total color difference (reduced by 15.6-62.3%), drip loss (reduced by 8.2-29.5%) and titration acidity (reduced by 1.3-9.4%), while increasing hardness (increased by 48.8-82.3%), vitamin C content (increased by 72.5-120.6%) and total soluble solids (increased by 21.8-53.7%) of frozen mangoes after 6 months. Dehydration with a sugar concentration of 40% was considered as the optimal pretreatment condition. In addition, a storage temperature of -55 °C provided better retention of quality than rubbery state storage at -18 °C. With prolonged storage time, the quality of frozen mangoes continued to change, even in the glassy state. However, the changes in quality of the osmotic-dehydrated samples were less than those of the untreated samples. The current work indicates that osmotic pretreatment and glassy state storage significantly improved the quality of frozen mangoes during long-term storage.

Mining the archives: a cross-platform analysis of gene ...

EPA Pesticide Factsheets

Formalin-fixed paraffin-embedded (FFPE) tissue samples represent a potentially invaluable resource for genomic research into the molecular basis of disease. However, use of FFPE samples in gene expression studies has been limited by technical challenges resulting from degradation of nucleic acids. Here we evaluated gene expression profiles derived from fresh-frozen (FRO) and FFPE mouse liver tissues using two DNA microarray protocols and two whole transcriptome sequencing (RNA-seq) library preparation methodologies. The ribo-depletion protocol outperformed the other three methods by having the highest correlations of differentially expressed genes (DEGs) and best overlap of pathways between FRO and FFPE groups. We next tested the effect of sample time in formalin (18 hours or 3 weeks) on gene expression profiles. Hierarchical clustering of the datasets indicated that test article treatment, and not preservation method, was the main driver of gene expression profiles. Meta- and pathway analyses indicated that biological responses were generally consistent for 18-hour and 3-week FFPE samples compared to FRO samples. However, clear erosion of signal intensity with time in formalin was evident, and DEG numbers differed by platform and preservation method. Lastly, we investigated the effect of age in FFPE block on genomic profiles. RNA-seq analysis of 8-, 19-, and 26-year-old control blocks using the ribo-depletion protocol resulted in comparable quality metrics, inc

Environmental Factors Related to a Semiarid Climate Influence the Freezability of Sperm from Collared Peccaries (Pecari tajacu Linnaeus, 1758).

PubMed

Maia, Keilla M; Souza, Ana L P; Praxedes, Erica C G; Bezerra, Luana G P; Silva, Andreia M; Campos, Livia B; Moreira, Samara S J; Apolinário, Carlos A C; Souza, João B F; Silva, Alexandre R

2018-04-30

The influence of environmental factors in a semiarid climate on characteristics of fresh and frozen/thawed sperm collected from collared peccaries (Pecari tajacu) was assessed. Semen from 11 male collared peccaries was collected by electroejaculation during the peaks of the dry and rainy periods while rainfall indices, air temperatures, relative humidity levels, and wind speeds were measured. The number, motility, morphology, osmotic response, and membrane integrity of sperm in the collected ejaculates were assessed. Samples were then frozen in liquid nitrogen, thawed, and reassessed. The rainfall index of the rainy period (73.2 mm) was significantly higher than that of the dry period (13.6 mm) and the relative humidity was significantly higher during the rainy period (74.6%) than it was during the dry period (66.8%). Air temperature and wind speed did not differ between the two periods. Characteristics of sperm in the fresh samples were not affected by environmental parameters. In contrast, computerized analysis revealed that sperm in samples frozen during the rainy period exhibited better post-thaw membrane integrity (28.6 ± 6%), motility (29.5 ± 7.7%), and rapid sperm population (13.7 ± 6.2%) than did sperm in samples frozen during the dry period (23.4 ± 3% membrane integrity, 14.6 ± 4.1% motility, and 4.1 ± 1.2% rapid sperm; p < 0.05). Other characteristics of the frozen/thawed sperm did not differ depending on the period in which they were collected. We demonstrated that environmental parameters did not affect the quality of fresh sperm, but could influence the freezability of sperm collected from collared peccaries raised under a semiarid climate.

Conventional freezing plus high pressure-low temperature treatment: Physical properties, microbial quality and storage stability of beef meat.

PubMed

Fernández, Pedro P; Sanz, Pedro D; Molina-García, Antonio D; Otero, Laura; Guignon, Bérengère; Vaudagna, Sergio R

2007-12-01

Meat high-hydrostatic pressure treatment causes severe decolouration, preventing its commercialisation due to consumer rejection. Novel procedures involving product freezing plus low-temperature pressure processing are here investigated. Room temperature (20°C) pressurisation (650MPa/10min) and air blast freezing (-30°C) are compared to air blast freezing plus high pressure at subzero temperature (-35°C) in terms of drip loss, expressible moisture, shear force, colour, microbial quality and storage stability of fresh and salt-added beef samples (Longissimus dorsi muscle). The latter treatment induced solid water transitions among ice phases. Fresh beef high pressure treatment (650MPa/20°C/10min) increased significantly expressible moisture while it decreased in pressurised (650MPa/-35°C/10min) frozen beef. Salt addition reduced high pressure-induced water loss. Treatments studied did not change fresh or salt-added samples shear force. Frozen beef pressurised at low temperature showed L, a and b values after thawing close to fresh samples. However, these samples in frozen state, presented chromatic parameters similar to unfrozen beef pressurised at room temperature. Apparently, freezing protects meat against pressure colour deterioration, fresh colour being recovered after thawing. High pressure processing (20°C or -35°C) was very effective reducing aerobic total (2-log(10) cycles) and lactic acid bacteria counts (2.4-log(10) cycles), in fresh and salt-added samples. Frozen+pressurised beef stored at -18°C during 45 days recovered its original colour after thawing, similarly to just-treated samples while their counts remain below detection limits during storage.

Microgravity

NASA Image and Video Library

1996-03-24

Astronaut Michael Clifford places a liquid nitrogen Dewar containing frozen protein solutions aboard Russia's space station Mir during a visit by the Space Shuttle (STS-76). The protein samples were flash-frozen on Earth and will be allowed to thaw and crystallize in the microgravity environment on Mir Space Station. A later crew will return the Dewar to Earth for sample analysis. Dr. Alexander McPherson of the University of California at Riverside is the principal investigator. Photo credit: NASA/Johnson Space Center.

Microgravity

NASA Image and Video Library

1996-09-20

Astronaut Tom Akers places a liquid nitrogen Dewar containing frozen protein solutions aboard Russia's space Station Mir during a visit by the Space Shuttle (STS-79). The protein samples were flash-frozen on Earth and will be allowed to thaw and crystallize in the microgravity environment on Mir Space Station. A later crew will return the Dewar to Earth for sample analysis. Dr. Alexander McPherson of the University of California at Riverside is the principal investigator. Photo credit: NASA/Johnson Space Center.

A cryopreservation method for Pasteurella multocida from wetland samples

USGS Publications Warehouse

Moore, Melody K.; Shadduck, D.J.; Goldberg, Diana R.; Samuel, M.D.

1998-01-01

A cryopreservation method and improved isolation techniques for detection of Pasteurella multocida from wetland samples were developed. Wetland water samples were collected in the field, diluted in dimethyl sulfoxide (DMSO, final concentration 10%), and frozen at -180 C in a liquid nitrogen vapor shipper. Frozen samples were transported to the laboratory where they were subsequently thawed and processed in Pasteurella multocida selective broth (PMSB) to isolate P. multocida. This method allowed for consistent isolation of 2 to 18 organisms/ml from water seeded with known concentrations of P. multocida. The method compared favorably with the standard mouse inoculation method and allowed for preservation of the samples until they could be processed in the laboratory.

Mechanics of fresh, frozen-thawed and heated porcine liver tissue.

PubMed

Wex, Cora; Stoll, Anke; Fröhlich, Marlen; Arndt, Susann; Lippert, Hans

2014-06-01

For a better understanding of the effects of thermally altered soft tissue, the biothermomechanics of these tissues need to be studied. Without the knowledge of the underlying physical processes and the parameters that can be controlled clinically, thermal treatment of cancerous hepatic tissue or the preservation of liver grafts are based primarily on trial and error. Thus, this study is concerned with the investigation of the influence of temperature on the rheological properties and the histological properties of porcine liver. Heating previously cooled porcine liver tissue above 40 °C leads to significant, irreversible stiffness changes observed in the amplitude sweep. The increase of the complex shear module of healthy porcine liver from room temperature to 70 °C is approximately 9-fold. Comparing the temperatures -20 °C and 20 °C, no significant difference of the mechanical properties was observed. Furthermore, there is a strong relation between the mechanical and histological properties of the porcine liver. Temperatures above 40 °C destroy the collagen matrix within the liver tissue. This results in the alteration of the biomechanical properties. The time-temperature superposition principle is applied to generate temperature-dependent shift factors that can be described by a two-part exponential function model with an inflection temperature of 45 °C. Tumor ablation techniques such as heating or freezing have a significant influence on the histology of liver tissue. However, only for temperatures above body temperature an influence on the mechanical properties of hepatic tissues was noticeable. Freezing up to -20 °C did not affect the liver mechanics.

Effect of sample handling on thiamine and thiaminolytic activity in alewife

USGS Publications Warehouse

Wright, G.M.; Brown, S.B.; Brown, L.R.; Moore, K.; Villella, M.; Zajicek, J.L.; Tillitt, D.E.; Fitzsimons, J.D.; Honeyfield, D.C.

2005-01-01

Alewives Alosa pseudoharengus were collected to evaluate handling and processing conditions that may affect the measurement of their thiamine-thiaminase content. Fish were captured by otter trawl, and reference samples of live fish were quick-frozen on dry ice immediately following capture. Other samples were placed on wet ice (4??C) or held in ambient lake water (21.5??C) for periods of up to 5 h before freezing. Total thiamine levels for reference samples averaged 26 nmol/g and consisted of 66, 15, and 19% thiamine pyrophosphate (TPP), thiamine monophosphate (TMP), and unphosphorylated thiamine (Th), respectively. After 120 min at either 4??C or 21.5??C, total thiamine concentrations were lower. At 21.5??C, the TPP proportion had decreased by 30 min and the proportion as Th increased after 60 min. In the groups sampled after 5 h, total thiamine concentrations were not altered but the proportion of TPP was lower and that of Th was higher than in reference samples. The stability of thiamine in thawed muscle samples from previously frozen alewives was poor (40% loss by 1 h at 22??C and 30% loss by 2 h at 4??C). Thiaminase activity averaged 5,975 pmol??g wet weight -1??min-1 in reference samples. In fresh-caught alewives, thiaminase activities were remarkably consistent throughout the sampling period. At 4??C, thiaminase activity in muscle tissue from previously frozen alewives was stable for the entire investigation period. At 25??C, the activity initially increased by 40% after 60 min but then decreased to 50% of initial value after 5 h. We conclude that sampling times greater than 25 min could cause some changes in the various thiamine forms and net loss in total thiamine. The thiamine content in previously frozen alewife samples is highly labile, requiring low temperatures during processing for analysis. ?? Copyright by the American Fisheries Society 2005.

Evaluation of motility, membrane status and DNA integrity of frozen-thawed bottlenose dolphin (Tursiops truncatus) spermatozoa after sex-sorting and recryopreservation.

PubMed

Montano, G A; Kraemer, D C; Love, C C; Robeck, T R; O'Brien, J K

2012-06-01

Artificial insemination (AI) with sex-sorted frozen-thawed spermatozoa has led to enhanced management of ex situ bottlenose dolphin populations. Extended distance of animals from the sorting facility can be overcome by the use of frozen-thawed, sorted and recryopreserved spermatozoa. Although one bottlenose dolphin calf had been born using sexed frozen-thawed spermatozoa derived from frozen semen, a critical evaluation of in vitro sperm quality is needed to justify the routine use of such samples in AI programs. Sperm motility parameters and plasma membrane integrity were influenced by stage of the sex-sorting process, sperm type (non-sorted and sorted) and freezing method (straw and directional) (P<0.05). After recryopreservation, sorted spermatozoa frozen with the directional freezing method maintained higher (P<0.05) motility parameters over a 24-h incubation period compared to spermatozoa frozen using straws. Quality of sperm DNA of non-sorted spermatozoa, as assessed by the sperm chromatin structure assay (SCSA), was high and remained unchanged throughout freeze-thawing and incubation processes. Though a possible interaction between Hoechst 33342 and the SCSA-derived acridine orange was observed in stained and sorted samples, the proportion of sex-sorted, recryopreserved spermatozoa exhibiting denatured DNA was low (6.6±4.1%) at 6 h after the second thawing step and remained unchanged (P>0.05) at 24 h. The viability of sorted spermatozoa was higher (P<0.05) than that of non-sorted spermatozoa across all time points after recryopreservation. Collective results indicate that bottlenose dolphin spermatozoa undergoing cryopreservation, sorting and recryopreservation are of adequate quality for use in AI.

Frozen section evaluation via dynamic real-time non-robotic Telepathology system in a university Cancer center by resident / faculty cooperation team.

PubMed

Vosoughi, Aram; Smith, Paul Taylor; Zeitouni, Joseph A; Sodeman, Gregori M; Jorda, Merce; Gomez-Fernandez, Carmen; Garcia-Buitrago, Monica; Petito, Carol K; Chapman, Jennifer R; Campuzano-Zuluaga, German; Rosenberg, Andrew E; Kryvenko, Oleksandr N

2018-04-30

Frozen section telepathology interpretation experience has been largely limited to practices with locations significantly distant from one another with sporadic need for frozen section diagnosis. In 2010 we established a real-time non-robotic telepathology system in a very active cancer center for daily frozen section service. Herein, we evaluate its accuracy compared to direct microscopic interpretation performed in the main hospital by the same faculty and its cost-efficiency over a 1-year period. From 643 (1416 parts) cases requiring intraoperative consultation, 333 cases (690 parts) were examined by telepathology and 310 cases (726 parts) by direct microscopy. Corresponding discrepancy rates were 2.6% (18 cases: 6 (0.9%) sampling and 12 (1.7%) diagnostic errors) and 3.2% (23 cases: 8 (1.1%) sampling and 15 (2.1%) diagnostic errors), P=.63. The sensitivity and specificity of intraoperative frozen diagnosis were 0.92 and 0.99, respectively, in telepathology, and 0.90 and 0.99, respectively, in direct microscopy. There was no correlation of error incidence with post graduate year level of residents involved in the telepathology service. Cost analysis indicated that the time saved by telepathology was $19691 over one year of the study period while the capital cost for establishing the system was $8924. Thus, real-time non-robotic telepathology is a reliable and easy to use tool for frozen section evaluation in busy clinical settings, especially when frozen section service involves more than one hospital, and it is cost efficient when travel is a component of the service. Copyright © 2018. Published by Elsevier Inc.

The effect of refrigerated and frozen storage on butter flavor and texture.

PubMed

Krause, A J; Miracle, R E; Sanders, T H; Dean, L L; Drake, M A

2008-02-01

Butter is often stored for extended periods of time; therefore, it is important for manufacturers to know the refrigerated and frozen shelf life. The objectives of this study were to characterize the effect of refrigerated and frozen storage on the sensory and physical characteristics of butter. Fresh butter was obtained on 2 occasions from 2 facilities in 113-g sticks and 4-kg bulk blocks (2 facilities, 2 package forms). Butters were placed into both frozen (-20 degrees C) and refrigerated storage (5 degrees C). Frozen butters were sampled after 0, 6, 12, 15, and 24 mo; refrigerated butters were sampled after 0, 3, 6, 9, 12, 15, and 18 mo. Every 3 mo, oxidative stability index (OSI) and descriptive sensory analysis (texture, flavor, and color) were conducted. Every 6 mo, peroxide value (PV), free fatty acid value (FFV), fatty acid profiling, vane, instrumental color, and oil turbidity were examined. A mixed-model ANOVA was conducted to characterize the effects of storage time, temperature, and package type. Storage time, temperature, and package type affected butter flavor, OSI, PV, and FFV. Refrigerated butter quarters exhibited refrigerator/stale off-flavors concurrent with increased levels of oxidation (lower oxidative stability and higher PV and FFV) within 6 mo of refrigerated storage, and similar trends were observed for refrigerated bulk butter after 9 mo. Off-flavors were not evident in frozen butters until 12 or 18 mo for quarters and bulk butters, respectively. Off-flavors in frozen butters were not correlated with instrumental oxidation measurements. Because butter is such a desirable fat source in terms of flavor and textural properties, it is important that manufacturers understand how long their product can be stored before negative attributes develop.

Presentation of frozen shoulder among diabetic and non-diabetic patients.

PubMed

Uddin, Mohammad Moin; Khan, Aminuddin A; Haig, Andrew J; Uddin, Mohammad Kafil

2014-12-01

The literature is inconsistent regarding the level of pain and disability in frozen shoulder patients with or without diabetes mellitus. The aim of this study is to evaluate some demographic features of frozen shoulder patients and to look into the disparity of information by comparing the level of pain and disability due to frozen shoulder between diabetic and non-diabetic people. This is a prospective comparative study. People with frozen shoulder attending an outpatient department were selected by consecutive sampling. Disability levels were assessed by the Shoulder Pain & Disability Index (SPADI). Means of pain and disability scores were compared using unpaired t-test. Among 140 persons with shoulder pain 99 (71.4%) had frozen shoulder. From the participating 40 frozen shoulder patients, 26 (65%) were males and 14 (35%) were females. Seventeen participants (42.5%) were diabetic, two (5%) had impaired glucose tolerance and 21 (52.5%) patients were non-diabetic. Mean disability scores (SPADI) were 51 ± 15.5 in diabetic and 57 ± 16 in non-diabetic persons. The differences in pain and disability level were not statistically significance (respectively, p = 0.24 and p = 0.13 at 95% confidence interval). No difference was found in level of pain and disability level between frozen shoulder patients with and without diabetes.

High-resolution scanning electron microscopy of frozen-hydrated cells.

PubMed

Walther, P; Chen, Y; Pech, L L; Pawley, J B

1992-11-01

Cryo-fixed yeast Paramecia and sea urchin embryos were investigated with an in-lens type field-emission SEM using a cold stage. The goal was to further develop and investigate the processing of frozen samples for the low-temperature scanning electron microscope (LTSEM). Uncoated frozen-hydrated samples were imaged with the low-voltage backscattered electron signal (BSE). Resolution and contrast were sufficient to visualize cross-fractured membranes, nuclear pores and small vesicles in the cytoplasm. It is assumed that the resolution of this approach is limited by the extraction depth of the BSE which depends upon the accelerating voltage of the primary beam (V0). In this study, the lowest possible V0 was 2.6 kV because below this value the sensitivity of the BSE detector is insufficient. It is concluded that the resolution of the uncoated specimen could be improved if equipment were available for high-resolution BSE imaging at 0.5-2 kV. Higher resolution was obtained with platinum cryo-coated samples, on which intramembranous particles were easily imaged. These images even show the ring-like appearance of the hexagonally arranged intramembranous particles known from high-resolution replica studies. On fully hydrated samples at high magnification, the observation time for a particular area is limited by mass loss caused by electron irradiation. Other potential sources of artefacts are the deposition of water vapour contamination and shrinkage caused by the sublimation of ice. Imaging of partially dehydrated (partially freeze-dried) samples, e.g. high-pressure frozen Paramecium and sea urchin embryos, will probably become the main application in cell biology. In spite of possible shrinkage problems, this approach has a number of advantages compared with any other electron microscopy preparation method: no chemical fixation is necessary, eliminating this source of artefacts; due to partial removal of the water additional structures in the cytoplasm can be investigated; and finally, the mass loss due to electron beam irradiation is greatly reduced compared to fully frozen-hydrated specimens.

Short communication: The effects of frozen storage on the survival of probiotic microorganisms found in traditionally and commercially manufactured kefir.

PubMed

O'Brien, K V; Aryana, K J; Prinyawiwatkul, W; Ordonez, K M Carabante; Boeneke, C A

2016-09-01

Kefir is a fermented milk traditionally made from a unique starter culture, which consists of numerous bacteria and yeast species bound together in an exopolysaccharide matrix produced by certain lactic acid bacteria. Many health benefits are associated with traditionally produced kefir; however, bulging and leaking packaging, caused by secondary yeast fermentation during storage, has limited large-scale manufacture. Commercial kefir products have been designed to reduce these effects by using a pure starter culture consisting of a mixture of bacteria and yeast species that give a flavor similar to traditional kefir, but some health benefits may be lost in commercial production due to reduced microbial diversity and lack of beneficial exopolysaccharides. In this study, traditional and commercial kefir was frozen to study the effects of frozen storage on the viability of probiotic bacteria over time. Traditional kefir was prepared by inoculating 1L of pasteurized whole goat milk with approximately 30g of kefir grains. Commercial kefir was prepared by inoculating 1L of full-fat, pasteurized goat milk with a commercial kefir starter. The milk was allowed to ferment at room temperature (24-28°C) until pH 4.6 was reached. Samples were frozen (-8 to -14°C) immediately following the completion of fermentation and were thawed and plated for lactobacilli, lactococci, and yeasts on d 0, 7, 14, and 30 of frozen storage. Lactobacilli, lactococci, and yeasts were significantly reduced in number during frozen storage; however, the traditionally produced kefir was shown to have significantly higher counts of bacteria and yeast at each sampling. We concluded that frozen storage and the development of frozen kefir products could eliminate most packaging concerns associated with the large-scale manufacture of traditionally produced kefir, resulting in increased production and marketability of this healthful product. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Impact of Frozen Storage on the Anthocyanin and Polyphenol Content of American Elderberry Fruit Juice

PubMed Central

Johnson, Mitch C.; Thomas, Andrew L.; Greenlief, C. Michael

2015-01-01

The effects of frozen storage on the anthocyanin and polyphenol content of elderberry fruit juice are investigated. Juice from three genotypes of American elderberry (Adams II, Bob Gordon, and Wyldewood) was screened for total phenolic (TP) and total monomeric anthocyanin (TMA) content with spectrophotometric methods. The individual anthocyanin content (IAC) of the juice was tested by coupling solid phase extraction with ultra-performance liquid chromatography/tandem mass spectrometry. Juice samples were tested initially upon harvest, then again after 3, 6, and 9 months of frozen storage. Juice from the three different genotypes had significantly different TP, TMA, and IAC profiles initially (p<0.05). The TP,, TMA, and IAC content of the juice from different genotypes were significantly affected (p<0.05) by the frozen storage time, suggesting that both genotype and length of frozen storage time can affect the anthocyanin content of elderberry fruit juice. PMID:26028422

Effects of ionizing radiation on sensorial, chemical, and microbiological quality of frozen corn and peas.

PubMed

Fan, Xuetong; Sokorai, Kimberly J B

2007-08-01

The effects of irradiation (0, 1.8, and 4.5 kGy) on the quality of frozen corn and peas were investigated during a 12month period of postirradiation storage at -18 degrees C. Irradiation of frozen corn and peas caused a reduction in ascorbic acid content of both vegetables and a loss of texture in peas but had no significant effects on instrumental color parameters (L*, a*, and b*), carotenoid and chlorophyll content, or antioxidant capacity of corn and peas. Irradiation reduced microbial loads of frozen peas and increased display life at 23 degrees C of thawed peas by preserving the green color, apparently because of slower increases in the population of acid-producing microorganisms in the irradiated samples. Overall, irradiation significantly reduced the microbial load and increased the display life of peas and had minimal detrimental effects on the quality of frozen corn and peas.

[Establishment of the experimental animal model of Babesia microti].

PubMed

Lu, Yan; Cai, Yu-Chun; Chen, Shao-Hong; Chen, Jia-Xu; Guo, Jian; Chen, Mu-Xin; Ai, Lin; Chu, Yan-Hong; Chen, Zhuo; Zhou, Xiao-Nong

2012-12-30

To establish the experimental animal model for the study of Babesia microti. BALB/c mice, immunosuppressive BALB/c mice, SCID mice and NOD-SCID mice were inoculated with B. microti-infected red blood cells (RBC) by intraperitoneal injection respectively. After inoculation, thin blood smears were prepared every day, stained with Giemsa staining and examined for the presence of parasitemia. Three mice were dissected to examine the infectivity in bone marrow, brain, spleen, heart, lung, kidney and liver tissues. The infection rate of erythrocytes in different tissues was recorded, and the relationship between the infectivity of tissues and infection rate in peripheral blood was analyzed. Blood samples infected with B. microti were preserved in liquid nitrogen with dimethyl sulfoxide (DMSO) for 2 months. The thawed parasitized blood was injected into the BALB/c mice by same route and the parasitemia was monitored. The four kinds of mice were all infected by B. microti with parasitemia. The percentage of parasitized red blood cells from peripheral blood were 82.4% (BALB/c mice, d7), 73.2% (immunosuppressive BALB/c mice, d5), 86.4% (SCID mice, d8) and 72.5% (NOD-SCID mice, d8) at the maximum, respectively. Parasitemia decreased rapidly in BALB/c mice, whereas decreased slowly in immunosuppressive BALB/c mice. Only the parasitemia in SCID mice and NOD-SCID mice decreased significantly and tended to picking up again. The parasites were observed in RBCs from bone marrow, brain, spleen, heart, lung, kidney and liver tissues. The infection rate of erythrocytes in tissues increased with an increase of infection in peripheral blood. After cryopreservation, the parasites proliferated in BALB/c mice. Parasitemia appeared after inoculation with frozen infected blood two days later than that of fresh infected blood. The infection rate reached its peak after inoculation with frozen infected blood one day later than that of fresh infected blood. The experimental animal model of B. microti has been established. The infection rate of erythrocytes is related to the immune status of the host mice.

Effects of type of freeze straw and thaw temperature on the fertilizing capacity of frozen chicken semen.

PubMed

Duplaix, M; Sexton, T J

1984-04-01

Two experiments were conducted to determine the relationship between thawing temperature and the type of straw in which chicken semen was frozen. In Experiment 1, semen was frozen in three different types of plastic straws: US (.5-ml capacity), French (.5-ml capacity), and French mini (.25-ml capacity). Experiment 1 was divided into two trials to compare semen packaged in the different straws and thawed at 15 (Trial 1) or .5 C (Trial 2). Although there were distinguishable features of the freeze and thaw curves between samples frozen in the different straws, the type of freeze straw had no effect on the fertilizing capacity of frozen semen when thaw temperature was held constant: fertility, Days 2 to 4 after artificial insemination, ranged from 16 to 27% for Trial 1 and 45 to 47% for Trial 2. In Experiment 2, semen was frozen in US straws and thawed at either .5 or 15 C to assess the effect of the thaw temperature. Fertility of frozen semen, Days 2 to 4 after artificial insemination, was significantly higher when semen was thawed at .5 than at 15 C (62 vs. 20%).

Effect of Chitin Isolated from Crustaceans and Cephalopods on Denaturation of Fish Myofibrillar Protein and the State of Water during Frozen Storage

NASA Astrophysics Data System (ADS)

Yamashita, Yasumitsu; Zhang, Nong; Nozaki, Yukinori

The effect of chitin hydrolysate made from shell of crustaceans and cartilage of cephalopods on the denaturation and the state of water of lizard fish myofibrillar protein (Mf) during frozen storage at -250°C for 120 days were investigated. 5.0% chitin hydrolysate (dried matter) added Mf samples were frozen at -25°C and storage for 120 days, and the change of inactivation of Mf Ca-ATPase and the change of unfrozen water in Mf were examined during the frozen storage. The decrease of Mf Ca-ATPase during frozen storage were slow by addition of chitin hydrolysate. The amount of unfrozen water in Mf were increased by addition of chitin hydrolysate, and decreaced gradually during frozen storage. On the other hand, the amount of unfrozen water of the control were decreaced suddenly during frozen storage. Above results suggest that chitin hydrolysate has cryoprotective effect on Mf. It is guessed that the mechanism of cryoprotective effect of chitin hydrolysate may be caused by an interaction between hydration sphere of Mf and equatorial OH groups in the molecular structure of chitin hydrolysate.

A Method for the Preparation of Chicken Liver Pâté that Reliably Destroys Campylobacters.

PubMed

Hutchison, Mike; Harrison, Dawn; Richardson, Ian; Tchórzewska, Monika

2015-04-28

This study devised a protocol for the manufacture of commercial quantities of chicken liver pâté that reliably destroyed campylobacters. A literature search identified 40 pâté manufacture recipes. Recipes stages with a potential to be antimicrobial were assembled to form a new protocol that included washing with organic acid, freeze-thaw and flambé in alcohol. Naturally-contaminated, high-risk livers were obtained from clearance flocks at slaughter and the effect of each stage of the protocol on Campylobacter populations was determined. Organic acid washing changed the color of the liver surfaces. However, there were no significant differences between liver surface color changes when a range of concentrations of lactic acid and ethanoic acid washes were compared by reflective spectrophotometry. A 5% (w/v) acid wash reduced numbers of indigenous campylobacters by around 1.5 log₁₀ CFU/g for both acids. The use of a Bain Marie was found to more reproducibly apply heat compared with pan-frying. Antimicrobial recipe stages reduced the numbers of campylobacters, but not significantly if thermal processing was ineffective. Cooking to 63°C was confirmed to be a critical control point for campylobacters cooked in a Bain Marie. Organoleptic and sensory assessment of pâté determined an overall preference for pâté made from frozen livers.

A Method for the Preparation of Chicken Liver Pâté that Reliably Destroys Campylobacters

PubMed Central

Hutchison, Mike; Harrison, Dawn; Richardson, Ian; Tchórzewska, Monika

2015-01-01

This study devised a protocol for the manufacture of commercial quantities of chicken liver pâté that reliably destroyed campylobacters. A literature search identified 40 pâté manufacture recipes. Recipes stages with a potential to be antimicrobial were assembled to form a new protocol that included washing with organic acid, freeze-thaw and flambé in alcohol. Naturally-contaminated, high-risk livers were obtained from clearance flocks at slaughter and the effect of each stage of the protocol on Campylobacter populations was determined. Organic acid washing changed the color of the liver surfaces. However, there were no significant differences between liver surface color changes when a range of concentrations of lactic acid and ethanoic acid washes were compared by reflective spectrophotometry. A 5% (w/v) acid wash reduced numbers of indigenous campylobacters by around 1.5 log10 CFU/g for both acids. The use of a Bain Marie was found to more reproducibly apply heat compared with pan-frying. Antimicrobial recipe stages reduced the numbers of campylobacters, but not significantly if thermal processing was ineffective. Cooking to 63°C was confirmed to be a critical control point for campylobacters cooked in a Bain Marie. Organoleptic and sensory assessment of pâté determined an overall preference for pâté made from frozen livers. PMID:25927478

Effect of sucrose and pectin addition on physical, chemical, thermal and rheological properties of frozen/thawed pineapple pulps

NASA Astrophysics Data System (ADS)

Conceição, Márcia Cavalcante; Fernandes, Tatiana Nunes; Prado, Mônica Elisabeth Torres; de Resende, Jaime Vilela

2012-09-01

Pectin (0-1.0 g/100 mL) and sucrose (0-20 g/100 mL) were added to pineapple pulp to improve their rheological properties, thermal properties and stability after freezing and thawing processes. The properties of the mixes were characterized before and after freezing and thawing. Samples were frozen at -20°C, and the freeze concentration was evaluated every 60 min. The thawing rate was evaluated at 19°C and quantified by photographic editing and image analysis software. The thawing rates and values for the freeze concentration were leveled out at pectin concentrations above 0.5 g/100 mL pectin, which indicated that pectin functions to maintain structural homogeneity during freezing. In the thawed samples, the plastic viscosity values were leveled out from pectin concentrations (0.25-0.75 g/100 mL) as the sucrose concentration increased when compared to unfrozen samples. The differences between the rheological parameters of the unfrozen and frozen/thawed pulps, the higher yield stress values after thawing were attributed to the size of suspended particles in the pulp. Applications can specify formulations of frozen products containing pectin, where these properties can be handled after thawing the product.

Treatment of rats with glucagon or mannoheptulose increases mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase activity and decreases succinyl-CoA content in liver.

PubMed Central

Quant, P A; Tubbs, P K; Brand, M D

1989-01-01

1. The activity of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase (EC 4.1.3.5) in extracts of rapidly frozen rat livers was doubled in animals treated in various ways to increase ketogenic flux. 2. Some 90% of the activity measured was mitochondrial, and changes in mitochondrial activity dominated changes in total enzyme activity. 3. The elevated HMG-CoA synthase activities persisted throughout the isolation of liver mitochondria. 4. Intramitochondrial succinyl-CoA content was lower in whole liver homogenates and in mitochondria isolated from animals treated with glucagon or mannoheptulose. 5. HMG-CoA synthase activity in mitochondria from both ox and rat liver was negatively correlated with intramitochondrial succinyl-CoA levels when these were manipulated artificially. Under these conditions, the differences between mitochondria from control and hormone-treated rats were abolished. 6. These findings show that glucagon can decrease intramitochondrial succinyl-CoA concentration, and that this in turn can regulate mitochondrial HMG-CoA synthase. They support the hypothesis that the formation of ketone bodies from acetyl-CoA may be regulated by the extent of succinylation of mitochondrial HMG-CoA synthase. PMID:2573345

[Liver transplantation preserving the vena cava and a temporary portocaval shunt].

PubMed

Hesse, U J; Berrevoet, F; Troisi, R; Mortier, E; Decruyenaere, J; Pattyn, P; de Hemptinne, B

1999-02-01

The experience with laterolateral cavocavostomy for hepatovenous reconstruction in liver transplantation is reviewed with and without the use of a temporary portocaval shunt. A total of 65 liver transplantations were analyzed. In 49 transplantations a laterolateral cavocaval anastomosis was performed (group I). In group II (n = 16) the same technique was used after a temporary portal caval shunt was constructed. Mean arterial pressure (mmHg): group I 128 +/- 34; group II 109 +/- 32. Cardiac output (l/min) decrease during the anhepatic phase was 2.3 +/- 1.9 and 1.2 +/- 1.5, respectively (P < 0.05). The peroperative blood loss measured as the number of packed cells transfused was 16.4 +/- 15.8 versus 1.2 +/- 2.3 (P < 0.04) and fresh frozen plasma 19.0 +/- 14.7 versus 3.7 +/- 4.0 (P < 0.02). Course on ICU (days), liver function tests, renal function and the need for reoperation because of bleeding were not statistically significantly different between the groups. One-year patient survival was 82.7 and 85.7%, respectively. In conclusion, we found that despite preservation of the caval flow during hepatectomy, the additional use of a temporary portocaval shunt was advantageous with regard to peroperative hemorrhage and hemodynamic stability and can potentially facilitate implantation of the liver graft.

Concentration of Nitrate near the Surface of Frozen Salt Solutions

NASA Astrophysics Data System (ADS)

Michelsen, R. R. H.; Marrocco, H. A.

2017-12-01

The photolysis of nitrate near the surface of snow and ice in Earth's environment results in the emission of nitrogen oxides (NO, NO2 and, in acidic snow, HONO) and OH radicals. As a result, nitrate photolysis affects the composition and oxidative capacity of the overlying atmosphere. Photolysis yields depend in part on how much nitrate is close enough to the surface to be photolyzed. These concentrations are assumed to be higher than the concentrations of nitrate that are measured in melted snow and ice samples. However, near-surface concentrations of nitrate have not been directly measured. In this work, laboratory studies of the concentration of nitrate in frozen aqueous solutions are described. Individual aqueous solutions of nitric acid, sodium nitrate, and magnesium nitrate were mixed. Attenuated total reflection infrared spectroscopy was utilized to measure the nitrate and liquid water signals within 200 - 400 nm of the lower surface of frozen samples. Temperature was varied from -18°C to -2°C. In addition to the amount of nitrate observed, changes to the frozen samples' morphology with annealing are discussed. Nitrate concentrations near the lower surface of these frozen solutions are high: close to 1 M at warmer temperatures and almost 4 M at the coldest temperature. Known freezing point depression data describe the observed concentrations better than ideal solution thermodynamics, which overestimate concentration significantly at colder temperatures. The implications for modeling the chemistry of snow are discussed. Extending and relating this work to the interaction of gas-phase nitric acid with the surfaces of vapor-deposited ice will also be explored.

76 FR 31575 - United States Standards for Grades of Frozen Onions

Federal Register 2010, 2011, 2012, 2013, 2014

2011-06-01

... processors of frozen onions in the United States. The petition provided information on style, sample size... change in the style designations for minced style, and a correction to the text. The members agreed with the proposed section concerning requirements for Styles, Type I, Whole onions and Type II, Pearl...

Effect of Chitin Isolated from Crustaceans and Cephalopods on Denaturation of Fish Myofibrillar Protein and the State of Water during Frozen Storage

NASA Astrophysics Data System (ADS)

Arredondo Romero, Eduardo; Nakamura, Yukio; Yamashita, Yasumitsu; Ichikawa, Hisashi; Goto, Shingi; Osatomi, Kiyoshi; Nozaki, Yukinori

From the point of view of utilization of shells as a waste product of fishery industry, the cryoprotective effect of chitin made from shell of crustaceans (Japanese fan lobster and Japanese swimming crab) and cartilage of cephalopods (spear squid) are studied. Chitin from the shells and cartilage were added to lizard fish myofibrils, and the changes of unfrozen water in myofibrils and ATPase activity of myofibrillar protein were observed during frozen storage at -250°C for 120days. The amount of unfrozen water were increased by addition of three kinds of chitin, and decreased moderately during forzen storage. Whereas, in the chitin free sample, the amount of unfrozen water were decreased rapidly during frozen storage. Changes of ATPase activity of samples showed similar tendency to that of the amount of unfrozen water. The present moderate cryoprotective effect of chitin and data of unfrozen water and ATPase activity of myofibrillar protein suggest the importance of the amount of unfrozen water in frozen matrix.

Effects of frozen storage on survival of Staphylococcus aureus and enterotoxin production in precooked tuna meat.

PubMed

Wu, Xulei; Su, Yi-Cheng

2014-08-01

This study investigated the survival of Staphylococcus aureus in precooked tuna meat for producing canned products during frozen storage (-20 ± 2 °C) as well as its growth and enterotoxin production at 35 to 37 °C after the storage. Samples (50 ± 5 g) of precooked albacore (loin, chunk, and flake) and skipjack (chunk and flake) tuna were inoculated with 5 enterotoxin-producing strains of S. aureus at a level of approximately 3.5 log CFU/g and individually packed in a vacuum bag after 3 h incubation at 35 to 37 °C. Vacuum-packed samples were stored in a freezer (-20 ± 2 °C) for 4 wk. The frozen samples were then thawed in 37 °C circulating water for 2 h and incubated at 35 to 37 °C for 22 h. Populations of S. aureus in all precooked tuna samples decreased slightly (<0.7 log CFU/g) after 4 wk of storage at -20 ± 2 °C, but increased rapidly once the samples were thawed and held at 35 to 37 °C. Total S. aureus counts in albacore and skipjack samples increased by greater than 3 log CFU/g after 6 and 8 h of exposure to 35 to 37 °C, respectively. All samples became spoiled after 10 h of exposure to 35 to 37 °C, while no enterotoxin was detected in any samples. However, enterotoxins were detected in albacore loin and other samples after 12 and 24 h of incubation at 35 to 37 °C, respectively. Frozen precooked tuna meat should be used for producing canned tuna within 6 to 8 h of thawing to avoid product spoilage and potential enterotoxin production by S. aureus in contaminated precooked tuna meat. © 2014 Institute of Food Technologists®

INFLUENCE OF INTRAMUSCULAR FAT LEVEL ON ORGANOLEPTIC, PHYSICAL, AND CHEMICAL CHARACTERISTICS OF IRRADIATED PORK. II. LOW-TEMPERATURE LONG-TIME PRE-IRRADIATION HEAT TREATMENT

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bray, R.W.; Weckel, K.G.; Evans, G.W.

1964-02-01

Pork muscle (longissimus dorsi) was graded in three marbling levels by both visual appraisal and ether-extraction analysis of total fat. A low- temperature (121 deg F, gradually increasing to 191 deg F), long-time (19-hr) heat treatment was used for enzyme inactivation. Samples were packed under vacuum in rigid containers and irradiated to 4.5 Mrad with gamma radiation. Irradiated and frozen control samples were evaluated at intervals over a 180-day period. The heat treatment caused excessive connective-tissue breakdown as evidenced by a soft, dry texture. Marbling level had no significant effect on consumer-panel judgments of irradiated or frozen control samples (servedmore » plain, without bread). Lower degrees of marbling were preferred in irradiated sandwich items. Irradiated samples were less preferred than frozen control samples in both plain and sandwich form. Hunter L (color) values a/sub L/ (redness), hue, and saturation attributes were increased by irradiation treatment. Hunter L and b/sub L/ (yellowness) values of highly marbled irradiated sample-s were elevated. Highly marbled samples displayed greater tenderness qualities, as evidenced by mechanical tenderness measurements. Expressible moisture values were decreased by marbling degree and radiation treatment and increased by advancing storage time. As a measure of oxidation rancidity, thiobarbituric acid values were increased by increasing levels of marbling and advancing storage time, but were not influenced by preservation method. No significant differences in pH values due to marbling level or preservation method were detected. Bacteriologic counts of randomly selected irradiated samples indicated that they were commercially sterile (average of 57 colonies/g of sample). Methylene blue stains of colonles revealed Micrococcus. Frozen (control) samples contained moderate numbers of colonies (av. 2300 colonies per g of sample). Micrococcus colonies were again predominant, but a few colonles of film yeasts were also seen. Irradiated samples were negative for thermophilic anaerobic spores producing gas. Control samples were positive, containing gram-negative rods, Escherichia coli, and gram- positive cocci. (BBB)« less

Single-layer centrifugation through PureSperm® 80 selects improved quality spermatozoa from frozen-thawed dog semen.

PubMed

Dorado, J; Alcaraz, L; Gálvez, M J; Acha, D; Ortiz, I; Urbano, M; Hidalgo, M

2013-08-01

The aim of this study was to investigate whether single-layer centrifugation (SLC) with PureSperm® 80 could select good quality spermatozoa, including those with specific motility patterns, from doses of frozen dog semen. Semen from 5 dogs was collected and cryopreserved following a standard protocol. After thawing, semen samples were divided into two aliquots: one of them was used as control and the other one processed by SLC. Assessment of sperm motility (assessed by computer-assisted semen analysis), morphology (Diff-Quick staining) and viability (triple fluorescent stain of propidium iodine/isothiocyanate-labeled peanut (Arachis hypogaea) agglutinin/Rhodamine 123), were performed on aliquots of fresh semen, frozen-thawed control and frozen-thawed SLC treated samples. A multivariate clustering procedure separated 26,051 motile spermatozoa into three subpopulations (sP): sP1 consisting of highly active but non-progressive spermatozoa (40.3%), sP2 consisting of spermatozoa with high velocity and progressive motility (30.0%), and sP3 consisting of poorly active and non-progressive spermatozoa (29.7%). SLC with PureSperm® 80 yielded sperm suspensions with improved motility, morphology, viability and acrosome integrity (P<0.001). The frozen-thawed SLC treated samples were enriched in sP2, reaching a proportion of 44.1% of the present spermatozoa. From these results, we concluded that SLC with PureSperm® 80 may be an alternative and successful method for improving the quality of frozen-thawed dog spermatozoa. Moreover, sP2 (high-speed and progressive spermatozoa) was more frequently observed after SLC. Finally, this study also demonstrated that the general motile sperm structure present in dogs remained constant despite the effect caused by either cryopreservation or separation by SLC through PureSperm® 80. Copyright © 2013 Elsevier B.V. All rights reserved.

Presence of intrahepatic (total and ccc) HBV DNA is not predictive of HBV recurrence after liver transplantation.

PubMed

Hussain, Munira; Soldevila-Pico, Consuelo; Emre, Sukru; Luketic, Velimir; Lok, Anna S F

2007-08-01

Previous studies reported that hepatitis B virus (HBV) deoxyribonucleic acid (DNA) can be detected in livers of patients who received transplants for hepatitis B despite the absence of serological markers of HBV recurrence. Quantification of HBV DNA was not performed and presence of covalently closed circular (ccc) DNA was not analyzed in most studies. We aimed to quantify total and ccc HBV DNA in explant liver and post-orthotopic liver transplantation (OLT) biopsies and to correlate the values with HBV recurrence post-OLT. Frozen liver tissue from 34 patients (9 with explant liver only, 9 with explant liver and post-OLT liver biopsies, and 16 with post-OLT biopsies only) in the National Institutes of Health HBV-OLT study was examined using real-time polymerase chain reaction (PCR). Among the 18 patients with explant liver, 7 were hepatitis B e antigen (HBeAg)-positive, 8 had detectable serum HBV DNA, and 10 received antiviral therapy prior to OLT. Total and ccc HBV DNA was detected in explant livers of 17 and 16 patients, respectively. Of the 10 patients who received antiviral therapy pre-OLT, serum HBV DNA was undetectable in 8 at transplantation but 7 had detectable total and ccc HBV DNA in their explant liver. Of the 25 patients with post-OLT biopsies, total HBV DNA was detected in 83% and ccc DNA in 17% of 47 biopsies, although only 2 patients had HBV recurrence. In conclusion, total and ccc HBV DNA could be detected in explant livers of most patients despite antiviral therapy pre-OLT. Total but not ccc HBV DNA could be detected in post-OLT liver biopsies of most patients despite undetectable serum HBV DNA and hepatitis B surface antigen (HBsAg). Our findings suggest that occult HBV reinfection occurs in most HBV patients after OLT and continued administration of appropriate prophylactic therapy is important in preventing overt HBV recurrence. Copyright (c) 2007 AASLD.

The impact of milk handling procedures on Ostertagia ostertagi antibody ELISA test results.

PubMed

Vanderstichel, Raphaël; Dohoo, Ian; Stryhn, Henrik

2010-04-19

The impact of various milk handling stressors were analyzed using a commercially available enzyme-linked immunosorbent assay (ELISA) test measuring Ostertagia ostertagi antibodies in milk from dairy cattle (Svanovir). An indirect ELISA has the ability to determine the amount of milk production losses related to intestinal parasitism. The ELISA test recommends fresh defatted milk, however, milk collected from Dairy Herd Improvement (DHI) programs in North America undergo many stressors, including, heating, freezing and are not defatted. Normalized optical density ratios (ODRs) were compared between fresh defatted milk and milk subjected to one or more stressors with a linear mixed model accounting for differences in variation between the fresh and the frozen samples. Concordance correlation coefficients were also analyzed for comparisons to other similar studies. After accounting for random cow and container effects, the treatment factors interacted with each other (p<0.001). Biologically interesting contrasts were created to explain the interaction. The estimated difference in ODR between the milk samples handled according to recommendations of the manufacturers of Svanovir and the whole milk samples that were subjected to the most extreme treatment (heated, frozen, thawed, and re-frozen for 4 weeks) was 0.062 (p<0.001). This difference represented less than 5% of the range, and was thus considered biologically negligible. Frozen whole milk processed by DHI programs, the most likely method of collecting on-farm samples in North America, will likely yield reliable results for the indirect ELISA tests, particularly, Svanovir.

Presentation of frozen shoulder among diabetic and non-diabetic patients☆

PubMed Central

Uddin, Mohammad Moin; Khan, Aminuddin A.; Haig, Andrew J.; Uddin, Mohammad Kafil

2014-01-01

Objective The literature is inconsistent regarding the level of pain and disability in frozen shoulder patients with or without diabetes mellitus. The aim of this study is to evaluate some demographic features of frozen shoulder patients and to look into the disparity of information by comparing the level of pain and disability due to frozen shoulder between diabetic and non-diabetic people. Design This is a prospective comparative study. People with frozen shoulder attending an outpatient department were selected by consecutive sampling. Disability levels were assessed by the Shoulder Pain & Disability Index (SPADI). Means of pain and disability scores were compared using unpaired t-test. Results Among 140 persons with shoulder pain 99 (71.4%) had frozen shoulder. From the participating 40 frozen shoulder patients, 26 (65%) were males and 14 (35%) were females. Seventeen participants (42.5%) were diabetic, two (5%) had impaired glucose tolerance and 21 (52.5%) patients were non-diabetic. Mean disability scores (SPADI) were 51 ± 15.5 in diabetic and 57 ± 16 in non-diabetic persons. The differences in pain and disability level were not statistically significance (respectively, p = 0.24 and p = 0.13 at 95% confidence interval). Conclusions No difference was found in level of pain and disability level between frozen shoulder patients with and without diabetes. PMID:25983497

Fusobacterium nucleatum in gastroenterological cancer: Evaluation of measurement methods using quantitative polymerase chain reaction and a literature review.

PubMed

Yamamura, Kensuke; Baba, Yoshifumi; Miyake, Keisuke; Nakamura, Kenichi; Shigaki, Hironobu; Mima, Kosuke; Kurashige, Junji; Ishimoto, Takatsugu; Iwatsuki, Masaaki; Sakamoto, Yasuo; Yamashita, Yoichi; Yoshida, Naoya; Watanabe, Masayuki; Baba, Hideo

2017-12-01

The human microbiome Fusobacterium nucleatum , which primarily inhabits the oral cavity, causes periodontal disease and has also been implicated in the development of colorectal cancer. However, whether F. nucleatum is present in other gastroenterological cancer tissues remains to be elucidated. The present study evaluated whether quantitative polymerase chain reaction (qPCR) assays were able to detect F. nucleatum DNA and measure the quantity of F. nucleatum DNA in esophageal, gastric, pancreatic and liver cancer tissues. The accuracy of the qPCR assay was determined from a calibration curve using DNA extracted from cells from the oral cavity. Formalin-fixed paraffin-embedded (FFPE) tumor tissues from 20 patients with gastroenterological [esophageal (squamous cell carcinoma), gastric, colorectal, pancreatic and liver] cancer and 20 matched normal tissues were evaluated for F. nucleatum DNA content. The cycle threshold values in the qPCR assay for F. nucleatum and solute carrier organic anion transporter family member 2A1 (reference sample) decreased linearly with the quantity of input DNA ( r 2 >0.99). The F. nucleatum detection rate in esophageal, gastric and colorectal cancer tissues were 20% (4/20), 10% (2/20) and 45% (9/20), respectively. F. nucleatum was not detected in liver and pancreatic cancer tissues. The qPCR results from the frozen and FFPE tissues were consistent. Notably, F. nucleatum was detected at a higher level in superficial areas compared with the invasive areas. F. nucleatum in esophageal, gastric and colorectal cancer tissues was evaluated by qPCR using FFPE tissues. F. nucleatum may be involved in the development of esophageal, gastric and colorectal cancer.

Enhancement of the spectral selectivity of complex samples by measuring them in a frozen state at low temperatures in order to improve accuracy for quantitative analysis. Part II. Determination of viscosity for lube base oils using Raman spectroscopy.

PubMed

Kim, Mooeung; Chung, Hoeil

2013-03-07

The use of selectivity-enhanced Raman spectra of lube base oil (LBO) samples achieved by the spectral collection under frozen conditions at low temperatures was effective for improving accuracy for the determination of the kinematic viscosity at 40 °C (KV@40). A collection of Raman spectra from samples cooled around -160 °C provided the most accurate measurement of KV@40. Components of the LBO samples were mainly long-chain hydrocarbons with molecular structures that were deformable when these were frozen, and the different structural deformabilities of the components enhanced spectral selectivity among the samples. To study the structural variation of components according to the change of sample temperature from cryogenic to ambient condition, n-heptadecane and pristane (2,6,10,14-tetramethylpentadecane) were selected as representative components of LBO samples, and their temperature-induced spectral features as well as the corresponding spectral loadings were investigated. A two-dimensional (2D) correlation analysis was also employed to explain the origin for the improved accuracy. The asynchronous 2D correlation pattern was simplest at the optimal temperature, indicating the occurrence of distinct and selective spectral variations, which enabled the variation of KV@40 of LBO samples to be more accurately assessed.

Interindividual Variability in Hepatic Organic Anion-Transporting Polypeptides and P-Glycoprotein (ABCB1) Protein Expression: Quantification by Liquid Chromatography Tandem Mass Spectroscopy and Influence of Genotype, Age, and Sex

PubMed Central

Prasad, Bhagwat; Evers, Raymond; Gupta, Anshul; Hop, Cornelis E. C. A.; Salphati, Laurent; Shukla, Suneet; Ambudkar, Suresh V.

2014-01-01

Interindividual variability in protein expression of organic anion-transporting polypeptides (OATPs) OATP1B1, OATP1B3, OATP2B1, and multidrug resistance-linked P-glycoprotein (P-gp) or ABCB1 was quantified in frozen human livers (n = 64) and cryopreserved human hepatocytes (n = 12) by a validated liquid chromatography tandem mass spectroscopy (LC-MS/MS) method. Membrane isolation, sample workup, and LC-MS/MS analyses were as described before by our laboratory. Briefly, total native membrane proteins, isolated from the liver tissue and cryopreserved hepatocytes, were trypsin digested and quantified by LC-MS/MS using signature peptide(s) unique to each transporter. The mean ± S.D. (maximum/minimum range in parentheses) protein expression (fmol/µg of membrane protein) in human liver tissue was OATP1B1- 2.0 ± 0.9 (7), OATP1B3- 1.1 ± 0.5 (8), OATP2B1- 1 1.7 ± 0.6 (5), and P-gp- 0.4 ± 0.2 (8). Transporter expression in the liver tissue was comparable to that in the cryopreserved hepatocytes. Most important is that livers with SLCO1B1 (encoding OATP1B1) haplotypes *14/*14 and *14/*1a [i.e., representing single nucleotide polymorphisms (SNPs), c.388A > G, and c.463C > A] had significantly higher (P < 0.0001) protein expression than the reference haplotype (*1a/*1a). Based on these genotype-dependent protein expression data, we predicted (using Simcyp) an up to ∼40% decrease in the mean area under the curve of rosuvastatin or repaglinide in subjects harboring these variant alleles compared with those harboring the reference alleles. SLCO1B3 (encoding OATP1B3) SNPs did not significantly affect protein expression. Age and sex were not associated with transporter protein expression. These data will facilitate the prediction of population-based human transporter-mediated drug disposition, drug-drug interactions, and interindividual variability through physiologically based pharmacokinetic modeling. PMID:24122874

Successful Application of Microarray Technology to Microdissected Formalin-Fixed, Paraffin-Embedded Tissue

PubMed Central

Coudry, Renata A.; Meireles, Sibele I.; Stoyanova, Radka; Cooper, Harry S.; Carpino, Alan; Wang, Xianqun; Engstrom, Paul F.; Clapper, Margie L.

2007-01-01

The establishment of a reliable method for using RNA from formalin-fixed, paraffin-embedded (FFPE) tissue would provide an opportunity to obtain novel gene expression data from the vast amounts of archived tissue. A custom-designed 22,000 oligonucleotide array was used in the present study to compare the gene expression profile of colonic epithelial cells isolated by laser capture microdissection from FFPE-archived samples with that of the same cell population from matched frozen samples, the preferred source of RNA. Total RNA was extracted from FFPE tissues, amplified, and labeled using the Paradise Reagent System. The quality of the input RNA was assessed by the Bioanalyzer profile, reverse transcriptase-polymerase chain reaction, and agarose gel electrophoresis. The results demonstrate that it is possible to obtain reliable microarray data from FFPE samples using RNA acquired by laser capture microdissection. The concordance between matched FFPE and frozen samples was evaluated and expressed as a Pearson’s correlation coefficient, with values ranging from 0.80 to 0.97. The presence of ribosomal RNA peaks in FFPE-derived RNA was reflected by a high correlation with paired frozen samples. A set of practical recommendations for evaluating the RNA integrity and quality in FFPE samples is reported. PMID:17251338

Mass spectrometric imaging and laser desorption ionization (LDI) with ice as a matrix using femtosecond laser pulses

NASA Astrophysics Data System (ADS)

Berry, Jamal Ihsan

The desorption of biomolecules from frozen aqueous solutions on metal substrates with femtosecond laser pulses is presented for the first time. Unlike previous studies using nanosecond pulses, this approach produces high quality mass spectra of biomolecules repeatedly and reproducibly. This novel technique allows analysis of biomolecules directly from their native frozen environments. The motivation for this technique stems from molecular dynamics computer simulations comparing nanosecond and picosecond heating of water overlayers frozen on Au substrates which demonstrate large water cluster formation and ejection upon substrate heating within ultrashort timescales. As the frozen aqueous matrix and analyte molecules are transparent at the wavelengths used, the laser energy is primarily absorbed by the substrate, causing rapid heating and explosive boiling of the ice overlayer, followed by the ejection of ice clusters and the entrained analyte molecule. Spectral characteristics at a relatively high fluence of 10 J/cm 2 reveal the presence of large molecular weight metal clusters when a gold substrate is employed, with smaller cluster species observed from frozen aqueous solutions on Ag, Cu, and Pb substrates. The presence of the metal clusters is indicative of an evaporative cooling mechanism which stabiles cluster ion formation and the ejection of biomolecules from frozen aqueous solutions. Solvation is necessary as the presence of metal clusters and biomolecular ion signals are not observed from bare metal substrates in absence of the frozen overlayer. The potential for mass spectrometric imaging with femtosecond LDI of frozen samples is also presented. The initial results for the characterization of peptides and peptoids linked to combinatorial beads frozen in ice and the assay of frozen brain tissue from the serotonin transporter gene knockout mouse via LDI imaging are discussed. Images of very good quality and resolution are obtained with 400 nm, 200 fs pulses at a fluence of 1.25 J/cm2 . An attractive feature of this technique is that images are acquired within minutes for large sample areas. Additionally, the images obtained with femtosecond laser desorption are high in lateral resolution with the laser capable of being focused to a spot size of 30 mum. Femtosecond laser desorption from ice is unique in that unlike matrix assisted laser desorption ionization mass spectrometry, it does not employ an organic UV absorbing matrix to desorb molecular ions. Instead, the laser energy is absorbed by the metal substrate causing explosive boiling and ejection of the frozen overlayer. This approach is significant in that femtosecond laser desorption possess the potential of analyzing and assaying biomolecules directly from their frozen native environments. This technique was developed to compliment existing ToF-SIMS imaging capability for analysis of tissue and cells, as well as other biological systems of interest.

Simultaneously Synchrotron X-ray Fluorescence and Ptychographic Imaging of Frozen Biological Single Cells

DOE PAGES

Chen, S.; Deng, J.; Nashed, Y. S. G.; ...

2016-07-25

Bionanoprobe (BNP), a hard x-ray fluorescence sample-scanning nanoprobe at the Advanced Photon Source of Argonne National Laboratory, has been used to quantitatively study elemental distributions in biological cells with sub-100 nm spatial resolution and high sensitivity. Cryogenic conditions enable biological samples to be studied in their frozen-hydrated state with both ultrastructure and elemental distributions more faithfully preserved compared to conventional chemical fixation or dehydration methods. Furthermore, radiation damage is reduced in two ways: the diffusion rate of free radicals is decreased at low temperatures; and the sample is embedded in vitrified ice, which reduces mass loss.

Conservation science in a terrorist age: the impact of airport security screening on the viability and DNA integrity of frozen felid spermatozoa.

PubMed

Gloor, Kayleen T; Winget, Doug; Swanson, William F

2006-09-01

In response to growing terrorism concerns, the Transportation Security Administration now requires that all checked baggage at U.S. airports be scanned through a cabinet x-ray system, which may increase risk of radiation damage to transported biologic samples and other sensitive genetic material. The objective of this study was to investigate the effect of these new airport security regulations on the viability and DNA integrity of frozen felid spermatozoa. Semen was collected from two domestic cats (Felis silvestris catus) and one fishing cat (Prionailurus viverrinus), cryopreserved in plastic freezing straws, and transferred into liquid nitrogen dry shippers for security screening. Treatment groups included frozen samples from each male scanned once or three times using a Transportation Security Administration-operated cabinet x-ray system, in addition to non-scanned samples (i.e., negative control) and samples previously scanned three times and exposed to five additional high-intensity x-ray bursts (i.e., positive control). Dosimeters placed in empty dry shippers were used to quantify radiation exposure. Following treatment, straws were thawed and spermatozoa analyzed for post-thaw motility (percentage motile and rate of progressive movement), acrosome status, and DNA integrity using single-cell gel electrophoresis (i.e., the comet assay). Dosimeter measurements determined that each airport screening procedure produced approximately 16 mrem of radiation exposure. Our results indicated that all levels of radiation exposure adversely affected (P < 0.05) post-thaw sperm motility, but the percentage of acrosome-intact spermatozoa did not differ (P > 0.05) among treatment groups. Results also showed that the amount of double-stranded DNA damage was greater (P < 0.05) in sperm samples from both cat species scanned three times compared to samples scanned once or negative controls. Findings suggest that new airport security measures may cause radiation-induced damage to frozen spermatozoa and other valuable biologic samples transported on passenger aircraft and that alternative modes of sample transportation should be used whenever possible.

The effect of Taro (Colocasia esculenta L.) and Lesser Yam flour (Dioscorea esculenta L.) as thickener agent on physical characteristics of frozen wheygurt

NASA Astrophysics Data System (ADS)

Nurhartadi, E.; Utami, R.; Widowati, E.; Karunawati, B. M.

2017-11-01

The results showed that the addition ratio of taro and lesser yam flour affected to the physical characteristics of frozen wheygurt. The addition of lesser yam flour increased total soluble solids until the addition ratio of 2:2 due to the higher ash content of lesser yam (2.87%) than taro (0.44%). Sample with addition ratio of 1:3 and 0:4 significantly different compared to other samples, due to the starch content difference between taro (70-80%) and lesser yam (51.34%). Addition ratio of taro and lesser yam flour do not have a significant effect on the viscosity of the frozen wheygurt, due to both starch have similar setback viscosity. Lesser yam setback viscosity was 684.8 cP, while taro was 838.3 cP. Setback viscosity showed a high tendency of retrogradation. The addition ratio of taro and lesser yam flour have a significant effect to the overrun of frozen wheygurt. Addition ratio of taro and lesser yam flour have a significant effect to melting rate of frozen wheygurt. This result was caused by higher peak viscosity of taro starch compared to lesser yam, thus produced thicker gel than lesser yam. This lead increased water contents in the mixtures entrapped and slows down water mobility, hence melting rate would decrease.

The Pyrolytic Profile of Lyophilized and Deep-Frozen Compact Part of the Human Bone

PubMed Central

Lodowska, Jolanta; Wolny, Daniel; Kurkiewicz, Sławomir; Węglarz, Ludmiła

2012-01-01

Background. Bone grafts are used in the treatment of nonunion of fractures, bone tumors and in arthroplasty. Tissues preserved by lyophilization or deep freezing are used as implants nowadays. Lyophilized grafts are utilized in the therapy of birth defects and bone benign tumors, while deep-frozen ones are applied in orthopedics. The aim of the study was to compare the pyrolytic pattern, as an indirect means of the analysis of organic composition of deep-frozen and lyophilized compact part of the human bone. Methods. Samples of preserved bone tissue were subjected to thermolysis and tetrahydroammonium-hydroxide- (TMAH-) associated thermochemolysis coupled with gas chromatography and mass spectrometry (Py-GC/MS). Results. Derivatives of benzene, pyridine, pyrrole, phenol, sulfur compounds, nitriles, saturated and unsaturated aliphatic hydrocarbons, and fatty acids (C12–C20) were identified in the pyrolytic pattern. The pyrolyzates were the most abundant in derivatives of pyrrole and nitriles originated from proteins. The predominant product in pyrolytic pattern of the investigated bone was pyrrolo[1,2-α]piperazine-3,6-dione derived from collagen. The content of this compound significantly differentiated the lyophilized graft from the deep-frozen one. Oleic and palmitic acid were predominant among fatty acids of the investigated samples. The deep-frozen implants were characterized by higher percentage of long-chain fatty acids than lyophilized grafts. PMID:22619606

High pressure treatments combined with sodium lactate to inactivate Escherichia coli O157:H7 and spoilage microbiota in cured beef carpaccio.

PubMed

Masana, Marcelo Oscar; Barrio, Yanina Ximena; Palladino, Pablo Martín; Sancho, Ana Maria; Vaudagna, Sergio Ramón

2015-04-01

High-pressure treatments (400 and 600 MPa) combined with the addition of sodium lactate (1 and 3%) were tested to reduce Escherichia coli O157:H7 (STEC O157) and spoilage microbiota contamination in a manufactured cured beef carpaccio in fresh or frozen conditions. Counts of spoilage microorganisms and STEC O157 were also examined during the curing step to prepare the carpaccio. STEC O157 counts remained almost unchanged through the curing process performed at 1 ± 1 °C for 12 days, with a small decrease in samples with 3% of sodium lactate. High-pressure treatments at 600 MPa for 5 min achieved an immediate reduction of up to 2 logarithmic units of STEC O157 in frozen carpaccio, and up to 1.19 log in fresh condition. Counts of spoilage bacteria diminished below detection limits in fresh or frozen carpaccio added with sodium lactate by the application of 400 and 600 MPa. Maximum injury on STEC O157 cells was observed at 600 MPa in carpaccio in fresh condition without added sodium lactate. Lethality of high-pressure treatments on STEC O157 was enhanced in frozen carpaccio, while the addition of sodium lactate at 3% reduced the lethality on STEC O157 in frozen samples, and the degree of injury in fresh carpaccio. Copyright © 2014 Elsevier Ltd. All rights reserved.

Effect of sample preparation method on sensory quality of cooked chicken breast fillets processed for food service

USDA-ARS?s Scientific Manuscript database

Chicken fillets (Pectoralis major) are one of popular items for food service. In the store, especially in fast food service stores, ready-to-cook meat products are commonly stored in freezers before use. The frozen meat can be cooked either directly from a frozen stage or after thawing. However, the...

Qualitative and quantitative assessment of DNA quality of frozen beef based on DNA yield, gel electrophoresis and PCR amplification and their correlations to beef quality.

PubMed

Zhao, Jing; Zhang, Ting; Liu, Yongfeng; Wang, Xingyu; Zhang, Lan; Ku, Ting; Quek, Siew Young

2018-09-15

Freezing is a practical method for meat preservation but the quality of frozen meat can deteriorate with storage time. This research investigated the effect of frozen storage time (up to 66 months) on changes in DNA yield, purity and integrity in beef, and further analyzed the correlation between beef quality (moisture content, protein content, TVB-N value and pH value) and DNA quality in an attempt to establish a reliable, high-throughput method for meat quality control. Results showed that frozen storage time influenced the yield and integrity of DNA significantly (p < 0.05). The DNA yield decreased as frozen storage time increased due to DNA degradation. The half-life (t 1/2  = ln2/0.015) was calculated as 46 months. The DNA quality degraded dramatically with the increased storage time based on gel electrophoresis results. Polymerase chain reaction (PCR) products from both mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) were observed in all frozen beef samples. Using real-time PCR for quantitative assessment of DNA and meat quality revealed that correlations could be established successfully with mathematical models to evaluate frozen beef quality. Copyright © 2018 Elsevier Ltd. All rights reserved.

[Histological and biochemical studies on the postnatal development of the guinea pig liver (author's transl)].

PubMed

Matthiesen, T h; Sallmann, H P; Heimann, W; Kunstýr, I

1976-01-01

In newborn guinea pigs the structural development of the liver was studied with emphasis on changes in the lipid content as related to biochemical parameters. The livers of 149 guinea pigs (strain PIRBRIGHT) up to an age of 21 days were examined. The mothers were grouped in cages and kept as usual (room temperature 22 degrees C, air humidity 60-70%, natural light-dark rhythm and commercial standard diet ad libitum, sawdust and straw as bedding). Pregnant animals were placed in individual cages being left there together with their offspring until weaning or sacrification. In this period additional uptake of solid diet was allowed to the guinea pigs. The animals were bleeded to death in anesthesia (by head stroke). For histological examination tissue samples from the Lobes sinister lateralis were fixed in Barker's formol or absolute alcohol. The following staining reactions were applied (paraffin embedding or frozen sections): haemalum-eosin stain; PAS-reaction; Sudan III; Sudan black; Best's carmine (for details on the techniques see ROMEIS 1968). In biochemical analysis the total lipids of the liver were determined by the isolation methods reported by FOLCH et al.(1957). Moreover, in modification of the analytical procedures earlier described (SALLMANN 1972 a, b) several fractions of the liver total lipids could be isolated. By dialysis against a rubber membrane a dialysable lipid component (hydrocarbons, triglycerides, incomplete glycerides, cholesterol and free lipid acids) was isolated from the phosphatide fractions. Further separation of the "dialysable" lipids on Florisil columns yielded purified glyceride fractions. The total lipids as well as all components isolated from these were determined gravimetrically after removal of the solvents concerned. In biochemical analysis the livers of fetuses 10-12 days ante partum and of young animals up to 53 days were included. Results (see also table 1): The relative liver weight--related to the body weight reduced for that of the gastrointestinal tract--is highest at birth. It is lowered independent of the development of the gastrointestinal tract within a short period, then it remains fairly constant during the rest of the time of observation. On the 1st day abundant lipids are regularly distributed throughout the whole liver lobule (fig. 1). Beginning with the 3rd day the central parts of the lobules are nearly free from lipids, only in the peripheral zones of the lobules lipid augmentation was still observed. Consequently the marked decrease in liver weight is due to the rapid reduction of lipids within the first 3 days of life. These observations are in accordance with the biochemical findings: about 10 days ante partum noticeable storage of lipids occurs in the liver reaching its peak 2 to 3 days after birth (physiological fatty infiltration of the liver during the last days of intrauterine life). In fetal liver tissue predominantly tryglycerides are augmented.

Effect of different concentrations of soybean lecithin and virgin coconut oil in Tris-based extender on the quality of chilled and frozen-thawed bull semen

PubMed Central

Tarig, A. A.; Wahid, H.; Rosnina, Y.; Yimer, N.; Goh, Y. M.; Baiee, F. H.; Khumran, A. M.; Salman, H.; Assi, M. A.; Ebrahimi, M.

2017-01-01

Aim: The objective of this study was to evaluate the effects of different concentrations of soybean lecithin (SL) and virgin coconut oil (VCO) in Tris-based extender on chilled and frozen-thawed bull semen quality parameters. Materials and Methods: A total of 24 ejaculates were collected from four bulls via an electroejaculator. Semen samples were diluted with 2% VCO in Tris-based extender which consists of various concentrations of SL (1, 1.25, 1.5, and 1.75%). A 20% egg yolk in Tris used as a positive control (C+). The diluted semen samples were divided into two fractions; one for chilling which were stored at 4°C for 24, 72, and 144 h before evaluated for semen quality parameters. The second fraction used for freezing was chilled for 3 h at 4°C, packed into 0.25 mL straws and then cryopreserved in liquid nitrogen. The samples were then evaluated after 7 and 14 days. Chilled and frozen semen samples were thawed at 37°C and assessed for general motility using computer-assisted semen analysis, viability, acrosome integrity and morphology (eosin-nigrosin stain), membrane integrity, and lipid peroxidation using thiobarbituric acid reaction test. Results: The results showed that all the quality parameters assessed were significantly (p<0.05) improved at 1.5% SL concentration in chilled semen. Treatment groups of 1, 1.25, 1.5, and 1.75% SL were higher in quality parameters than the control group (C+) in chilled semen. However, all the quality parameters in frozen-thawed semen were significantly higher in the C+ than the treated groups. Conclusion: In conclusion, supplementation of 1.5% SL in 2% VCO Tris-based extender enhanced the chilled bull semen. However, there was no marked improvement in the frozen-thawed quality parameters after treatment. PMID:28717321

Effect of different concentrations of soybean lecithin and virgin coconut oil in Tris-based extender on the quality of chilled and frozen-thawed bull semen.

PubMed

Tarig, A A; Wahid, H; Rosnina, Y; Yimer, N; Goh, Y M; Baiee, F H; Khumran, A M; Salman, H; Assi, M A; Ebrahimi, M

2017-06-01

The objective of this study was to evaluate the effects of different concentrations of soybean lecithin (SL) and virgin coconut oil (VCO) in Tris-based extender on chilled and frozen-thawed bull semen quality parameters. A total of 24 ejaculates were collected from four bulls via an electroejaculator. Semen samples were diluted with 2% VCO in Tris-based extender which consists of various concentrations of SL (1, 1.25, 1.5, and 1.75%). A 20% egg yolk in Tris used as a positive control (C+). The diluted semen samples were divided into two fractions; one for chilling which were stored at 4°C for 24, 72, and 144 h before evaluated for semen quality parameters. The second fraction used for freezing was chilled for 3 h at 4°C, packed into 0.25 mL straws and then cryopreserved in liquid nitrogen. The samples were then evaluated after 7 and 14 days. Chilled and frozen semen samples were thawed at 37°C and assessed for general motility using computer-assisted semen analysis, viability, acrosome integrity and morphology (eosin-nigrosin stain), membrane integrity, and lipid peroxidation using thiobarbituric acid reaction test. The results showed that all the quality parameters assessed were significantly (p<0.05) improved at 1.5% SL concentration in chilled semen. Treatment groups of 1, 1.25, 1.5, and 1.75% SL were higher in quality parameters than the control group (C+) in chilled semen. However, all the quality parameters in frozen-thawed semen were significantly higher in the C+ than the treated groups. In conclusion, supplementation of 1.5% SL in 2% VCO Tris-based extender enhanced the chilled bull semen. However, there was no marked improvement in the frozen-thawed quality parameters after treatment.

Breed differences of bull frozen-thawed semen.

PubMed

Ntemka, A; Tsousis, G; Brozos, C; Kiossis, E; Boscos, C M; Tsakmakidis, I A

2016-12-01

The objective of this study was to investigate the quality of frozen-thawed semen from different bull breeds. Commercial frozen-thawed bull semen samples (26 per breed, 130 totally) of five breeds (Holstein [Η], Brown Swiss [BS], Limousin [L], Belgian Blue [BB], Blonde d' Aquitaine [BA]) were used. After thawing, each semen sample was subjected to thermal resistance test (TR) for 0.5 and 1 hr at 38°C and hypo-osmotic swelling test (HOST) for 1 hr at 150 mOsm at 37°C. Additionally, all samples were evaluated at times 0 hr (thawing), 0.5 hr (TR), 1 hr (TR) for kinetics by CASA [progressive, immotile, rapid, medium, slow moving spermatozoa, curvilinear velocity (VCL), average path velocity (VAP), straight line velocity (VSL), linearity (LIN), straightness (STR), beat cross-frequency (BCF), amplitude of lateral head displacement (ALH), wobble (WOB)]. Moreover, directly after thawing, all semen samples were evaluated for morphometry, morphology, viability and DNA fragmentation. Statistical analysis was conducted using a mixed model for repeated measures. The results showed (a) higher VCL after thawing in H, L breeds compared to BB and BA, (b) higher VAP after thawing in L compared to BB, BA, (c) higher values of progressive spermatozoa after TR in H, BS compared to BB, BA, (d) higher values of rapid spermatozoa after thawing and 0.5 hr of TR in H, BS, L compared to BB, BA, (e) lower viability in BA after thawing compared to H, BS, BB, (f) lower morphological abnormalities in H compared to L, BB, (g) higher head length in Η compared to BB. No significant differences were observed in the results from HOST and DNA fragmentation between breeds. In conclusion, quality characteristics of frozen-thawed bull semen are dependent on the breed. Frozen semen from BB and BA breeds should be handled more carefully after thawing, as it is more sensitive to stress. © 2016 Blackwell Verlag GmbH.

On the compressibility and temperature boundary of warm frozen soils

NASA Astrophysics Data System (ADS)

Qi, Jilin; Dang, Boxiang; Guo, Xueluan; Sun, Xiaoyu; Yan, Xu

2017-04-01

A silty-clay obtained along the Qinghai-Tibetan railway and a standard Chinese sand were taken as study objects. Saturated frozen soil samples were prepared for testing. Step-load was used and confined compression was carried out on the soils under different temperatures. Compression index and pseudo-preconsolidation pressure (PPC) were obtained. Unlike unfrozen soils, PPC is not associated with stress history. However, it is still the boundary of elastic and plastic deformations. Different compression indexes can be obtained from an individual compression curve under pressures before and after PPC. The parameters at different thermal and stress conditions were analyzed. It is found that temperature plays a critical role in mechanical behaviours of frozen soils. Efforts were then made on the silty-clay in order to suggest a convincing temperature boundary in defining warm frozen soil. Three groups of ice-rich samples with different ice contents were prepared and tested under confined compression. The samples were compressed under a constant load and with 5 stepped temperatures. Strain rates at different temperatures were examined. It was found that the strain rate at around -0.6°C increased abruptly. Analysis of compression index was performed on the data both from our own testing program and from the literature, which showed that at about -1°C was a turning point in the curves for compression index against temperature. Based on both our work and taking into account the unfrozen water content vs. temperature, the range of -1°C to -0.5°C seems to be the temperature where the mechanical properties change greatly. For convenience, -1.0°C can be defined as the boundary for warm frozen soils.

Effect of cooling rate on sperm quality of cryopreserved Andalusian donkey spermatozoa.

PubMed

Demyda-Peyrás, S; Bottrel, M; Acha, D; Ortiz, I; Hidalgo, M; Carrasco, J J; Gómez-Arrones, V; Gósalvez, J; Dorado, J

2018-06-01

The aim of this study was to evaluate the effect of different cooling rates on post-thaw quality of cryopreserved donkey spermatozoa. Eighteen ejaculates from six adult Andalusian donkeys (three ejaculates per donkey) were collected using an artificial vagina. Pooled semen samples (two ejaculates per pool) were divided into three aliquots, and frozen in Gent freezing extender using three different cryopreservation protocols (P): P1 (conventional slow freezing, as control): semen pre-cooled in an Equitainer for 2 h and frozen in liquid nitrogen (LN 2 ) vapour; P2 (controlled pre-freeze cooling rate): semen pre-cooled at a controlled rate for 73 min and frozen in LN 2 vapour; and P3 (rapid freezing) semen frozen immediately in LN 2 vapour. After thawing at 37 °C for 30 s, semen samples were assessed for motility, morphology, acrosome and plasma membrane integrity; spermatozoa were also tested for DNA integrity. Significant (P < 0.01) differences were found between the cryopreservation protocols for all sperm parameters evaluated, except for DNA integrity. Semen samples frozen using P2 showed significantly (P < 0.01) higher values for sperm motility, morphology, sperm membrane integrity, and acrosome integrity. On the contrary, P3 reduced sperm motility (P < 0.01) and increased the percentage of spermatozoa with damaged plasma membrane (P < 0.001). In our study, we demonstrated that the sperm of Andalusian donkey is particularly sensitive to the cooling rate used before freezing. Furthermore, Andalusian donkey semen can be successfully cryopreserved using controlled cooling rates combined with freezing in LN 2 vapour. Copyright © 2018 Elsevier B.V. All rights reserved.

Spawn viability in edible mushrooms after freezing in liquid nitrogen without a cryoprotectant.

PubMed

Mata, Gerardo; Pérez-Merlo, Rosalía

2003-08-01

Five strains of edible mushrooms (Lentinula boryana, Lentinula edodes, Pleurotus djamor, Pleurotus pulmonarius, and Volvariella volvacea) were studied. Spawn were prepared from sorghum seeds and then incubated for 14 days under optimum conditions for each species. Once covered by mycelia, the sorghum seeds were placed in polycarbonate vials for freezing in liquid nitrogen. The effect of adding a cryoprotective solution before freezing (either 10% glycerol v/v or 5% dimethylsulfoxide v/v) was evaluated as a function of mycelial growth and percent viability. Three main treatments were undertaken: (1) freezing with a glycerol or dimethylsulfoxide cryoprotectant, (2) freezing with water and (3) freezing without cryoprotectant or water. Samples were maintained frozen for a week, after which time they were thawed (10 min at 30 degrees C) and the seeds placed in Petri dishes with a culture medium. A recovery rate of 96.8% was obtained for the total number of samples summed over all strains and treatments. In contrast, 99.2% of the samples frozen without cryoprotectant were recovered. The recovery of frozen mycelia was delayed with respect to a control group, which was not frozen. However, no difference was observed in percent recovery and mycelial diameter when a new series of spawn was prepared from mycelia that had been previously frozen. Results obtained from this experiment demonstrate that an adequate recovery of mycelia can be obtained without using a cryoprotectant. This capacity might enable large quantities of commercial mushroom strains to be handled at reduced production costs. It is suggested that the mycelia survived freezing without cryoprotectants because they were embedded and protected within the sorghum seeds used to elaborate the spawn.

Quality Evaluation of Pork with Various Freezing and Thawing Methods

PubMed Central

2014-01-01

In this study, the physicochemical and sensory quality characteristics due to the influence of various thawing methods on electro-magnetic and air blast frozen pork were examined. The packaged pork samples, which were frozen by air blast freezing at −45℃ or electro-magnetic freezing at −55℃, were thawed using 4 different methods: refrigeration (4±1℃), room temperature (RT, 25℃), cold water (15℃), and microwave (2450 MHz). Analyses were carried out to determine the drip and cooking loss, water holding capacity (WHC), moisture content and sensory evaluation. Frozen pork thawed in a microwave indicated relatively less thawing loss (0.63-1.24%) than the other thawing methods (0.68-1.38%). The cooking loss after electro-magnetic freezing indicated 37.4% by microwave thawing, compared with 32.9% by refrigeration, 36.5% by RT, and 37.2% by cold water in ham. The thawing of samples frozen by electro-magnetic freezing showed no significant differences between the methods used, while the moisture content was higher in belly thawed by microwave (62.0%) after electro-magnetic freezing than refrigeration (54.8%), RT (61.3%), and cold water (61.1%). The highest overall acceptability was shown for microwave thawing after electro-magnetic freezing but there were no significant differences compared to that of the other samples. PMID:26761493

Impact of water extractable arabinoxylan from rye bran on the frozen steamed bread dough quality.

PubMed

Wang, Pei; Tao, Han; Jin, Zhengyu; Xu, Xueming

2016-06-01

Impact of water extractable arabinoxylan from rye bran on frozen steamed bread dough quality was investigated in terms of the bread characteristics, ice crystallization, yeast activity as well as the gluten molecular weight distribution and glutenin macropolymer content in the present study. Results showed that water extractable arabinoxylan significantly improved bread characteristics during the 60-day frozen storage. Less water was crystallized in the water extractable arabinoxylan dough during storage, which could explain the alleviated yeast activity loss. For all the frozen dough samples, more soluble high molecular weight (Mw ≈ 91,000-688,000) and low molecular weight (Mw ≈ 91,000-16,000) proteins were derived from glutenin macropolymer depolymerization. Nevertheless, water extractable arabinoxylan dough developed higher glutenin macropolymer content with lowered level of soluble low molecular weight proteins throughout the storage. This study suggested water extractable arabinoxylan from rye bran had great potential to be served as an effective frozen steamed bread dough improver. Copyright © 2016 Elsevier Ltd. All rights reserved.

Intracytoplasmic sperm injection outcomes with cryopreserved testicular sperm aspiration samples.

PubMed

Roque, M; Valle, M; Marques, F; Sampaio, M; Geber, S

2016-04-01

Intracytoplasmic sperm injection (ICSI) may be performed with testicular frozen-thawed spermatozoa in patients with nonobstructive azoospermia (NOA). Sperm retrieval can be performed in advance of oocyte aspiration, as it may avoid the possibility of no recovery of spermatozoa on the day of oocyte pickup. There are few studies available in the literature concerning the use of frozen-thawed spermatozoa obtained from testicular sperm aspiration (TESA). To evaluate the effects and the outcomes of ICSI with frozen-thawed spermatozoa obtained by TESA, we performed a retrospective analysis of 43 ICSI cycles using frozen-thawed TESA. We obtained acceptable results with a fertilisation rate of 67.9%, an implantation rate (IR) of 17.1%, and clinical and ongoing pregnancy rates of 41.9% and 37.2% respectively. The results of this study suggest that performing ICSI using cryopreserved frozen-thawed testicular spermatozoa with TESA as a first option is a viable, safe, economic and effective method for patients with NOA. © 2015 Blackwell Verlag GmbH.

Effect of freezing of sputum samples on flow cytometric analysis of lymphocyte subsets.

PubMed

Jaksztat, E; Holz, O; Paasch, K; Kelly, M M; Hargreave, F E; Cox, G; Magnussen, H; Jörres, R A

2004-08-01

Sputum samples should be processed shortly after induction to prevent cell degradation. For intermediate storage, freezing of homogenised samples or immediate fixation have been shown to be suitable for cytospins. The aim of this study was to investigate whether freezing or immediate fixation of sputum affect the analysis of lymphocyte subsets by flow cytometry. Selected plugs from 24 sputum samples were homogenised. One aliquot was processed immediately and analysed by flow cytometry. A second aliquot was homogenised, frozen at -20 C after addition of dimethylsulfoxide and stored for a median time of 6 days. In six samples a third aliquot was fixed in formalin after induction and stored for up to 72 h before further processing. Compared to immediate processing, percentages of total lymphocytes and T-suppressor cells were elevated after being frozen, with a minor decrease in the T4/T8 ratio. Proportions of total lymphocytes, T-helper and T-suppressor cells correlated between native and frozen samples, intra-class correlation coefficients being 0.74, 0.85 and 0.70, respectively. The formalin-fixed aliquots could not be analysed with the antibodies used. In conclusion, freezing seems to be a suitable technique to store sputum samples for flow cytometry of CD3, CD4 and CD8 lymphocyte subsets. Its effects were minor compared to the variation between subjects.

Hepatic immunohistochemical localization of the tight junction protein ZO-1 in rat models of cholestasis.

PubMed Central

Anderson, J. M.; Glade, J. L.; Stevenson, B. R.; Boyer, J. L.; Mooseker, M. S.

1989-01-01

Structural alterations in hepatocyte tight junctions accompanying cholestasis were investigated using immunolocalization of ZO-1, the first known protein component of the tight junction. Disruption in the paracellular barrier function of the tight junction has been proposed to allow reflux of bile into the blood. Cholestasis was induced in 210 to 235 g male Sprague-Dawley rats either by five consecutive daily subcutaneous injections of 17-alpha-ethinyl estradiol (0.5 mg/kg/d in propylene glycol) or ligation of the common bile duct for 72 hours. The structural organization of the tight junction was assessed in each model by indirect immunofluorescent and immunoperoxidase staining for ZO-1 on frozen sections of liver and compared with controls. In control, sham-operated, and estradiol-injected animals, ZO-1 localizes in a uniform continuous manner along the margins of the canaliculi. In contrast, bile duct ligation results in the appearance of numerous discontinuities in ZO-1 staining accompanied by dilation or collapse of the lumenal space. Tissue content of the ZO-1 protein, as determined by quantitative immunoblotting, was unaffected in either cholestatic model compared with controls. These findings indicate that the molecular organization of the tight junction can be assessed from immunostaining patterns of ZO-1 in frozen sections of cholestatic livers. Under these experimental conditions, the organization of the tight junction at the level of the ZO-1 protein is altered by bile duct obstruction but not by ethinyl estradiol. Images Figure 1 Figure 2 PMID:2719075

Training effect of the exchange bias in sputter deposited Fe3O4 thin films with varying thickness

NASA Astrophysics Data System (ADS)

Muhammed Shameem, P. V.; Senthil Kumar, M.

2018-07-01

The training effect property of the exchange bias in the reactively sputtered polycrystalline Fe3O4 thin films of varying thicknesses in the range 25-200 nm are studied. Structural studies by X-ray diffraction, X-ray photoelectron spectroscopy and selected area electron diffraction confirm the formation of single phase Fe3O4. The scanning electron spectroscopy images show that the grains are uniformly distributed. All the samples show clear and consistent exchange bias training behaviour due to the dynamics of the spins at the interface of the ferrimagnetic core and the spin glass-like surface of the grains. The analysis of the training effect data of the exchange bias field HE measured at 2 K by using three different models show that the model based on the relaxation of the frozen and rotatable spin components at the interface gives the best description for all the samples. From this model, it is found that the reversible interface spins relax around 7 times faster than the frozen interface spins at 2 K for all the samples and that their relative relaxation rates are independent of the sample thickness. This constancy show that the relative relaxation rates of the interfacial frozen and rotatable spin components is a material dependent property. The frozen component of the interfacial spins of each sample is found to be dominated at the initial stage of the training. A direct equivalence between the HE and remanence asymmetry ME is observed. Above the spin freezing temperature, the training effect measurements at 75 K show that the HE decreases sharply with successive field cycling as compared to the measurements made at 2 K and the HE vanishes after first few cycles.

Comparison of cook loss, shear force, and sensory descriptive profiles of boneless skinless white meat cooked from a frozen or thawed state.

PubMed

Zhuang, Hong; Savage, Elizabeth M

2013-11-01

Four replications were conducted to compare quality measurements, cook loss, shear force, and sensory quality attributes of cooked boneless skinless white meat, broiler breast fillets (pectoralis major) prepared directly from a frozen state or prepared from a thawed state. In each replication, fresh broiler fillets (removed from carcasses 6-8 h postmortem) were procured from a local commercial processing plant and stored in a -20°C freezer until use. On the sensory evaluation date, fillets were cooked to an endpoint temperature of 78°C either directly from the frozen state (thawing during cooking) or after the frozen samples were thawed in a refrigerator (2°C) overnight (thawing before cooking). Cook loss and Warner-Bratzler (WB) shear force were used as indicators for instrumental quality measurements. Sensory quality measurements were conducted by trained descriptive panelists using 0 to 15 universal intensity scales for 8 texture and 10 flavor attributes. Results show that there were no differences (P > 0.05) in measurements for sensory descriptive flavor attributes of cooked fillets between the 2 sample thawing methods, indicating that the sensory flavor profiles of both methods were similar to each other. However, WB shear force (36.98 N), cook loss (21.2%), sensory texture attributes of cohesiveness (intensity score was 5.59), hardness (5.14), rate of breakdown (5.50), and chewiness (5.21) of the breast fillets cooked directly from the frozen state were significantly higher (P < 0.05) than those of the breast meat cooked after being thawed (30.56 N, 19.0%, 5.19, 4.78, 5.29, and 5.02, respectively). These results indicate that cookery directly from frozen boneless skinless white meat can result in different measurement values of cook loss, shear force, and sensory descriptive texture attributes compared with cookery after frozen fillets are thawed.

Detection of Cryptosporidium oocysts in fresh and frozen cattle faeces: comparison of three methods.

PubMed

Brook, E J; Christley, R M; French, N P; Hart, C A

2008-01-01

The aim of this study was to compare the performance of three commonly used screening tests for Cryptosporidium oocysts in fresh and frozen cattle faeces. Twenty-nine freshly voided faecal samples were collected from calves from three farms in the northwest of England. Three diagnostic tests for Cryptosporidium were carried out on each sample both before and after freezing - the modified Ziehl-Neelsen (MZN) and auramine phenol (APh) stains and a commercial enzyme immunoassay (EIA) kit, the ProSpecT Cryptosporidium Microplate assay (Remel, Lenexa, KS). Twelve samples were deemed positive by the reference standard (polymerase chain reaction, PCR). There were some discrepancies between the results of the screening tests and the levels of agreement were quantified. The sensitivity and specificity of each method was determined, with PCR as the gold standard. Sensitivity and specificity of the MZN stain was optimized when samples with fewer than two oocyst-like bodies were classified as negative. All three screening methods used were effective in detecting Cryptosporidium infection in both fresh and frozen calf faeces. This study has highlighted the value of determining characteristics of tests used for diagnosis and epidemiological studies.

Influence of freeze-thawing on hyaluronic acid binding of human spermatozoa.

PubMed

Nijs, Martine; Creemers, Eva; Cox, Annemie; Janssen, Mia; Vanheusden, Elke; Castro-Sanchez, Yovanna; Thijs, Herbert; Ombelet, Willem

2009-08-01

Mature human spermatozoa have at least three specific hyaluronic acid (HA) binding proteins present on their sperm membrane. These receptors play a role in the acrosome reaction, hyaluronidase activity, hyaluronan-mediated motility and sperm-zona and sperm-oolemmal binding. Cryopreservation of spermatozoa can cause ultrastructural and even molecular damage. The aim of this study was to investigate if HA binding receptors of human spermatozoa remain functional after freeze-thawing. Forty patients were enrolled in the study. Semen samples were analysed before and after cryopreservation. Parameters analysed included concentration, motility, morphology and hyaluronan binding. Samples were frozen in CBS straws using a glycerol-glucose-based cryoprotectant. HA binding was studied using the sperm-hyaluronan binding assay. Freeze-thawing resulted in a significant decline in motility: the percentage of motile spermatozoa reduced from 50.6 to 30.3% (P < 0.001). HA binding properties of frozen-thawed spermatozoa remained unchanged after the freeze-thawing process: 68.5 +/- 17.1% spermatozoa of the neat sample were bound to HA, as were 71.3 +/- 20.4 of the frozen-thawed sample. This study indicates that freeze-thawing did not alter the functional hyaluronan binding sites of mature motile spermatozoa, and therefore will not alter their fertilizing potential.

An evaluation of Brix refractometry instruments for measurement of colostrum quality in dairy cattle.

PubMed

Bielmann, V; Gillan, J; Perkins, N R; Skidmore, A L; Godden, S; Leslie, K E

2010-08-01

Acquisition of high quality colostrum is an important factor influencing neonatal calf health. Many methods have been used to assess the Ig concentration of colostrum; however, improved, validated evaluation tools are needed. The aims of this study were to evaluate both optical and digital Brix refractometer instruments for the measurement of Ig concentration of colostrum as compared with the gold standard radial immunodiffusion assay laboratory assessment and to determine the correlation between Ig measurements taken from fresh and frozen colostrum samples for both Brix refractometer instruments. This research was completed using 288 colostrum samples from 3 different farms. It was concluded that the optical and digital Brix refractometers were highly correlated for both fresh and frozen samples (r=0.98 and r=0.97, respectively). Correlation between both refractometer instruments for fresh and frozen samples and the gold standard radial immunodiffusion assay were determined to be very similar, with a correlation coefficient between 0.71 and 0.74. Both instruments exhibited excellent test characteristics, indicating an appropriate cut-off point of 22% Brix score for the identification of good quality colostrum. Copyright (c) 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

The Change of Total Anthocyanins in Blueberries and Their Antioxidant Effect After Drying and Freezing

PubMed Central

Srzednicki, George

2004-01-01

This study examined the effects of freezing, storage, and cabinet drying on the anthocyanin content and antioxidant activity of blueberries (Vaccinium corymbosum L). Fresh samples were stored for two weeks at 5°C while frozen samples were kept for up to three months at −20°C. There were two drying treatments, one including osmotic pretreatment followed by cabinet drying and the other involving only cabinet drying. Total anthocyanins found in fresh blueberries were 7.2 ± 0.5 mg/g dry matter, expressed as cyanidin 3-rutinoside equivalents. In comparison with fresh samples, total anthocyanins in untreated and pretreated dried blueberries were significantly reduced to 4.3 ± 0.1 mg/g solid content, 41% loss, and 3.7 ± 0.2 mg/g solid content, 49% loss, respectively. Osmotic treatment followed by a thermal treatment had a greater effect on anthocyanin loss than the thermal treatment alone. In contrast, the frozen samples did not show any significant decrease in anthocyanin level during three months of storage. Measurement of the antioxidant activity of anthocyanin extracts from blueberries showed there was no significant difference between fresh, dried, and frozen blueberries. PMID:15577185

Biologically active antimicrobial and antioxidant substances in the Helianthus annuus L. bee pollen.

PubMed

Fatrcová-Šramková, Katarína; Nôžková, Janka; Máriássyová, Magda; Kačániová, Miroslava

2016-01-01

The objective of this study was to measure the content of flavonoids, polyphenols, and carotenoids in the Helianthus annuus L. bee pollen. It was also to evaluate the ability of the dried, frozen, and freeze-dried extracts of sunflower (H. annuus) pollen, its scavenged free radicals and reducing action. Another aim of this study was to investigate the antimicrobial in vitro action of the H. annuus pollen extracts against the Gram-positive and Gram-negative bacteria and fungi. All pollen extracts showed medium antiradical activity and reductive ability. The most effective was the freeze-dried extract in both evaluation systems. The evaluation of the protective effects of DNA using a biosensor showed an opposite trending-frozen ˃ dried ˃ freeze-dried pollen. For the evaluation of antiradical activity, the DPPH method was used, and reductive ability was assessed by means of phosphomolybdic complex formation. The comparison of the polyphenols content shows higher values in freeze-dried bee pollen than in the dried and frozen pollen. The highest content of flavonoids was found in the frozen samples and the most carotenoids were present in the dried samples. In our study, the best antibacterial effects of the dried sunflower bee pollen extracts were found against Paenibacillus larvae, Pseudomonas aeruginosa, and Enterococcus raffinosus. The best inhibitory properties of the frozen sunflower bee pollen extracts were found against Escherichia coli, Pseudomonas aeruginosa, and Paenibacillus larvae. Very good inhibitory effects of freeze-dried sunflower bee pollen were found against Paenibacillus larvae, Brochotrix thermosphacta, and Enterococcus raffinosus. The best antifungal activity of the sunflower bee pollen was found in the frozen bee pollen extracts against Aspergillus ochraceus and freeze-dried bee pollen extracts against Aspergillus niger.

Effect of different procedures of ejaculate collection, extenders and packages on DNA integrity of boar spermatozoa following freezing-thawing.

PubMed

Fraser, L; Strzezek, J

2007-06-01

Whole ejaculate or sperm-rich fraction, collected from four sexually mature boars, was frozen in an extender containing lactose-hen egg yolk with glycerol (lactose-HEY-G) or extender containing lactose, lyophilized lipoprotein fractions isolated from ostrich egg yolk and glycerol (lactose-LPFo-G), and Orvus Es Paste, respectively. The sperm samples were also frozen in a standard boar semen extender (Kortowo-3), without the addition of cryoprotective substances. Sperm DNA integrity was assessed using a modified neutral comet assay. Sperm characteristics such as motility, plasma membrane integrity (SYBR-14/PI), mitochondrial function (rhodamine 123) and acrosome integrity were monitored. Freezing-thawing caused a significant increase (P<0.05) in sperm DNA fragmentation, irrespective of the procedures of ejaculate collection and extender type. Sperm DNA fragmentation was significantly lower (P<0.05) in the whole ejaculate compared with the sperm-rich fraction, indicating that spermatozoa maintained in the whole seminal plasma prior to its removal for freezing-thawing procedure were less vulnerable to cryo-induced DNA fragmentation. Furthermore, spermatozoa frozen in lactose-HEY-G or lactose-LPFo-G extender exhibited lower (P<0.05) DNA fragmentation than those frozen in the absence of cryoprotective substances. The levels of sperm DNA damage, as expressed by comet tail length and tail moment values, were significantly higher (P<0.05) in sperm samples frozen in the absence of cryoprotective substances. The deterioration in post-thaw sperm DNA integrity was concurrent with reduced sperm characteristics. It can be suggested that evaluation of DNA integrity, coupled with different sperm characteristics such as motility, plasma membrane integrity and mitochondrial function, may aid in determining the quality of frozen-thawed boar semen.

Inactivation kinetics of Vibrio vulnificus in phosphate-buffered saline at different freezing and storage temperatures and times.

PubMed

Seminario, Diana M; Balaban, Murat O; Rodrick, Gary

2011-03-01

Vibrio vulnificus (Vv) is a pathogen that can be found in raw oysters. Freezing can reduce Vv and increase the shelf life of oysters. The objective of this study was to develop predictive inactivation kinetic models for pure cultures of Vv at different frozen storage temperatures and times. Vv was diluted in phosphate-buffered saline (PBS) to obtain about 10(7) CFU/mL. Samples were frozen at -10, -35, and -80 °C (different freezing rates), and stored at different temperatures. Survival of Vv was followed after freezing and storage at -10 °C (0, 3, 6, and 9 d) and at -35 and -80 °C (every week for 6 wk). For every treatment, time-temperature data was obtained using thermocouples in blank vials. Predictive models were developed using first-order, Weibull and Peleg inactivation kinetics. Different freezing temperatures did not significantly (α = 0.05) affect survival of Vv immediately after freezing. The combined effect of freezing and 1 wk frozen storage resulted in 1.5, 2.6, and 4.9 log10 reductions for samples stored at -80, -35, and -10 °C, respectively. Storage temperature was the critical parameter in survival of Vv. A modified Weibull model successfully predicted Vv survival during frozen storage: log10 Nt = log 10No - 1.22 - ([t/10{-1.163-0.0466T}][0.00025T(2) + 0.049325]). N(o) and N(t) are initial and time t (d) survival counts, T is frozen storage temperature, Celsius degree. Vibrio vulnificus can be inactivated by freezing. Models to predict survival of V. vulnificus at different freezing temperatures and times were developed. This is the first step towards the prediction of V. vulnificus related safety of frozen oysters.

Effect of Holder pasteurization and frozen storage on macronutrients and energy content of breast milk.

PubMed

García-Lara, Nadia Raquel; Vieco, Diana Escuder; De la Cruz-Bértolo, Javier; Lora-Pablos, David; Velasco, Noelia Ureta; Pallás-Alonso, Carmen Rosa

2013-09-01

The aim of this study was to explore the effect of Holder pasteurization and frozen storage at -20°C after pasteurization on fat, total nitrogen, lactose, and energy content of breast milk. Both procedures are routinely practiced in human milk banks. A total of 34 samples of frozen breast milk, donated by 28 women, were collected. Once thawed, an aliquot of each sample was analyzed before pasteurization; the remaining milk was pasteurized (Holder method) and split into 8 aliquots. One aliquot was analyzed after pasteurization and the remainder frozen at -20°C and analyzed 30, 60, 90, 120, and 180 days later. For every aliquot, fat, total nitrogen, lactose, and energy content were determined using the device human Milk Analyzer. We observed a significant reduction in fat (3.5%; -0.17 (-0.29; -0.04) g/dL) and energy content (2.8%; -2.03 (-3.60; -0.46) g/dL) after pasteurization. A significant decrease over time was observed for fat, lactose and energy content. No significant changes were observed for nitrogen content. Mean differences between day 0 postpasteurization and day 180 were -0.13 (-0.21; -0.06) g/dL for fat, -0.08 (-0.13; -0.03) g/dL for lactose, and -1.55 (-2.38; -0.71) kcal/dL for energy content. The relative decreases were 2.8%, 1.7%, and 2.2%, respectively. Overall (postpasteurization + frozen storage), a 6.2% and 5% decrease were observed for fat and energy, respectively. Holder pasteurization decreased fat and energy content of human milk. Frozen storage at -20°C of pasteurized milk significantly reduced fat, lactose, and energy content of human milk.

Hepatic stellate cell-targeted imatinib nanomedicine versus conventional imatinib: A novel strategy with potent efficacy in experimental liver fibrosis.

PubMed

El-Mezayen, Nesrine S; El-Hadidy, Wessam F; El-Refaie, Wessam M; Shalaby, Th I; Khattab, Mahmoud M; El-Khatib, Aiman S

2017-11-28

Liver fibrosis is a global health problem without approved treatment. Imatinib inhibits two key profibrotic pathways; platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-β) and thus can be used to treat liver fibrosis. However, conventional imatinib therapy is hampered by low concentration at target tissue and increased toxicity to other tissues especially heart, lung and liver. Since hepatic stellate cells (HSCs) are the main contributors to liver fibrosis pathogenesis and sole hepatic vitamin A (V A ) storage cells, they can be actively targeted by coupling liposomes to V A . In this study, novel V A -coupled imatinib-loaded liposomes (ILC) were prepared and optimized regarding V A -coupling efficiency, imatinib entrapment efficiency, and particle size. Preferential accumulation of the selected formula in liver was proved by tracing intraperitoneally (i.p.)-injected V A -coupled liposomes loaded with Nile Red (LCNR) to rats with CCl 4 -induced liver fibrosis using live animal imaging. Co-localization of LCNR with immunofluorescently-labeled PDGFR-β in frozen liver tissue sections confirmed HSCs targeting. ILC bio-distribution, following single i.p. injection, revealed 13.5 folds higher hepatic accumulation than conventional imatinib in addition to limited bio-distribution to other organs including heart and lung reflecting diminished adverse effects. ILC therapy resulted in a potent inhibition of phosphorylated PDGFR-β expression when compared to conventional imatinib. Subsequently, there was a statistically significant improvement in liver function tests and reversal of hepatotoxicity along with liver fibrosis. Anti-fibrotic effect was evident from histopathologic Ishak score reduction as well as normalization of the level of profibrotic mediators (hydroxyproline, TGF-B and matrix metalloproteinase-2). Thus, HSC-targeted imatinib therapy shows outstanding anti-fibrotic effects with reduced cytotoxicity compared to conventional imatinib. It can represent a promising novel approach for liver fibrosis treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

Influence of Freezing and Storage Procedure on Human Urine Samples in NMR-Based Metabolomics

PubMed Central

Rist, Manuela J.; Muhle-Goll, Claudia; Görling, Benjamin; Bub, Achim; Heissler, Stefan; Watzl, Bernhard; Luy, Burkhard

2013-01-01

It is consensus in the metabolomics community that standardized protocols should be followed for sample handling, storage and analysis, as it is of utmost importance to maintain constant measurement conditions to identify subtle biological differences. The aim of this work, therefore, was to systematically investigate the influence of freezing procedures and storage temperatures and their effect on NMR spectra as a potentially disturbing aspect for NMR-based metabolomics studies. Urine samples were collected from two healthy volunteers, centrifuged and divided into aliquots. Urine aliquots were frozen either at −20 °C, on dry ice, at −80 °C or in liquid nitrogen and then stored at −20 °C, −80 °C or in liquid nitrogen vapor phase for 1–5 weeks before NMR analysis. Results show spectral changes depending on the freezing procedure, with samples frozen on dry ice showing the largest deviations. The effect was found to be based on pH differences, which were caused by variations in CO2 concentrations introduced by the freezing procedure. Thus, we recommend that urine samples should be frozen at −20 °C and transferred to lower storage temperatures within one week and that freezing procedures should be part of the publication protocol. PMID:24957990

Influence of Freezing and Storage Procedure on Human Urine Samples in NMR-Based Metabolomics.

PubMed

Rist, Manuela J; Muhle-Goll, Claudia; Görling, Benjamin; Bub, Achim; Heissler, Stefan; Watzl, Bernhard; Luy, Burkhard

2013-04-09

It is consensus in the metabolomics community that standardized protocols should be followed for sample handling, storage and analysis, as it is of utmost importance to maintain constant measurement conditions to identify subtle biological differences. The aim of this work, therefore, was to systematically investigate the influence of freezing procedures and storage temperatures and their effect on NMR spectra as a potentially disturbing aspect for NMR-based metabolomics studies. Urine samples were collected from two healthy volunteers, centrifuged and divided into aliquots. Urine aliquots were frozen either at -20 °C, on dry ice, at -80 °C or in liquid nitrogen and then stored at -20 °C, -80 °C or in liquid nitrogen vapor phase for 1-5 weeks before NMR analysis. Results show spectral changes depending on the freezing procedure, with samples frozen on dry ice showing the largest deviations. The effect was found to be based on pH differences, which were caused by variations in CO2 concentrations introduced by the freezing procedure. Thus, we recommend that urine samples should be frozen at -20 °C and transferred to lower storage temperatures within one week and that freezing procedures should be part of the publication protocol.

Use of eculizumab and plasma exchange in successful combined liver-kidney transplantation in a case of atypical HUS associated with complement factor H mutation.

PubMed

Tran, Ha; Chaudhuri, Abanti; Concepcion, Waldo; Grimm, Paul C

2014-03-01

Atypical hemolytic uremic syndrome (aHUS) evolves into end-stage renal failure in nearly half of affected patients and is associated with defective regulation of the alternative complement pathway. Patients with a complement factor H (CFH) mutation have a 30-100% risk of graft loss due to aHUS recurrence or graft thrombosis. Since CFH is produced predominantly by the liver, combined liver-kidney transplant is a curative treatment option. One major unexpected risk includes liver failure secondary to uncontrolled complement activation. We report a successful combined liver-kidney transplantation with perioperative plasma exchange and use of the humanized anti-C5 monoclonal antibody eculizumab. An 11-month-old female presented with oliguric renal failure after 3 weeks of flu-like symptoms in the absence of diarrhea. Following the identification of Escherichia coli 0157:H7 in her stool, she was discharged home on peritoneal dialysis with a diagnosis of Shiga toxin-associated HUS. Three months later, she developed severe anemia, thrombocytopenia, and neurological involvement. aHUS was diagnosed and confirmed, and genetic testing revealed a mutation in CFH SCR20. Once donor organs became available, she received preoperative plasma exchange followed by eculizumab infusion with intra-operative fresh frozen plasma prior to combined liver-kidney transplant. At 19 months post-transplant, she continues to have excellent allograft and liver function without signs of disease recurrence. Perioperative use of eculizumab in conjunction with plasma exchange during simultaneous liver-kidney transplant can be used to inhibit terminal complement activity, thereby optimizing successful transplantation by reducing the risk of graft thrombosis.

Comparison of Collection Methods for Fecal Samples in Microbiome Studies

PubMed Central

Vogtmann, Emily; Chen, Jun; Amir, Amnon; Shi, Jianxin; Abnet, Christian C.; Nelson, Heidi; Knight, Rob; Chia, Nicholas; Sinha, Rashmi

2017-01-01

Prospective cohort studies are needed to assess the relationship between the fecal microbiome and human health and disease. To evaluate fecal collection methods, we determined technical reproducibility, stability at ambient temperature, and accuracy of 5 fecal collection methods (no additive, 95% ethanol, RNAlater Stabilization Solution, fecal occult blood test cards, and fecal immunochemical test tubes). Fifty-two healthy volunteers provided fecal samples at the Mayo Clinic in Rochester, Minnesota, in 2014. One set from each sample collection method was frozen immediately, and a second set was incubated at room temperature for 96 hours and then frozen. Intraclass correlation coefficients (ICCs) were calculated for the relative abundance of 3 phyla, 2 alpha diversity metrics, and 4 beta diversity metrics. Technical reproducibility was high, with ICCs for duplicate fecal samples between 0.64 and 1.00. Stability for most methods was generally high, although the ICCs were below 0.60 for 95% ethanol in metrics that were more sensitive to relative abundance. When compared with fecal samples that were frozen immediately, the ICCs were below 0.60 for the metrics that were sensitive to relative abundance; however, the remaining 2 alpha diversity and 3 beta diversity metrics were all relatively accurate, with ICCs above 0.60. In conclusion, all fecal sample collection methods appear relatively reproducible, stable, and accurate. Future studies could use these collection methods for microbiome analyses. PMID:27986704

Indocyanine green plasma disappearance rate during the anhepatic phase of orthotopic liver transplantation.

PubMed

Bruegger, Lukas; Studer, Peter; Schmid, Stefan W; Pestel, Gunther; Reichen, Juerg; Seiler, Christian; Candinas, Daniel; Inderbitzin, Daniel

2008-01-01

Non-invasive pulse spectrophotometry to measure indocyanine green (ICG) elimination correlates well with the conventional invasive ICG clearance test. Nevertheless, the precision of this method remains unclear for any application, including small-for-size liver remnants. We therefore measured ICG plasma disappearance rate (PDR) during the anhepatic phase of orthotopic liver transplantation using pulse spectrophotometry. Measurements were done in 24 patients. The median PDR after exclusion of two outliers and two patients with inconstant signal was 1.55%/min (95% confidence interval [CI]=0.8-2.2). No correlation with patient age, gender, body mass, blood loss, administration of fresh frozen plasma, norepinephrine dose, postoperative albumin (serum), or difference in pre and post transplant body weight was detected. In conclusion, we found an ICG-PDR different from zero in the anhepatic phase, an overestimation that may arise in particular from a redistribution into the interstitial space. If ICG pulse spectrophotometry is used to measure functional hepatic reserve, the verified average difference from zero (1.55%/min) determined in our study needs to be taken into account.

Implications of the pH and temperature of diluted, cooled boar semen on fresh and frozen-thawed sperm motility characteristics.

PubMed

Purdy, P H; Tharp, N; Stewart, T; Spiller, S F; Blackburn, H D

2010-10-15

Boar semen is typically collected, diluted and cooled for AI use over numerous days, or frozen immediately after shipping to capable laboratories. The storage temperature and pH of the diluted, cooled boar semen could influence the fertility of boar sperm. Therefore, the purpose of this study was to determine the effects of pH and storage temperature on fresh and frozen-thawed boar sperm motility end points. Semen samples (n = 199) were collected, diluted, cooled and shipped overnight to the National Animal Germplasm Program laboratory for freezing and analysis from four boar stud facilities. The temperature, pH and motility characteristics, determined using computer automated semen analysis, were measured at arrival. Samples were then cryopreserved and post-thaw motility determined. The commercial stud was a significant source of variation for mean semen temperature and pH, as well as total and progressive motility, and numerous other sperm motility characteristics. Based on multiple regression analysis, pH was not a significant source of variation for fresh or frozen-thawed boar sperm motility end points. However, significant models were derived which demonstrated that storage temperature, boar, and the commercial stud influenced sperm motility end points and the potential success for surviving cryopreservation. We inferred that maintaining cooled boar semen at approximately 16 °C during storage will result in higher fresh and frozen-thawed boar sperm quality, which should result in greater fertility. Copyright © 2010 Elsevier Inc. All rights reserved.

7 CFR 91.20 - Shipping.

Code of Federal Regulations, 2013 CFR

2013-01-01

... refrigerated samples should contain ice or ice packs to maintain temperatures of 0° to 5 °C, unless a different temperature is required for the sample to be tested. (d) Containers for frozen samples should contain dry ice...

7 CFR 91.20 - Shipping.

Code of Federal Regulations, 2012 CFR

2012-01-01

... refrigerated samples should contain ice or ice packs to maintain temperatures of 0° to 5 °C, unless a different temperature is required for the sample to be tested. (d) Containers for frozen samples should contain dry ice...

7 CFR 91.20 - Shipping.

Code of Federal Regulations, 2014 CFR

2014-01-01

... refrigerated samples should contain ice or ice packs to maintain temperatures of 0° to 5 °C, unless a different temperature is required for the sample to be tested. (d) Containers for frozen samples should contain dry ice...

7 CFR 91.20 - Shipping.

Code of Federal Regulations, 2011 CFR

2011-01-01

... refrigerated samples should contain ice or ice packs to maintain temperatures of 0° to 5 °C, unless a different temperature is required for the sample to be tested. (d) Containers for frozen samples should contain dry ice...

7 CFR 91.20 - Shipping.

Code of Federal Regulations, 2010 CFR

2010-01-01

... refrigerated samples should contain ice or ice packs to maintain temperatures of 0° to 5 °C, unless a different temperature is required for the sample to be tested. (d) Containers for frozen samples should contain dry ice...

High-Explosive Cratering a Frozen and Unfrozen Soils in Alaska

DTIC Science & Technology

1980-02-01

SAMPLES I DEPTH TO .... T 24 DATE HOLEIROUN - S]T AR ED adO~ ________ F __24_ad_2____________7 EL . ITHOE (4IQ1 ~.0oioy Section ChlofRrndotboelI a...MotorIoI. Bronco Dno 20 f t.D. FREDRICKSON DEPTH %WATER SAI.IPLE SOIL MAAX FEET .4CtENT NO LEGEND CLASSIFICATION IZE F G i Silty Sandy Gravel Brown, Frozen

Annealing to optimize the primary drying rate, reduce freezing-induced drying rate heterogeneity, and determine T(g)' in pharmaceutical lyophilization.

PubMed

Searles, J A; Carpenter, J F; Randolph, T W

2001-07-01

In a companion paper we show that the freezing of samples in vials by shelf-ramp freezing results in significant primary drying rate heterogeneity because of a dependence of the ice crystal size on the nucleation temperature during freezing.1 The purpose of this study was to test the hypothesis that post-freezing annealing, in which the product is held at a predetermined temperature for a specified duration, can reduce freezing-induced heterogeneity in sublimation rates. In addition, we test the impact of annealing on primary drying rates. Finally, we use the kinetics of relaxations during annealing to provide a simple measurement of T(g)', the glass transition temperature of the maximally freeze-concentrated amorphous phase, under conditions and time scales most appropriate for industrial lyophilization cycles. Aqueous solutions of hydroxyethyl starch (HES), sucrose, and HES:sucrose were either frozen by placement on a shelf while the temperature was reduced ("shelf-ramp frozen") or by immersion into liquid nitrogen. Samples were then annealed for various durations over a range of temperatures and partially lyophilized to determine the primary drying rate. The morphology of fully dried liquid nitrogen-frozen samples was examined using scanning electron microscopy. Annealing reduced primary drying rate heterogeneity for shelf-ramp frozen samples, and resulted in up to 3.5-fold increases in the primary drying rate. These effects were due to increased ice crystal sizes, simplified amorphous structures, and larger and more numerous holes on the cake surface of annealed samples. Annealed HES samples dissolved slightly faster than their unannealed counterparts. Annealing below T(g)' did not result in increased drying rates. We present a simple new annealing-lyophilization method of T(g)' determination that exploits this phenomenon. It can be carried out with a balance and a freeze-dryer, and has the additional advantage that a large number of candidate formulations can be evaluated simultaneously.

High-speed shaking of frozen blood clots for extraction of human and malaria parasite DNA.

PubMed

Lundblom, Klara; Macharia, Alex; Lebbad, Marianne; Mohammed, Adan; Färnert, Anna

2011-08-08

Frozen blood clots remaining after serum collection is an often disregarded source of host and pathogen DNA due to troublesome handling and suboptimal outcome. High-speed shaking of clot samples in a cell disruptor manufactured for homogenization of tissue and faecal specimens was evaluated for processing frozen blood clots for DNA extraction. The method was compared to two commercial clot protocols based on a chemical kit and centrifugation through a plastic sieve, followed by the same DNA extraction protocol. Blood clots with different levels of parasitaemia (1-1,000 p/μl) were prepared from parasite cultures to assess sensitivity of PCR detection. In addition, clots retrieved from serum samples collected within two epidemiological studies in Kenya (n = 630) were processed by high speed shaking and analysed by PCR for detection of malaria parasites and the human α-thalassaemia gene. High speed shaking succeeded in fully dispersing the clots and the method generated the highest DNA yield. The level of PCR detection of P. falciparum parasites and the human thalassaemia gene was the same as samples optimally collected with an anticoagulant. The commercial clot protocol and centrifugation through a sieve failed to fully dissolve the clots and resulted in lower sensitivity of PCR detection. High speed shaking was a simple and efficacious method for homogenizing frozen blood clots before DNA purification and resulted in PCR templates of high quality both from humans and malaria parasites. This novel method enables genetic studies from stored blood clots.

Analytical study of comet nucleus samples

NASA Technical Reports Server (NTRS)

Albee, A. L.

1989-01-01

Analytical procedures for studying and handling frozen (130 K) core samples of comet nuclei are discussed. These methods include neutron activation analysis, x ray fluorescent analysis and high resolution mass spectroscopy.

A preliminary result of three-dimensional microarray technology to gene analysis with endoscopic ultrasound-guided fine-needle aspiration specimens and pancreatic juices

PubMed Central

2010-01-01

Background Analysis of gene expression and gene mutation may add information to be different from ordinary pathological tissue diagnosis. Since samples obtained endoscopically are very small, it is desired that more sensitive technology is developed for gene analysis. We investigated whether gene expression and gene mutation analysis by newly developed ultra-sensitive three-dimensional (3D) microarray is possible using small amount samples from endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) specimens and pancreatic juices. Methods Small amount samples from 17 EUS-FNA specimens and 16 pancreatic juices were obtained. After nucleic acid extraction, the samples were amplified with labeling and analyzed by the 3D microarray. Results The analyzable rate with the microarray was 46% (6/13) in EUS-FNA specimens of RNAlater® storage, and RNA degradations were observed in all the samples of frozen storage. In pancreatic juices, the analyzable rate was 67% (4/6) in frozen storage samples and 20% (2/10) in RNAlater® storage. EUS-FNA specimens were classified into cancer and non-cancer by gene expression analysis and K-ras codon 12 mutations were also detected using the 3D microarray. Conclusions Gene analysis from small amount samples obtained endoscopically was possible by newly developed 3D microarray technology. High quality RNA from EUS-FNA samples were obtained and remained in good condition only using RNA stabilizer. In contrast, high quality RNA from pancreatic juice samples were obtained only in frozen storage without RNA stabilizer. PMID:20416107

Effects of yeast, fermentation time, and preservation methods on tarhana.

PubMed

Gurbuz, Ozan; Gocmen, Duygu; Ozmen, Nese; Dagdelen, Fatih

2010-01-01

The physicochemical properties of tarhana soup produced with different dough treatments, fermentation times, and preservation methods were examined. Tarhana doughs were prepared with yogurt (control) or baker's yeast (Saccharomyces cerevisiae) and fermented for 3 days. Samples were taken at 24, 48, and 72 hr. Samples were then preserved via one of four methods: sun dried, dried in the shade, vacumn dried, and frozen. Frozen samples produced lower organic acid levels after 72 hr of fermentation in both control (0.68 g/100 g) and yeast (0.61 g/100 g) applications than samples that were dried (0.94 g/100 g control samples; 0.81 g/100 g samples with yeast). Increasing fermentation time resulted in a significant effect on the formation of organic acid in the tarhana (p < .01). At 72 hr of fermentation, total acidity increased 11%, 17%, and 23% for tarhana samples vacumn-dried, sun-dried, and dried in the shade, respectively. Preservation methods also affected the moisture, ash, crude protein, total acidity, pH, salt, fat, reducing sugar levels, and the sensory assestment of tarhana soup (p < .01). Sensory characteristics were not significantly affected by baker's yeast in any of the preservation methods used (p > .01). However, sensory scores for tarhana prepared from the samples dried in a sheltered area showed a reduction in color desireablilty as the fermentation time increased. The soup prepared from frozen tarhana (72 hr fermentation, with yeast) had the highest scores with respect to color, mouth feel, flavor, and overall acceptability. Vacuum-dried samples' scores in these areas were also high in comparison to the two other drying methods.

Comparison of biochemical values in serum and plasma, fresh and frozen plasma, and hemolyzed samples from orange-winged Amazon parrots (Amazona amazonica).

PubMed

Hawkins, Michelle G; Kass, Philip H; Zinkl, Joseph G; Tell, Lisa A

2006-06-01

To the authors' knowledge, on the basis of sample type, storage condition, or hemolysis, differences in serum and plasma biochemical values have not been evaluated in orange-winged Amazon parrots (Amazona amazonica). The purpose of this study was to compare values for biochemical analytes in serum vs plasma, fresh vs frozen plasma, and nonhemolyzed vs hemolyzed samples in orange-winged Amazon parrots. We also compared differences in serum and plasma yield from whole-blood aliquots. Fifteen biochemical analytes were evaluated in paired serum and plasma, fresh and frozen plasma, nonhemolyzed and hemolyzed serum and plasma samples from orange-winged Amazon parrots (n = 10) using a wet reagent analyzer. Hemolysis was assessed qualitatively (visually) and quantitatively (hemoglobin [Hgb] measured spectrophotometrically). Serum and plasma yields from 500-microl whole-blood aliquots were determined from centrifuged samples. Analyte values significantly differed among sample groups, but were still within published reference intervals, with the exception of increases in potassium concentration in markedly hemolyzed serum and plasma samples. Clinically important changes in hemolyzed serum and plasma samples included increases in potassium, phosphorus, and albumin concentrations and lactate dehydrogenase activity. The degree of hemolysis assigned qualitatively did not correlate with quantitative Hgb concentration. A significantly greater yield of plasma (288 +/- 13 microL) than serum (241 +/- 44 microL) was obtained. Significant differences may occur in different sample types, however, only changes in potassium, phosphorus, albumin, and lactate dehydrogenase values in hemolyzed samples were considered clinically relevant. Lack of agreement between qualitative and quantitative Hgb concentration indicates the unreliability of visual estimation. Based on higher sample yield, and lack of clinically relevant differences from serum, plasma is a better sample choice for clinical chemistry analysis in birds.

A Taphonomic Study Exploring the Differences in Decomposition Rate and Manner between Frozen and Never Frozen Domestic Pigs (Sus scrofa).

PubMed

Roberts, Lindsey G; Dabbs, Gretchen R

2015-05-01

This research examined differences in decomposition rate and manner of domestic pig subjects (Sus scrofa) in never frozen (control) and previously frozen (experimental) research conditions. Eight control and experimental subjects were placed in an identical outdoor research environment. Daily quantitative and qualitative measurements were collected: abdominal circumference, total body score (TBS), temperature, photographs, descriptive decomposition stages, and visual observations. Field necropsies were performed at accumulated degree days (ADD) between 50 and 300 (Celsius). Paired samples t-tests of ADD to TBS >3.0, TBS >9.5, and TBS >16.0 indicate the rate of decomposition of experimental subjects was significantly slower than controls at both TBS >3 and >9.5 (p = 0.003 and p = 0.002, respectively). A suite of qualitative indicators of predecomposition freezing is also reported. The differences between experimental and control subjects suggest previously frozen subjects should not be used in taphonomic research, as results do not accurately reflect the "normal" taphonomic condition. © 2015 American Academy of Forensic Sciences.

Presence and dehydration of ikaite, calcium carbonate hexahydrate, in frozen shrimp shell.

PubMed

Mikkelsen, A; Andersen, A B; Engelsen, S B; Hansen, H C; Larsen, O; Skibsted, L H

1999-03-01

Ikaite, calcium carbonate hexahydrate, has by means of X-ray diffraction analyses of frozen samples been identified as the mineral component of the white spots formed in the shell of frozen shrimp during storage. When the shrimp thaw and the shell material is dried and kept at room temperature, ikaite rapidly transforms into a mixture of anhydrous calcium carbonate forms. X-ray diffraction analyses and Raman spectra of synthetic ikaite as well as the dehydration product confirm the assignments, and the rate constant for dehydration is approximately 7 x 10(-)(4) s(-)(1) at ambient temperature. Differential scanning calorimetry showed that dehydration of synthetic ikaite is an entropy-driven, athermal process and confirms that a single first-order reaction is rate-determining. Ikaite is found to be stable in aqueous solution at temperatures below 5 degrees C and in the shell of frozen shrimps but decomposes on thawing to form anhydrous calcium carbonates.

Abdominal intra-compartment syndrome - a non-hydraulic model of abdominal compartment syndrome due to post-hepatectomy hemorrhage in a man with a localized frozen abdomen due to extensive adhesions: a case report.

PubMed

Bressan, Alexsander K; Kirkpatrick, Andrew W; Ball, Chad G

2016-09-15

Postoperative hemorrhage is a significant cause of morbidity and mortality following liver resection. It typically presents early within the postoperative period, and conservative management is possible in the majority of cases. We present a case of late post-hepatectomy hemorrhage associated with overt abdominal compartment syndrome resulting from a localized functional compartment within the abdomen. A 68-year-old white man was readmitted with sudden onset of upper abdominal pain, vomiting, and hemodynamic instability 8 days after an uneventful hepatic resection for metachronous colon cancer metastasis. A frozen abdomen with adhesions due to complicated previous abdominal surgeries was encountered at the first intervention, but the surgery itself and initial recovery were otherwise unremarkable. Prompt response to fluid resuscitation at admission was followed by a computed tomography of his abdomen that revealed active arterial hemorrhage in the liver resection site and hemoperitoneum (estimated volume <2 L). Selective arteriography successfully identified and embolized a small bleeding branch of his right hepatic artery. He remained hemodynamically stable, but eventually developed overt abdominal compartment syndrome. Surgical exploration confirmed a small volume of ascites and blood clots (1.2 L) under significant pressure in his supramesocolic region, restricted by his frozen lower abdomen, which we evacuated. Dramatic improvement in his ventilatory pressure was immediate. His abdomen was left open and a negative pressure device was placed for temporary abdominal closure. The fascia was formally closed after 48 hours. He was discharged home at postoperative day 6. Intra-abdominal pressure and radiologic findings of intra-abdominal hemorrhage should be carefully interpreted in patients with extensive intra-abdominal adhesions. A high index of suspicion and detailed understanding of abdominal compartment mechanics are paramount for the timely diagnosis of abdominal compartment syndrome in these patients. Clinicians should be aware that abnormal anatomy (such as adhesions) coupled with localized pathophysiology (such as hemorrhage) can create a so-named abdominal intra-compartment syndrome requiring extra vigilance to diagnose.

High post-thaw survival of ram sperm after partial freeze-drying.

PubMed

Arav, Amir; Idda, Antonella; Nieddu, Stefano Mario; Natan, Yehudit; Ledda, Sergio

2018-03-14

Recrystallization damages occur when a frozen sample is held at high subzero temperatures and when the warming process is too slow. In this work, ram semen diluted in two different concentrations of sugar solutions (Lyo A consisted of 0.4 M sorbitol and 0.25 M trehalose, and the second, Lyo B composed of 0.26 M sorbitol and 0.165 M trehalose) in egg yolk and Tris medium were compared after freezing 10 μL samples to: (1) - 10, - 25, and - 35 °C and thawing. (2) Freezing to - 10 and - 25 °C, holding for 1 h and then thawing, and (3) freezing to - 10 and - 25 °C and drying for 1 h at these temperatures at a vacuum of 80 mTorr, prior thawing. For drying, we used a new freeze-drying apparatus (Darya, FertileSafe, Israel) having a condensation temperature below - 110 °C and a vacuum pressure of 10-100 mTorr that is reached in less than 10s. Results showed that samples in Lyo B solution frozen at - 25 °C had significantly higher sperm motility in partially freeze-dried samples than frozen samples (46.6 ± 2.8% vs 1.2 ± 2.5%, P < 0.001). Moreover, partially dried samples in Lyo B showed higher motility than Lyo A at - 25 °C (46.6 ± 2.8% vs 35 ± 4%). Cryomicroscopy and low-temperature/low-pressure environmental scanning electronic microscope demonstrated that the amount of the ice crystals present in partially dried samples was lower than in the frozen samples. Holding the sperm at high subzero temperatures is necessary for the primary drying of cells during the freeze-drying process. Rapid freeze-drying can be achieved using this new device, which enables to reduce recrystallization damages.

Freezing and calcium chloride marination effects on beef tenderness and calpastatin activity.

PubMed

Whipple, G; Koohmaraie, M

1992-10-01

Because freezing samples decreases calpastatin activity and the application of exogenous calcium activates the calpain proteolytic system, thereby improving tenderness, the objective of this study was to determine whether freezing would enhance the effects of CaCl2 marination on the tenderness of beef steaks. Longissimus steaks were obtained from 10 beef steers 6 d postmortem. One-half of the steaks were frozen at -30 degrees C for 6 wk. The remaining steaks were treated fresh; one-half were subjected to a 150 mM CaCl2 marinade for 48 h. Frozen steaks were thawed and subjected to the same treatment. Treatments consisted of 1) fresh control, 2) fresh marinated, 3) frozen control, and 4) frozen marinated. Samples were taken before and after treatment (6 and 8 d) for calpastatin activity determination and d 8 for SDS-PAGE. Warner-Bratzler shear force values were measured 8 d postmortem. Data were analyzed using a paired comparison t-test procedure. Results showed that freezing and marination significantly decreased calpastatin activity. A .35-kg improvement (P = .07) in Warner-Bratzler shear force was observed with freezing, whereas a .78-kg improvement (P less than .01) in tenderness was observed with marination. However, prior freezing enhanced the effects of marination. Therefore, the decrease in calpastatin activity seemed to allow greater proteolysis by the calpains with the application of Ca2+. The SDS-PAGE of myofibril preparations indicated that more small polypeptide fragments (28 to 32 kDa) appeared and a 95-kDa fragment was more intense in the marinated samples than in control samples, indicating that proteolysis was enhanced.(ABSTRACT TRUNCATED AT 250 WORDS)

Frozen human cells can record radiation damage accumulated during space flight: mutation induction and radioadaptation.

PubMed

Yatagai, Fumio; Honma, Masamitsu; Takahashi, Akihisa; Omori, Katsunori; Suzuki, Hiromi; Shimazu, Toru; Seki, Masaya; Hashizume, Toko; Ukai, Akiko; Sugasawa, Kaoru; Abe, Tomoko; Dohmae, Naoshi; Enomoto, Shuichi; Ohnishi, Takeo; Gordon, Alasdair; Ishioka, Noriaki

2011-03-01

To estimate the space-radiation effects separately from other space-environmental effects such as microgravity, frozen human lymphoblastoid TK6 cells were sent to the "Kibo" module of the International Space Station (ISS), preserved under frozen condition during the mission and finally recovered to Earth (after a total of 134 days flight, 72 mSv). Biological assays were performed on the cells recovered to Earth. We observed a tendency of increase (2.3-fold) in thymidine kinase deficient (TK(-)) mutations over the ground control. Loss of heterozygosity (LOH) analysis on the mutants also demonstrated a tendency of increase in proportion of the large deletion (beyond the TK locus) events, 6/41 in the in-flight samples and 1/17 in the ground control. Furthermore, in-flight samples exhibited 48% of the ground-control level in TK(-) mutation frequency upon exposure to a subsequent 2 Gy dose of X-rays, suggesting a tendency of radioadaptation when compared with the ground-control samples. The tendency of radioadaptation was also supported by the post-flight assays on DNA double-strand break repair: a 1.8- and 1.7-fold higher efficiency of in-flight samples compared to ground control via non-homologous end-joining and homologous recombination, respectively. These observations suggest that this system can be used as a biodosimeter, because DNA damage generated by space radiation is considered to be accumulated in the cells preserved frozen during the mission, Furthermore, this system is also suggested to be applicable for evaluating various cellular responses to low-dose space radiation, providing a better understanding of biological space-radiation effects as well as estimation of health influences of future space explores. © Springer-Verlag 2010

Variability of measurements of sweat sodium using the regional absorbent-patch method.

PubMed

Dziedzic, Christine E; Ross, Megan L; Slater, Gary J; Burke, Louise M

2014-09-01

There is interest in including recommendations for the replacement of the sodium lost in sweat in individualized hydration plans for athletes. Although the regional absorbent-patch method provides a practical approach to measuring sweat sodium losses in field conditions, there is a need to understand the variability of estimates associated with this technique. Sweat samples were collected from the forearms, chest, scapula, and thigh of 12 cyclists during 2 standardized cycling time trials in the heat and 2 in temperate conditions. Single measure analysis of sodium concentration was conducted immediately by ion-selective electrodes (ISE). A subset of 30 samples was frozen for reanalysis of sodium concentration using ISE, flame photometry (FP), and conductivity (SC). Sweat samples collected in hot conditions produced higher sweat sodium concentrations than those from the temperate environment (P = .0032). A significant difference (P = .0048) in estimates of sweat sodium concentration was evident when calculated from the forearm average (mean ± 95% CL; 64 ± 12 mmol/L) compared with using a 4-site equation (70 ± 12 mmol/L). There was a high correlation between the values produced using different analytical techniques (r2 = .95), but mean values were different between treatments (frozen FP, frozen SC > immediate ISE > frozen ISE; P < .0001). Whole-body sweat sodium concentration estimates differed depending on the number of sites included in the calculation. Environmental testing conditions should be considered in the interpretation of results. The impact of sample freezing and subsequent analytical technique was small but statistically significant. Nevertheless, when undertaken using a standardized protocol, the regional absorbent-patch method appears to be a relatively robust field test.

Rapid detection of frozen-then-thawed minced beef using multispectral imaging and Fourier transform infrared spectroscopy.

PubMed

Ropodi, Athina I; Panagou, Efstathios Z; Nychas, George-John E

2018-01-01

In recent years, fraud detection has become a major priority for food authorities, as fraudulent practices can have various economic and safety consequences. This work explores ways of identifying frozen-then-thawed minced beef labeled as fresh in a rapid, large-scale and cost-effective way. For this reason, freshly-ground beef was purchased from seven separate shops at different times, divided in fifteen portions and placed in Petri dishes. Multi-spectral images and FTIR spectra of the first five were immediately acquired while the remaining were frozen (-20°C) and stored for 7 and 32days (5 samples for each time interval). Samples were thawed and subsequently subjected to similar data acquisition. In total, 105 multispectral images and FTIR spectra were collected which were further analyzed using partial least-squares discriminant analysis and support vector machines. Two meat batches (30 samples) were reserved for independent validation and the remaining five batches were divided in training and test set (75 samples). Results showed 100% overall correct classification for test and external validation MSI data, while FTIR data yielded 93.3 and 96.7% overall correct classification for FTIR test set and external validation set respectively. Copyright © 2017 Elsevier Ltd. All rights reserved.

Deep Sequencing to Identify the Causes of Viral Encephalitis

PubMed Central

Chan, Benjamin K.; Wilson, Theodore; Fischer, Kael F.; Kriesel, John D.

2014-01-01

Deep sequencing allows for a rapid, accurate characterization of microbial DNA and RNA sequences in many types of samples. Deep sequencing (also called next generation sequencing or NGS) is being developed to assist with the diagnosis of a wide variety of infectious diseases. In this study, seven frozen brain samples from deceased subjects with recent encephalitis were investigated. RNA from each sample was extracted, randomly reverse transcribed and sequenced. The sequence analysis was performed in a blinded fashion and confirmed with pathogen-specific PCR. This analysis successfully identified measles virus sequences in two brain samples and herpes simplex virus type-1 sequences in three brain samples. No pathogen was identified in the other two brain specimens. These results were concordant with pathogen-specific PCR and partially concordant with prior neuropathological examinations, demonstrating that deep sequencing can accurately identify viral infections in frozen brain tissue. PMID:24699691

Examination of core samples from the Mount Elbert Gas Hydrate Stratigraphic Test Well, Alaska North Slope: Effects of retrieval and preservation

USGS Publications Warehouse

Kneafsey, T.J.; Lu, H.; Winters, W.; Boswell, R.; Hunter, R.; Collett, T.S.

2011-01-01

Collecting and preserving undamaged core samples containing gas hydrates from depth is difficult because of the pressure and temperature changes encountered upon retrieval. Hydrate-bearing core samples were collected at the BPXA-DOE-USGS Mount Elbert Gas Hydrate Stratigraphic Test Well in February 2007. Coring was performed while using a custom oil-based drilling mud, and the cores were retrieved by a wireline. The samples were characterized and subsampled at the surface under ambient winter arctic conditions. Samples thought to be hydrate bearing were preserved either by immersion in liquid nitrogen (LN), or by storage under methane pressure at ambient arctic conditions, and later depressurized and immersed in LN. Eleven core samples from hydrate-bearing zones were scanned using x-ray computed tomography to examine core structure and homogeneity. Features observed include radial fractures, spalling-type fractures, and reduced density near the periphery. These features were induced during sample collection, handling, and preservation. Isotopic analysis of the methane from hydrate in an initially LN-preserved core and a pressure-preserved core indicate that secondary hydrate formation occurred throughout the pressurized core, whereas none occurred in the LN-preserved core, however no hydrate was found near the periphery of the LN-preserved core. To replicate some aspects of the preservation methods, natural and laboratory-made saturated porous media samples were frozen in a variety of ways, with radial fractures observed in some LN-frozen sands, and needle-like ice crystals forming in slowly frozen clay-rich sediments. Suggestions for hydrate-bearing core preservation are presented.

Rotational Thromboelastometry or Conventional Coagulation Tests in Liver Transplantation: Comparing Blood Loss, Transfusions, and Cost.

PubMed

Smart, Laura; Mumtaz, Khalid; Scharpf, Danielle; Gray, Nicole O'Bleness; Traetow, Daniel; Black, Sylvester; Michaels, Anthony J; Elkhammas, Elmahdi; Kirkpatrick, Robert; Hanje, A James

Orthotopic liver transplantation (OLT) can be associated with significant bleeding requiring multiple blood product transfusions. Rotational thromboelastometry (ROTEM) is a point-of-care device that has been used to monitor coagulation during OLT. Whether it reduces blood loss/transfusions during OLT remains controversial. We aim to compare ROTEM with conventional coagulation tests (aPTT, PT, INR, platelet count, fibrinogen) to guide transfusion of platelets, cryoprecipitate, and fresh frozen plasma (FFP) during OLT over 3 years. Thirty-four patients who had transfusions guided by ROTEM were compared to 34 controls who received transfusions guided by conventional coagulation tests (CCT). Intraoperative blood loss, type/ amount of blood products transfused, and direct costs were compared between the two groups. The ROTEM group had significantly less intra-operative blood loss (2.0 vs. 3.0 L, p = 0.04) and fresh frozen plasma (FFP) transfusion (4 units vs. 6.5 units, p = 0.015) compared to the CCT group (2.0L vs. 3.0L, p = 0.04). However, total number of patients transfused cryoprecipitate was increased in ROTEM (n = 25;73%) as compared to CCT (n = 19; 56%), p = 0.033. The direct cost of blood products plus testing was reduced in the ROTEM group ($113,142.89 vs. $127,814.77). In conclusion implementation of a ROTEM-guided transfusion algorithm resulted in a reduction in intra-operative blood loss, FFP transfusion and a decrease in direct cost during OLT. ROTEM is a useful and safe point of care device in OLT setting.

CO₂ processing and hydration of fruit and vegetable tissues by clathrate hydrate formation.

PubMed

Takeya, Satoshi; Nakano, Kohei; Thammawong, Manasikan; Umeda, Hiroki; Yoneyama, Akio; Takeda, Tohoru; Hyodo, Kazuyuki; Matsuo, Seiji

2016-08-15

CO2 hydrate can be used to preserve fresh fruits and vegetables, and its application could contribute to the processing of carbonated frozen food. We investigated water transformation in the frozen tissue of fresh grape samples upon CO2 treatment at 2-3 MPa and 3°C for up to 46 h. Frozen fresh bean, radish, eggplant and cucumber samples were also investigated for comparison. X-ray diffraction indicated that after undergoing CO2 treatment for several hours, structure I CO2 hydrate formed within the grape tissue. Phase-contrast X-ray imaging using the diffraction-enhanced imaging technique revealed the presence of CO2 hydrate within the intercellular spaces of these tissues. The carbonated produce became effervescent because of the dissociation of CO2 hydrate through the intercellular space, especially above the melting point of ice. In addition, suppressed metabolic activity resulting from CO2 hydrate formation, which inhibits water and nutrient transport through intercellular space, can be expected. Copyright © 2016 Elsevier Ltd. All rights reserved.

Cross-contamination during the preparation of frozen chickens in the kitchen.

PubMed Central

de Wit, J. C.; Broekhuizen, G.; Kampelmacher, E. H.

1979-01-01

A study was made of the extent to which frozen broilers, contaminated with indicator organisms, can cause cross-contamination in the kitchen. In 60 kitchens a number of relevant objects were sampled during the preparation of contaminated frozen broilers. The results show that cross-contamination occurred in a high proportion of the kitchens examined. In many instances the indicator organism was still present on various objects even after rinsing, 'clearing' or washing up. In view of the possible risk of a cross-contamination with Salmonella spp. the importance of instructing food preparers is emphasized. No salmonellas could be found in the sinks of the 60 kitchens examined. PMID:379210

Detection and prevalence of pathogenic Yersinia enterocolitica in refrigerated and frozen dairy products by duplex PCR and dot hybridization targeting the virF and ail genes.

PubMed

Ye, Y W; Ling, N; Han, Y J; Wu, Q P

2014-11-01

Pathogenic Yersinia enterocolitica is involved in yersiniosis through expression of chromosome-borne or plasmid-borne virulence factors. Yersinia enterocolitica is a cold-tolerant pathogen frequently isolated from refrigerated or frozen foods. However, little attention has been focused on the prevalence of pathogenic Y. enterocolitica in refrigerated or frozen dairy samples in China. In this study, we developed a new duplex PCR targeting the plasmid-borne virF gene and chromosome-borne ail gene for detection of pathogenic Y. enterocolitica isolates. We established a detection limit for the duplex PCR of 6.5 × 10(2)cfu/mL in artificially contaminated dairy samples. In addition, the duplex PCR could detect directly 4.5 to 5.7 cfu of Y. enterocolitica in 5 mL of brain heart infusion broth after 6 h of enrichment at 28 °C. A newly developed dot hybridization assay further confirmed specificity of the duplex PCR for detection of virulent Y. enterocolitica. Furthermore, 13 Y. enterocolitica and 5 pathogenic strains, from 88 commercial frozen or refrigerated dairy products, were detected successfully by the China National Standard method (GB/T4789.8-2008) and the duplex PCR, respectively. Finally, biotypes and serotypes of pathogenic Y. enterocolitica strains were further characterized. The duplex PCR developed here is reliable for large-scale screening, routine monitoring, and risk assessment of pathogenic Y. enterocolitica in refrigerated or frozen dairy products. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Storage of Unfed and Leftover Mothers' Own Milk.

PubMed

Fogleman, April D; Meng, Ting; Osborne, Jason; Perrin, Maryanne T; Jones, Frances; Allen, Jonathan C

The objective was to examine the bacteriological and immunological properties of freshly expressed, previously frozen, and leftover mothers' own milk during storage. In the first of two pilot studies, 12 mother-infant dyads participated. The milk studied included freshly expressed unfed and freshly expressed leftover milk. Milk samples were stored at 24°C, 4°C, or -20°C. In the second pilot study, 11 mother-infant dyads participated. The milk studied included milk that had been previously frozen, including previously frozen leftover milk. Milk samples were stored at 24°C and 4°C. After storage in both studies, the milk was analyzed for bacteriological and immunological properties. Bacteriological and immunological characteristics of freshly expressed unfed and freshly expressed leftover milk and previously frozen unfed and previously frozen leftover milk remained stable during storage at 4°C for at least 6 days. The quality of all groups of mothers' milk declined when stored at 24°C for longer than 3 hours. While this study provides evidence that human milk might be safe at longer storage times, storage guidelines should not be revised until more research is performed. This study serves as a call to action for more research on the topic of human milk storage, specifically leftover human milk. The study provides information to inform future study designs on the topic of unpasteurized human milk storage. More research is needed regarding leftover human milk storage with a greater number of participants, determination of the quality of human milk, and the storage of human milk in a real-life setting.

Optimizing Frozen Sample Preparation for Laser Microdissection: Assessment of CryoJane Tape-Transfer System®

PubMed Central

Golubeva, Yelena G.; Smith, Roberta M.; Sternberg, Lawrence R.

2013-01-01

Laser microdissection is an invaluable tool in medical research that facilitates collecting specific cell populations for molecular analysis. Diversity of research targets (e.g., cancerous and precancerous lesions in clinical and animal research, cell pellets, rodent embryos, etc.) and varied scientific objectives, however, present challenges toward establishing standard laser microdissection protocols. Sample preparation is crucial for quality RNA, DNA and protein retrieval, where it often determines the feasibility of a laser microdissection project. The majority of microdissection studies in clinical and animal model research are conducted on frozen tissues containing native nucleic acids, unmodified by fixation. However, the variable morphological quality of frozen sections from tissues containing fat, collagen or delicate cell structures can limit or prevent successful harvest of the desired cell population via laser dissection. The CryoJane Tape-Transfer System®, a commercial device that improves cryosectioning outcomes on glass slides has been reported superior for slide preparation and isolation of high quality osteocyte RNA (frozen bone) during laser dissection. Considering the reported advantages of CryoJane for laser dissection on glass slides, we asked whether the system could also work with the plastic membrane slides used by UV laser based microdissection instruments, as these are better suited for collection of larger target areas. In an attempt to optimize laser microdissection slide preparation for tissues of different RNA stability and cryosectioning difficulty, we evaluated the CryoJane system for use with both glass (laser capture microdissection) and membrane (laser cutting microdissection) slides. We have established a sample preparation protocol for glass and membrane slides including manual coating of membrane slides with CryoJane solutions, cryosectioning, slide staining and dissection procedure, lysis and RNA extraction that facilitated efficient dissection and high quality RNA retrieval from CryoJane preparations. CryoJane technology therefore has the potential to facilitate standardization of laser microdissection slide preparation from frozen tissues. PMID:23805281

Histology of a Woolly Mammoth (Mammuthus primigenius) Preserved in Permafrost, Yamal Peninsula, Northwest Siberia.

PubMed

Papageorgopoulou, Christina; Link, Karl; Rühli, Frank J

2015-06-01

In 2007, the baby woolly mammoth (Mammuthus primigenius) named Lyuba was found frozen in the Siberian tundra permafrost along the Yuribey River. She was proclaimed the best-preserved mammoth discovery. As part of the endoscopic examination of Lyuba, tissue samples of hair, muscle, and internal organs were taken. The sectioned biopsies were stained using standard and special histological stains. In general, the microscopic preservation of the tissue was good although no clearly identifiable cell nuclei were found by standard staining methods. Only a few cell nuclei could be identified in some samples when fluorescence stained with DAPI. The best-preserved structures were collagen fibers and muscle tissue, which gave some structural resemblance to the organs. In the hairs, evidence of pigmentation, a scaly surface, diagonal intra-hair structures, and a medulla were seen. Fat droplets could be identified with Sudan Red in the subcutaneous fat sample and in several organs. Bacteria were seen on the lumen side of the small intestine and caecum, and in the liver and lung tissue. In addition, fungi and pollen were seen in the lung sample. In the wall of the caecum and small intestine, blood vessels and nerves were visualized. Iron was identified in the vivianite sample. Some biopsies compared well structurally with the African elephant tissue sections. The histological findings support the theory that Lyuba drowned in muddy water. The microscopic tissue preservation and cell nuclei destruction indicate that Lyuba's body underwent at least one freeze-thaw cycle. © 2015 Wiley Periodicals, Inc.

Survival of cold-stressed Campylobacter jejuni on ground chicken and chicken skin during frozen storage.

PubMed

Bhaduri, Saumya; Cottrell, Bryan

2004-12-01

Campylobacter jejuni is prevalent in poultry, but the effect of combined refrigerated and frozen storage on its survival, conditions relevant to poultry processing and storage, has not been evaluated. Therefore, the effects of refrigeration at 4 degrees C, freezing at -20 degrees C, and a combination of refrigeration and freezing on the survival of C. jejuni in ground chicken and on chicken skin were examined. Samples were enumerated using tryptic soy agar containing sheep's blood and modified cefoperazone charcoal deoxycholate agar. Refrigerated storage alone for 3 to 7 days produced a reduction in cell counts of 0.34 to 0.81 log10 CFU/g in ground chicken and a reduction in cell counts of 0.31 to 0.63 log10 CFU/g on chicken skin. Declines were comparable for each sample type using either plating medium. Frozen storage, alone and with prerefrigeration, produced a reduction in cell counts of 0.56 to 1.57 log10 CFU/g in ground chicken and a reduction in cell counts of 1.38 to 3.39 log10 CFU/g on chicken skin over a 2-week period. The recovery of C. jejuni following freezing was similar on both plating media. The survival following frozen storage was greater in ground chicken than on chicken skin with or without prerefrigeration. Cell counts after freezing were lower on chicken skin samples that had been prerefrigerated for 7 days than in those that had been prerefrigerated for 0, 1, or 3 days. This was not observed for ground chicken samples, possibly due to their composition. C. jejuni survived storage at 4 and -20 degrees C with either sample type. This study indicates that, individually or in combination, refrigeration and freezing are not a substitute for safe handling and proper cooking of poultry.

Successful Validation of Sample Processing and Quantitative Real-Time PCR Capabilities on the International Space Station

NASA Technical Reports Server (NTRS)

Parra, Macarena; Jung, Jimmy; Almeida, Eduardo; Boone, Travis; Schonfeld, Julie; Tran, Luan

2016-01-01

The WetLab-2 system was developed by NASA Ames Research Center to offer new capabilities to researchers. The system can lyse cells and extract RNA (Ribonucleic Acid) on-orbit from different sample types ranging from microbial cultures to animal tissues. The purified RNA can then either be stabilized for return to Earth or can be used to conduct on-orbit quantitative Reverse Transcriptase PCR (Polymerase Chain Reaction) (qRT-PCR) analysis without the need for sample return. The qRT-PCR results can be downlinked to the ground a few hours after the completion of the run. The validation flight of the WetLab-2 system launched on SpaceX-8 on April 8, 2016. On orbit operations started on April 15th with system setup and was followed by three quantitative PCR runs using an E. coli genomic DNA template pre-loaded at three different concentrations. These runs were designed to discern if quantitative PCR functions correctly in microgravity and if the data is comparable to that from the ground control runs. The flight data showed no significant differences compared to the ground data though there was more variability in the values, this was likely due to the numerous small bubbles observed. The capability of the system to process samples and purify RNA was then validated using frozen samples prepared on the ground. The flight data for both E. coli and mouse liver clearly shows that RNA was successfully purified by our system. The E. coli qRT-PCR run showed successful singleplex, duplex and triplex capability. Data showed high variability in the resulting Cts (Cycle Thresholds [for the PCR]) likely due to bubble formation and insufficient mixing during the procedure run. The mouse liver qRT-PCR run had successful singleplex and duplex reactions and the variability was slightly better as the mixing operation was improved. The ability to purify and stabilize RNA and to conduct qRT-PCR on-orbit is an important step towards utilizing the ISS as a National Laboratory facility. The ability to get on-orbit data will provide investigators with the opportunity to adjust experimental parameters in real time without the need for sample return and re-flight. The WetLab-2 Project is supported by the Research Integration Office in the ISS Program.

Robust Image Restoration for Ground-Based Space Surveillance

DTIC Science & Technology

2013-09-01

systems can be characterized by well-separated layers of frozen turbulence with different velocity vectors (the frozen flow model, FFM ) [5[. Studies...of the atmosphere at Mt. Haleakala have suggested that there are typically 2-3 such layers [6]. The FFM requires that we know the wind velocities...as a sum of independent static turbulent layers: where denotes the velocity of the ith layer. Using the FFM results in better sampling of the

Predictors of renal function recovery among patients undergoing renal replacement therapy following orthotopic liver transplantation.

PubMed

Andreoli, Maria Claudia Cruz; Souza, Nádia Karina Guimarães de; Ammirati, Adriano Luiz; Matsui, Thais Nemoto; Carneiro, Fabiana Dias; Ramos, Ana Claudia Mallet de Souza; Iizuca, Ilson Jorge; Coelho, Maria Paula Vilela; Afonso, Rogério Carballo; Ferraz-Neto, Ben-Hur; Almeida, Marcio Dias de; Durão, Marcelino; Batista, Marcelo Costa; Monte, Julio Cesar; Pereira, Virgílio Gonçalves; Santos, Oscar Pavão Dos; Santos, Bento Cardoso Dos

2017-01-01

Renal dysfunction frequently occurs during the periods preceding and following orthotopic liver transplantation (OLT), and in many cases, renal replacement therapy (RRT) is required. Information regarding the duration of RRT and the rate of kidney function recovery after OLT is crucial for transplant program management. We evaluated a sample of 155 stable patients undergoing post-intensive care hemodialysis (HD) from a patient population of 908 adults who underwent OLT. We investigated the average time to renal function recovery (duration of RRT required) and determined the risk factors for remaining on dialysis > 90 days after OLT. Log-rank tests were used for univariate analysis, and Cox proportional hazards models were used to identify factors associated with the risk of remaining on HD. The results of our analysis showed that of the 155 patients, 28% had pre-OLT diabetes mellitus, 21% had pre-OLT hypertension, and 40% had viral hepatitis. Among the patients, the median MELD (Model for End-Stage Liver Disease) score was 27 (interquartile range [IQR] 22-35). When they were listed for liver transplantation, 32% of the patients had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD, and 50% had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD at the time of OLT. Of the transplanted patients, 25% underwent pre-OLT intermittent HD, and 14% and 41% underwent continuous renal replacement therapy (CRRT) pre-OLT and post-OLT, respectively. At 90 days post-OLT, 118 (76%) patients had been taken off dialysis, and 16 (10%) patients had died while undergoing HD. The median recovery time of these post-OLT patients was 33 (IQR 27-39) days. In the multivariate analysis, fulminant hepatic failure as the cause of liver disease (p<0.001), the absence of pre-OLT hypertension (p = 0.016), a lower intraoperative fresh-frozen plasma (FFP) transfusion volume (p = 0.019) and not undergoing pre-OLT intermittent HD (p = 0.032) were associated with performing RRT for less than 90 days. Therefore, a high proportion of OLT patients showed improved renal function after OLT, and those who were diagnosed with fulminant hepatic failure, had no pre-OLT hypertension, received a lower transfused volume of intraoperative FFP and did not undergo pre-OLT intermittent HD had a higher probability of recovery.

Predictors of renal function recovery among patients undergoing renal replacement therapy following orthotopic liver transplantation

PubMed Central

de Souza, Nádia Karina Guimarães; Ammirati, Adriano Luiz; Matsui, Thais Nemoto; Carneiro, Fabiana Dias; Ramos, Ana Claudia Mallet de Souza; Iizuca, Ilson Jorge; Afonso, Rogério Carballo; Ferraz-Neto, Ben-Hur; de Almeida, Marcio Dias; Durão, Marcelino; Batista, Marcelo Costa; Monte, Julio Cesar; Pereira, Virgílio Gonçalves; dos Santos, Oscar Pavão

2017-01-01

Renal dysfunction frequently occurs during the periods preceding and following orthotopic liver transplantation (OLT), and in many cases, renal replacement therapy (RRT) is required. Information regarding the duration of RRT and the rate of kidney function recovery after OLT is crucial for transplant program management. We evaluated a sample of 155 stable patients undergoing post-intensive care hemodialysis (HD) from a patient population of 908 adults who underwent OLT. We investigated the average time to renal function recovery (duration of RRT required) and determined the risk factors for remaining on dialysis > 90 days after OLT. Log-rank tests were used for univariate analysis, and Cox proportional hazards models were used to identify factors associated with the risk of remaining on HD. The results of our analysis showed that of the 155 patients, 28% had pre-OLT diabetes mellitus, 21% had pre-OLT hypertension, and 40% had viral hepatitis. Among the patients, the median MELD (Model for End-Stage Liver Disease) score was 27 (interquartile range [IQR] 22-35). When they were listed for liver transplantation, 32% of the patients had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD, and 50% had serum creatinine (Scr) levels > 1.5 mg/dL or were on HD at the time of OLT. Of the transplanted patients, 25% underwent pre-OLT intermittent HD, and 14% and 41% underwent continuous renal replacement therapy (CRRT) pre-OLT and post-OLT, respectively. At 90 days post-OLT, 118 (76%) patients had been taken off dialysis, and 16 (10%) patients had died while undergoing HD. The median recovery time of these post-OLT patients was 33 (IQR 27–39) days. In the multivariate analysis, fulminant hepatic failure as the cause of liver disease (p<0.001), the absence of pre-OLT hypertension (p = 0.016), a lower intraoperative fresh-frozen plasma (FFP) transfusion volume (p = 0.019) and not undergoing pre-OLT intermittent HD (p = 0.032) were associated with performing RRT for less than 90 days. Therefore, a high proportion of OLT patients showed improved renal function after OLT, and those who were diagnosed with fulminant hepatic failure, had no pre-OLT hypertension, received a lower transfused volume of intraoperative FFP and did not undergo pre-OLT intermittent HD had a higher probability of recovery. PMID:28574999

A new insight into male fertility preservation for patients with completely immotile spermatozoa.

PubMed

Chen, Huanhua; Feng, Guixue; Zhang, Bo; Zhou, Hong; Wang, Caizhu; Shu, Jinhui; Gan, Xianyou; Lin, Ruoyun; Huang, Dongmei; Huang, Yingqin

2017-09-18

Sperm cryopreservation is the most effective method to preserve male fertility but this is normally used for motile spermatozoa. Thus, only motile spermatozoa are used for cryopreservation in most reproductive medicine centers worldwide. The immotile spermatozoa from some problematic patients are usually discarded, resulting in a missed opportunity of sterility cryopreservation for future assisted reproductive treatments. Many studies have shown that successful fertilization can be obtained after selection of viable sperm from the completely immotile spermatozoa before ICSI. Whether the completely immotile spermatozoa are worth of freezing has not been realized The aim of this study is to explore the clinical value of cryopreservation of immotile spermatozoa. Completely immotile spermatozoa were collected and frozen, and subsequently viable but immotile frozen-thawed spermatozoa were selected by laser plus for ICSI. Main outcomes included spermatozoa survival index, fertilization rate and good quality embryo rate. After identification by laser, the fresh samples of spermatozoa presented with a mean survival rate of 54.86% and 26.05%, and this was reduced to 44.13% and 18.13% in frozen-thawed spermatozoa samples, which showed a frozen-thawed spermatozoa survival index of 0.80 and 0.70 in the testicular and ejaculate sperm, respectively. There were no statistically differences in fertilization rate (80% vs80.51%, 75.00% vs 81.48%), cleavage rate (95.45% vs 98.95%, 100.00% vs 95.45%) and good quality embryo rate (40.48% vs 52.13%, 33.33%vs38.10%) between the frozen-thawed immotile spermatozoa group and the routine fresh immotile spermatozoa ICSI group in both testicular and ejaculate sperm, respectively. The results of the study show that completely immotile spermatozoa can be frozen in order to preserve male fertility as long as viable spermatozoa are present. This procedure provides a further possibility for fertility preservation for patients with completely immotile spermatozoa.

Flow Cytometric Human Leukocyte Antigen-B27 Typing with Stored Samples for Batch Testing

PubMed Central

Seo, Bo Young

2013-01-01

Background Flow cytometry (FC) HLA-B27 typing is still used extensively for the diagnosis of spondyloarthropathies. If patient blood samples are stored for a prolonged duration, this testing can be performed in a batch manner, and in-house cellular controls could easily be procured. In this study, we investigated various methods of storing patient blood samples. Methods We compared four storage methods: three methods of analyzing lymphocytes (whole blood stored at room temperature, frozen mononuclear cells, and frozen white blood cells [WBCs] after lysing red blood cells [RBCs]), and one method using frozen platelets (FPLT). We used three ratios associated with mean fluorescence intensities (MFI) for HLAB27 assignment: the B27 MFI ratio (sample/control) for HLA-B27 fluorescein-5-isothiocyanate (FITC); the B7 MFI ratio for HLA-B7 phycoerythrin (PE); and the ratio of these two ratios, B7/B27 ratio. Results Comparing the B27 MFI ratios of each storage method for the HLA-B27+ samples and the B7/B27 ratios for the HLA-B7+ samples revealed that FPLT was the best of the four methods. FPLT had a sensitivity of 100% and a specificity of 99.3% for HLA-B27 assignment in DNA-typed samples (N=164) when the two criteria, namely, B27 MFI ratio >4.0 and B7/B27 ratio <1.5, were used. Conclusions The FPLT method was found to offer a simple, economical, and accurate method of FC HLA-B27 typing by using stored patient samples. If stored samples are used, this method has the potential to replace the standard FC typing method when used in combination with a complementary DNA-based method. PMID:23667843

INFLUENCE OF INTRAMUSCULAR FAT LEVEL ON ORGANOLEPTIC, PHYSICAL, AND CHEMICAL CHARACTERISTICS OF IRRADIATED PORK. I. HIGH-TEMPERATURE SHORT-TIME PRE-IRRADIATION HEAT TREATMENT

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bray, R.W.; Weckel, K.G.; Evans, G.W.

1964-02-01

The influence of intramuscular fat (degree of marbling) on characteristics of precooked and irradiated pork muscle was studied. Loins were selected and categorized into three marbling levels by visual appraisal. A relatively high temperature (325 deg F) and short time (2 hr) heat treatment was used for enzyme inactivation. Samples were packed under vacuum in rigid containers and irradiated to 4.5 Mrad with gamma radiation. Irradiated and frozen control samples were evaluated up to 2l0 days later. Degree of marbling had no apparent influence on organoleptic properties of either irradiated or frozen control longissimus dorsi muscle samples. Frozen control samplesmore » were preferred in general appearance, flavor, and over-all acceptability by panelists. Irradiated samples were preferred in texture qualities. Storage time was not a major factor in organoleptic acceptability; however, acceptability of irradiated samples declined between 150 and 210 days of storage. Hunter color attributes were not affected by marbling level. L, a/sub L/ hue, and saturation were increased by radiation treatment. Mechanical tenderness values were decreased due to higher marbling level and radiation treatment. Expressible-moisture values were lowered by radiation treatment and increased with storage time. Iodine numbers were decreased by radiation. Degree of marbling did not affect thiobarbituric acid values but they were significantly lower for irradiated samples. pH values increased with higher levels of intramuscular fat, were significantly higher in irradiated samples than controls, and tended to increase steadily with advancing storage time. (BBB)« less

High-speed shaking of frozen blood clots for extraction of human and malaria parasite DNA

PubMed Central

2011-01-01

Background Frozen blood clots remaining after serum collection is an often disregarded source of host and pathogen DNA due to troublesome handling and suboptimal outcome. Methods High-speed shaking of clot samples in a cell disruptor manufactured for homogenization of tissue and faecal specimens was evaluated for processing frozen blood clots for DNA extraction. The method was compared to two commercial clot protocols based on a chemical kit and centrifugation through a plastic sieve, followed by the same DNA extraction protocol. Blood clots with different levels of parasitaemia (1-1,000 p/μl) were prepared from parasite cultures to assess sensitivity of PCR detection. In addition, clots retrieved from serum samples collected within two epidemiological studies in Kenya (n = 630) were processed by high speed shaking and analysed by PCR for detection of malaria parasites and the human α-thalassaemia gene. Results High speed shaking succeeded in fully dispersing the clots and the method generated the highest DNA yield. The level of PCR detection of P. falciparum parasites and the human thalassaemia gene was the same as samples optimally collected with an anticoagulant. The commercial clot protocol and centrifugation through a sieve failed to fully dissolve the clots and resulted in lower sensitivity of PCR detection. Conclusions High speed shaking was a simple and efficacious method for homogenizing frozen blood clots before DNA purification and resulted in PCR templates of high quality both from humans and malaria parasites. This novel method enables genetic studies from stored blood clots. PMID:21824391

A Multistate Investigation of Antibiotic-Resistant Salmonella enterica Serotype I 4,[5],12:i:- Infections as Part of an International Outbreak Associated with Frozen Feeder Rodents

PubMed Central

Cartwright, E. J.; Nguyen, T.; Melluso, C.; Ayers, T.; Lane, C.; Hodges, A.; Li, X.; Quammen, J.; Yendell, S. J.; Adams, J.; Mitchell, J.; Rickert, R.; Klos, R.; Williams, I. T.; Behravesh, C. Barton; Wright, J.

2015-01-01

While most human Salmonella infections result from exposure to contaminated foods, an estimated 11% of all Salmonella infections are attributed to animal exposures, including both direct animal handling and indirect exposures such as cleaning cages and handling contaminated pet food. This report describes the epidemiologic, environmental and laboratory investigations conducted in the United States as part of the response to an international outbreak of tetracycline-resistant Salmonella enterica serotype I 4,[5],12:i:- infections with over 500 illnesses occurring from 2008 to 2010. This investigation found that illness due to the outbreak strain was significantly associated with exposure to pet reptiles and frozen feeder rodents used as food for pet reptiles. Salmonella isolates indistinguishable from the outbreak strain were isolated from a frozen feeder mice-fed reptile owned by a case patient, as well as from frozen feeder mice and environmental samples collected from a rodent producing facility (Company A). An international voluntary recall of all Company A produced frozen feeder animals sold between May 2009 and July 2010 occurred. Only 13% of cases in our investigation were aware of the association between Salmonella infection and mice or rats. Consumers, the pet industry, healthcare providers and veterinarians need to be aware of the potential health risk posed by feeder rodents, whether live or frozen. Frozen feeder rodent producers, suppliers and distributors should follow the animal food labelling requirements as described in 21 CFR §501.5, and all packages of frozen feeder rodents should include safe handling instructions. Persons should wash their hands thoroughly with soap and water after handling live or frozen feeder rodents, as well as reptiles or anything in the area where the animals live. Continued opportunities exist for public health officials, the pet industry, veterinarians and consumers to work together to prevent salmonellosis associated with pet food, pets and other animals. PMID:25996458

A Multistate Investigation of Antibiotic-Resistant Salmonella enterica Serotype I 4,[5],12:i:- Infections as Part of an International Outbreak Associated with Frozen Feeder Rodents.

PubMed

Cartwright, E J; Nguyen, T; Melluso, C; Ayers, T; Lane, C; Hodges, A; Li, X; Quammen, J; Yendell, S J; Adams, J; Mitchell, J; Rickert, R; Klos, R; Williams, I T; Barton Behravesh, C; Wright, J

2016-02-01

While most human Salmonella infections result from exposure to contaminated foods, an estimated 11% of all Salmonella infections are attributed to animal exposures, including both direct animal handling and indirect exposures such as cleaning cages and handling contaminated pet food. This report describes the epidemiologic, environmental and laboratory investigations conducted in the United States as part of the response to an international outbreak of tetracycline-resistant Salmonella enterica serotype I 4,[5],12:i:- infections with over 500 illnesses occurring from 2008 to 2010. This investigation found that illness due to the outbreak strain was significantly associated with exposure to pet reptiles and frozen feeder rodents used as food for pet reptiles. Salmonella isolates indistinguishable from the outbreak strain were isolated from a frozen feeder mice-fed reptile owned by a case patient, as well as from frozen feeder mice and environmental samples collected from a rodent producing facility (Company A). An international voluntary recall of all Company A produced frozen feeder animals sold between May 2009 and July 2010 occurred. Only 13% of cases in our investigation were aware of the association between Salmonella infection and mice or rats. Consumers, the pet industry, healthcare providers and veterinarians need to be aware of the potential health risk posed by feeder rodents, whether live or frozen. Frozen feeder rodent producers, suppliers and distributors should follow the animal food labelling requirements as described in 21 CFR §501.5, and all packages of frozen feeder rodents should include safe handling instructions. Persons should wash their hands thoroughly with soap and water after handling live or frozen feeder rodents, as well as reptiles or anything in the area where the animals live. Continued opportunities exist for public health officials, the pet industry, veterinarians and consumers to work together to prevent salmonellosis associated with pet food, pets and other animals. © 2015 Blackwell Verlag GmbH.

A comparative analysis of toluidine blue with frozen section in oral squamous cell carcinoma

PubMed Central

2012-01-01

Background Surgical excision of the primary tumor with safe margins remains the mainstay of treatment for oral cavity squamous cell carcinoma (OSCC). The standard of care for assessment of intraoperative margins is frozen section histopathology. Unfortunately the facility is not available at most centers in limited resource countries. Toluidine blue, a metachromatic dye, has been well described in clinical identification of malignant and premalignant lesion in the oral cavity. Considering this we decided to explore intraoperative use of toluidine blue staining, in comparison with frozen sections, for the assessment of tumor-free margins. Methods After obtaining clearance from the in-house ethical review committee, a prospective study was conducted at Aga Khan University Hospital, Karachi, from August 15, 2009 to March 14, 2010. A sample of 56 consenting patients with biopsy-proven OSCC were included in the study, giving us 280 tumor margins. Margins were analyzed using toluidine blue staining and frozen section histopathology. A receiver operator curve (ROC) was then applied to compare assessment of margin status by toluidine blue and frozen section. Results Of the 280 examined margins 11 stained positive with toluidine blue, three were positive on frozen section biopsy, and three were positive on final histopathology. Toluidine blue staining had sensitivity and specificity of 100% and 97%, respectively. The diagnostic accuracy of toluidine blue was found to be 97.1% with a positive predictive value (PPV) of 27.2% and a negative predictive value (NPV) of 100%. Conclusions Toluidine blue can be used as an effective screening modality for the assessment of intraoperative margins in resource limited environments and reducing the number of frozen section biopsies performed. Further by providing real-time clinical information within minutes it can reduce indirect costs such as operating room time. It may also be used as an ad hoc for frozen section biopsies where frozen section facilities are available. PMID:22500814

Four studies on effects of environmental factors on the quality of National Atmospheric Deposition Program measurements

USGS Publications Warehouse

Wetherbee, Gregory A.; Latysh, Natalie E.; Lehmann, Christopher M.B.; Rhodes, Mark F.

2011-01-01

Selected aspects of National Atmospheric Deposition Program / National Trends Network (NADP/NTN) protocols are evaluated in four studies. Meteorological conditions have minor impacts on the error in NADP/NTN sampling. Efficiency of frozen precipitation sample collection is lower than for liquid precipitation samples. Variability of NTN measurements is higher for relatively low-intensity deposition of frozen precipitation than for higher-intensity deposition of liquid precipitation. Urbanization of the landscape surrounding NADP/NTN sites is not affecting trends in wet-deposition chemistry data to a measureable degree. Five NADP siting criteria intended to preserve wet-deposition sample integrity have varying degrees of effectiveness. NADP siting criteria for objects within the 90 degrees cones and trees within the 120 degrees cones projected from the collector bucket to sky are important for protecting sample integrity. Tall vegetation, fences, and other objects located within 5 meters of the collectors are related to the frequency of visible sample contamination, indicating the importance of these factors in NADP siting criteria.

Height-resolved large-sample INAA of a 1 m long, 13 cm diameter ditch-bottom sample

NASA Astrophysics Data System (ADS)

Blaauw, M.; Baas, H. W.; Donze, M.

2003-06-01

A facility for instrumental neutron activation analysis (INAA) of large samples (up to 1 m long and 15 cm diameter) has been built. Correction methods for the simultaneous occurrence of neutron self-shielding and gamma-ray self-attenuation effects have been implemented and tested with a variety of samples. Now, the method has been extended to allow for the interpretation of scanned, collimated measurements, where results are obtained for individual voxels. As a validation and demonstration, a ditch-bottom sample of the maximum size was taken in a frozen condition. It was cut in 2 cm slices, still frozen, and put together again with each slice in a 2 cm height Petri dish divided in three sections. This allowed for verification of the results by ordinary INAA. Possible explanations for the discrepancies we observed between ordinary and large-sample INAA in the region where the concentration gradients are the steepest are discussed.

Effects of chilled-then-frozen storage (up to 52weeks) on an indicator of protein oxidation and indices of protein degradation in lamb M. longissimus lumborum.

PubMed

Coombs, Cassius E O; Holman, Benjamin W B; Collins, Damian; Kerr, Matthew J; Friend, Michael A; Hopkins, David L

2018-01-01

This study investigated the protein oxidation properties of lamb following chilled-then-frozen storage. Experimental (n=360) M. longissimus lumborum (LL) were randomly sampled from the boning room of a commercial Australian abattoir, at 24h post mortem, and assigned to five chilled storage periods (0, 2, 4, 6 and 8weeks) and six subsequent frozen storage periods (0, 4, 8, 12, 24 and 52weeks). Upon completion of each storage treatment combination, corresponding LL were sub-sectioned and analysed for carbonyl content, protein solubility, nitrate/nitrite content, particle size analysis and estimated myoglobin fractions. The association between these protein measures and shear force was also explored. During chilled storage, particle size and sarcoplasmic protein solubility decreased which indicated protein degradation, while frozen storage only affected myoglobin oxidation. Tenderness was best explained by decreased particle size, decreased deoxymyoglobin and increased oxymyoglobin. No carbonyl effects were observed. It can be concluded that, according to these analyses, that in chilled-then-frozen lamb carbonyl formation was negligible. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

A comparison of liver sampling techniques in dogs.

PubMed

Kemp, S D; Zimmerman, K L; Panciera, D L; Monroe, W E; Leib, M S; Lanz, O I

2015-01-01

The liver sampling technique in dogs that consistently provides samples adequate for accurate histopathologic interpretation is not known. To compare histopathologic results of liver samples obtained by punch, cup, and 14 gauge needle to large wedge samples collected at necropsy. Seventy dogs undergoing necropsy. Prospective study. Liver specimens were obtained from the left lateral liver lobe with an 8 mm punch, a 5 mm cup, and a 14 gauge needle. After sample acquisition, two larger tissue samples were collected near the center of the left lateral lobe to be used as a histologic standard for comparison. Histopathologic features and numbers of portal triads in each sample were recorded. The mean number of portal triads obtained by each sampling method were 2.9 in needle samples, 3.4 in cup samples, 12 in punch samples, and 30.7 in the necropsy samples. The diagnoses in 66% of needle samples, 60% of cup samples, and 69% of punch samples were in agreement with the necropsy samples, and these proportions were not significantly different from each other. The corresponding kappa coefficients were 0.59 for needle biopsies, 0.52 for cup biopsies, and 0.62 for punch biopsies. The histopathologic interpretation of a liver sample in the dog is unlikely to vary if the liver biopsy specimen contains at least 3-12 portal triads. However, in comparison large necropsy samples, the accuracy of all tested methods was relatively low. Copyright © 2014 by the American College of Veterinary Internal Medicine.

Texture of Frozen Food

NASA Astrophysics Data System (ADS)

Wani, Kohmei

Quantitative determination of textural quality of frozen food due to freezing and storage conditions is complicated,since the texture is consisted of multi-dimensiona1 factors. The author reviewed the importance of texture in food quality and the factors which is proposed by a priori estimation. New classification of expression words of textural properties by subjective evaluation and an application of four elements mechanical model for analysis of physical characteristics was studied on frozen meat patties. Combination of freezing-thawing condition on the subjective properties and physiochemical characteristics of beef lean meat and hamachi fish (Yellow-tail) meat was studied. Change of the plasticity and the deformability of these samples differed by freezing-thawing rate and cooking procedure. Also optimum freezing-thawing condition was differed from specimens.

[Association between the use of blood components and the five-year mortality after liver transplant].

PubMed

de Morais, Bruno Salomé; Sanches, Marcelo Dias; Ribeiro, Daniel Dias; Lima, Agnaldo Soares; de Abreu Ferrari, Teresa Cristina; Duarte, Malvina Maria de Freitas; Cançado, Guilherme Henrique Gomes Moreira

2011-01-01

Liver transplant (LT) surgery is associated with significant bleeding in 20% of cases, and several authors have demonstrated the risks related to blood components. The objective of the present study was to evaluate the impact of using blood components during hospitalization in five-year survival of patients undergoing LT. One hundred and thirteen patients were evaluated retrospectively. Several variables, including the use of blood components intraoperatively and throughout hospitalization, were categorized and evaluated by univariate analysis using Fisher's test. A level of significance of 5% was adopted. Results with p < 0.2 underwent multivariate analysis using multinomial logistic regression. Parenchymal diseases, preoperative renal dysfunction, and longer stay in hospital and ICU are associated with greater five-year mortality after LT (p < 0.05). Unlike the intraoperative use of blood components, the accumulated transfusion of packed red blood cell, frozen fresh plasma, and platelets during the entire hospitalization was associated with greater five-year mortality after liver transplantation (p < 0.01). This study emphasizes the relationship between the use of blood components during hospitalization and increased mortality in five years after LT. 2011 Elsevier Editora Ltda. All rights reserved.

Localization of cytochrome P4501A in liver and extrahepatic organs of the pilot whale, Globiocephala melaena

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stegeman, J.; Miller, C.; Moore, M.

1994-12-31

Marine mammals may be important indicators of effects of contaminants that are globally distributed. Recently the authors described apparent environmental induction of hepatic CYPLA in beluga whales. Here they describe the localization and extent of CYPLA expression in organs of the pilot whale. Tissues from 18 pilot whales stranded on Cape Cod in 1990/91 were frozen in liquid N2 or fixed in formalin and embedded. Liver microsomal EROD activity were comparable to results with other cetaceans. Immunohistochemical analysis showed a periportal localization of CYPLA in liver parenchyma, and staining in the endothelium. Renal staining was strong in brush border andmore » endothelium. Testis, ovary, and spleen showed CYPLA staining only in endothelium. Adrenal zona fasciculata and zona reticularis stained more weakly than did endothelium. In lung there was mild staining of bronchiolar epithelium and strong staining of endothelium. The results indicate that active concentrations of inducer have penetrated throughout the body. CYPLA stained in dermal endothelium, indicating that analysis of skin biopsies could allow nondestructive analysis of CYPLA induction in marine mammals. CYPLA expression in these whales was surprisingly strong, suggesting the possibility of chemical effects related to CYPLA induction.« less

Comparison of enterotomy leak pressure among fresh, cooled, and frozen-thawed porcine jejunal segments.

PubMed

Aeschlimann, Kimberly A; Mann, F A; Middleton, John R; Belter, Rebecca C

2018-05-01

OBJECTIVE To determine whether stored (cooled or frozen-thawed) jejunal segments can be used to obtain dependable leak pressure data after enterotomy closure. SAMPLE 36 jejunal segments from 3 juvenile pigs. PROCEDURES Jejunal segments were harvested from euthanized pigs and assigned to 1 of 3 treatment groups (n = 12 segments/group) as follows: fresh (used within 4 hours after collection), cooled (stored overnight at 5°C before use), and frozen-thawed (frozen at -12°C for 8 days and thawed at room temperature [23°C] for 1 hour before use). Jejunal segments were suspended and 2-cm enterotomy incisions were made on the antimesenteric border. Enterotomies were closed with a simple continuous suture pattern. Lactated Ringer solution was infused into each segment until failure at the suture line was detected. Leak pressure was measured by use of a digital transducer. RESULTS Mean ± SD leak pressure for fresh, cooled, and frozen-thawed segments was 68.3 ± 23.7 mm Hg, 55.3 ± 28.1 mm Hg, and 14.4 ± 14.8 mm Hg, respectively. Overall, there were no significant differences in mean leak pressure among pigs, but a significant difference in mean leak pressure was detected among treatment groups. Mean leak pressure was significantly lower for frozen-thawed segments than for fresh or cooled segments, but mean leak pressure did not differ significantly between fresh and cooled segments. CONCLUSIONS AND CLINICAL RELEVANCE Fresh porcine jejunal segments or segments cooled overnight may be used for determining intestinal leak pressure, but frozen-thawed segments should not be used.

Validation of the shake test for detecting freeze damage to adsorbed vaccines.

PubMed

Kartoglu, Umit; Ozgüler, Nejat Kenan; Wolfson, Lara J; Kurzatkowski, Wiesław

2010-08-01

To determine the validity of the shake test for detecting freeze damage in aluminium-based, adsorbed, freeze-sensitive vaccines. A double-blind crossover design was used to compare the performance of the shake test conducted by trained health-care workers (HCWs) with that of phase contrast microscopy as a "gold standard". A total of 475 vials of 8 different types of World Health Organization prequalified freeze-sensitive vaccines from 10 different manufacturers were used. Vaccines were kept at 5 degrees C. Selected numbers of vials from each type were then exposed to -25 degrees C and -2 degrees C for 24-hour periods. There was complete concordance between HCWs and phase-contrast microscopy in identifying freeze-damaged vials and non-frozen samples. Non-frozen samples showed a fine-grain structure under phase contrast microscopy, but freeze-damaged samples showed large conglomerates of massed precipitates with amorphous, crystalline, solid and needle-like structures. Particles in the non-frozen samples measured from 1 microm (vaccines against diphtheria-tetanus-pertussis; Haemophilus influenzae type b; hepatitis B; diphtheria-tetanus-pertussis-hepatitis B) to 20 microm (diphtheria and tetanus vaccines, alone or in combination). By contrast, aggregates in the freeze-damaged samples measured up to 700 microm (diphtheria-tetanus-pertussis) and 350 microm on average. The shake test had 100% sensitivity, 100% specificity and 100% positive predictive value in this study, which confirms its validity for detecting freeze damage to aluminium-based freeze-sensitive vaccines.

Effect of refrigeration and frozen storage on the Campylobacter jejuni recovery from naturally contaminated broiler carcasses

PubMed Central

Maziero, Maike T.; de Oliveira, Tereza Cristina R. M.

2010-01-01

Campylobacter jejuni is the most common thermophilic Campylobacter associated with human enteritis in many countries. Broilers and their by-products are the main sources for human enteritis. Refrigeration and freezing are used to control bacterial growth in foods. The effect of these interventions on survival of Campylobacter jejuni is yet not quite understood. This study evaluated the effect of storage temperature on the survival of C. jejuni in chicken meat stored for seven days at 4°C and for 28 days at -20°C. The influence of selective enrichment on recovery of Campylobacter was also evaluated. Thirty fresh chicken meat samples were analyzed and 93.3% was contaminated with termotolerant Campylobacter spp. with average count of 3.08 Log10 CFU/g on direct plating. After refrigeration, 53.3% of the analyzed samples tested positive for Campylobacter and the average count was 1.19 Log10 CFU/g. After storage at -20°C, 36.6% of the samples were positive with a verage count of 0.75 Log10 CFU/g. C. jejuni was detected after enrichment, respectively, in 50% of the fresh, 36.7% of the refrigerated and 33.3% of the frozen meat samples analyzed. No difference was detected for the recovery of C. jejuni from fresh, refrigerated or frozen samples after selective enrichment, showing that this microorganism can survive under the tested storage conditions. PMID:24031523

Use of lactic acid with electron beam irradiation for control of Escherichia coli O157:H7, non-O157 VTEC E. coli, and Salmonella serovars on fresh and frozen beef.

PubMed

Li, Shuliu; Kundu, Devapriya; Holley, Richard A

2015-04-01

Lactic acid pre-treatment was examined to enhance the antimicrobial action of electron (e-) beam irradiation of beef trim. Meat samples were inoculated with Escherichia coli O157:H7, non-O157 VTEC E. coli or Salmonella cocktails and treated with 5% lactic acid at 55 °C. Samples were packaged aerobically or vacuum-packed, kept at 4 °C and treated with 1 kGy e-beam energy. Frozen samples were treated with 1, 3 or 7 kGy and stored at -20 °C for ≤ 5 d. Lactic acid enhanced the antimicrobial action of 1 kGy e-beam treatment against Salmonella by causing an additional <1.8 log CFU/g reduction. One kGy treatment of refrigerated samples reduced VTEC E. coli viability by 4.5 log CFU/g, and while lactic acid did not improve the reduction, after freezing additive effects were found. After 3 kGy irradiation, Salmonella was reduced by 2 and 4 log CFU/g in the irradiated and lactic acid plus irradiated samples, respectively. Lactic acid pre-treatment was of limited value with 1 kGy treatment for improving control of toxigenic E. coli in fresh beef trim, however, it would be useful with low dose irradiation for controlling both VTEC E. coli and Salmonella in frozen product. Copyright © 2014 Elsevier Ltd. All rights reserved.

Detection of somatic mutations by high-resolution DNA melting (HRM) analysis in multiple cancers.

PubMed

Gonzalez-Bosquet, Jesus; Calcei, Jacob; Wei, Jun S; Garcia-Closas, Montserrat; Sherman, Mark E; Hewitt, Stephen; Vockley, Joseph; Lissowska, Jolanta; Yang, Hannah P; Khan, Javed; Chanock, Stephen

2011-01-17

Identification of somatic mutations in cancer is a major goal for understanding and monitoring the events related to cancer initiation and progression. High resolution melting (HRM) curve analysis represents a fast, post-PCR high-throughput method for scanning somatic sequence alterations in target genes. The aim of this study was to assess the sensitivity and specificity of HRM analysis for tumor mutation screening in a range of tumor samples, which included 216 frozen pediatric small rounded blue-cell tumors as well as 180 paraffin-embedded tumors from breast, endometrial and ovarian cancers (60 of each). HRM analysis was performed in exons of the following candidate genes known to harbor established commonly observed mutations: PIK3CA, ERBB2, KRAS, TP53, EGFR, BRAF, GATA3, and FGFR3. Bi-directional sequencing analysis was used to determine the accuracy of the HRM analysis. For the 39 mutations observed in frozen samples, the sensitivity and specificity of HRM analysis were 97% and 87%, respectively. There were 67 mutation/variants in the paraffin-embedded samples, and the sensitivity and specificity for the HRM analysis were 88% and 80%, respectively. Paraffin-embedded samples require higher quantity of purified DNA for high performance. In summary, HRM analysis is a promising moderate-throughput screening test for mutations among known candidate genomic regions. Although the overall accuracy appears to be better in frozen specimens, somatic alterations were detected in DNA extracted from paraffin-embedded samples.

Detection of Somatic Mutations by High-Resolution DNA Melting (HRM) Analysis in Multiple Cancers

PubMed Central

Gonzalez-Bosquet, Jesus; Calcei, Jacob; Wei, Jun S.; Garcia-Closas, Montserrat; Sherman, Mark E.; Hewitt, Stephen; Vockley, Joseph; Lissowska, Jolanta; Yang, Hannah P.; Khan, Javed; Chanock, Stephen

2011-01-01

Identification of somatic mutations in cancer is a major goal for understanding and monitoring the events related to cancer initiation and progression. High resolution melting (HRM) curve analysis represents a fast, post-PCR high-throughput method for scanning somatic sequence alterations in target genes. The aim of this study was to assess the sensitivity and specificity of HRM analysis for tumor mutation screening in a range of tumor samples, which included 216 frozen pediatric small rounded blue-cell tumors as well as 180 paraffin-embedded tumors from breast, endometrial and ovarian cancers (60 of each). HRM analysis was performed in exons of the following candidate genes known to harbor established commonly observed mutations: PIK3CA, ERBB2, KRAS, TP53, EGFR, BRAF, GATA3, and FGFR3. Bi-directional sequencing analysis was used to determine the accuracy of the HRM analysis. For the 39 mutations observed in frozen samples, the sensitivity and specificity of HRM analysis were 97% and 87%, respectively. There were 67 mutation/variants in the paraffin-embedded samples, and the sensitivity and specificity for the HRM analysis were 88% and 80%, respectively. Paraffin-embedded samples require higher quantity of purified DNA for high performance. In summary, HRM analysis is a promising moderate-throughput screening test for mutations among known candidate genomic regions. Although the overall accuracy appears to be better in frozen specimens, somatic alterations were detected in DNA extracted from paraffin-embedded samples. PMID:21264207

Highly Pathogenic Avian Influenza Virus (H5N1) in Frozen Duck Carcasses, Germany, 2007

PubMed Central

Harder, Timm C.; Teuffert, Jürgen; Starick, Elke; Gethmann, Jörn; Grund, Christian; Fereidouni, Sasan; Durban, Markus; Bogner, Karl-Heinz; Neubauer-Juric, Antonie; Repper, Reinhard; Hlinak, Andreas; Engelhardt, Andreas; Nöckler, Axel; Smietanka, Krzysztof; Minta, Zenon; Kramer, Matthias; Globig, Anja; Mettenleiter, Thomas C.; Conraths, Franz J.

2009-01-01

We conducted phylogenetic and epidemiologic analyses to determine sources of outbreaks of highly pathogenic avian influenza virus (HPAIV), subtype H5N1, in poultry holdings in 2007 in Germany, and a suspected incursion of HPAIV into the food chain through contaminated deep-frozen duck carcasses. In summer 2007, HPAIV (H5N1) outbreaks in 3 poultry holdings in Germany were temporally, spatially, and phylogenetically linked to outbreaks in wild aquatic birds. Detection of HPAIV (H5N1) in frozen duck carcass samples of retained slaughter batches of 1 farm indicated that silent infection had occurred for some time before the incidental detection. Phylogenetic analysis established a direct epidemiologic link between HPAIV isolated from duck meat and strains isolated from 3 further outbreaks in December 2007 in backyard chickens that had access to uncooked offal from commercial deep-frozen duck carcasses. Measures that will prevent such undetected introduction of HPAIV (H5N1) into the food chain are urgently required. PMID:19193272

Evaluation of reactive oxygen metabolites in frozen serum samples. Effect of storage and repeated thawing.

PubMed

Cavalleri, A; Colombo, C; Venturelli, E; Miceli, R; Mariani, L; Cornelli, U; Pala, V; Berrino, F; Secreto, G

2004-01-01

Measuring the free radical activity in serum samples from prospective studies is the best way to investigate the association between oxidative stress and human diseases. Prospective studies require the analysis of serum samples that have often been stored for a long time. Our study was designed to determine the effect of storage at -30 degrees C and -80 degrees C for two years on free radical activity. We analyzed the free radical activity by measuring circulating hydroperoxides in a pool of sera at baseline and after one day, one week, one month and 25 months of storage, using a photometric method (d-ROMs test). Measurements were performed in aliquots thawed only once at each time point and in aliquots frozen and thawed repeatedly over the study period. After two years we observed a small but statistically significant 4% decrease in the hydroperoxide concentration, which was substantially unaffected by storage temperatures and repeated freeze-thaw cycles. We also carried out the d-ROMs test in sera from ten apparently healthy volunteers at 2, 8, 24, and 48 hours after collection and storage at 4 degrees C and did not observe any significant variation. In conclusion, the d-ROMs test is a simple method suitable to evaluate the free radical activity in frozen serum samples after long-term storage.

Artificial insemination of cows with semen in vitro contaminated with Neospora caninum tachyzoites failed to induce neosporosis.

PubMed

Canada, Nuno; Meireles, Carla Sofia; Ferreira, Paulo; Correia da Costa, José Manuel; Rocha, António

2006-06-30

Neosporosis is a major cause of abortion in cattle all over the world. Congenital transmission as well as horizontal transmission by ingestion of oocysts has been described. The detection of Neospora caninum DNA in bull semen warrants the investigation of possible transmission through the use of contaminated semen. In this experiment four cows were artificially inseminated with frozen-thawed semen contaminated in vitro with viable N. caninum tachyzoites (group A) and four control cows were inseminated with tachyzoites-free frozen-thawed semen, from the same bull (group B). Serum samples were collected 15 days before the artificial insemination (AI) and at days 10, 14, 21, 28, 45, 60 and 75 post-insemination. All sera samples were tested for neosporosis by direct agglutination test (DAT). Three of the cows from group A had negative DAT titers (< or =1:20) in all of the samples, while the fourth cow from this group had a low titer of antibodies (1:80) at day 10, and became negative at day 45, suggesting a stimulation of the immune system by the tachyzoites placed in uterus, rather than the induction of an infection. All of the cows from group B had negative DAT titers (< or =1:20) in all of the samples. These results suggest that transmission of neosporosis by artificial insemination with frozen-thawed semen is an unlikely event.

Evaluation of a dried blood and plasma collection device, SampleTanker(®), for HIV type 1 drug resistance genotyping in patients receiving antiretroviral therapy.

PubMed

Diallo, Karidia; Lehotzky, Erica; Zhang, Jing; Zhou, Zhiyong; de Rivera, Ivette Lorenzana; Murillo, Wendy E; Nkengasong, John; Sabatier, Jennifer; Zhang, Guoqing; Yang, Chunfu

2014-01-01

Whatman 903 filter paper is the only filter paper that has been used for HIV drug resistance (HIVDR) genotyping in resource-limited settings. In this study, we evaluated another dried blood specimen collection device, termed SampleTanker(®) (ST), for HIVDR genotyping. Blood specimens from 123 antiretroviral therapy (ART)-experienced patients were used to prepare ST whole blood and ST plasma specimens; they were then stored at ambient temperature for 2 or 4 weeks. The remaining plasma specimens were stored at -80°C and used as frozen plasma controls. Frozen plasma viral load (VL) was determined using the Roche Amplicor HIV-1 Monitor test, v.1.5 and 50 specimens with VL ≥3.00 log10 copies/ml were genotyped using the broadly sensitive genotyping assay. The medium VL for the 50 frozen plasma specimens with VL ≥3.00 log10 was 3.58 log10 copies/ml (IQR: 3.32-4.11) and 96.0% (48/50) of them were genotyped. Comparing to frozen plasma specimens, significantly lower genotyping rates were obtained from ST whole blood (48.98% and 42.85%) and ST plasma specimens (36.0% and 36.0%) stored at ambient temperature for 2 and 4 weeks, respectively (p<0.001). Nucleotide sequence identity and resistance profile analyses between the matched frozen plasma and ST whole blood or ST plasma specimens revealed high nucleotide sequence identities and concordant resistance profiles (98.1% and 99.0%, and 96.6% and 98.9%, respectively). Our results indicate that with the current design, the ST may not be the ideal dried blood specimen collection device for HIVDR monitoring for ART patients in resource-limited settings.

Epidemiological evaluation of hepatic trauma victims undergoing surgery.

PubMed

Kalil, Mitre; Amaral, Isaac Massaud Amim

2016-02-01

to evaluate the epidemiological variables and diagnostic and therapeutic modalities related to hepatic trauma patients undergoing laparotomy in a public referral hospital in the metropolitan region of Vitória-ES. we conducted a retrospective study, reviewing charts of trauma patients with liver injuries, whether isolated or in association with other organs, who underwent exploratory laparotomy, from January 2011 to December 2013. We studied 392 patients, 107 of these with liver injury. The male: female ratio was 6.6 : 1 and the mean age was 30.12 years. Penetrating liver trauma occurred in 78.5% of patients, mostly with firearms. Associated injuries occurred in 86% of cases and intra-abdominal injuries were more common in penetrating trauma (p <0.01). The most commonly used operative technique was hepatorrhaphy and damage control surgery was applied in 6.5% of patients. The average amounts of blood products used were 6.07 units of packed red blood cells and 3.01 units of fresh frozen plasma. The incidence of postoperative complications was 29.9%, the most frequent being infectious, including pneumonia, peritonitis and intra-abdominal abscess. The survival rate of patients suffering from blunt trauma was 60%, and penetrating trauma, 87.5% (p <0.05). despite technological advances in diagnosis and treatment, mortality rates in liver trauma remain high, especially in patients suffering from blunt trauma in relation to penetrating one.

Mycobiota and mycotoxins in bee pollen collected from different areas of Slovakia.

PubMed

Kačániová, Miroslava; Juráček, Miroslav; Chlebo, Róbert; Kňazovická, Vladimíra; Kadasi-Horáková, Miriam; Kunová, Simona; Lejková, Jadža; Haščík, Peter; Mareček, Ján; Simko, Milan

2011-01-01

Contamination by microscopic fungi and mycotoxins in different bee pollen samples, which were stored under three different ways of storing as freezing, drying and UV radiation, was investigated. During spring 2009, 45 samples of bee-collected pollen were gathered from beekeepers who placed their bee colonies on monocultures of sunflower, rape and poppy fields within their flying distance. Bee pollen was collected from bees' legs by special devices placed at the entrance to hives. Samples were examined for the concentration and identification of microscopic fungi able to grow on Malt and Czapek-Dox agar and mycotoxins content [deoxynivalenol (DON), T-2 toxin (T-2), zearalenone (ZON) and total aflatoxins (AFL), fumonisins (FUM), ochratoxins (OTA)] by direct competitive enzyme-linked immunosorbent assays (ELISA). The total number of microscopic fungi in this study ranged from 2.98 ± 0.02 in frozen sunflower bee pollen to 4.06 ± 0.10 log cfu.g(-1) in sunflower bee pollen after UV radiation. In this study, 449 isolates belonging to 21 fungal species representing 9 genera were found in 45 samples of bee pollen. The total isolates were detected in frozen poppy pollen 29, rape pollen 40, sunflower pollen 80, in dried poppy pollen 12, rape pollen 36, sunflower 78, in poppy pollen after UV radiation treatment 54, rape 59 and sunflower 58. The most frequent isolates of microscopic fungi found in bee pollen samples of all prevalent species were Mucor mucedo (49 isolates), Alternaria alternata (40 isolates), Mucor hiemalis (40 isolates), Aspergillus fumigatus (33 isolates) and Cladosporium cladosporioides (31 isolates). The most frequently found isolates were detected in sunflower bee pollen frozen (80 isolates) and the lowest number of isolates was observed in poppy bee pollen dried (12 isolates). The most prevalent mycotoxin of poppy bee pollen was ZON (361.55 ± 0.26 μg.kg(-1)), in rape bee pollen T-2 toxin (265.40 ± 0.18 μg.kg(-1)) and in sunflower bee pollen T-2 toxin (364.72 ± 0.13 μg.kg(-1)) in all cases in frozen samples.

Biomarkers for liver fibrosis

DOEpatents

Jacobs, Jon M.; Burnum-Johnson, Kristin E.; Baker, Erin M.; Smith, Richard D.; Gritsenko, Marina A.; Orton, Daniel

2017-05-16

Methods and systems for diagnosing or prognosing liver fibrosis in a subject are provided. In some examples, such methods and systems can include detecting liver fibrosis-related molecules in a sample obtained from the subject, comparing expression of the molecules in the sample to controls representing expression values expected in a subject who does not have liver fibrosis or who has non-progressing fibrosis, and diagnosing or prognosing liver fibrosis in the subject when differential expression of the molecules between the sample and the controls is detected. Kits for the diagnosis or prognosis of liver fibrosis in a subject are also provided which include reagents for detecting liver fibrosis related molecules.

Biomarkers for liver fibrosis

DOEpatents

Jacobs, Jon M.; Burnum-Johnson, Kristin E.; Baker, Erin M.; Smith, Richard D.; Gritsenko, Marina A.; Orton, Daniel

2015-09-15

Methods and systems for diagnosing or prognosing liver fibrosis in a subject are provided. In some examples, such methods and systems can include detecting liver fibrosis-related molecules in a sample obtained from the subject, comparing expression of the molecules in the sample to controls representing expression values expected in a subject who does not have liver fibrosis or who has non-progressing fibrosis, and diagnosing or prognosing liver fibrosis in the subject when differential expression of the molecules between the sample and the controls is detected. Kits for the diagnosis or prognosis of liver fibrosis in a subject are also provided which include reagents for detecting liver fibrosis related molecules.

Importance of 18F-FDG PET/CT to select patients with nonresectable colorectal liver metastases for liver transplantation.

PubMed

Grut, Harald; Revheim, Mona-Elisabeth; Line, Pål-Dag; Dueland, Svein

2018-04-20

The aim of this study was to evaluate fluorine-18-fluorodeoxyglucose (F-FDG) PET/CT for the selection of patients with nonresectable colorectal liver metastases (NCLM) for liver transplantation (LT). In the secondary cancer study, we reported an improved 5-year overall survival in patients treated with LT for NCLM (56%) compared with chemotherapy (9%). However, many patients were rejected for LT owing to the detection of extrahepatic disease at preoperative imaging. F-FDG PET/CT and contrast-enhanced computed tomography (ceCT) examinations before tentative LT for NCLM were assessed, and findings contraindicating LT were registered. Maximum, mean and peak standardized uptake values; tumor-to-background ratio; metabolic tumor volume; and total lesion glycolysis were measured and calculated for all liver metastases. Overall survival was calculated by the Kaplan-Meier method. Thirty-two patients excluded by F-FDG PET/CT and/or ceCT before tentative LT for NCLM were identified. F-FDG PET/CT from 20 of the 32 excluded patients revealed extrahepatic disease. Eight of the other 12 patients had a negative F-FDG PET/CT finding but were excluded by ceCT. Ten patients were excluded by F-FDG PET/CT only. Four patients were excluded owing to detected malignancy from frozen sections at the start of the intended transplant operation. Tumor-to-background ratio of the liver metastases was significantly higher in patients where F-FDG PET/CT detected extrahepatic disease (P=0.03). The median (range) survival after exclusion was 16 (0-52) months. The ability of F-FDG PET/CT to detect extrahepatic disease before LT for NCLM is vital to establish LT as a treatment option.

The classification of secondary colorectal liver cancer in human biopsy samples using angular dispersive x-ray diffraction and multivariate analysis

NASA Astrophysics Data System (ADS)

Theodorakou, Chrysoula; Farquharson, Michael J.

2009-08-01

The motivation behind this study is to assess whether angular dispersive x-ray diffraction (ADXRD) data, processed using multivariate analysis techniques, can be used for classifying secondary colorectal liver cancer tissue and normal surrounding liver tissue in human liver biopsy samples. The ADXRD profiles from a total of 60 samples of normal liver tissue and colorectal liver metastases were measured using a synchrotron radiation source. The data were analysed for 56 samples using nonlinear peak-fitting software. Four peaks were fitted to all of the ADXRD profiles, and the amplitude, area, amplitude and area ratios for three of the four peaks were calculated and used for the statistical and multivariate analysis. The statistical analysis showed that there are significant differences between all the peak-fitting parameters and ratios between the normal and the diseased tissue groups. The technique of soft independent modelling of class analogy (SIMCA) was used to classify normal liver tissue and colorectal liver metastases resulting in 67% of the normal tissue samples and 60% of the secondary colorectal liver tissue samples being classified correctly. This study has shown that the ADXRD data of normal and secondary colorectal liver cancer are statistically different and x-ray diffraction data analysed using multivariate analysis have the potential to be used as a method of tissue classification.

Prevalence of Campylobacter spp. in poultry and poultry products for sale on the Bulgarian retail market.

PubMed

Stoyanchev, Todor; Vashin, Ivan; Ring, Christian; Atanassova, Viktoria

2007-10-01

The aim of this study was to investigate the presence of Campylobacter spp. in poultry and poultry products available for the consumers at retail markets in Bulgaria. Samples (n = 210) of poultry carcasses and poultry products for sale at the retail market in Bulgaria were analysed for the presence of Campylobacter spp., of these 35 frozen whole carcasses, 135 chilled poultry cuts (45 wing cuts, 45 thigh cuts and 45 fillet) and 40 thermally treated (ready-to-eat) poultry products. The results obtained showed that 35.2% of the frozen poultry carcasses for sale in the markets were Campylobacter contaminated. In the chilled poultry cuts Campylobacter was isolated at the highest percentage in wing- and thigh cuts, 91.1% and 88.9%, respectively. The fillet samples were contaminated by Campylobacter in 48.9% of cases. In the chilled poultry products as well as in the frozen carcasses C. jejuni (74.8%/70.3%) was the most commonly isolated Campylobacter species, with the remainder being C. coli (25.2%/29.7%). Campylobacter spp. were not detected in the thermally treated poultry products.

Twenty-first century brain banking. Processing brains for research: the Columbia University methods

PubMed Central

del Amaya, Maria Pilar; Keller, Christian E.

2007-01-01

Carefully categorized postmortem human brains are crucial for research. The lack of generally accepted methods for processing human postmortem brains for research persists. Thus, brain banking is essential; however, it cannot be achieved at the cost of the teaching mission of the academic institution by routing brains away from residency programs, particularly when the autopsy rate is steadily decreasing. A consensus must be reached whereby a brain can be utilizable for diagnosis, research, and teaching. The best diagnostic categorization possible must be secured and the yield of samples for basic investigation maximized. This report focuses on integrated, novel methods currently applied at the New York Brain Bank, Columbia University, New York, which are designed to reach accurate neuropathological diagnosis, optimize the yield of samples, and process fresh-frozen samples suitable for a wide range of modern investigations. The brains donated for research are processed as soon as possible after death. The prosector must have a good command of the neuroanatomy, neuropathology, and the protocol. One half of each brain is immersed in formalin for performing the thorough neuropathologic evaluation, which is combined with the teaching task. The contralateral half is extensively dissected at the fresh state. The anatomical origin of each sample is recorded using the map of Brodmann for the cortical samples. The samples are frozen at −160°C, barcode labeled, and ready for immediate disbursement once categorized diagnostically. A rigorous organization of freezer space, coupled to an electronic tracking system with its attached software, fosters efficient access for retrieval within minutes of any specific frozen samples in storage. This report describes how this achievement is feasible with emphasis on the actual processing of brains donated for research. PMID:17985145

Potential of DNA methylation in rectal cancer as diagnostic and prognostic biomarkers

PubMed Central

Exner, Ruth; Pulverer, Walter; Diem, Martina; Spaller, Lisa; Woltering, Laura; Schreiber, Martin; Wolf, Brigitte; Sonntagbauer, Markus; Schröder, Fabian; Stift, Judith; Wrba, Fritz; Bergmann, Michael; Weinhäusel, Andreas; Egger, Gerda

2015-01-01

Background: Aberrant DNA methylation is more prominent in proximal compared with distal colorectal cancers. Although a number of methylation markers were identified for colon cancer, yet few are available for rectal cancer. Methods: DNA methylation differences were assessed by a targeted DNA microarray for 360 marker candidates between 22 fresh frozen rectal tumour samples and 8 controls and validated by microfluidic high-throughput and methylation-sensitive qPCR in fresh frozen and formalin-fixed paraffin-embedded (FFPE) samples, respectively. The CpG island methylator phenotype (CIMP) was assessed by MethyLight in FFPE material from 78 patients with pT2 and pT3 rectal adenocarcinoma. Results: We identified and confirmed two novel three-gene signatures in fresh frozen samples that can distinguish tumours from adjacent tissue as well as from blood with a high sensitivity and specificity of up to 1 and an AUC of 1. In addition, methylation of individual CIMP markers was associated with specific clinical parameters such as tumour stage, therapy or patients' age. Methylation of CDKN2A was a negative prognostic factor for overall survival of patients. Conclusions: The newly defined methylation markers will be suitable for early disease detection and monitoring of rectal cancer. PMID:26335606

A Multivariate Evaluation of Factors Affecting the Quality of Freshly Frozen Tissue Specimens.

PubMed

Wang, Tong-Hong; Chen, Chin-Chuan; Liang, Kung-Hao; Chen, Chi-Yuan; Chuang, Wen-Yu; Ueng, Shir-Hwa; Chu, Pao-Hsien; Huang, Chung-Guei; Chen, Tse-Ching; Hsueh, Chuen

2017-08-01

Well-prepared and preserved freshly frozen specimens are indispensable materials for clinical studies. To manage specimen quality and to understand the factors potentially affecting specimen quality during preservation processes, we analyzed the quality of RNA and genomic DNA of various tissues collected between 2002 and 2011 in Linkou Chang Gung Memorial Hospital, Taiwan. During this period, a total of 1059 freshly frozen specimens from eight major cancer categories were examined. It was found that preservation duration, organ origin, and tissue type could all influence the quality of RNA samples. The increased preservation period correlated with decreased RNA quality; the brain, breast, and stomach RNA specimens displayed faster degradation rates than those of other organs, and RNA specimens isolated from tumor tissues were apparently more stable than those of other tissues. These factors could all be used as quality predictors of RNA quality. In contrast, almost all analyses revealed that the genomic DNA samples had good quality, which was not influenced by the aforementioned factors. The results assisted us in determining preservation factors that affect specimen quality, which could provide evidence for improving processes of sample collection and preservation. Furthermore, the results are also useful for researchers to adopt as the evaluation criteria for choosing specimen collection and preservation strategies.

In vitro efficacy of pro- and anticoagulant strategies in compensated and acutely ill patients with cirrhosis.

PubMed

Lisman, Ton; Kleiss, Simone; Patel, Vishal C; Fisher, Caleb; Adelmeijer, Jelle; Bos, Sarah; Singanayagam, Arjuna; Stoy, Sidsel Hyldgaard; Shawcross, Debbie L; Bernal, William

2018-05-16

A simultaneous decline in pro- and anticoagulant drivers in patients with liver diseases results in a 'rebalanced' hemostatic system, even in acutely ill patients. Nevertheless, both bleeding and thrombotic events are common. Here, we explored efficacy of pro- and antihemostatic strategies in compensated and acutely ill cirrhotics which may be unpredictable given the profound hemostatic changes. We tested the effects in vitro of the addition of clinically relevant doses of commonly used pro- and antihemostatic strategies in plasma from healthy individuals (n=30) and patients with compensated (n=18) and acutely decompensated cirrhosis (n=18), and acute-on-chronic liver failure (n=10). We used thrombin generation tests and fibrin clot permeability assays to assess potency of various approaches. Fresh frozen plasma and recombinant factor VIIa modestly increased thrombin generation (10-20%). Prothrombin complex concentrate increased thrombin generation 2-fold in controls and 2-4-fold in patients. Clot permeability decreased after addition of fibrinogen concentrate by 51% in controls and by 50-60% in patients. Low molecular weight heparin decreased thrombin generation by 18% in controls and by 23-54% in patients. Similarly, dabigatran decreased thrombin generation by 33% in controls and by 47-100% in patients. In contrast, rivaroxaban decreased thrombin generation by 55% in controls, but only by 11-38% in patients. These in vitro data suggest little prohemostatic effect of fresh frozen plasma and recombinant factor VIIa in acutely ill cirrhotics, whereas prothrombin complex concentrate and fibrinogen concentrate clearly improved hemostasis. Furthermore, our data suggest the requirement for dose-adjustments of commonly used anticoagulants in these patients. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Veg-01 Plant Harvest

NASA Image and Video Library

2014-06-10

Commander Steve Swanson harvests plants for the VEG-01 investigation. He is harvesting them on the Maintenance Work Area (MWA) in the Node 2/Harmony. The Veg-01 hardware validation test investigation utilizes the Veggie facility on ISS. This investigation will assess on-orbit function and performance of the Veggie,and focus on the growth and development of Outredgeous Lettuce (Lactuca sativa ) seedlings in the spaceflight environment and the effects of the spaceflight environment on composition of microbial flora on the Veggie-grown plants and the Veggie facility. Lettuce plants are harvested on-orbit, frozen at <-80oC and returned to the ground for post-flight evaluation. Microbial sampling swabs will be taken of the Veggie facility and plant material, frozen and returned to the ground for environmental microbiological examination. Rooting pillows and water sample syringes will also be returned for microbial sampling and root analysis.

Cryo-scanning electron microscopy discloses differences in dehydration of frozen boar semen stored in large containers.

PubMed

Ekwall, H

2009-02-01

In general, freezing in flat plastic polyethylene terephthalate (PET) bags (FlatPacks) at 50 degrees C/min gives better post-thaw viability, in terms of sperm motility and membrane integrity, than does freezing in plastic maxi-straws, probably owing to differences in cryobiology. To test the hypothesis that this better survival post-thaw relates to the degree of sperm dehydration during freezing, the present study investigated the structure of boar semen in a frozen state using cryo-scanning electron microscopy (cryo-SEM) to compare two different packages (FlatPacks and maxi-straws) for single artificial insemination (AI) doses, and three different freezing rates. The semen was split-sample frozen in maxi-straws or FlatPacks (both holding 5 ml) using 3% glycerol as cryoprotectant. Three freezing rates were applied from -5 degrees C to -100 degrees C, namely 2 degrees C/min, 50 degrees C/min and 1200 degrees C/min, the lattermost by plunging the samples into liquid nitrogen (LN(2)). The samples were thereafter fractured into LN(2) and larger areas of extra-cellular, unbound frozen water ('ice lakes') were measured to determine the degree of dehydration of the spermatozoa. These areas decreased in size with an increase in cooling rate, the differences in size being more dramatic for maxi-straws than for FlatPacks. Size of ice lakes was also influenced by location within package in relation to cooling rate, the central values being always smaller in maxi-straws than in Flatpacks (p < 0.05 at 2 degrees C/min and 50 degrees C/min) but not at 1200 degrees C/min, which suggested the FlatPack allows for more homogenous freezing of boar semen.

Validation of the Lung Subtyping Panel in Multiple Fresh-Frozen and Formalin-Fixed, Paraffin-Embedded Lung Tumor Gene Expression Data Sets.

PubMed

Faruki, Hawazin; Mayhew, Gregory M; Fan, Cheng; Wilkerson, Matthew D; Parker, Scott; Kam-Morgan, Lauren; Eisenberg, Marcia; Horten, Bruce; Hayes, D Neil; Perou, Charles M; Lai-Goldman, Myla

2016-06-01

Context .- A histologic classification of lung cancer subtypes is essential in guiding therapeutic management. Objective .- To complement morphology-based classification of lung tumors, a previously developed lung subtyping panel (LSP) of 57 genes was tested using multiple public fresh-frozen gene-expression data sets and a prospectively collected set of formalin-fixed, paraffin-embedded lung tumor samples. Design .- The LSP gene-expression signature was evaluated in multiple lung cancer gene-expression data sets totaling 2177 patients collected from 4 platforms: Illumina RNAseq (San Diego, California), Agilent (Santa Clara, California) and Affymetrix (Santa Clara) microarrays, and quantitative reverse transcription-polymerase chain reaction. Gene centroids were calculated for each of 3 genomic-defined subtypes: adenocarcinoma, squamous cell carcinoma, and neuroendocrine, the latter of which encompassed both small cell carcinoma and carcinoid. Classification by LSP into 3 subtypes was evaluated in both fresh-frozen and formalin-fixed, paraffin-embedded tumor samples, and agreement with the original morphology-based diagnosis was determined. Results .- The LSP-based classifications demonstrated overall agreement with the original clinical diagnosis ranging from 78% (251 of 322) to 91% (492 of 538 and 869 of 951) in the fresh-frozen public data sets and 84% (65 of 77) in the formalin-fixed, paraffin-embedded data set. The LSP performance was independent of tissue-preservation method and gene-expression platform. Secondary, blinded pathology review of formalin-fixed, paraffin-embedded samples demonstrated concordance of 82% (63 of 77) with the original morphology diagnosis. Conclusions .- The LSP gene-expression signature is a reproducible and objective method for classifying lung tumors and demonstrates good concordance with morphology-based classification across multiple data sets. The LSP panel can supplement morphologic assessment of lung cancers, particularly when classification by standard methods is challenging.

An improved ATAC-seq protocol reduces background and enables interrogation of frozen tissues.

PubMed

Corces, M Ryan; Trevino, Alexandro E; Hamilton, Emily G; Greenside, Peyton G; Sinnott-Armstrong, Nicholas A; Vesuna, Sam; Satpathy, Ansuman T; Rubin, Adam J; Montine, Kathleen S; Wu, Beijing; Kathiria, Arwa; Cho, Seung Woo; Mumbach, Maxwell R; Carter, Ava C; Kasowski, Maya; Orloff, Lisa A; Risca, Viviana I; Kundaje, Anshul; Khavari, Paul A; Montine, Thomas J; Greenleaf, William J; Chang, Howard Y

2017-10-01

We present Omni-ATAC, an improved ATAC-seq protocol for chromatin accessibility profiling that works across multiple applications with substantial improvement of signal-to-background ratio and information content. The Omni-ATAC protocol generates chromatin accessibility profiles from archival frozen tissue samples and 50-μm sections, revealing the activities of disease-associated DNA elements in distinct human brain structures. The Omni-ATAC protocol enables the interrogation of personal regulomes in tissue context and translational studies.

Hepatitis A Outbreak in Europe: Imported Frozen Berry Mix Suspected to be the Source of At least One Infection in Austria in 2013.

PubMed

Wenzel, J J; Schemmerer, M; Oberkofler, H; Kerschner, H; Sinha, P; Koidl, C; Allerberger, F

2014-12-01

We tested 19 sera from Austrian patients with acute hepatitis A. A serum from a 48-year-old female patient yielded HAV-nucleic acid that showed 99.7% homology to the HAV-sequence obtained from samples taken during the current outbreak in several European countries, which is associated with consumption of frozen berries. So far, Austria was considered not to be affected by this hepatitis A outbreak.

Assessment of the hygienic performances of hamburger patty production processes.

PubMed

Gill, C O; Rahn, K; Sloan, K; McMullen, L M

1997-05-20

The hygienic conditions of the hamburger patties collected from three patty manufacturing plants and six retail outlets were examined. At each manufacturing plant a sample from newly formed, chilled patties and one from frozen patties were collected from each of 25 batches of patties selected at random. At three, two or one retail outlet, respectively, 25 samples from frozen, chilled or both frozen and chilled patties were collected at random. Each sample consisted of 30 g of meat obtained from five or six patties. Total aerobic, coliform and Escherichia coli counts per gram were enumerated for each sample. The mean log (x) and standard deviation (s) were calculated for the log10 values for each set of 25 counts, on the assumption that the distribution of counts approximated the log normal. A value for the log10 of the arithmetic mean (log A) was calculated for each set from the values of x and s. A chi2 statistic was calculated for each set as a test of the assumption of the log normal distribution. The chi2 statistic was calculable for 32 of the 39 sets. Four of the sets gave chi2 values indicative of gross deviation from log normality. On inspection of those sets, distributions obviously differing from the log normal were apparent in two. Log A values for total, coliform and E. coli counts for chilled patties from manufacturing plants ranged from 4.4 to 5.1, 1.7 to 2.3 and 0.9 to 1.5, respectively. Log A values for frozen patties from manufacturing plants were between < 0.1 and 0.5 log10 units less than the equivalent values for chilled patties. Log A values for total, coliform and E. coli counts for frozen patties on retail sale ranged from 3.8 to 8.5, < 0.5 to 3.6 and < 0 to 1.9, respectively. The equivalent ranges for chilled patties on retail sale were 4.8 to 8.5, 1.8 to 3.7 and 1.4 to 2.7, respectively. The findings indicate that the general hygienic condition of hamburgers patties could be improved by their being manufactured from only manufacturing beef of superior hygienic quality, and by the better management of chilled patties at retail outlets.

Effect of controlled and uncontrolled rate of cooling, prior to controlled rate of freezing, on motion characteristics and acrosomal integrity of cryopreserved ram spermatozoa.

PubMed

Joshi, Anil; Kumar, Davendra; Naqvi, S M K; Maurya, V P

2008-12-01

A programmable cell freezer provides ideal cryobiological conditions for controlled-rate cooling and freezing of ram spermatozoa. The purpose of this study was to investigate the effects of controlled (Group 1) and uncontrolled (Group 2) cooling conditions prior to programmable freezing of ram semen on post-thaw sperm motion characteristics and acrosomal integrity of ram spermatozoa. Semen samples of good initial motility obtained from adult Malpura rams were pooled, diluted to 1 × 10(9) spermatozoa per milliliter with Egg yolk-TEST-glycerol extender, and packaged in 0.25 mL straws. Straws representing Group 1 were cooled in a programmable cell freezer from 25°C to 5°C at the rate of -0.15°C per minute followed by a holding time of 2 h for equilibration, while straws of Group 2 were allowed to cool slowly up to 5°C and equilibrate for 2 h in the cold cabinet. After equilibration, straws of Group 2 were also loaded in the cell freezer for freezing straws of both the treatment groups simultaneously from 5°C to -125°C at the rate of -25°C per minute. Thawing of straws was done at 50°C for 10 s and the quality of frozen-thawed spermatozoa was objectively assessed by using sperm motility analyzer. Thawed samples were also evaluated for acrosomal integrity after staining the dried semen smears with Giemsa stain. The average post-thaw motility of straws was significantly higher (P < 0.05) in samples frozen after controlled cooling, compared with samples frozen after uncontrolled rate of cooling. The percent of spermatozoa with normal acrosome was also significantly (P < 0.05) higher in Group 1, compared to Group 2. The results indicate that controlled-rate cooling has a significant effect on post-thaw motility and acrosomal integrity of frozen-thawed ram spermatozoa, compared to uncontrolled-rate cooling prior to programmable freezing.

Repurposing Archival Samples for Investigating Toxicological Modes of Action

EPA Science Inventory

Little is known about formalin fixation induced genomic artifacts, limiting the use of formalin-fixed paraffin-embedded (FFPE) samples in toxicological and clinical studies. Previously, we identified a consistent shift in transcriptional profiles between paired frozen and FFPE sa...

Spatial Systems Lipidomics Reveals Nonalcoholic Fatty Liver Disease Heterogeneity

PubMed Central

2018-01-01

Hepatocellular lipid accumulation characterizes nonalcoholic fatty liver disease (NAFLD). However, the types of lipids associated with disease progression are debated, as is the impact of their localization. Traditional lipidomics analysis using liver homogenates or plasma dilutes and averages lipid concentrations, and does not provide spatial information about lipid distribution. We aimed to characterize the distribution of specific lipid species related to NAFLD severity by performing label-free molecular analysis by mass spectrometry imaging (MSI). Fresh frozen liver biopsies from obese subjects undergoing bariatric surgery (n = 23) with various degrees of NAFLD were cryosectioned and analyzed by matrix-assisted laser desorption/ionization (MALDI)-MSI. Molecular identification was verified by tandem MS. Tissue sections were histopathologically stained, annotated according to the Kleiner classification, and coregistered with the MSI data set. Lipid pathway analysis was performed and linked to local proteome networks. Spatially resolved lipid profiles showed pronounced differences between nonsteatotic and steatotic tissues. Lipid identification and network analyses revealed phosphatidylinositols and arachidonic acid metabolism in nonsteatotic regions, whereas low–density lipoprotein (LDL) and very low–density lipoprotein (VLDL) metabolism was associated with steatotic tissue. Supervised and unsupervised discriminant analysis using lipid based classifiers outperformed simulated analysis of liver tissue homogenates in predicting steatosis severity. We conclude that lipid composition of steatotic and nonsteatotic tissue is highly distinct, implying that spatial context is important for understanding the mechanisms of lipid accumulation in NAFLD. MSI combined with principal component–linear discriminant analysis linking lipid and protein pathways represents a novel tool enabling detailed, comprehensive studies of the heterogeneity of NAFLD. PMID:29570976

Skin temperature response to cryotherapy.

PubMed

Chesterton, Linda S; Foster, Nadine E; Ross, Lesley

2002-04-01

To compare the localized skin-cooling effects of 2 cryotherapy modalities and to review the clinical relevance of the results. Randomized controlled trial with repeated measures. Laboratory experiment. Convenience sample of 20 volunteers (13 women, 7 men), ages 21.3 to 44 years (mean, 31.3 +/- 6.8 y). A flexible frozen gel pack, a 454 g packet of frozen peas, or a control applied to the anterior thigh. No blinding was undertaken. Surface skin temperature under the modality at baseline and 10 and 20 minutes after application. Significant effects were recorded for modality (F(2) = 290.56, P <.0001), time (F(1.27) = 1868.07, P <.0001), and their interaction (F(2.09) = 305.47, P <.0001). After 20 minutes, frozen peas produced the lowest mean skin temperature +/- standard deviation of 10.8 degrees C +/- 2.28 degrees C compared with 14.4 degrees C +/- 2.53 degrees C from the gel pack and 26.1 degrees C +/- 1.75 degrees C from the control. Skin temperature fell between both time periods with the application of frozen peas but stabilized after 10 minutes of gel pack and control application. Application of frozen peas produced mean skin temperatures adequate to induce localized skin analgesia, to reduce nerve conduction velocity, and to reduce metabolic enzyme activity to clinically relevant levels. Flexible frozen gel packs did not cool skin sufficiently to achieve these levels. Copyright 2002 by the American Congress of Rehabilitation Medicine and the American Academy of Physical Medicine and Rehabilitation

Ultrastructure in frozen/etched saline solutions: on the internal cleansing of ice.

PubMed

Menger, Fredric M; Galloway, Ashley L; Chlebowski, Mary E; Apkarian, Robert P

2004-05-19

Seawater, with its 3.5% salt content, freezes into hexagonal ice (Ih) that encloses concentrated brine within its matrix. When unsubmerged sea ice reaches a certain height and temperature, the brine drains downward through narrow channels. This mechanism was now modeled by frozen 2-3.5% saline as investigated by cryo-etch high-resolution secondary electron microscopy. Thus, saline was either plunge-frozen in liquid ethane at -183 degrees C or else high-pressure frozen to -105 degrees C in 5-6 ms. Ice from a freshly exposed surface was then subjected to a high-vacuum sublimation ("etching"), a procedure that removes pure bulk ice in preference to ice from frozen hydrated salt. After chromium-coating the etched surface with a 2-nm film, the sample was examined by cryo-HRSEM. Granular icy "fences" were seen surrounding empty areas where amorphous ice had originally resided. Since the fences, about 1-2 mum high, survived the etching, it is likely that they consist of frozen brine. The presence of such fences suggests that, during freezing, saline can purge itself of salt with remarkable speed (5-6 ms). Alternatively, channels (perhaps routed around submicroscopic crystallites of cubic ice (Ic) embedded in the amorphous ice at -105 degrees C) can guide the migration of salt to the periphery of ice patches. Macromolecules fail to form fences because they diffuse too slowly or because they are too large to pass through the channels.

A comparison of digitized frozen section and smear preparations for intraoperative neurotelepathology.

PubMed

Gould, Peter V; Saikali, Stephan

2012-01-01

Intraoperative consultations in neuropathology are often assessed by smear preparations rather than by frozen sections. Both techniques are standard practice for light microscopic examination on site, but there is little data comparing these techniques in a telepathology setting. Thirty cases of brain tumours submitted for intraoperative consultation at our institution between July and December 2010 were identified in which both frozen section and tissue smear preparations were available for digitization at 20× magnification. Slides were digitized using a Hamamatsu Nanozoomer 2.0 HT whole slide scanner, and resulting digital images were visualized at 1680 × 1050 pixel resolution with NDP. view software. The original intraoperative diagnosis was concordant with the sign out diagnosis in 29/30 cases; one tumeur was initially interpreted as a high grade glioma but proved to be a lymphoma at sign out. Digitized frozen section slides were sufficient for diagnosis at 10× magnification in 27/30 cases. Digitized tissue smears were sufficient for diagnosis at 10× magnification in 28/30 cases. In two cases tumour was present on the tissue smear but not the frozen section (one case of recurrent astrocytoma, one case of meningeal carcinomatosis). In one case of lymphoma, tumour was present on frozen section only. These discrepancies were attributed to tissue sampling rather than image quality. Examination of digitized slides at higher magnfication (20×) permitted confirmation of mitoses and Rosenthal fibers on tissue smear preparations, but did not change the primary diagnosis. Intra-slide variations in tissue thickness on smear preparations led to variable loss of focus in digitized images, but did not affect image quality in thinner areas of the smear or impede diagnosis. Digitized tissue smears are suitable for intraoperative neurotelepathology and provide comparable information to digitized frozen sections at medium power magnification.

Comparing the acidities of aqueous, frozen, and freeze-dried phosphate buffers: Is there a "pH memory" effect?

PubMed

Vetráková, Ľubica; Vykoukal, Vít; Heger, Dominik

2017-09-15

The concept of "pH memory" has been established in the literature for the correlation between the pH of a pre-lyophilization solution and the ionization state of freeze-dried powder (lyophile). In this paper, the concept of "pH memory" is explored for the system of an aqueous solution, a frozen solution, and a lyophile. Sodium and potassium phosphate buffers in the pH range of 5-9 were frozen and lyophilized with sulfonephthalein indicators as acidity probes, and their Hammett acidity functions were compared to the initial pH of the aqueous solution. The results show that the acidities of the lyophiles are somewhat changed compared to the initial pHs, but the acidities in the frozen state differ more substantially. The Hammett acidity functions of the frozen buffers were found to be markedly dissimilar from the initial pH, especially in the sodium phosphate frozen at 233K, where an increase in the initial pH led to a decrease in the Hammett acidity function of the frozen state at a certain pH range. The large acidification observed after freezing the sodium phosphate buffer was not detected in the lyophiles after the sample had been dried; the phenomenon is explained considering the formed crystals analyzed by X-ray powder diffraction. The results suggest that monitoring the final acidity of a lyophile is not sufficient to predict all the acidity changes throughout the whole lyophilization process. The importance of well-controlled freezing and lyophilization conditions follows from the results of the research. Copyright © 2017 Elsevier B.V. All rights reserved.

Image-guided intervention in the coagulopathic patient.

PubMed

Kohli, Marc; Mayo-Smith, William; Zagoria, Ronald; Sandrasegaran, Kumar

2016-04-01

Determining practice parameters for interventional procedures is challenging due to many factors including unreliable laboratory tests to measure bleeding risk, variable usage of standardized terminology for adverse events, poorly defined standards for administration of blood products, and the growing numbers of anticoagulant and antiplatelet medications. We aim to address these and other issues faced by radiologists performing invasive procedures through a review of available literature, and experiential guidance from three academic medical centers. We discuss the significant limitations with respect to using prothrombin-time and international normalized ratio to measure bleeding risk, especially in patients with synthetic defects due to liver function. Factors affecting platelet function including the impact of uremia; recent advances in laboratory testing, including platelet function testing; and thromboelastography are also discussed. A review of the existing literature of fresh-frozen plasma replacement therapy is included. The literature regarding comorbidities affecting coagulation including malignancy, liver failure, and uremia are also reviewed. Finally, the authors present a set of recommendations for laboratory thresholds, corrective transfusions, as well as withholding and restarting medications.

Suicidal intoxication with potassium chlorate successfully treated with renal replacement therapy and extracorporeal liver support.

PubMed

Sein Anand, Jacek; Barwina, Małgorzata; Zajac, Maciej; Kaletha, Krystian

2012-01-01

We present a case of a 22-year-old male who, in a suicide attempt, ingested approximately 200 g of potassium chlorate. Upon admission to the hospital, he presented in full respiratory failure with cyanosis. Methylene blue antidote was given but found to be ineffective. The patient was intubated and mechanical ventilation was initiated. Because of renal failure with anuria, intermittent haemodialysis (iHD) followed by continuous venovenous hemodiafiltration (CVVHDF) was performed. His hospital stay was also complicated by hemolysis, disseminated intravascular coagulation, and atrial fibrillation. Transfusions of packed red blood cells, platelets, and fresh frozen plasma were necessary to correct the deficits. He also developed liver failure and required two sessions of molecular adsorbent recirculating system (MARS) therapy. On day 14 of his hospitalization, he regained consciousness, as well as full respiratory and circulatory function. There are no controlled studies addressing management of potassium chlorate poisoning. We suggest that early renal replacement therapy should be strongly considered.

HCV-RNA quantification in liver bioptic samples and extrahepatic compartments, using the abbott RealTime HCV assay.

PubMed

Antonucci, FrancescoPaolo; Cento, Valeria; Sorbo, Maria Chiara; Manuelli, Matteo Ciancio; Lenci, Ilaria; Sforza, Daniele; Di Carlo, Domenico; Milana, Martina; Manzia, Tommaso Maria; Angelico, Mario; Tisone, Giuseppe; Perno, Carlo Federico; Ceccherini-Silberstein, Francesca

2017-08-01

We evaluated the performance of a rapid method to quantify HCV-RNA in the hepatic and extrahepatic compartments, by using for the first time the Abbott RealTime HCV-assay. Non-tumoral (NT), tumoral (TT) liver samples, lymph nodes and ascitic fluid from patients undergoing orthotopic-liver-transplantation (N=18) or liver resection (N=4) were used for the HCV-RNA quantification; 5/22 patients were tested after or during direct acting antivirals (DAA) treatment. Total RNA and DNA quantification from tissue-biopsies allowed normalization of HCV-RNA concentrations in IU/μg of total RNA and IU/10 6 liver-cells, respectively. HCV-RNA was successfully quantified with high reliability in liver biopsies, lymph nodes and ascitic fluid samples. Among the 17 untreated patients, a positive and significant HCV-RNA correlation between serum and NT liver-samples was observed (Pearson: rho=0.544, p=0.024). Three DAA-treated patients were HCV-RNA "undetectable" in serum, but still "detectable" in all tested liver-tissues. Differently, only one DAA-treated patient, tested after sustained-virological-response, showed HCV-RNA "undetectability" in liver-tissue. HCV-RNA was successfully quantified with high reliability in liver bioptic samples and extrahepatic compartments, even when HCV-RNA was "undetectable" in serum. Abbott RealTime HCV-assay is a good diagnostic tool for HCV quantification in intra- and extra-hepatic compartments, whenever a bioptic sample is available. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of sanitation, freezing and frozen storage on enteric viruses in berries and herbs.

PubMed

Butot, S; Putallaz, T; Sánchez, G

2008-08-15

Norovirus (NV) and hepatitis A virus (HAV) are foodborne enteric viruses associated with outbreaks of disease following consumption of fresh or frozen produce. Model experiments were performed to determine the effectiveness of certain commercial processes for the removal of enteric viruses that might be present in berries and herbs. The survival and persistence of HAV, NV, rotavirus (RV) and feline calicivirus (FCV), a surrogate for NV, in frozen produce over time were determined. Survival and inactivation of HAV, RV and FCV were assessed by viral culture and quantitative reverse transcription-PCR (RT-PCR), whereas NV persistence was determined by quantitative RT-PCR only. Freezing did not significantly reduce the viability of any of the viruses except the infectivity of FCV in strawberries. Frozen storage for 3 months had limited effects on HAV and RV survival in all tested food products, whereas in frozen raspberries and strawberries FCV infectivity showed the highest decay rate due to acid pH. To simulate postharvesting conditions, fresh berries and herbs were rinsed with tap, warm or chlorinated water or with a chlorine dioxide (ClO(2)) solution. Available chlorine at a concentration of 200 ppm and ClO(2) at 10 ppm reduced measurable enteric viruses in raspberry and parsley samples by less than 2 log(10) units.

MRI-determined liver proton density fat fraction, with MRS validation: Comparison of regions of interest sampling methods in patients with type 2 diabetes.

PubMed

Vu, Kim-Nhien; Gilbert, Guillaume; Chalut, Marianne; Chagnon, Miguel; Chartrand, Gabriel; Tang, An

2016-05-01

To assess the agreement between published magnetic resonance imaging (MRI)-based regions of interest (ROI) sampling methods using liver mean proton density fat fraction (PDFF) as the reference standard. This retrospective, internal review board-approved study was conducted in 35 patients with type 2 diabetes. Liver PDFF was measured by magnetic resonance spectroscopy (MRS) using a stimulated-echo acquisition mode sequence and MRI using a multiecho spoiled gradient-recalled echo sequence at 3.0T. ROI sampling methods reported in the literature were reproduced and liver mean PDFF obtained by whole-liver segmentation was used as the reference standard. Intraclass correlation coefficients (ICCs), Bland-Altman analysis, repeated-measures analysis of variance (ANOVA), and paired t-tests were performed. ICC between MRS and MRI-PDFF was 0.916. Bland-Altman analysis showed excellent intermethod agreement with a bias of -1.5 ± 2.8%. The repeated-measures ANOVA found no systematic variation of PDFF among the nine liver segments. The correlation between liver mean PDFF and ROI sampling methods was very good to excellent (0.873 to 0.975). Paired t-tests revealed significant differences (P < 0.05) with ROI sampling methods that exclusively or predominantly sampled the right lobe. Significant correlations with mean PDFF were found with sampling methods that included higher number of segments, total area equal or larger than 5 cm(2) , or sampled both lobes (P = 0.001, 0.023, and 0.002, respectively). MRI-PDFF quantification methods should sample each liver segment in both lobes and include a total surface area equal or larger than 5 cm(2) to provide a close estimate of the liver mean PDFF. © 2015 Wiley Periodicals, Inc.

Growth hormone (GH) hypersecretion and GH receptor resistance in streptozotocin diabetic mice in response to a GH secretagogue.

PubMed

Johansen, Peter B; Segev, Yael; Landau, Daniel; Phillip, Moshe; Flyvbjerg, Allan

2003-01-01

The growth hormone (GH) and insulin-like growth factor I (IGF-I) axis were studied in streptozotocin (STZ) diabetic and nondiabetic female mice following intravenous (IV) injection of the GH secretagogue (GHS) ipamorelin or saline. On day 14, blood samples were obtained before and 10 minutes after the injection. Livers were removed and frozen for determination of the mRNA expressions of the GH receptor, GH-binding protein, and IGF-I, and hepatic IGF-I peptide. Serum samples were analyzed for GH and IGF-I. Following ipamorelin injection, the GH levels were found to be 150 +/- 35 microg/L and 62 +/- 11 microg/L in the diabetic compared to the nondiabetic mice (P <.05). Serum IGF-I levels were lower in diabetic than in nondiabetic animals, and rose after stimulation only in the nondiabetic animals. Furthermore, hepatic GH resistance and IGF-I mRNA levels and IGF-I peptide were increased in nondiabetic animals in response to GH stimulation, whereas the low levels per se of all these parameters in diabetic mice were unaffected. The study shows that STZ diabetic mice demonstrate a substantial part of the clinical features of type 1 diabetes in humans, including GH hypersecretion and GH resistance. Accordingly, it is proposed that STZ diabetic mice may be a better model of the perturbations of the GH/IGF-I axis in diabetes than STZ diabetic rats.

Determination and assessment of total mercury levels in local, frozen and canned fish in Lebanon.

PubMed

Obeid, Pierre J; El-Khoury, Bilal; Burger, Joanne; Aouad, Samer; Younis, Mira; Aoun, Amal; El-Nakat, John Hanna

2011-01-01

Fish is an important constituent of the Lebanese diet. However, very little attention in our area is given to bring awareness regarding the effect of the toxicity of mercury (Hg) mainly through fish consumption. This study aimed to report analytical data on total mercury levels in several fish species for the first time in thirty years and to also made individuals aware of the presence and danger from exposure to mercury through fish consumption. Fish samples were selected from local Lebanese markets and fisheries and included 94 samples of which were fresh, frozen, processed, and canned fish. All values were reported as microgram of mercury per gram of fish based on wet weight. The level of mercury ranged from 0.0190 to 0.5700 microg/g in fresh samples, 0.0059 to 0.0665 microg/g in frozen samples, and 0.0305 to 0.1190 microg/g in canned samples. The data clearly showed that higher levels of mercury were detected in local fresh fish as opposed to other types thus placing consumers at higher risk from mercury exposure. Moreover, the data revealed that Mallifa (yellowstripe barracuda/Sphyraena chrysotaenia), Sargous (white seabream/Diplodus sargus), Ghobbos (bogue/Boops boops), and shrimp (Penaeus sp.) were among the types containing the highest amounts of mercury. On the other hand, processed fish such as fish fillet, fish burger, small shrimp and crab are found to contain lower levels of mercury and are associated with lower exposure risks to mercury. Lebanese population should therefore, be aware to consume limited amounts of fresh local fish to minimize exposure to mercury.

The effect of post-exsanguination infusion on the composition, exudation, color and post-mortem metabolic changes in lamb.

PubMed

Farouk, M M; Price, J F

1994-01-01

Twenty-four lamb carcasses were assigned to three treatment groups: (1) control (Ctr), (2) infused with 10% (vol/wt) of a tenderizing blend (NCa), and (3) NCa plus 0·015 m CaCl(2) (WCa). Results indicated that the infused carcass solution was retained in the following order: shoulder > lion > leg. Infusion had no effect (P > 0·05) on drip and cooking losses in refrigerated samples. Samples frozen and then thawed from infused carcasses had greater thaw drip (P < 0·05) and cooking losses (P < 0·01) than control samples. The amounts of drip and cooking losses were in the order: WCa > NCa > Ctr. Frozen storage preserved the red color but lowered the lightness and yellowness of ovine muscles; the opposite effect was observed following refrigerated storage. Infused samples were lighter and yellower than control in both fresh and frozen samples (P < 0·01). WCa had less red color (P < 0·01) than NCa and Ctr at all times and storage conditions. Infusion lowered (P < 0·05) the temperature of carcasses over the first 3 h postmortem (pm) compared with Ctr. The rate of glycolysis was higher in infraspinatus (IS) than in longissimus thoracis et lumborum muscle (LTL or longissimus). In both IS and LTL, glycolysis was completed within the first 6 h postmortem in NCa, whereas in Ctr and WCa, it took 12-24 h for glycolysis to be completed. The rate of glycolysis was in the order: NCa > WCa > Ctr. Copyright © 1994. Published by Elsevier Ltd.

Evaluation of Nucleic Acid Stabilization Products for Ambient Temperature Shipping and Storage of Viral RNA and Antibody in a Dried Whole Blood Format

PubMed Central

Dauner, Allison L.; Gilliland, Theron C.; Mitra, Indrani; Pal, Subhamoy; Morrison, Amy C.; Hontz, Robert D.; Wu, Shuenn-Jue L.

2015-01-01

Loss of sample integrity during specimen transport can lead to false-negative diagnostic results. In an effort to improve upon the status quo, we used dengue as a model RNA virus to evaluate the stabilization of RNA and antibodies in three commercially available sample stabilization products: Whatman FTA Micro Cards (GE Healthcare Life Sciences, Pittsburgh, PA), DNAstāble Blood tubes (Biomātrica, San Diego, CA), and ViveST tubes (ViveBio, Alpharetta, GA). Both contrived and clinical dengue-positive specimens were stored on these products at ambient temperature or 37°C for up to 1 month. Antibody and viral RNA levels were measured by enzyme-linked immunosorbent assay (ELISA) and quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays, respectively, and compared with frozen unloaded controls. We observed reduced RNA and antibody levels between stabilized contrived samples and frozen controls at our earliest time point, and this was particularly pronounced for the FTA cards. However, despite some time and temperature dependent loss, a 94.6–97.3% agreement was observed between stabilized clinical specimens and their frozen controls for all products. Additional considerations such as cost, sample volume, matrix, and ease of use should inform any decision to incorporate sample stabilization products into a diagnostic testing workflow. We conclude that DNAstāble Blood and ViveST tubes are useful alternatives to traditional filter paper for ambient temperature shipment of clinical specimens for downstream molecular and serological testing. PMID:25940193

Procedures for cryogenic X-ray ptychographic imaging of biological samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yusuf, M.; Zhang, F.; Chen, B.

Biological sample-preparation procedures have been developed for imaging human chromosomes under cryogenic conditions. A new experimental setup, developed for imaging frozen samples using beamline I13 at Diamond Light Source, is described. This paper describes the equipment and experimental procedures as well as the authors' first ptychographic reconstructions using X-rays.

Procedures for cryogenic X-ray ptychographic imaging of biological samples

DOE PAGES

Yusuf, M.; Zhang, F.; Chen, B.; ...

2017-01-12

Biological sample-preparation procedures have been developed for imaging human chromosomes under cryogenic conditions. A new experimental setup, developed for imaging frozen samples using beamline I13 at Diamond Light Source, is described. This paper describes the equipment and experimental procedures as well as the authors' first ptychographic reconstructions using X-rays.

Compact cold stage for micro-computerized tomography imaging of chilled or frozen samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hullar, Ted; Anastasio, Cort, E-mail: canastasio@ucdavis.edu; Paige, David F.

2014-04-15

High resolution X-ray microCT (computerized tomography) can be used to image a variety of objects, including temperature-sensitive materials. In cases where the sample must be chilled or frozen to maintain sample integrity, either the microCT machine itself must be placed in a refrigerated chamber, or a relatively expensive commercial cold stage must be purchased. We describe here the design and construction of a low-cost custom cold stage suitable for use in a microCT imaging system. Our device uses a boron nitride sample holder, two-stage Peltier cooler, fan-cooled heat sink, and electronic controller to maintain sample temperatures as low as −25 °Cmore » ± 0.2 °C for the duration of a tomography acquisition. The design does not require modification to the microCT machine, and is easily installed and removed. Our custom cold stage represents a cost-effective solution for refrigerating CT samples for imaging, and is especially useful for shared equipment or machines unsuitable for cold room use.« less

Validation of the Complexity INdex in SARComas prognostic signature on formalin-fixed, paraffin-embedded, soft tissue sarcomas.

PubMed

Le Guellec, S; Lesluyes, T; Sarot, E; Valle, C; Filleron, T; Rochaix, P; Valentin, T; Pérot, G; Coindre, J-M; Chibon, F

2018-05-31

Prediction of metastatic outcome in sarcomas is challenging for clinical management since they are aggressive and carry a high metastatic risk. A 67-gene expression signature, the Complexity INdex in SARComas (CINSARC), has been identified as a better prognostic factor than the reference pathological grade. Since it cannot be applied easily in standard laboratory practice, we assessed its prognostic value using nanoString on formalin-fixed, paraffin-embedded (FFPE) blocks to evaluate its potential in clinical routine practice and guided therapeutic management. A code set consisting of 67 probes derived from the 67 genes of the CINSARC signature was built and named NanoCind®. To compare the performance of RNA-seq and nanoString (NanoCind®), we used expressions of various sarcomas (n=124, frozen samples) using both techniques and compared predictive values based on CINSARC risk groups and clinical annotations. We also used nanoString on FFPE blocks (n=67) and matching frozen and FFPE samples (n=45) to compare their level of agreement. Metastasis-free survival and agreement values in classification groups were evaluated. CINSARC strongly predicted metastatic outcome using nanoString on frozen samples (HR = 2.9, 95% CI 1.23-6.82) with similar risk-group classifications (86%). While more than 50% of FFPE blocks were not analyzable by RNA-seq owing to poor RNA quality, all samples were analyzable with nanoString. When similar (risk-group) classifications were measured with frozen tumors (RNA-seq) compared to FFPE blocks (84% agreement), the CINSARC signature was still a predictive factor of metastatic outcome with nanoString on FFPE samples (HR = 4.43, 95% CI 1.25-15.72). CINSARC is a material-independent prognostic signature for metastatic outcome in sarcomas and outperforms histological grade. Unlike RNA-seq, nanoString is not influenced by the poor quality of RNA extracted from FFPE blocks. The CINSARC signature can potentially be used in combination with nanoString (NanoCind®) in routine clinical practice on FFPE blocks to predict metastatic outcome.

Non-heat-treated frozen raspberries the most likely vehicle of a norovirus outbreak in Oslo, Norway, November 2013.

PubMed

Einöder-Moreno, M; Lange, H; Grepp, M; Osborg, E; Vainio, K; Vold, L

2016-10-01

In November 2013, the Norwegian Institute of Public Health was notified of a gastroenteritis outbreak following two meetings held at a conference centre. Identical food and beverages were served during the meetings. We investigated in order to identify the vehicle of infection and implement control measures. Meeting participants completed an online questionnaire on consumption of foods and beverages. We asked symptomatic participants to provide a stool sample. We defined a case as diarrhoea and/or vomiting in a participant who became ill within 3 days after the meeting. We calculated attack rates (AR) and adjusted risk ratios (aRR) with 95% confidence intervals (CI) using binomial regression. We conducted environmental investigations. Overall, 147/168 (88%) participants responded, of which 74 (50%) met the case definition. All five stool samples provided were norovirus positive. No kitchen staff reported being sick. Risk of illness was higher in those who consumed raspberry mousse (aRR 3·4, 95% CI 1·4-8·2) and sliced fresh fruit (aRR 1·9, 95% CI 1·3-2·8). Seventy cases (95%) ate raspberry mousse. Frozen raspberries used for the mousse were imported and not heat-treated before consumption. Non-heat-treated frozen raspberries were the most likely outbreak vehicle. Contamination by a food handler could not be excluded. We recommend heat-treatment of imported frozen berries before consumption.

Effects of semen storage and separation techniques on sperm DNA fragmentation.

PubMed

Jackson, Robert E; Bormann, Charles L; Hassun, Pericles A; Rocha, André M; Motta, Eduardo L A; Serafini, Paulo C; Smith, Gary D

2010-12-01

To determine the effect of semen storage and separation techniques on sperm DNA fragmentation. Controlled clinical study. An assisted reproductive technology laboratory. Thirty normoozospermic semen samples obtained from patients undergoing infertility evaluation. One aliquot from each sample was immediately prepared (control) for the sperm chromatin dispersion assay (SCD). Aliquots used to assess storage techniques were treated in the following ways: snap frozen by liquid nitrogen immersion, slow frozen with Tris-yolk buffer and glycerol, kept on ice for 24 hours or maintained at room temperature for 4 and 24 hours. Aliquots used to assess separation techniques were processed by the following methods: washed and centrifuged in media, swim-up from washed sperm pellet, density gradient separation, density gradient followed by swim-up. DNA integrity was then measured by SCD. DNA fragmentation as measured by SCD. There was no significant difference in fragmentation among the snap frozen, slow frozen, and wet-ice groups. Compared to other storage methods short-term storage at room temperature did not impact DNA fragmentation yet 24 hours storage significantly increased fragmentation. Swim-up, density gradient and density gradient/swim-up had significantly reduced DNA fragmentation levels compared with washed semen. Postincubation, density gradient/swim-up showed the lowest fragmentation levels. The effect of sperm processing methods on DNA fragmentation should be considered when selecting storage or separation techniques for clinical use. Copyright © 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Efficient quantification of the health-relevant anthocyanin and phenolic acid profiles in commercial cultivars and breeding selections of blueberries ( Vaccinium spp.).

PubMed

Yousef, Gad G; Brown, Allan F; Funakoshi, Yayoi; Mbeunkui, Flaubert; Grace, Mary H; Ballington, James R; Loraine, Ann; Lila, Mary A

2013-05-22

Anthocyanins and phenolic acids are major secondary metabolites in blueberry with important implications for human health maintenance. An improved protocol was developed for the accurate, efficient, and rapid comparative screening for large blueberry sample sets. Triplicates of six commercial cultivars and four breeding selections were analyzed using the new method. The compound recoveries ranged from 94.2 to 97.5 ± 5.3% when samples were spiked with commercial standards prior to extraction. Eighteen anthocyanins and 4 phenolic acids were quantified in frozen and freeze-dried fruits. Large variations for individual and total anthocyanins, ranging from 201.4 to 402.8 mg/100 g, were assayed in frozen fruits. The total phenolic acid content ranged from 23.6 to 61.7 mg/100 g in frozen fruits. Across all genotypes, freeze-drying resulted in minor reductions in anthocyanin concentration (3.9%) compared to anthocyanins in frozen fruits. However, phenolic acids increased by an average of 1.9-fold (±0.3) in the freeze-dried fruit. Different genotypes frequently had comparable overall levels of total anthocyanins and phenolic acids, but differed dramatically in individual profiles of compounds. Three of the genotypes contained markedly higher concentrations of delphinidin 3-O-glucoside, cyanidin 3-O-glucoside, and malvidin 3-O-glucoside, which have previously been implicated as bioactive principles in this fruit. The implications of these findings for human health benefits are discussed.

Semen of spinal cord injured men freezes reliably.

PubMed

Padron, O F; Brackett, N L; Weizman, M S; Lynne, C M

1994-01-01

The objectives of the present study were to: 1) determine the effect of cryopreservation on the percent and the grade of motility of sperm from spinal cord injured (SCI) men and 2) determine which method of freezing yields the best post-thaw motility in sperm from SCI men. Antegrade semen samples were obtained from 9 SCI subjects and 10 age-matched healthy control subjects. Motility in fresh samples was determined and cryopreservative medium was added to each sample. Aliquots of each sample were frozen according to three methods: 1) liquid nitrogen vapor only (V); 2) vapor for 12 minutes followed by submersion into liquid nitrogen (V+N2); and 3) direct submersion into liquid nitrogen (N2). Samples were frozen for 1 week, then thawed. The post-thaw percent and grade of motility was determined. The mean percent motility of fresh samples for SCI subjects (21.0%) was significantly lower than for control subjects (55.7%). After thawing, the mean percent drop in motility for V, V+N2, and N2 for controls was 65.2%, 73.5%, and 79.4%, respectively, and for SCI subjects, it was 64.7%, 74.5%, and 81.6%, respectively. There was no statistically significant difference between control and SCI subjects by method of freezing. Vapor only as a freezing method was superior to all other methods for retention of sperm motility in both control and SCI subjects. We conclude that the semen of SCI men may be frozen reliably and that their sperm retain motility similar to that of normal men. Vapor only, being the most gentle method used, gives the best recovery of sperm motility in either group.

High-volume plasma exchange in patients with acute liver failure: An open randomised controlled trial.

PubMed

Larsen, Fin Stolze; Schmidt, Lars Ebbe; Bernsmeier, Christine; Rasmussen, Allan; Isoniemi, Helena; Patel, Vishal C; Triantafyllou, Evangelos; Bernal, William; Auzinger, Georg; Shawcross, Debbie; Eefsen, Martin; Bjerring, Peter Nissen; Clemmesen, Jens Otto; Hockerstedt, Krister; Frederiksen, Hans-Jørgen; Hansen, Bent Adel; Antoniades, Charalambos G; Wendon, Julia

2016-01-01

Acute liver failure (ALF) often results in cardiovascular instability, renal failure, brain oedema and death either due to irreversible shock, cerebral herniation or development of multiple organ failure. High-volume plasma exchange (HVP), defined as exchange of 8-12 or 15% of ideal body weight with fresh frozen plasma in case series improves systemic, cerebral and splanchnic parameters. In this prospective, randomised, controlled, multicentre trial we randomly assigned 182 patients with ALF to receive either standard medical therapy (SMT; 90 patients) or SMT plus HVP for three days (92 patients). The baseline characteristics of the groups were similar. The primary endpoint was liver transplantation-free survival during hospital stay. Secondary-endpoints included survival after liver transplantation with or without HVP with intention-to-treat analysis. A proof-of-principle study evaluating the effect of HVP on the immune cell function was also undertaken. For the entire patient population, overall hospital survival was 58.7% for patients treated with HVP vs. 47.8% for the control group (hazard ratio (HR), with stratification for liver transplantation: 0.56; 95% confidence interval (CI), 0.36-0.86; p=0.0083). HVP prior to transplantation did not improve survival compared with patients who received SMT alone (CI 0.37 to 3.98; p=0.75). The incidence of severe adverse events was similar in the two groups. Systemic inflammatory response syndrome (SIRS) and sequential organ failure assessment (SOFA) scores fell in the treated group compared to control group, over the study period (p<0.001). Treatment with HVP improves outcome in patients with ALF by increasing liver transplant-free survival. This is attributable to attenuation of innate immune activation and amelioration of multi-organ dysfunction. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Liver Zonation Index of Drug Transporter and Metabolizing Enzyme Protein Expressions in Mouse Liver Acinus.

PubMed

Tachikawa, Masanori; Sumiyoshiya, Yuna; Saigusa, Daisuke; Sasaki, Kazunari; Watanabe, Michitoshi; Uchida, Yasuo; Terasaki, Tetsuya

2018-05-01

The purpose of the present study was to clarify the molecular basis of zonated drug distributions in mouse liver based on the protein expression levels of transporters and metabolizing enzymes in periportal (PP) and pericentral (PC) vein regions of mouse hepatic lobules. The distributions of sulforhodamine 101 (SR-101), a substrate of organic anion transporting polypeptides (Oatps), and ribavirin, a substrate of equilibrative nucleoside transporter 1 (Ent1), were elucidated in frozen liver sections of mice, to which each compound had been intravenously administered. Regions strongly positive for SR-101 (SR-101 + ) and regions weakly positive or negative for SR-101 (SR-101 - ) were separated by laser microdissection. The zonated distribution of protein expression was quantified in terms of the liver zonation index. Quantitative targeted absolute proteomics revealed the selective expression of glutamine synthetase in the SR-101 + region, indicating predominant distribution of SR-101 in hepatocytes of the PC vein region. The protein levels of Oatp1a1, Oatp1b2, organic cation transporter 1 (Oct1), and cytochrome P450 (P450) 2e1 were greater in the PC vein regions, whereas the level of organic anion transporter 2 (Oat2) was greater in the PP vein regions. Mouse Oatp1a1 mediated SR-101 transport. On the other hand, there were no statistically significant differences in expression of Ent1, Na + -taurocholate cotransporting polypeptide, several canalicular transporters, P450 enzymes, and UDP-glucuronosyltransferases between the PP and PC vein regions. This is consistent with the almost uniform distribution of ribavirin in the liver. In conclusion, sinusoidal membrane transporters such as Oatp1a1, Oatp1b2, Oct1, and Oat2 appear to be determinants of the zonated distribution of drugs in the liver. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

Histochemistry of leucine aminoaphthylamidase (LAN) in rainbow trout (Salmo gairdneri)

USGS Publications Warehouse

Bouck, Gerald R.

1979-01-01

The histochemistry of leucine aminonaphthylamidase (LAN) was studied in frozen tissue sections of rainbow trout both in yearling and adult fish. Age of fish had relatively little effect upon the results. The most intense LAN color production was in epithelial cells of midgut, pyloric ceca, hindgut, and in some segments of kidney tubules. Lower levels of LAN were evident in liver cells of Kupffer, and still lower or slight levels of LAN activity were found in blood cells, muscle, nerve, connective tissue, gonad, and pancreas. The results indicate that LAN might be useful in assessing histotoxicity to LAN-rich areas of the body.

Histochemistry of leucine aminonaphthylamidase (LAN) in rainbow trout (Salmo gairdneri)

USGS Publications Warehouse

Bouck, Gerald R.

1979-01-01

The histochemistry of leucine aminonaphthylamidase (LAN) was studied in frozen tissue sections of rainbow trout both in yearling and adult fish. Age of fish had relatively little effect upon the results. The most intense LAN color production was in epithelial cells of midgut, pyloric ceca, hindgut, and in some segments of kidney tubules. Lower levels of LAN were evident in liver cells of Kupffer, and still lower or slight levels of LAN activity were found in blood cells, muscle, nerve, connective tissue, gonad, and pancreas. The results indicate that LAN might be useful in assessing histotoxicity to LAN-rich areas of the body.

Effects of chilled-then-frozen storage (up to 52weeks) on lamb M. longissimus lumborum quality and safety parameters.

PubMed

Coombs, Cassius E O; Holman, Benjamin W B; Collins, Damian; Friend, Michael A; Hopkins, David L

2017-12-01

This study evaluated the effect of chilled followed by frozen storage on lamb quality and safety parameters. Experimental (n=360) M. longissimus lumborum (LL) were randomly sampled from the boning room of a commercial Australian abattoir, at 24 h post-mortem, and assigned to five chilled storage periods (0, 2, 4, 6 and 8 weeks) and six subsequent frozen storage periods (0, 4, 8, 12, 24 and 52 weeks). Upon completion of each storage treatment combination, corresponding LL were sub-sectioned and analysed for colour stability (0, 1, 2 and 3 days), shear force, fluid losses (purge, thaw and cooking losses), intramuscular fat content, sarcomere length, water activity and microbial load (lactic acid bacteria, Enterobacteriaceae sp., Brochothrix thermosphacta, Clostridium perfringens and Escherichia coli). LL stored chilled for 2-4 weeks prior to freezing presented superior results for shear force, display colour and low levels of spoilage microbes, correlating with good eating quality and safety following more than one year of frozen storage. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

[In vitro activity of human bone marrow cells after cryopreservation in liquid nitrogen for 21 - 25 years].

PubMed

Huang, You-Zhang; Shen, Jian-Liang; Gong, Li-Zhong; Zheng, Pei-Hao; Liu, Yi; Yin, Wen-Jie; Cen, Jian; Wang, Ning; Zhao, De-Feng

2010-02-01

The aim of this study was to investigate the best method to preserve human bone marrow cells and the effectiveness of long term cryopreservation at -80 degrees C. The human bone marrow cells in 20 samples were firstly frozen by a programmed freezer or -80 degrees C refrigerator, and then were preserved in liquid nitrogen with DMSO-AuP (10% dimethylsulfonamide, 10% autologous plasma) or DMSO-HES-HuA (5% dimethylsulfonamide, 6% hydroxyethyl starch, 4% human serum albumin) as cryoprotectant for 21 to 25 years. They were thawed in 38 degrees C. The cell sample frozen in -80 degrees C refrigerator was frozen at a low frozen speed of 1 degrees C/min which was the same as the programmed freezer before -30 degrees C. Before detection the bone marrow cells were taken from liquid nitrogen and were thawed in 38 degrees C, then the suspension of bone marrow cells was prepared for detection. The cell morphology and recovery rate of erythrocytes, nucleocytes and platelets; the recovery rate of hematopoietic stem progenitors cells, as well as mesenchymal stem cells were determined. The results showed that the protective effectiveness of DMSO-HES-HuA was better than DMSO-AuP. The mature erythrocytes were destroyed lightly [(3.5 +/- 1.5)% versus (12.6 +/- 4.8)%], the hemolysis rate was lower [(3.3 +/- 1.6)% versus (23.1 +/- 5.1)%]. Osmotic fragility of erythrocytes in the former was not changed, but was dropped in the latter. The recovery rates of red cell, platelet, granulocyte-macrophage colony forming units and long term culture-initiating cells were higher in the former than that in the latter [(96.1 +/- 1.8)%, (70.0 +/- 9.5)%, (49.2 +/- 10.9)%, (54.2 +/- 13.8)% versus (76.3 +/- 5.6)%, (52.7 +/- 8.1)%, (43.5 +/- 12.3)%, (47.2 +/- 13.6)% respectively]. With each kind of cryoprotectant or frozen method, the frozen MSC could keep the original growth properties. With the same cryoprotectant and different frozen method, the cryopreservative effectiveness was not different. The influence of the cryoprotectant prescriptions and the frozen methods on the cryopreservative effectiveness was little. It is concluded that the human bone marrow cells with DMSO-AuP or DMSO-HES-HuA as cryoprotectant, frozen by a programmed freezer or -80 degrees C refrigerator, could be then preserved in liquid nitrogen for long time. When the preserving time was as long as 21 to 25 years, the morphology, the recovery rate and the activity of various kinds of cells were still good. The method of freezing by -80 degrees C refrigerator with 5% DMSO-6% HES-4% HuA and preserving in liquid nitrogen would be convenient, cheap and easily-manipulated for preservation of the human bone marrow cells.

Wetlab-2 - Quantitative PCR Tools for Spaceflight Studies of Gene Expression Aboard the International Space Station

NASA Technical Reports Server (NTRS)

Schonfeld, Julie E.

2015-01-01

Wetlab-2 is a research platform for conducting real-time quantitative gene expression analysis aboard the International Space Station. The system enables spaceflight genomic studies involving a wide variety of biospecimen types in the unique microgravity environment of space. Currently, gene expression analyses of space flown biospecimens must be conducted post flight after living cultures or frozen or chemically fixed samples are returned to Earth from the space station. Post-flight analysis is limited for several reasons. First, changes in gene expression can be transient, changing over a timescale of minutes. The delay between sampling on Earth can range from days to months, and RNA may degrade during this period of time, even in fixed or frozen samples. Second, living organisms that return to Earth may quickly re-adapt to terrestrial conditions. Third, forces exerted on samples during reentry and return to Earth may affect results. Lastly, follow up experiments designed in response to post-flight results must wait for a new flight opportunity to be tested.

Expression of Enzymes that Metabolize Medications

NASA Technical Reports Server (NTRS)

Wotring, V. E.; Peters, C. P.

2011-01-01

INTRODUCTION: Increased exposure to radiation is one physiological stressor associated with spaceflight and it is feasible to conduct ground experiments using known radiation exposures. The health of the liver, especially the activity rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. While radiation is known to alter normal physiological function, how radiation affects liver metabolism of administered medications is unclear. Crew health could be affected if the actions of medications used in spaceflight deviated from expectations formed during terrestrial medication use. This study is an effort to identify liver metabolic enzymes whose expression is altered by spaceflight or by radiation exposures that mimic features of the spaceflight environment. METHODS: Using procedures approved by the Animal Care and Use Committee, mice were exposed to either 137Cs (controls, 50 mGy, 6Gy, or 50 mGy + 6Gy separated by 24 hours) or 13 days of spaceflight on STS 135. Animals were anesthetized and sacrificed at several time points (4 hours, 24 hours or 7 days) after their last radiation exposure, or within 6 hours of return to Earth for the STS 135 animals. Livers were removed immediately and flash-frozen in liquid nitrogen. Tissue was homogenized, RNA extracted, purified and quality-tested. Complementary DNA was prepared from high-quality RNA samples, and used in RT-qPCR experiments to determine relative expression of a wide variety of genes involved in general metabolism and drug metabolism. RESULTS: Results of the ground radiation exposure experiments indicated 65 genes of the 190 tested were significantly affected by at least one of the radiation doses. Many of the affected genes are involved in the metabolism of drugs with hydrophobic or steroid-like structures, maintenance of redox homeostasis and repair of DNA damage. Most affected genes returned to near control expression levels by 7 days post-treatment. Not all recovered completely by the final time point tested: with 6 Gy exposure, metallothionein expression was 132-fold more than control at the 4 hr time point, and fell at each later time point (11-fold at 24 hrs, and 8-fold at 7 days). In contrast, there were other genes whose expression was altered and remained relatively constant through the 7 day period we tested. One examples is Cyp17a1, which showed a 4-fold elevation at 4 hrs after exposure and remained constant for 7 days after the last treatment. Spaceflight samples evaluated with similar methods and comparisons will be made between the radiation-treated groups and the spaceflight samples. CONCLUSION It seems likely that radiation exposure triggers homeostatic mechanisms, which could include alterations of gene expression. Better understanding of these pathways could aid in optimizing medications doses given to crewmembers who require treatment and eventually, to development of new countermeasures to ameliorate or prevent radiation-induced damage to cells and tissues.

Detection of Hepatic Fibrosis in Ex Vivo Liver Samples Using an Open-Photoacoustic-Cell Method: Feasibility Study

NASA Astrophysics Data System (ADS)

Stolik, S.; Fabila, D. A.; de la Rosa, J. M.; Escobedo, G.; Suárez-Álvarez, K.; Tomás, S. A.

2015-09-01

Design of non-invasive and accurate novel methods for liver fibrosis diagnosis has gained growing interest. Different stages of liver fibrosis were induced in Wistar rats by intraperitoneally administering different doses of carbon tetrachloride. The liver fibrosis degree was conventionally determined by means of histological examination. An open-photoacoustic-cell (OPC) technique for the assessment of liver fibrosis was developed and is reported here. The OPC technique is based on the fact that the thermal diffusivity can be accurately measured by photoacoustics taking into consideration the photoacoustic signal amplitude versus the modulation frequency. This technique measures directly the heat generated in a sample, due to non-radiative de-excitation processes, following the absorption of light. The thermal diffusivity was measured with a home-made open-photoacoustic-cell system that was specially designed to perform the measurement from ex vivo liver samples. The human liver tissue showed a significant increase in the thermal diffusivity depending on the fibrosis stage. Specifically, liver samples from rats exhibiting hepatic fibrosis showed a significantly higher value of the thermal diffusivity than for control animals.

Nondestructive cryomicro-CT imaging enables structural and molecular analysis of human lung tissue.

PubMed

Vasilescu, Dragoş M; Phillion, André B; Tanabe, Naoya; Kinose, Daisuke; Paige, David F; Kantrowitz, Jacob J; Liu, Gang; Liu, Hanqiao; Fishbane, Nick; Verleden, Stijn E; Vanaudenaerde, Bart M; Lenburg, Marc; Stevenson, Christopher S; Spira, Avrum; Cooper, Joel D; Hackett, Tillie-Louise; Hogg, James C

2017-01-01

Micro-computed tomography (CT) enables three-dimensional (3D) imaging of complex soft tissue structures, but current protocols used to achieve this goal preclude cellular and molecular phenotyping of the tissue. Here we describe a radiolucent cryostage that permits micro-CT imaging of unfixed frozen human lung samples at an isotropic voxel size of (11 µm) 3 under conditions where the sample is maintained frozen at -30°C during imaging. The cryostage was tested for thermal stability to maintain samples frozen up to 8 h. This report describes the methods used to choose the materials required for cryostage construction and demonstrates that whole genome mRNA integrity and expression are not compromised by exposure to micro-CT radiation and that the tissue can be used for immunohistochemistry. The new cryostage provides a novel method enabling integration of 3D tissue structure with cellular and molecular analysis to facilitate the identification of molecular determinants of disease. The described micro-CT cryostage provides a novel way to study the three-dimensional lung structure preserved without the effects of fixatives while enabling subsequent studies of the cellular matrix composition and gene expression. This approach will, for the first time, enable researchers to study structural changes of lung tissues that occur with disease and correlate them with changes in gene or protein signatures. Copyright © 2017 the American Physiological Society.

Liquid and Frozen Storage of Agouti (Dasyprocta leporina) Semen Extended with UHT Milk, Unpasteurized Coconut Water, and Pasteurized Coconut Water

PubMed Central

Mollineau, W. M.; Adogwa, A. O.; Garcia, G. W.

2011-01-01

This study evaluated the effects of semen extension and storage on forward progressive motility % (FPM%) in agouti semen. Three extenders were used; sterilized whole cow's milk (UHT Milk), unpasteurized (CW) and pasteurized coconut water (PCW), and diluted to 50, 100, 150, and 200 × 106 spermatozoa/ml. Experiment 1: 200 ejaculates were extended for liquid storage at 5∘C and evaluated every day for 5 days to determine FPM% and its rate of deterioration. Experiment 2: 150 ejaculates were extended for storage as frozen pellets in liquid nitrogen at −195∘C, thawed at 30∘ to 70∘C for 20 to 50 seconds after 5 days and evaluated for FPM% and its rate of deterioration. Samples treated with UHT milk and storage at concentrations of 100 × 106 spermatozoa/ml produced the highest means for FPM% and the slowest rates of deterioration during Experiment 1. During Experiment 2 samples thawed at 30∘C for 20 seconds exhibited the highest means for FPM% (12.18 ± 1.33%), 85% rate of deterioration. However, samples were incompletely thawed. This was attributed to the diameter of the frozen pellets which was 1 cm. It was concluded that the liquid storage method was better for short term storage. PMID:20871831

Quality evaluation of frozen guava and yellow passion fruit pulps by NIR spectroscopy and chemometrics.

PubMed

Alamar, Priscila D; Caramês, Elem T S; Poppi, Ronei J; Pallone, Juliana A L

2016-07-01

The present study investigated the application of near infrared spectroscopy as a green, quick, and efficient alternative to analytical methods currently used to evaluate the quality (moisture, total sugars, acidity, soluble solids, pH and ascorbic acid) of frozen guava and passion fruit pulps. Fifty samples were analyzed by near infrared spectroscopy (NIR) and reference methods. Partial least square regression (PLSR) was used to develop calibration models to relate the NIR spectra and the reference values. Reference methods indicated adulteration by water addition in 58% of guava pulp samples and 44% of yellow passion fruit pulp samples. The PLS models produced lower values of root mean squares error of calibration (RMSEC), root mean squares error of prediction (RMSEP), and coefficient of determination above 0.7. Moisture and total sugars presented the best calibration models (RMSEP of 0.240 and 0.269, respectively, for guava pulp; RMSEP of 0.401 and 0.413, respectively, for passion fruit pulp) which enables the application of these models to determine adulteration in guava and yellow passion fruit pulp by water or sugar addition. The models constructed for calibration of quality parameters of frozen fruit pulps in this study indicate that NIR spectroscopy coupled with the multivariate calibration technique could be applied to determine the quality of guava and yellow passion fruit pulp. Copyright © 2016 Elsevier Ltd. All rights reserved.

Non-Destructive Sampling of Ancient Insect DNA

PubMed Central

Thomsen, Philip Francis; Elias, Scott; Gilbert, M. Thomas P.; Haile, James; Munch, Kasper; Kuzmina, Svetlana; Froese, Duane G.; Holdaway, Richard N.; Willerslev, Eske

2009-01-01

Background A major challenge for ancient DNA (aDNA) studies on insect remains is that sampling procedures involve at least partial destruction of the specimens. A recent extraction protocol reveals the possibility of obtaining DNA from past insect remains without causing visual morphological damage. We test the applicability of this protocol on historic museum beetle specimens dating back to AD 1820 and on ancient beetle chitin remains from permafrost (permanently frozen soil) dating back more than 47,000 years. Finally, we test the possibility of obtaining ancient insect DNA directly from non-frozen sediments deposited 3280-1800 years ago - an alternative approach that also does not involve destruction of valuable material. Methodology/Principal Findings The success of the methodological approaches are tested by PCR and sequencing of COI and 16S mitochondrial DNA (mtDNA) fragments of 77–204 base pairs (-bp) in size using species-specific and general insect primers. Conclusion/Significance The applied non-destructive DNA extraction method shows promising potential on insect museum specimens of historical age as far back as AD 1820, but less so on the ancient permafrost-preserved insect fossil remains tested, where DNA was obtained from samples up to ca. 26,000 years old. The non-frozen sediment DNA approach appears to have great potential for recording the former presence of insect taxa not normally preserved as macrofossils and opens new frontiers in research on ancient biodiversity. PMID:19337382

Oxidative stability of cooked, frozen, reheated beef patties: effect of antioxidants.

PubMed

Colindres, Paola; Brewer, M Susan

2011-03-30

The effect of selected antioxidants (grape seed extract (GS), oleoresin rosemary (OR), water-soluble oregano extract (WO), propyl gallate (PG), butylated hydroxyanisole (BHA)), butylated hydroxytoluene (BHT)) on sensory, color and oxidative stability of cooked, frozen, reheated ground beef patties was evaluated. Beef lean and trim were ground; antioxidants and salt were added. Patties were cooked (71 °C), overwrapped in commercial polyvinyl chloride film, and stored frozen (-18 °C), then evaluated monthly for 6 months. Flavor, odor and color were determined using a descriptive panel. Instrumental color was determined by a spectrocolorimeter. Lipid oxidation was determined using thiobarbituric acid reactive substances (TBARS). After 6 months of storage, PG and GS samples had lower rancid odor scores and TBARS than controls. Control samples and those containing BHT did not differ statistically in sensory grassy or rancid odor, indicating that they were the most oxidized. TBARS correlated with grassy, rancid, cardboard and beef odors during the 6-month storage period. Based on TBARS, the order of effectiveness of the antioxidants was PG and GS > OR > BHA > WO and BHT > control. TBARS were well correlated with sensory evaluations of odor and flavor. Antioxidants also protected a* values during storage. Copyright © 2011 Society of Chemical Industry.

An Assessment of macro-scale in situ Raman and ultraviolet-induced fluorescence spectroscopy for rapid characterization of frozen peat and ground ice

NASA Astrophysics Data System (ADS)

Laing, Janelle R.; Robichaud, Hailey C.; Cloutis, Edward A.

2016-04-01

The search for life on other planets is an active area of research. Many of the likeliest planetary bodies, such as Europa, Enceladus, and Mars are characterized by cold surface environments and ice-rich terrains. Both Raman and ultraviolet-induced fluorescence (UIF) spectroscopies have been proposed as promising tools for the detection of various kinds of bioindicators in these environments. We examined whether macro-scale Raman and UIF spectroscopy could be applied to the analysis of unprocessed terrestrial frozen peat and clear ground ice samples for detection of bioindicators. It was found that this approach did not provide unambiguous detection of bioindicators, likely for a number of reasons, particularly due to strong broadband induced fluorescence. Other contributing factors may include degradation of organic matter in frozen peat to the point that compound-specific emitted fluorescence or Raman peaks were not resolvable. Our study does not downgrade the utility of either UIF or Raman spectroscopy for astrobiological investigations (which has been demonstrated in previous studies), but does suggest that the choice of instrumentation, operational conditions and sample preparation are important factors in ensuring the success of these techniques.

Short communication: Preliminary investigation into the effect of freezing and a cryopreservant on the recovery of mastitis pathogens from ewe milk.

PubMed

Smith, E M; Monaghan, E M; Huntley, S J; Green, L E

2011-10-01

The objective of this study was to investigate the recovery of bacteria from ewe milk after freezing for 4 or 8 wk with and without the addition of glycerol as a cryopreservant. A total of 50 udder-half milk samples with a known range of bacterial species were selected, stored, and analyzed in 5 treatment groups: time zero; frozen for 4 wk with, and without, glycerol; and frozen for 8 wk with, and without, glycerol. A lower recovery was observed in all bacterial species studied after freezing. Samples containing fewer than 100 cfu/mL came from ewes with a lower somatic cell count and were more likely to be bacteriologically negative after freezing than those above this threshold. The addition of glycerol increased recovery of gram-negative bacteria after freezing, although this requires further study to draw strong conclusions. The effects on gram-positive species were inconsistent. We conclude that although the addition of glycerol had a small beneficial effect on the sensitivity of detection of bacteria from frozen sheep milk, sensitivity was highest in cultures from fresh milk. Copyright © 2011 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Estimate of colostral immunoglobulin G concentration using refractometry without or with caprylic acid fractionation.

PubMed

Morrill, K M; Conrad, E; Polo, J; Lago, A; Campbell, J; Quigley, J; Tyler, H

2012-07-01

Our objectives were to evaluate the use of refractometry as a means of estimating immunoglobulin G (IgG) concentration of bovine maternal colostrum (MC) and determine if fractionation of MC using caprylic acid (CA) improved estimates of IgG. Samples (n=85) of MC were collected from a single dairy in California and used to determine the method of CA fraction that produced the best prediction of IgG based on CA fractionation followed by refractometry. Subsequently, samples of MC (n=827) were collected from 67 farms in 12 states to compare refractometry with or without CA fractionation as methods to estimate IgG concentration. Samples were collected from the feeding pool and consisted of fresh (n=196), previously frozen (n=479), or refrigerated (n=152) MC. Samples were further classified by the number freeze-thaw cycles before analysis. Fractionation with CA was conducted by adding 1 mL of MC to a tube containing 75 μL of CA and 1 mL of 0.06 M acetic acid. The tube was shaken and allowed to react for 1 min. Refractive index of the IgG-rich supernatant (nDf) was determined using a digital refractometer. Whole, nonfractionated MC was analyzed for IgG by radial immunodiffusion (RID) and refractive index (nDw). The relationship between nDf and IgG (r=0.53; n=805) was weak, whereas that between nDw and IgG was stronger (r=0.73; n=823). Fresh samples analyzed by refractometry that subsequently went through 1 freeze-thaw cycle before RID analysis resulted in the strongest relationship between IgG and nDf or nDw (r=0.93 and 0.90, respectively). The MC samples collected fresh on the farm but frozen 2 or more times before analysis by refractometry or RID had low correlations between IgG and nDf and nDw (r=0.09 and 0.01). Samples refrigerated or frozen on the farm before analysis had weaker relationships between RID and nDf or nDw (r=0.38 to 0.80), regardless of the number of freeze-thaw cycles. Breed and lactation number did not affect the accuracy of either test. These results indicated that refractometry, without or with CA fractionation, was an accurate and rapid method to determine IgG concentration when samples of MC were not previously stored before refractometry and frozen only once before RID analysis. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Targeted or whole genome sequencing of formalin fixed tissue samples: potential applications in cancer genomics.

PubMed

Munchel, Sarah; Hoang, Yen; Zhao, Yue; Cottrell, Joseph; Klotzle, Brandy; Godwin, Andrew K; Koestler, Devin; Beyerlein, Peter; Fan, Jian-Bing; Bibikova, Marina; Chien, Jeremy

2015-09-22

Current genomic studies are limited by the poor availability of fresh-frozen tissue samples. Although formalin-fixed diagnostic samples are in abundance, they are seldom used in current genomic studies because of the concern of formalin-fixation artifacts. Better characterization of these artifacts will allow the use of archived clinical specimens in translational and clinical research studies. To provide a systematic analysis of formalin-fixation artifacts on Illumina sequencing, we generated 26 DNA sequencing data sets from 13 pairs of matched formalin-fixed paraffin-embedded (FFPE) and fresh-frozen (FF) tissue samples. The results indicate high rate of concordant calls between matched FF/FFPE pairs at reference and variant positions in three commonly used sequencing approaches (whole genome, whole exome, and targeted exon sequencing). Global mismatch rates and C · G > T · A substitutions were comparable between matched FF/FFPE samples, and discordant rates were low (<0.26%) in all samples. Finally, low-pass whole genome sequencing produces similar pattern of copy number alterations between FF/FFPE pairs. The results from our studies suggest the potential use of diagnostic FFPE samples for cancer genomic studies to characterize and catalog variations in cancer genomes.

Effects of salinity on physicochemical properties of Alaska pollock surimi after repeated freeze-thaw cycles.

PubMed

Kang, E J; Hunt, A L; Park, J W

2008-06-01

The effects of residual salt in surimi on physicochemical properties as affected by various freeze and thaw (FT) cycles were examined. Fresh Alaska pollock surimi was mixed with 4.0% sugar and 5.0% sorbitol, along with 8 combinations of salt (0.4%, 0.6%, 0.8%, and 1.0% NaCl) and sodium polyphosphate (0.25% and 0.5%), vacuum-packed, and stored at -18 degrees C until used. FT cycles (0, 6, and 9) were used to mimic long-term frozen storage. At the time of gel preparation, each treatment was appropriately adjusted to maintain 2% salt and 78% moisture. The pH decreased as residual salt increased during frozen storage. Salt extractable protein (SEP) decreased (P < 0.05) as FT cycles extended from 0 to 9. Regardless of residual salt and phosphate concentration during frozen storage, whiteness value (L*- 3b*) decreased (P < 0.05) as FT cycles extended, except for samples with 0.4% salt/0.5% phosphate and 0.6% salt/0.25% phosphate. Water retention ability (WRA) and texture significantly (P < 0.05) decreased at higher salt content (0.8% and 1.0%) after 9 FT cycles, indicating higher residual salt concentration can shorten the shelf life of frozen surimi. Our study revealed lower residual salt concentration and higher phosphate concentration are likely to extend the shelf life of frozen surimi.

Application of Lactobacillus acidophilus (LA 5) strain in fruit-based ice cream

PubMed Central

Senanayake, Suraji A; Fernando, Sirimali; Bamunuarachchi, Arthur; Arsekularatne, Mariam

2013-01-01

A study was performed to apply a probiotic strain into fermented ice cream mix with suitable fruit bases to develop a value-added product with a substantial level of viable organisms for a sufficient shelf life. Pure direct vat strain culture of Lactobacillus acidophilus (LA 5) in freeze-dried form was inoculated into a mixture of ice cream, frozen, and the number of viable organisms during frozen storage for a period of time was enumerated, using turbidity measurements with a spectrophotometer. An ice cream sample prepared without the probiotic culture was compared with the test sample for quality, by testing the basic quality parameters for ice cream. Results show a reduction in the over run of the probiotic ice cream compared to the nonprobiotic ice cream. Significantly high level (P < 0.05) of total solids (42%), proteins (16.5%), and titratable acidity (2.2%) was observed in the test sample compared to the nonprobiotic ice cream. Significantly low pH level in the probiotic sample may be due to the lactic acid produced by the probiotic culture. No significant difference (P > 0.05) in the fat content in the two types of ice cream was observed. A significantly low level (P < 0.05) of melting in the probiotic one may have resulted from less over run, than the nonprobiotic sample. Rapid reduction in the viable cells during frozen storage occurred at −18°C and gradual adaptation occurred over the first 4 weeks. At the 10th week, 1.0 × 107 numbers of viable organisms were present in 1 g of the probiotic ice cream. Results show the presence of a sufficient number of viable organisms in the product for the 10-week period, which would be beneficial to consumers. PMID:24804052

Application of Lactobacillus acidophilus (LA 5) strain in fruit-based ice cream.

PubMed

Senanayake, Suraji A; Fernando, Sirimali; Bamunuarachchi, Arthur; Arsekularatne, Mariam

2013-11-01

A study was performed to apply a probiotic strain into fermented ice cream mix with suitable fruit bases to develop a value-added product with a substantial level of viable organisms for a sufficient shelf life. Pure direct vat strain culture of Lactobacillus acidophilus (LA 5) in freeze-dried form was inoculated into a mixture of ice cream, frozen, and the number of viable organisms during frozen storage for a period of time was enumerated, using turbidity measurements with a spectrophotometer. An ice cream sample prepared without the probiotic culture was compared with the test sample for quality, by testing the basic quality parameters for ice cream. Results show a reduction in the over run of the probiotic ice cream compared to the nonprobiotic ice cream. Significantly high level (P < 0.05) of total solids (42%), proteins (16.5%), and titratable acidity (2.2%) was observed in the test sample compared to the nonprobiotic ice cream. Significantly low pH level in the probiotic sample may be due to the lactic acid produced by the probiotic culture. No significant difference (P > 0.05) in the fat content in the two types of ice cream was observed. A significantly low level (P < 0.05) of melting in the probiotic one may have resulted from less over run, than the nonprobiotic sample. Rapid reduction in the viable cells during frozen storage occurred at -18°C and gradual adaptation occurred over the first 4 weeks. At the 10th week, 1.0 × 10(7) numbers of viable organisms were present in 1 g of the probiotic ice cream. Results show the presence of a sufficient number of viable organisms in the product for the 10-week period, which would be beneficial to consumers.

Evaluation of two 4th generation point-of-care assays for the detection of Human Immunodeficiency Virus infection.

PubMed

Stafylis, Chrysovalantis; Klausner, Jeffrey D

2017-01-01

Fourth generation assays detect simultaneously antibodies for HIV and the p24 antigen, identifying HIV infection earlier than previous generation tests. Previous studies have shown that the Alere Determine HIV-1/2 Combo has lower than anticipated performance in detecting antibodies for HIV and the p24 antigen. Furthermore, there are currently very few studies evaluating the performance of Standard Diagnostics BIOLINE HIV Ag/Ab Combo. To evaluate the performance of the Alere Determine HIV-1/2 Combo and the Standard Diagnostics BIOLINE HIV Ag/Ab Combo in a panel of frozen serum samples. The testing panel included 133 previously frozen serum specimens from the UCLA Clinical Microbiology & Immunoserology laboratory. Reference testing included testing for HIV antibodies by a 3rd generation enzyme immunoassay followed by HIV RNA detection. Antibody negative and RNA positive sera were also tested by a laboratory 4th generation HIV Ab/Ag enzyme immunoassay. Reference testing yielded 97 positives for HIV infection and 36 negative samples. Sensitivity of the Alere test was 95% (88-98%), while the SD Bioline sensitivity was 91% (83-96%). Both assays showed 100% (90-100%) specificity. No indeterminate or invalid results were recorded. Among 13 samples with acute infection (HIV RNA positive, HIV antibody negative), 12 were found positive by the first assay and 8 by the second. The antigen component of the Alere assay detected 10 acute samples, while the SD Bioline assay detected only one. Both rapid assays showed very good overall performance in detecting HIV infection in frozen serum samples, but further improvements are required to improve the performance in acute infection.

Stability of hemostatic proteins in canine fresh-frozen plasma thawed with a modified commercial microwave warmer or warm water bath.

PubMed

Pashmakova, Medora B; Barr, James W; Bishop, Micah A

2015-05-01

To compare stability of hemostatic proteins in canine fresh-frozen plasma (FFP) thawed with a modified commercial microwave warmer (MCM) or warm water bath (37°C; WWB) or at room temperature (22°C). Fresh-frozen plasma obtained from 8 canine donors of a commercial blood bank. A commercial microwave warmer was modified with a thermocouple to measure surface temperature of bags containing plasma. The MCM and a WWB were each used to concurrently thaw a 60-mL bag of plasma obtained from the same donor. Two 3-mL control aliquots of FFP from each donor were thawed to room temperature without use of a heating device. Concentrations of hemostatic proteins, albumin, and D-dimers; prothrombin time (PT); and activated partial thromboplastin time (aPTT) were determined for all samples. Significant decreases in concentrations of factors II, IX, X, XI, fibrinogen, von Willebrand factor, antithrombin, protein C, and albumin and significant increases in PT and aPTT were detected for plasma thawed with the MCM, compared with results for samples thawed with the WWB. Concentrations of factors VII, VIII, and XII were not significantly different between plasma thawed with the MCM and WWB. Concentrations of D-dimers were above the reference range for all thawed samples regardless of thawing method. No significant differences in factor concentrations were detected between control and WWB-thawed samples. Significant differences in hemostatic protein concentrations and coagulation times were detected for plasma thawed with an MCM but not between control and WWB-thawed samples. Clinical importance of these changes should be investigated.

Characterization of HBV integration patterns and timing in liver cancer and HBV-infected livers.

PubMed

Furuta, Mayuko; Tanaka, Hiroko; Shiraishi, Yuichi; Unida, Takuro; Imamura, Michio; Fujimoto, Akihiro; Fujita, Masahi; Sasaki-Oku, Aya; Maejima, Kazuhiro; Nakano, Kaoru; Kawakami, Yoshiiku; Arihiro, Koji; Aikata, Hiroshi; Ueno, Masaki; Hayami, Shinya; Ariizumi, Shun-Ichi; Yamamoto, Masakazu; Gotoh, Kunihito; Ohdan, Hideki; Yamaue, Hiroki; Miyano, Satoru; Chayama, Kazuaki; Nakagawa, Hidewaki

2018-05-18

Integration of Hepatitis B virus (HBV) into the human genome can cause genetic instability, leading to selective advantages for HBV-induced liver cancer. Despite the large number of studies for HBV integration into liver cancer, little is known about the mechanism of initial HBV integration events owing to the limitations of materials and detection methods. We conducted an HBV sequence capture, followed by ultra-deep sequencing, to screen for HBV integrations in 111 liver samples from human-hepatocyte chimeric mice with HBV infection and human clinical samples containing 42 paired samples from non-tumorous and tumorous liver tissues. The HBV infection model using chimeric mice verified the efficiency of our HBV-capture analysis and demonstrated that HBV integration could occur 23 to 49 days after HBV infection via microhomology-mediated end joining and predominantly in mitochondrial DNA. Overall HBV integration sites in clinical samples were significantly enriched in regions annotated as exhibiting open chromatin, a high level of gene expression, and early replication timing in liver cells. These data indicate that HBV integration in liver tissue was biased according to chromatin accessibility, with additional selection pressures in the gene promoters of tumor samples. Moreover, an integrative analysis using paired non-tumorous and tumorous samples and HBV-related transcriptional change revealed the involvement of TERT and MLL4 in clonal selection. We also found frequent and non-tumorous liver-specific HBV integrations in FN1 and HBV-FN1 fusion transcript. Extensive survey of HBV integrations facilitates and improves the understanding of the timing and biology of HBV integration during infection and HBV-related hepatocarcinogenesis.

iss055e043245

NASA Image and Video Library

2018-04-30

iss055e043245 (April 30, 2018) --- NASA astronaut Ricky Arnold transfers frozen biological samples from science freezers aboard the International Space Station to science freezers inside the SpaceX Dragon resupply ship. The research samples were returned to Earth aboard Dragon for retrieval by SpaceX engineers and analysis by NASA scientists.

Digital PCR for Quantifying Norovirus in Oysters Implicated in Outbreaks, France.

PubMed

Polo, David; Schaeffer, Julien; Fournet, Nelly; Le Saux, Jean-Claude; Parnaudeau, Sylvain; McLeod, Catherine; Le Guyader, Françoise S

2016-12-01

Using samples from oysters clearly implicated in human disease, we quantified norovirus levels by using digital PCR. Concentrations varied from 43 to 1,170 RNA copies/oyster. The analysis of frozen samples from the production area showed the presence of norovirus 2 weeks before consumption.

[Measurement of the status of trace elements in cattle using liver biopsy samples].

PubMed

Ouweltjes, W; de Zeeuw, A C; Moen, A; Counotte, G H M

2007-02-01

Serum, plasma, or urine samples are usually used for the measurement of the trace elements copper; zinc, iron, selenium, because these samples are easy to obtain; however; these samples are not always appropriate. For example, it is not possible to measure molybdenum, the major antagonist of copper; in blood or urine. Therefore measurement of trace elements in liver tissue is considered the gold standard. For the assessment of selenium the method of choice remains determination of glutathion peroxidase in erythrocytes and for the assessment of magnesium determination of magnesium in urine. We determined the accuracy and repeatability of measuring trace elements in liver biopsies and whole liver homogenates. The levels of trace elements measured were similar in both preparations (92% agreement). Liver biopsy in live animals is a relatively simple procedure but not common in The Netherlands. Reference levels of trace elements, classified as too low, low, adequate, high, and too high, were established on the basis of our research and information in the literature. In a second study we investigated the practical aspects of obtaining liver tissue samples and their use. Samples were collected from cattle on a commercial dairy farm. Liver biopsy provided additional information to that obtained from serum and urine samples. We prepared a biopsy protocol and a test package, which we tested on 14 farms where an imbalance of trace minerals was suspected. Biopsy samples taken from 4 to 6 animals revealed extreme levels of trace elements.

Gene expression of leptin, resistin, and adiponectin in the white adipose tissue of obese patients with non-alcoholic fatty liver disease and insulin resistance.

PubMed

Baranova, Ancha; Gowder, Shobha J; Schlauch, Karen; Elariny, Hazem; Collantes, Rochelle; Afendy, Arian; Ong, Janus P; Goodman, Zachary; Chandhoke, Vikas; Younossi, Zobair M

2006-09-01

Adipose tissue is an active endocrine organ that secretes a variety of metabolically important substances including adipokines. These factors affect insulin sensitivity and may represent a link between obesity, insulin resistance, type 2 diabetes (DM), and nonalcoholic fatty liver disease (NAFLD). This study uses real-time polymerase chain reaction (PCR) quantification of mRNAs encoding adiponectin, leptin, and resistin on snap-frozen samples of intra-abdominal adipose tissue of morbidly obese patients undergoing bariatric surgery. Morbidly obese patients undergoing bariatric surgery were studied. Patients were classified into two groups: Group A (with insulin resistance) (N=11; glucose 149.84 +/- 40.56 mg/dL; serum insulin 8.28 +/- 3.52 microU/mL), and Group B (without insulin resistance) (N=10; glucose 102.2 +/- 8.43 mg/dL; serum insulin 3.431 +/- 1.162 microU/mL). Adiponectin mRNA in intra-abdominal adipose tissue and serum adiponectin levels were significantly lower in Group A compared to Group B patients (P<0.016 and P<0.03, respectively). Although serum resistin was higher in Group A than in Group B patients (P<0.005), resistin gene expression was not different between the two groups. Finally, for leptin, neither serum level nor gene expression was different between the two groups. Serum adiponectin level was the only predictor of nonalcoholic steatohepatitis (NASH) in this study (P=0.024). Obese patients with insulin resistance have decreased serum adiponectin and increased serum resistin. Additionally, adiponectin gene expression is also decreased in the adipose tissue of these patients. This low level of adiponectin expression may predispose patients to the progressive form of NAFLD or NASH.

THE USE OF GAMMA RADIATION TO DESTROY SALMONELLAE IN FROZEN WHOLE EGG

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brooks, J.; Hannan, R.S.; Hobbs, B.C.

Frozen whole egg (an intimate mixture of the white and yolk) is the most important egg product in international commerce. Average imports into the United Kingdom alone have amounted to about 22,000 tons per annum over the last ten years. It has been known for some time that egg products may contain members of the Salmonella group capable of causing food poisoning, but the position has been rendered more serious by the recent discovery of S. paratyphi B in egg products imported. In fact, the proved association of egg products with paratyphoid B fever has created a new situation inmore » the epidemiology of the disease. After treatment of small samples of frozen whole egg with 2-Mev cathode rays, it was concluded that a dose of about 300,000 to 500,000 rads would destroy the numbers of salmonellas normally encountered in the product without impairing the baking qualities of the material. Whole tins, each containing 22 pounds, were therefore irradiated in the frozen state with cobalt-80 gamma rays. Two tins were treated at each of three dose levels of 300,000, 400,000, and 500,000 rads. The contaminants present before irradiation were S. paratyphi B (three tins), S. newport (two tins), and S. thompson (one tin). No salmonellas were detected in duplicate samples of material taken from each of the tins after irradiation. If the effectiveness of the treatment is confirmed by further work, the process has obvious attractions since it dispenses with the need to thaw or otherwise to handle the product. (auth)« less

The Impact of Multiple Freeze-Thaw Cycles on the Microstructure of Aggregates from a Soddy-Podzolic Soil: A Microtomographic Analysis

NASA Astrophysics Data System (ADS)

Skvortsova, E. B.; Shein, E. V.; Abrosimov, K. N.; Romanenko, K. A.; Yudina, A. V.; Klyueva, V. V.; Khaidapova, D. D.; Rogov, V. V.

2018-02-01

With the help of computed X-ray microtomography with a resolution of 2.75 μm, changes in the microstructure and pore space of aggregates of 3 mm in diameter from the virgin soddy-podzolic soil (Glossic Retisol (Loamic)) in the air-dry, capillary-moistened, and frozen states after five freeze-thaw cycles were studied in a laboratory experiment. The freezing of the samples was performed at their capillary moistening. It was shown that capillary moistening of initially air-dry samples from the humus (AY), eluvial (EL), and illuvial (BT1) horizons at room temperature resulted in the development of the platy, fine vesicular, and angular blocky microstructure, respectively. The total volume of tomographically visible pores >10 μm increased by 1.3, 2.2, and 3.4 times, respectively. After freeze-thaw cycles, frozen aggregates partly preserved the structural arrangement formed during the capillary moistening. At the same time, in the frozen aggregate from the AY horizon, the total tomographic porosity decreased to the initial level of the air-dry soil. In the frozen aggregate from the EL horizon, large vesicular pores were formed, owing to which the total pore volume retained its increased values. The resistance of aggregate shape to the action of freeze-thaw cycles differed. The aggregate from the EL horizon completely lost its original configuration by the end of the experiment. The aggregate from the AY horizon displayed definite features of sagging after five freeze-thaw cycles, whereas the aggregate from the BT1 horizon preserved its original configuration.

Effective freezing rate for semen cryopreservation in endangered Mediterranean brown trout (Salmo trutta macrostigma) inhabiting the Biferno river (South Italy).

PubMed

Iaffaldano, Nicolaia; Di Iorio, Michele; Manchisi, Angelo; Esposito, Stefano; Gibertoni, Pier Paolo

2016-10-01

This study was designed to determine: (i) the in vitro effects of different freezing rates on post-thaw semen quality of Mediterranean brown trout (Salmo trutta macrostigma) from the Biferno river; and (ii) the in vivo fertilization and hatching percentage of freezing rate giving rise to the best post-thaw semen quality. Pooled semen samples were diluted 1:3 (v:v) in a freezing extender composed of 300 mM glucose, 10% egg yolk and 10% dimethyl sulfoxide (DMSO). The extended semen was packaged in 0.25 ml plastic straws and frozen at different heights above the liquid nitrogen surface (1, 5 or 10 cm) for 10 min to give three different freezing rates. Semen samples were thawed at 30°C for 10 s. The variables assessed after thawing were sperm motility, duration of motility and viability. Our results clearly indicate a significant effect of freezing rate on post-thaw semen quality. Semen frozen 5 cm above the liquid nitrogen surface showed the best quality after freezing/thawing. Based on these in vitro data, 2 groups of 200 eggs were fertilized with fresh semen or semen frozen 5 cm above the liquid nitrogen surface. Fertilization and hatching rates recorded for eggs fertilized with frozen semen were significantly lower (25.4% and 22.5%, respectively) than the ones obtained using fresh semen (87.8% and 75.5%, respectively). An effective freezing protocol will allow for the creation of a sperm cryobank to recover the original population of Mediterranean brown trout in the Biferno river.

Polonium-210 and Caesium-137 in lynx (Lynx lynx), wolverine (Gulo gulo) and wolves (Canis lupus).

PubMed

Gjelsvik, Runhild; Holm, Elis; Kålås, John Atle; Persson, Bertil; Asbrink, Jessica

2014-12-01

Wolves, lynx and wolverines are on the top of the food-chain in northern Scandinavia and Finland. (210)Po and (137)Cs have been analysed in samples of liver, kidney and muscle from 28 wolves from Sweden. In addition blood samples were taken from 27 wolves. In 9 of the wolves, samples of muscle, liver and blood were analysed for (210)Po. Samples of liver and muscle were collected from 16 lynx and 16 wolverines from Norway. The liver samples were analysed for (210)Po and (137)Cs. Only (137)Cs analyses were carried out for the muscle samples. The wolves were collected during the winter 2010 and 2011, while the samples for lynx and wolverines were all from 2011. The activity concentrations of (210)Po in wolves were higher for liver (range 20-523 Bq kg(-1) d.w.) and kidney (range 24-942 Bq kg(-1) d.w.) than muscle (range 1-43 Bq kg(-1) d.w.) and blood (range 2-54 Bq kg(-1) d.w.). Activity ratios, (210)Po/(210)Pb, in wolf samples of muscle, liver and blood were in the ranges 2-77, 9-56 and 2-54. Using a wet weight ratio of 3.8 the maximal absorbed dose from (210)Po to wolf liver was estimated to 3500 μGy per year. Compared to wolf, the ranges of (210)Po in liver samples were lower in lynx (range 22-211 Bq kg(-1) d.w.) and wolverine (range16-160 Bq kg(-1) d.w.). Concentration of (137)Cs in wolf samples of muscle, liver, kidney and blood were in the ranges 70-8410 Bq kg(-1) d.w., 36-4050 Bq kg(-1) d.w., 31-3453 Bq kg(-1) d.w. and 4-959 Bq kg(-1) d.w., respectively. (137)Cs in lynx muscle and liver samples were in the ranges 44-13393 Bq kg(-1) d.w. and 125-10260 Bq kg(-1) d.w. The corresponding values for (137)Cs in wolverine were 22-3405 Bq kg(-1) d.w. for liver and 53-4780 Bq kg(-1) d.w. for muscle. The maximal absorbed dose from (137)Cs to lynx was estimated to 3000 μGy per year. Copyright © 2014 Elsevier Ltd. All rights reserved.

Lipid class and fatty acid composition of a little-known and rarely collected alga Exophyllum wentii Weber-van Bosse from Bali Island, Indonesia.

PubMed

Illijas, Muhammad I; Indy, Jeane R; Yasui, Hajime; Itabashi, Yutaka

2009-01-01

The lipid class and fatty acid composition of a little-known and rarely collected alga Exophyllum wentii from Bali Island, Indonesia were determined for fresh and frozen-thawed samples using thin-layer chromatography, gas-liquid chromatography, and high-performance liquid chromatography. Glycoglycerolipids, which mainly consisted of mongalactosyldiacylglycerols (MGDG) and digalactosyldiacylglycerols (DGDG), were the predominant lipid components, accounting for 67% and 56% of the total polar lipid content in the fresh and frozen-thawed samples, respectively. Phospholipids, including phosphatidylcholines (PC) and phosphatidylglycerols (PG), were detected with lesser amounts in both samples (16-17% of the total polar lipid content). Free fatty acids (FFA), sterols and triacylglycerols (TAG) were also detected in minor quantities; however, the FFA content in the frozen-thawed sample increased to up to 20% of the total lipid content, suggesting that hydrolysis of the membrane lipids had occurred. A crude enzyme preparation from the alga showed activities for hydrolyzing the acyl groups of the phospholipids and glycoglycerolipids. Palmitic acid (16:0) and arachidonic acid (20:4n-6) were the major fatty acids in both the total lipid and in individual polar lipid classes as well as the dominant fatty acids released from the membrane lipids by enzymatic hydrolysis. The high level of 20:4n-6 (29%) in the total lipid and the presence of considerable amounts of PC (11% of the total polar lipid) and PG (6.2%) support classification of E. wentii into the Division Rhodophyta.

The influence of low temperatures on dynamic mechanical properties of animal bone

NASA Astrophysics Data System (ADS)

Mardas, Marcin; Kubisz, Leszek; Mielcarek, Slawomir; Biskupski, Piotr

2009-01-01

Different preservation methods are currently used in bone banks, even though their effects on allograft quality are not fully understood. Freezing is one of the most popular methods of preservation in tissue banking. Yet, there is not a lot of data on dynamic mechanical properties of frozen bone. Material used in this study was femoral bones from adult bovine that were machine cut and frozen to the temperature 140°C. Both elastic modulus and loss modulus were measured at 1, 3, 5, 10, and 20 Hz in the temperature range of 30-200°C. Differences between frozen and control samples were observed. The frequency increase always led to the increase in elastic modulus values and decrease in loss modulus values. Freezing reduced the elastic modulus values of about 25% and the loss modulus values of about 45% when measured at 20°C.

Detection of human norovirus from frozen raspberries in a cluster of gastroenteritis outbreaks.

PubMed

Maunula, L; Roivainen, M; Keränen, M; Mäkela, S; Söderberg, K; Summa, M; von Bonsdorff, C H; Lappalainen, M; Korhonen, T; Kuusi, M; Niskanen, T

2009-12-10

We describe a cluster of norovirus outbreaks affecting about 200 people in Southern Finland in September and October 2009. All outbreaks occurred after consumption of imported raspberries from the same batch intended for the catering sector. Human norovirus genotype GI.4 was found in frozen raspberries. The berries were served in toppings of cakes in separate catering settings or mixed in curd cheese as a snack for children in a daycare center. The relative risk for consumption of the berry dish was 3.0 (p

Cryopreservation of epididymal sperm.

PubMed

Patrizio, P

2000-11-27

The advent of ICSI and the perfecting of freezing protocols for sperm samples that in the pre-ICSI era would not have been frozen, allows now routine cryopreservation of epididymal sperm regardless of their quality and quantity. There are two methods to retrieve epididymal sperm: microsurgical epididymal sperm aspiration (MESA) and percutaneous epididymal sperm aspiration (PESA). The majority of the literature has focused on the technique of MESA to obtain sperm on the claim that the amount of sperm retrieved with PESA might not be sufficient to allow cryopreservation. However, there are no data on cryopreservation and ICSI with epididymal sperm collected with PESA technique. In this study, a total of 68 consecutive cycles of PESA, of which 46 were performed with fresh epididymal sperm and 22 with frozen/thawed specimens were retrospectively analyzed. In the fresh epididymal group (n = 46), 446 eggs were injected and 207 cleaving embryos were obtained (fertilization rate of 46%). In the cryopreserved epididymal sperm group (n = 22), 216 eggs were injected and 115 cleaving embryos were obtained (fertilization rate of 53%, P = NS). There were 18 pregnancies (39%) with 17 (37%) delivered/ongoing in the fresh group, while there were 11 (50%) with 9 (41%) delivered/ongoing in the frozen group (P = NS). Epididymal sperm for cryopreservation was available in 44 of the 46 PESA cycles. Additionally, in the fresh group, 19 couples had excess embryos for cryopreservation while in the frozen group, ten couples had excess embryos for cryopreservation. A total of 17 frozen embryo transfer with epididymal sperm from PESA were analyzed. Of these, 12 FET were from embryos from the fresh epididymal group and three pregnancies with livebirths (25%) were recorded. Five FET were performed with extra embryos from frozen epididymal sperm and two (40%) pregnancies with livebirths were obtained. In summary, these data show that epididymal sperm obtained by PESA can be successfully cryopreserved in order to avoid future retrievals procedures and fertilization and pregnancy rates are similar between fresh and cryopreserved epididymal sperm. It is also reported for the first time that the transfer of frozen embryos obtained with either fresh or frozen thawed epididymal sperm leads to the same pregnancy'and delivery rate.

Multiplex transcriptional analysis of paraffin-embedded liver needle biopsy from patients with liver fibrosis

PubMed Central

2012-01-01

Background The possibility of extracting RNA and measuring RNA expression from paraffin sections can allow extensive investigations on stored paraffin samples obtained from diseased livers and could help with studies of the natural history of liver fibrosis and inflammation, and in particular, correlate basic mechanisms to clinical outcomes. Results To address this issue, a pilot study of multiplex gene expression using branched-chain DNA technology was conducted to directly measure mRNA expression in formalin-fixed paraffin-embedded needle biopsy samples of human liver. Twenty-five genes were selected for evaluation based on evidence obtained from human fibrotic liver, a rat BDL model and in vitro cultures of immortalized human hepatic stellate cells. The expression levels of these 25 genes were then correlated with liver fibrosis and inflammation activity scores. Statistical analysis revealed that three genes (COL3A1, KRT18, and TUBB) could separate fibrotic from non-fibrotic samples and that the expression of ten genes (ANXA2, TIMP1, CTGF, COL4A1, KRT18, COL1A1, COL3A1, ACTA2, TGFB1, LOXL2) were positively correlated with the level of liver inflammation activity. Conclusion This is the first report describing this multiplex technique for liver fibrosis and has provided the proof of concept of the suitability of RNA extracted from paraffin sections for investigating the modulation of a panel of proinflammatory and profibrogenic genes. This pilot study suggests that this technique will allow extensive investigations on paraffin samples from diseased livers and possibly from any other tissue. Using identical or other genes, this multiplex expression technique could be applied to samples obtained from extensive patient cohorts with stored paraffin samples in order to correlate gene expression with valuable clinically relevant information. This method could be used to provide a better understanding of the mechanisms of liver fibrosis and inflammation, its progression, and help development of new therapeutic approaches for this indication. PMID:23270325

Prior frozen storage enhances the effect of edible coatings against Listeria monocytogenes on cold-smoked salmon during subsequent refrigerated storage.

PubMed

Ye, M; Neetoo, H; Chen, H

2011-10-01

Listeria monocytogenes is a major safety concern for ready-to-eat foods. The overall objective of this study was to investigate whether prior frozen storage could enhance the efficacy of edible coatings against L. monocytogenes on cold-smoked salmon during subsequent refrigerated storage. A formulation consisting of sodium lactate (SL, 1·2-2·4%) and sodium diacetate (SD, 0·125-0·25%) or 2·5% Opti.Form (a commercial formulation of SL and SD) was incorporated into each of five edible coatings: alginate, κ-carrageenan, pectin, gelatin and starch. The coatings were applied onto the surface of cold-smoked salmon slices inoculated with L. monocytogenes at a level of 500 CFU cm⁻². In the first phase, the slices were first frozen at -18°C for 6 days and stored at 22°C for 6 days. Alginate, gelatin and starch appeared to be the most effective carriers. In the second phase, cold-smoked salmon slices were inoculated with L. monocytogenes, coated with alginate, gelatin or starch with or without the antimicrobials and stored frozen at -18°C for 12 months. Every 2 months, samples were removed from the freezer and kept at 4°C for 30 days. Prior frozen storage at -18°C substantially enhanced the antilisterial efficacy of the edible coatings with or without antimicrobials during the subsequent refrigerated storage. Plain coatings with ≥ 2 months frozen storage and antimicrobial edible coatings represent an effective intervention to inhibit the growth of L. monocytogenes on cold-smoked salmon. This study demonstrates the effectiveness of the conjunct application of frozen storage and edible coatings to control the growth of L. monocytogenes to enhance the microbiological safety of cold-smoked salmon. © 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.

Microbiological evaluation of South Australian rock lobster meat.

PubMed

Yap, A S

1977-12-01

Samples of frozen precooked rock lobster meat from five South Australian fish-processing plants situated in the West Coast and south-east regions were tested over a period of six months during the 1974/5 lobster fishing season. The most probable number (MPN) of E. coli and coliforms, Staphylococcus aureus and Salmonella, as well as total plate count (TPC) were determined in 480 samples. Monthly geometric mean TPC ranged from 1600/g to 25,000/g. The highest geometric mean of the MPN of coliforms and E. coli were 4.9/g and 1.8/g respectively. The highest geometric mean number of staphylococci was 18.6/g. Salmonella was not detected in the 480 units tested. Only 0.4% of the samples had TPC exceeding 100,000/g. Coliforms and E. coli were not present in 76.1% and 92.7% respectively of the samples tested. Staphylococcus aureus was not detected in 67.7% of the samples. The numbers of organisms in 82% of the samples fall within the microbiological standards proposed by the National Health and Medical Research Council of Australia for frozen precooked foods. The results of this study demonstrate the microbial quality of precooked lobster meat attainable when good manufacturing practices are used.

Determination of the methylation status of MGMT in different regions within glioblastoma multiforme.

PubMed

Hamilton, Mark G; Roldán, Gloria; Magliocco, Anthony; McIntyre, John B; Parney, Ian; Easaw, Jacob C

2011-04-01

Epigenetic silencing of the MGMT gene through promoter methylation correlates with improved survival in Glioblastoma Multiforme (GBM) patients receiving concurrent chemoradiotherapy. Although the clinical benefit is primarily seen in patients with methylated MGMT promoter, some unmethylated patients also respond to Temozolomide. One possible explanation may be intratumoral heterogeneity. This study was designed to assess the methylation status of the MGMT promoter in different areas of GBM and determine if methylation status varied depending on the fixation technique (paraffin-embedding versus fresh frozen) used to store tissue. Using intraoperative navigation, biopsies were obtained from three distinct regions: the enhancing outer area, the non-enhancing inner core, and an area immediately outside the enhancing region. Only patients with GBM were included for evaluation and analysis. Samples taken from each area were divided with half stored by flash freezing and the other half stored using paraffin fixation. Methylation Specific-PCR (MS-PCR) was used for analysis of MGMT promoter methylation. Thirteen patients were included. Ten were male with a median age of 62 years. In each patient, samples were taken from the enhancing rim and the necrotic centre. However, it was not considered safe or feasible to obtain samples from the area immediately adjacent to the enhancing tumor rim in one case. All patients were homogeneous for methylation status throughout their tumor and tissue taken adjacent to it when frozen tissue was used. However, four patients had discrepancies in the MGMT promoter status between the frozen and paraffin-embedded blocks and one patient was not homogeneous within the tumor when paraffin-embedded tissue was used. MGMT promoter methylation status was homogeneous in all GBM tumors. Our observation that methylation status varied depending if the DNA was extracted from paraffin-embedded versus frozen tissue is concerning. Although the reason for this is unclear, we postulate that the timing from resection to fixation or the process of fixation itself may potentially alter methylation status in paraffin-embedded tumors.

Immersion freezing of birch pollen washing water

NASA Astrophysics Data System (ADS)

Augustin, S.; Wex, H.; Niedermeier, D.; Pummer, B.; Grothe, H.; Hartmann, S.; Tomsche, L.; Clauss, T.; Voigtländer, J.; Ignatius, K.; Stratmann, F.

2013-11-01

Birch pollen grains are known to be ice nucleating active biological particles. The ice nucleating activity has previously been tracked down to biological macromolecules that can be easily extracted from the pollen grains in water. In the present study, we investigated the immersion freezing behavior of these ice nucleating active (INA) macromolecules. Therefore we measured the frozen fractions of particles generated from birch pollen washing water as a function of temperature at the Leipzig Aerosol Cloud Interaction Simulator (LACIS). Two different birch pollen samples were considered, with one originating from Sweden and one from the Czech Republic. For the Czech and Swedish birch pollen samples, freezing was observed to start at -19 and -17 °C, respectively. The fraction of frozen droplets increased for both samples down to -24 °C. Further cooling did not increase the frozen fractions any more. Instead, a plateau formed at frozen fractions below 1. This fact could be used to determine the amount of INA macromolecules in the droplets examined here, which in turn allowed for the determination of nucleation rates for single INA macromolecules. The main differences between the Swedish birch pollen and the Czech birch pollen were obvious in the temperature range between -17 and -24 °C. In this range, a second plateau region could be seen for Swedish birch pollen. As we assume INA macromolecules to be the reason for the ice nucleation, we concluded that birch pollen is able to produce at least two different types of INA macromolecules. We were able to derive parameterizations for the heterogeneous nucleation rates for both INA macromolecule types, using two different methods: a simple exponential fit and the Soccer ball model. With these parameterization methods we were able to describe the ice nucleation behavior of single INA macromolecules from both the Czech and the Swedish birch pollen.

Changes in the quality of surimi made from thornback ray (Raja clavata, L. 1758) during frozen storage.

PubMed

Turan, Hülya; Sönmez, Gülşah

2007-11-01

Surimi was prepared from the thornback ray (Raja clavata L. 1758) and divided into two groups. The first group was prepared with 4% sorbitol, 4% sucrose and 0.3% sodium tripolyphosphate as a cryoprotectant, while surimi in second group was prepared with 8% sorbitol and 0.3% sodium tripolyphosphate. The frozen surimi samples were stored at 23.8 +/- 2 degrees C for 6 months. The total volatile basic nitrogen (8.40 mg/100 g for group A, 6.30 mg/100 g for group B), trimethylamine nitrogen (2.55 mg/100 g for group A, 2.38 mg/100 g for group B), thiobarbituric acid (1.29 mg malondialdehyde/100 g for group A, 1.17 mg malondialdehyde/ 100 g for group B), and pH values (7.34 for group A, 6.98 for group B) of surimi increased during frozen storage but remained within the acceptable limits. Total psychrophilic aerobic bacteria counts and sensory evaluation points in both groups decreased during frozen storage. The results of this study showed that thornback ray was found to be suitable for surimi production and the surimis were still acceptable at the end of the 6-month storage period.

Bis(trifluoromethylsulfonyl)imide-based frozen ionic liquid for the hollow-fiber solid-phase microextraction of dichlorodiphenyltrichloroethane and its main metabolites.

PubMed

Pang, Long; Yang, Peijie; Pang, Rong; Li, Shunyi

2017-08-01

1-Hexadecyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide is a solid-phase ionic organic material under ambient temperature and is considered as a kind of "frozen" ionic liquid. Because of their solid-state and ultra-hydrophobicity, "frozen" ionic liquids are able to be confined in the pores of hollow fiber, based on which a simple method was developed for the hollow-fiber solid-phase microextraction of dichlorodiphenyltrichloroethane and its main metabolites. Under optimized conditions, the proposed method results in good linearity (R 2 > 0.9965) over the range of 0.5-50 μg/L, with low limits of detection and quantification in the range of 0.33-0.38 and 1.00-1.25 μg/L, respectively. Intra- and interday precisions evaluated by relative standard deviation were 3-6 and 1-6%, respectively. The spiked recoveries of dichlorodiphenyltrichloroethane and its main metabolites from real water samples were in the range of 64-113 and 79-112%, respectively, at two different concentration levels. The results suggest that "frozen" ionic liquids are promising for use as a class of novel sorbents. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Association of birth defects with the mode of assisted reproductive technology in a Chinese data-linkage cohort.

PubMed

Yu, Hui-Ting; Yang, Qing; Sun, Xiao-Xi; Chen, Guo-Wu; Qian, Nai-Si; Cai, Ren-Zhi; Guo, Han-Bing; Wang, Chun-Fang

2018-05-01

To evaluate the impact of assisted reproductive technology (ART) on the offspring of Chinese population. Retrospective, data-linkage cohort. Not applicable. Live births resulting from ART or natural conception. None. Birth defects coded according to ICD-10. Births after ART were more likely to be female and multiple births, especially after intracytoplasmic sperm injection (ICSI). ART was associated with a significantly increased risk of birth defects, especially, among singleton births, a significantly increased risk in fresh-embryo cycles after in vitro fertilization (IVF) and frozen-embryo cycles after ICSI. Associations between ART and multiple defects, between ART and gastrointestinal malformation, genital organs malformation, and musculoskeletal malformation among singleton births, and between ART and cardiac septa malformation among multiple births were observed. This study suggests that ART increases the risk of birth defects. Subgroup analyses indicate higher risk for both fresh and frozen embryos, although nonsignificantly for frozen embryos after IVF and for fresh embryos were presented with low power. Larger sample size research is needed to clarify effects from fresh- or frozen-embryo cycles after IVF and ICSI. Copyright © 2018 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Evaluation and utilization of preassembled frozen commercial fast real-time qPCR master mixes for detection of cytomegalovirus and BK virus.

PubMed

Glover, William A; Atienza, Ederlyn E; Nesbitt, Shannon; Kim, Woo J; Castor, Jared; Cook, Linda; Jerome, Keith R

2016-01-01

Quantitative DNA detection of cytomegalovirus (CMV) and BK virus (BKV) is critical in the management of transplant patients. Quantitative laboratory-developed procedures for CMV and BKV have been described in which much of the processing is automated, resulting in rapid, reproducible, and high-throughput testing of transplant patients. To increase the efficiency of such assays, the performance and stability of four commercial preassembled frozen fast qPCR master mixes (Roche FastStart Universal Probe Master Mix with Rox, Bio-Rad SsoFast Probes Supermix with Rox, Life Technologies TaqMan FastAdvanced Master Mix, and Life Technologies Fast Universal PCR Master Mix), in combination with in-house designed primers and probes, was evaluated using controls and standards from standard CMV and BK assays. A subsequent parallel evaluation using patient samples was performed comparing the performance of freshly prepared assay mixes versus aliquoted frozen master mixes made with two of the fast qPCR mixes (Life Technologies TaqMan FastAdvanced Master Mix, and Bio-Rad SsoFast Probes Supermix with Rox), chosen based on their performance and compatibility with existing PCR cycling conditions. The data demonstrate that the frozen master mixes retain excellent performance over a period of at least 10 weeks. During the parallel testing using clinical specimens, no difference in quantitative results was observed between the preassembled frozen master mixes and freshly prepared master mixes. Preassembled fast real-time qPCR frozen master mixes perform well and represent an additional strategy laboratories can implement to reduce assay preparation times, and to minimize technical errors and effort necessary to perform clinical PCR. © 2015 Wiley Periodicals, Inc.

Effect of cryopreservation on IL-4, IFNgamma and IL-6 production of porcine peripheral blood lymphocytes.

PubMed

Li, Xiujin; Zhong, Zhenyu; Liang, Shuang; Wang, Xingxing; Zhong, Fei

2009-12-01

Cryopreservation of animal or human peripheral blood mononuclear cells (PBMC) is a commonly used technique. Effects of cryopreservation on functional capacity, especially the cytokine production of human PBMCs, have been extensively defined. However, certain animals, such as livestock, are a shortage of these information. Here we investigated the effects of cryopreservation on cytokine (IL-4, IFNgamma and IL-6) production of porcine PBMC. The porcine PBMCs were cryopreserved at -196 degrees C for a variety time periods for 2, 5, 25 and 50 days. Viability and cytokine production of the porcine PBMCs were measured before and after cryopreservation. The results showed that about 90% cell recovery rate was obtained at each storage time, indicating that about 10% loss of PBMCs in this short-term cryopreservation was due to the freezing process rather than the duration of cryopreservation. The fresh or frozen resting porcine PBMCs produced little cytokines in the absence of stimulation. However, three cytokines were apparently increased after PMA stimulation in both fresh and frozen porcine PBMCs. The sensitivity of frozen cells to PMA simulation for IFNgamma and IL-6 production was different from that of the fresh ones. IFNgamma production from the frozen PBMCs was significantly higher than that from the fresh ones (P<0.01). In contrast, IL-6 level from the frozen sample was significantly lower than that from the fresh one (P<0.05). Those results indicate that cryopreservation can increase the sensitivity of porcine PBMCs stimulated by PMA for IFNgamma production but not for IL-6 production. There was no significant difference of IL-4 production between fresh and frozen cells either stimulated (P>0.05) or un-stimulated (P>0.05).

Trend analysis of Trichinella in a red fox population from a low endemic area using a validated artificial digestion and sequential sieving technique.

PubMed

Franssen, Frits; Deksne, Gunita; Esíte, Zanda; Havelaar, Arie; Swart, Arno; van der Giessen, Joke

2014-11-28

Freezing of fox carcasses to minimize professional hazard of infection with Echinococcus multilocularis is recommended in endemic areas, but this could influence the detection of Trichinella larvae in the same host species. A method based on artificial digestion of frozen fox muscle, combined with larva isolation by a sequential sieving method (SSM), was validated using naturally infected foxes from Latvia. The validated SSM was used to detect dead Trichinella muscle larvae (ML) in frozen muscle samples of 369 red foxes from the Netherlands, of which one fox was positive (0.067 larvae per gram). This result was compared with historical Trichinella findings in Dutch red foxes. Molecular analysis using 5S PCR showed that both T. britovi and T. nativa were present in the Latvian foxes, without mixed infections. Of 96 non-frozen T. britovi ML, 94% was successfully sequenced, whereas this was the case for only 8.3% of 72 frozen T. britovi ML. The single Trichinella sp. larva that was recovered from the positive Dutch fox did not yield PCR product, probably due to severe freeze-damage. In conclusion, the SSM presented in this study is a fast and effective method to detect dead Trichinella larvae in frozen meat. We showed that the Trichinella prevalence in Dutch red fox was 0.27% (95% CI 0.065-1.5%), in contrast to 3.9% in the same study area fifteen years ago. Moreover, this study demonstrated that the efficacy of 5S PCR for identification of Trichinella britovi single larvae from frozen meat is not more than 8.3%.

Effect of semen extenders on frozen-thawed boar sperm characteristics and distribution in the female genital tract after deep intrauterine insemination in sows.

PubMed

Noguchi, Michiko; Yoshioka, Koji; Hikono, Hirokazu; Suzuki, Chie; Kikuchi, Kazuhiro

2015-12-01

We compared the effects of extenders of frozen-thawed semen on post-thaw sperm characteristics and the distribution of frozen-thawed spermatozoa in the female genital tract after fixed-timed deep intrauterine insemination (DIUI) in sows. Frozen semen samples were thawed and diluted in either modified Modena solution (mMS) or porcine fertilization medium (PFM) containing theophylline, adenosine and cysteine. Sperm quality, assessed in vitro based on motility using a computer-assisted sperm analyzer and the integrity of the plasma and acrosomal membranes using flow cytometry, was evaluated at 0.5, 1.5, 3 and 6h after thawing. Progressive motility and the percentage of spermatozoa with damaged acrosomal membranes in PFM were significantly better than in mMS throughout the 6h. Sows with estrus synchronized using prostaglandin F2 alpha, equine chorionic gonadotropin and human chorionic gonadotropin (hCG) were inseminated once with mMS- or PFM-diluted 5 × 10(8) frozen-thawed spermatozoa by DIUI at 34 h after the hCG injection. At 4h after DIUI, reproductive tracts were recovered from 30 sows. There were significantly fewer polymorphonuclear leukocytes (PMNs) and more spermatozoa outside PMNs in the uterine horn after PFM treatment than with mMS. When 22 sows were administered DIUI with 10 × 10(8) frozen-thawed spermatozoa at 36 h after hCG, the pregnancy rates did not differ significantly between the mMS- (36%) and PFM- (64%) treated groups. Thus, PFM enhanced progressive sperm motility but increased sperm membrane damage compared with mMS; it also suppressed the migration of PMNs into the uterine lumen. Copyright © 2015 Elsevier B.V. All rights reserved.

A score model for the continuous grading of early allograft dysfunction severity.

PubMed

Pareja, Eugenia; Cortes, Miriam; Hervás, David; Mir, José; Valdivieso, Andrés; Castell, José V; Lahoz, Agustín

2015-01-01

Early allograft dysfunction (EAD) dramatically influences graft and patient outcomes. A lack of consensus on an EAD definition hinders comparisons of liver transplant outcomes and management of recipients among and within centers. We sought to develop a model for the quantitative assessment of early allograft function [Model for Early Allograft Function Scoring (MEAF)] after transplantation. A retrospective study including 1026 consecutive liver transplants was performed for MEAF score development. Multivariate data analysis was used to select a small number of postoperative variables that adequately describe EAD. Then, the distribution of these variables was mathematically modeled to assign a score for each actual variable value. A model, based on easily obtainable clinical parameters (ie, alanine aminotransferase, international normalized ratio, and bilirubin) and scoring liver function from 0 to 10, was built. The MEAF score showed a significant association with patient and graft survival at 3-, 6- and 12-month follow-ups. Hepatic steatosis and age for donors; cold/warm ischemia times and postreperfusion syndrome for surgery; and intensive care unit and hospital stays, Model for End-Stage Liver Disease and Child-Pugh scores, body mass index, and fresh frozen plasma transfusions for recipients were factors associated significantly with EAD. The model was satisfactorily validated by its application to an independent set of 200 patients who underwent liver transplantation at a different center. In conclusion, a model for the quantitative assessment of EAD severity has been developed and validated for the first time. The MEAF provides a more accurate graft function assessment than current categorical classifications and may help clinicians to make early enough decisions on retransplantation benefits. Furthermore, the MEAF score is a predictor of recipient and graft survival. The standardization of the criteria used to define EAD may allow reliable comparisons of recipients' treatments and transplant outcomes among and within centers. © 2014 American Association for the Study of Liver Diseases.

Coherent three-dimensional X-ray cryo-imaging.

PubMed

Robinson, Ian

2015-09-01

The combination of cryogenic sample temperatures with three-dimensional coherent diffractive imaging for the case of whole frozen-hydrated cells is discussed in the light of theoretical predictions of the achievable resolution.

Automated liver sampling using a gradient dual-echo Dixon-based technique.

PubMed

Bashir, Mustafa R; Dale, Brian M; Merkle, Elmar M; Boll, Daniel T

2012-05-01

Magnetic resonance spectroscopy of the liver requires input from a physicist or physician at the time of acquisition to insure proper voxel selection, while in multiecho chemical shift imaging, numerous regions of interest must be manually selected in order to ensure analysis of a representative portion of the liver parenchyma. A fully automated technique could improve workflow by selecting representative portions of the liver prior to human analysis. Complete volumes from three-dimensional gradient dual-echo acquisitions with two-point Dixon reconstruction acquired at 1.5 and 3 T were analyzed in 100 subjects, using an automated liver sampling algorithm, based on ratio pairs calculated from signal intensity image data as fat-only/water-only and log(in-phase/opposed-phase) on a voxel-by-voxel basis. Using different gridding variations of the algorithm, the average correct liver volume samples ranged from 527 to 733 mL. The average percentage of sample located within the liver ranged from 95.4 to 97.1%, whereas the average incorrect volume selected was 16.5-35.4 mL (2.9-4.6%). Average run time was 19.7-79.0 s. The algorithm consistently selected large samples of the hepatic parenchyma with small amounts of erroneous extrahepatic sampling, and run times were feasible for execution on an MRI system console during exam acquisition. Copyright © 2011 Wiley Periodicals, Inc.

78 FR 2946 - United States Standards for Grades of Frozen Vegetables

Federal Register 2010, 2011, 2012, 2013, 2014

2013-01-15

... these proposed revisions are: frozen asparagus, frozen lima beans, frozen speckled butter beans, frozen... document. United States Standards for Grades of Frozen Lima Beans Update address for AMS. Change ``U.S... obtain color standards for frozen lima beans. United States Standards for Grades of Frozen Mixed...

Bacteriological survey of frozen meat and gravy produced at establishments under federal inspection.

PubMed

Surkiewicz, B F; Harris, M E; Johnston, R W

1973-10-01

During visits to 34 federally inspected establishments producing frozen meat and gravy, 541 production line samples and 535 finished product units were collected for bacteriological analyses. It was found that more than 70% of the sets of finished product (10 units/set) produced under good manufacturing practices had: (i) four or fewer coliform-positive units, (ii) two or fewer Escherichia coli-positive units, (iii) three or fewer Staphylococcus aureus-positive units, and (iv) an aerobic plate count of fewer than 50,000/g (geometric mean of 10 units). All finished product units were negative for salmonellae.

Bacteriological Survey of Frozen Meat and Gravy Produced at Establishments Under Federal Inspection

PubMed Central

Surkiewicz, Bernard F.; Harris, Marshall E.; Johnston, Ralph W.

1973-01-01

During visits to 34 federally inspected establishments producing frozen meat and gravy, 541 production line samples and 535 finished product units were collected for bacteriological analyses. It was found that more than 70% of the sets of finished product (10 units/set) produced under good manufacturing practices had: (i) four or fewer coliform-positive units, (ii) two or fewer Escherichia coli-positive units, (iii) three or fewer Staphylococcus aureus-positive units, and (iv) an aerobic plate count of fewer than 50,000/g (geometric mean of 10 units). All finished product units were negative for salmonellae. PMID:4584595

Multiphoton microscopy for the in-situ investigation of cellular processes and integrity in cryopreservation.

PubMed

Doerr, Daniel; Stark, Martin; Ehrhart, Friederike; Zimmermann, Heiko; Stracke, Frank

2009-08-01

In this study we demonstrate a new noninvasive imaging method to monitor freezing processes in biological samples and to investigate life in the frozen state. It combines a laser scanning microscope with a computer-controlled cryostage. Nearinfrared (NIR) femtosecond laser pulses evoke the fluorescence of endogenous fluorophores and fluorescent labels due to multiphoton absorption.The inherent optical nonlinearity of multiphoton absorption allows 3D fluorescence imaging for optical tomography of frozen biological material in-situ. As an example for functional imaging we use fluorescence lifetime imaging (FLIM) to create images with chemical and physical contrast.

The Effect of Volcanic Ash Composition on Ice Nucleation Affinity

NASA Astrophysics Data System (ADS)

Genareau, K. D.; Cloer, S.; Primm, K.; Woods, T.; Tolbert, M. A.

2017-12-01

Understanding the role that volcanic ash plays in ice nucleation is important for knowledge of lightning generation in both volcanic plumes and in clouds developing downwind from active volcanoes. Volcanic ash has long been suggested to influence heterogeneous ice nucleation following explosive eruptions, but determining precisely how composition and mineralogy affects ice nucleation affinity (INA) is poorly constrained. For the study presented here, volcanic ash samples with different compositions and mineral/glass contents were tested in both the deposition and immersion modes, following the methods presented in Schill et al. (2015). Bulk composition was determined with X-ray fluorescence (XRF), grain size distribution was determined with laser diffraction particle size analysis (LDPSA), and mineralogy was determined with X-ray diffraction (XRD) and scanning electron microscopy (SEM). Results of the deposition-mode experiments reveal that there is no relationship between ice saturation ratios (Sice) and either mineralogy or bulk ash composition, as all samples have similar Sice ratios. In the immersion-mode experiments, frozen fractions were determined from -20 °C to -50 °C using three different amounts of ash (0.5, 1.0, and 2.0 wt% of slurry). Results from the immersion freezing reveal that the rhyolitic samples (73 wt% SiO2) nucleate ice at higher temperatures compared to the basaltic samples (49 wt% SiO2). There is no observed correlation between frozen fractions and mineral content of ash samples, but the two most efficient ice nuclei are rhyolites that contain the greatest proportion of amorphous glass (> 90 %), and are enriched in K2O relative to transition metals (MnO and TiO2), the latter of which show a negative correlation with frozen fraction. Higher ash abundance in water droplets increases the frozen fraction at all temperatures, indicating that ash amount plays the biggest role in ice nucleation. If volcanic ash can reach sufficient abundance (≥ 2 wt%) in hydrometeors, and be compositionally enriched in K2O relative to MnO and TiO2, the nucleation of ice should efficiently occur. These chemical relationships are not only important for understanding ice nucleation in volcanic plumes, but also for constraining the effect of composition on the INA of other atmospheric aerosols.

Traumatic Hemothorax Blood Contains Elevated Levels of Microparticles that are Prothrombotic but Inhibit Platelet Aggregation.

PubMed

Mitchell, Thomas A; Herzig, Maryanne C; Fedyk, Chriselda G; Salhanick, Marc A; Henderson, Aaron T; Parida, Bijaya K; Prat, Nicolas J; Dent, Daniel L; Schwacha, Martin G; Cap, Andrew P

2017-06-01

Autotransfusion of shed blood from traumatic hemothorax is an attractive option for resuscitation of trauma patients in austere environments. However, previous analyses revealed that shed hemothorax (HX) blood is defibrinated, thrombocytopenic, and contains elevated levels of D-dimer. Mixing studies with normal pooled plasma demonstrated hypercoagulability, evoking concern for potentiation of acute traumatic coagulopathy. We hypothesized that induction of coagulopathic changes by shed HX blood may be due to increases in cellular microparticles (MP) and that these may also affect recipient platelet function. Shed HX blood was obtained from 17 adult trauma patients under an Institutional Review Board approved prospective observational protocol. Blood samples were collected every hour up to 4 h after thoracostomy tube placement. The corresponding plasma was isolated and frozen for analysis. The effects of shed HX frozen plasma (HFP) and isolated HX microparticles (HMP) on coagulation and platelet function were assessed through mixing studies with platelet-rich plasma at various dilutions followed by analysis with thromboelastometry (ROTEM), platelet aggregometry (Multiplate), enzyme-linked immunosorbent assays, and flow cytometry. Furthermore, HFP was assessed for von Willebrand factor antigen levels and multimer content, and plasma-free hemoglobin. ROTEM analysis demonstrated that diluted HFP and isolated HMP samples decreased clotting time, clotting formation time, and increased α angle, irrespective of sample concentrations, when compared with diluted control plasma. Isolated HMP inhibited platelet aggregation in response to adenosine diphosphate, arachidonic acid, and collagen. HFP contained elevated levels of fibrin-degradation products and tissue factor compared with control fresh frozen plasma samples. MP concentrations in HFP were significantly increased and enriched in events positive for phosphatidylserine, tissue factor, CD235, CD45, CD41a, and CD14. von Willebrand factor (vWF) multimer analysis revealed significant loss of high molecular weight multimers in HFP samples. Plasma-free hemoglobin levels were 8-fold higher in HFP compared with fresh frozen plasma. HFP induces plasma hypercoagulability that is likely related to increased tissue factor and phosphatidylserine expression originating from cell-derived MP. In contrast, platelet dysfunction is induced by HMP, potentially aggravated by depletion of high molecular weight multimers of vWF. Thus, autologous transfusion of shed traumatic hemothorax blood may induce a range of undesirable effects in patients with acute traumatic coagulopathy.

Development and validation of a UPLC-MS/MS method for the simultaneous determination of paritaprevir and ritonavir in rat liver.

PubMed

Ocque, Andrew J; Hagler, Colleen E; Difrancesco, Robin; Woolwine-Cunningham, Yvonne; Bednasz, Cindy J; Morse, Gene D; Talal, Andrew H

2016-07-01

Determination of paritaprevir and ritonavir in rat liver tissue samples. We successfully validated a UPLC-MS/MS method to measure paritaprevir and ritonavir in rat liver using deuterated internal standards (d8-paritapervir and d6-ritonavir). The method is linear from 20 to 20,000 and 5 to 10,000 pg on column for paritaprevir and ritonavir, respectively, and is normalized per milligram tissue. Interday and intraday variability ranged from 0.591 to 5.33% and accuracy ranged from -6.68 to 10.1% for quality control samples. The method was then applied to the measurement of paritaprevir and ritonavir in rat liver tissue samples from a pilot study. The validated method is suitable for measurement of paritaprevir and ritonavir within rat liver tissue samples for PK studies.

7 CFR 70.105 - Procedures for appeal gradings.

Code of Federal Regulations, 2011 CFR

2011-01-01

... PRODUCTS AND RABBIT PRODUCTS Grading of Poultry Products and Rabbit Products Appeal of A Grading Or... sample size for the lot shall consist of double the samples required in § 70.80. (c) Poultry or rabbits... is performed. (d) Overwraps on frozen poultry or rabbits shall be removed from all birds or rabbits...

A Study of Some Problems in Network Information Theory

ERIC Educational Resources Information Center

Kamath, Sudeep Uday

2013-01-01

Red snapper, "Lutjanus campechanus," were sampled with hook and line at natural (n = 33) and artificial (n = 27) reef sites in the northern Gulf of Mexico from 2009-2011. Stomachs (n = 708) were extracted and their contents preserved for gut content analysis, and muscle tissue samples (n = 200) were dissected and frozen for stable…

Evaluation of the qualitative and quantitative effectiveness of three media of centrifugation (Maxifreeze, Cushion Fluid Equine, and PureSperm 100) in preparation of fresh or frozen-thawed brown bear spermatozoa.

PubMed

Nicolas, M; Alvarez, M; Borragán, S; Martinez-Pastor, F; Chamorro, C A; Alvarez-Rodriguez, M; de Paz, P; Anel, L

2012-04-01

Centrifugation is a crucial procedure in sperm cryopreservation protocols of brown bear (Ursus arctos), because the semen must be processed to increase sperm concentration and/or clean urine-contaminated samples. The efficacy of three media for centrifugation (Maxifreeze [IMV technologies, L'Aigle, France], Cushion Fluid Equine (Minitübe, Tiefenbach, Germany), and PureSperm [Nidacon, Gothenburg, Sweden]) on the quality of bear spermatozoa was evaluated. In experiment one, two cushioned media used for protecting against mechanical stress during centrifugation were analyzed. In experiment two, a density gradient based on PureSperm was assessed in relation to the maximum retrieval and the quality of fresh spermatozoa, and the freezability of the spermatozoa selected in this density gradient was studied in experiment three. Finally, the selection of frozen-thawed sperm using PureSperm was analyzed in experiment four. Our results indicate that the use of dense isotonic cushion solutions (Maxifreeze, Cushion Fluid Equine) in centrifugation did not improve the quality of recovered spermatozoa compared with standard centrifugation. However, a density gradient prepared with PureSperm improved the quality of spermatozoa in fresh semen and frozen-thawed semen, but the spermatozoa selected from the fresh sample with this density gradient did not show a better resistance to freezing with this density gradient in comparison with the control sample. Copyright © 2012 Elsevier Inc. All rights reserved.

IRRADIATION OF EGGS AND EGG PRODUCTS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brooks, J.; Hannan, R.S.; Hobbs, B.C.

It has been known for some time that egg products may contain members of the Salmonella group capable of causing food poisoning, and the position has been rendered more serious by the recent discovery of S. paratyphi B in some of the samples of egg products. Attempts to destroy salmonellae in liquid egg by pasteurization before freezing or drying have met with considerable success but the process requires careful control. The tins in which the frozen product is distributed hold up to 20 kg, and are therefore of a size suitable for treatment with gamma -radiation. After treatment of smallmore » samples of frozen whole egg with 2 Mev cathode rays it was concluded that a dose of about O.3 to 0.5 Mrad would destroy the numbers of salmonellae normally encountered in the product without impairing the baking qualities of the material. Whole tins, each containing 10 kg of infected material, were therefore irradiated in the frozen state with Co/sup 60/ gamma -rays. Two tins were treated at each of three dose levels of 0.3, 0.4, and 0.5 Mrad. No salmonellae were detected in duplicate samples of 25 g of material taken from each of the tins after irradiation. If the effectiveness of the treatment is confirmed, the process has obvious attractions since it dispenses with the need to thaw or otherwise to handle the product. The paper also contains a general discussion on the irradiation of eggs and egg products. (auth)« less

Evaluation of green tea extract as a glazing material for shrimp frozen by cryogenic freezing.

PubMed

Sundararajan, Srijanani; Prudente, Alfredo; Bankston, J David; King, Joan M; Wilson, Paul; Sathivel, Subramaniam

2011-09-01

Solutions of green tea (Camellia sinensis) extract (GTE) in distilled water were evaluated as a glazing material for shrimp frozen by cryogenic freezing. Total of 2%, 3%, and/or 5% GTE solutions (2GTE, 3GTE, 5GTE) were used for glazing. Distilled water glazed (GDW) and nonglazed shrimp (NG) served as controls. The GTE was characterized by measuring color, pH, (o) Brix, total phenols, and % antiradical activity. Individual catechins were identified by HPLC. The freezing time, freezing rate, and energy removal rate for freezing shrimp by cryogenic freezing process were estimated. The frozen shrimp samples were stored in a freezer at -21 °C for 180 d. Samples were analyzed for pH, moisture content, glazing yield, thaw yield, color, cutting force, and thiobarbituric acid reactive substances (TBARS) after 1, 30, 90, and 180 d. The HPLC analysis of GTE revealed the presence of catechins and their isomers and the total polyphenol content was 148.10 ± 2.49 g/L. The freezing time (min) and energy removal rate (J/s) were 48.67 ± 2.3 and 836.67 ± 78.95, respectively. Glazed samples had higher moisture content compared to NG shrimp after 180 d storage. GTE was effective in controlling the lipid oxidation in shrimp. Glazing with GTE affected a* and b* color values, but had no significant effect on the L* values of shrimp. © 2011 Institute of Food Technologists®

[Surgical Treatment of Right Atrial Malignant Tumor Thrombus Under Deep Hypothermia with Intermittent Circulatory Arrest;Report of Two Cases].

PubMed

Fujita, Akira; Kobayashi, Toshiro; Hironaka, Hideharu; Jinbou, Mitsutaka; Uesugi, Naomasa; Saito, Satoshi; Takahashi, Tsuyoshi; Gohra, Hidenori

2016-12-01

Right atrial tumor thrombus is rare in patients with visceral malignant tumors and can cause right heart failure or sudden death. We present 2 cases of right atrial tumor thrombus treated under deep hypothermic intermittent circulatory arrest (DHICA). A 45-year-old man with right heart failure was diagnosed with right renal cancer extending to the right atrium. Computed tomography revealed no metastasis. He underwent right nephrectomy and tumor thrombus resection under DHICA. He was discharged on postoperative day 11 in good clinical course. A 67-year-old woman with hepatitis C virus liver cirrhosis( Child-Pugh A) was diagnosed with hepatocellular carcinoma and right atrial tumor. She underwent S8 and tumor thrombus resection under DHICA. Hemorrhagic diathesis was controlled using fresh frozen plasma transfusion. She was discharged on postoperative day 24 without liver failure. In cases of atrial tumor thrombus resection, DIHCA may be useful to achieve a bloodless operation field because the procedure is relatively simple and the primary disease need not be considered.

Comparison of rapid diagnostic tests to detect Mycobacterium avium subsp. paratuberculosis disseminated infection in bovine liver.

PubMed

Zarei, Mehdi; Ghorbanpour, Masoud; Tajbakhsh, Samaneh; Mosavari, Nader

2017-08-01

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease, a chronic enteritis in cattle and other domestic and wild ruminants. The presence of MAP in tissues other than intestines and associated lymph nodes, such as meat and liver, is a potential public health concern. In the present study, the relationship between the results of rapid diagnostic tests of the Johne's disease, such as serum ELISA, rectal scraping PCR, and acid-fast staining, and the presence of MAP in liver was evaluated. Blood, liver, and rectal scraping samples were collected from 200 slaughtered cattle with unknown Johne's disease status. ELISA was performed to determine the MAP antibody activity in the serum. Acid-fast staining was performed on rectal scraping samples, and PCR was performed on rectal scraping and liver samples. PCR-positive liver samples were used for mycobacterial culture. Overall, the results of this study demonstrated that MAP can be detected and cultured from liver of slaughtered cattle and rapid diagnostic tests of Johne's disease have limited value in detecting cattle with MAP infection in liver. These findings show that the presence of MAP in liver tissue may occur in cows with negative results for rapid diagnostic tests and vice versa. Hence, liver might represent another possible risk of human exposure to MAP. Given concerns about a potential zoonotic role for MAP, these results show the necessity to find new methods for detecting cattle with MAP disseminated infection.

Biomolecular solid state NMR with magic-angle spinning at 25K.

PubMed

Thurber, Kent R; Tycko, Robert

2008-12-01

A magic-angle spinning (MAS) probe has been constructed which allows the sample to be cooled with helium, while the MAS bearing and drive gases are nitrogen. The sample can be cooled to 25K using roughly 3 L/h of liquid helium, while the 4-mm diameter rotor spins at 6.7 kHz with good stability (+/-5 Hz) for many hours. Proton decoupling fields up to at least 130 kHz can be applied. This helium-cooled MAS probe enables a variety of one-dimensional and two-dimensional NMR experiments on biomolecular solids and other materials at low temperatures, with signal-to-noise proportional to 1/T. We show examples of low-temperature (13)C NMR data for two biomolecular samples, namely the peptide Abeta(14-23) in the form of amyloid fibrils and the protein HP35 in frozen glycerol/water solution. Issues related to temperature calibration, spin-lattice relaxation at low temperatures, paramagnetic doping of frozen solutions, and (13)C MAS NMR linewidths are discussed.

Growth Hormone (GH) Hypersecretion and GH Receptor Resistance in Streptozotocin Diabetic Mice in Response to a GH Secretagogue

PubMed Central

Segev, Yael; Landau, Daniel; Phillip, Moshe; Flyvbjerg, Allan

2003-01-01

The growth hormone (GH) and insulin-like growth factor I (IGF-I) axis were studied in streptozotocin (STZ) diabetic and nondiabetic female mice following intravenous (IV) injection of the GH secretagogue (GHS) ipamorelin or saline. On day 14, blood samples were obtained before and 10 minutes after the injection. Livers were removed and frozen for determination of the mRNA expressions of the GH receptor, GH-binding protein, and IGF-I, and hepatic IGF-I peptide. Serum samples were analyzed for GH and IGF-I. Following ipamorelin injection, the GH levels were found to be 150 ± 35 μg/L and 62 ± 11 μg/L in the diabetic compared to the nondiabetic mice (P < .05). Serum IGF-I levels were lower in diabetic than in nondiabetic animals, and rose after stimulation only in the nondiabetic animals. Furthermore, hepatic GH resistance and IGF-I mRNA levels and IGF-I peptide were increased in nondiabetic animals in response to GH stimulation, whereas the low levels per se of all these parameters in diabetic mice were unaffected. The study shows that STZ diabetic mice demonstrate a substantial part of the clinical features of type 1 diabetes in humans, including GH hypersecretion and GH resistance. Accordingly, it is proposed that STZ diabetic mice may be a better model of the perturbations of the GH/IGF-I axis in diabetes than STZ diabetic rats. PMID:14630569

Frequency of the HFE C282Y and H63D mutations in Danish patients with clinical haemochromatosis initially diagnosed by phenotypic methods.

PubMed

Milman, Nils; Koefoed, Pernille; Pedersen, Palle; Nielsen, Finn Cilius; Eiberg, Hans

2003-12-01

To assess the frequency of the C282Y and H63D mutations on the HFE gene in Danish patients with clinical hereditary haemochromatosis initially diagnosed by phenotypic methods. In the period 1950-1985, an epidemiological survey in Denmark identified 179 patients with clinical idiopathic haemochromatosis diagnosed by phenotypic methods (serum transferrin saturation, serum ferritin, liver biopsy and mobilisable body iron stores). In 32 unrelated patients, frozen blood samples were available for genetic analysis. In a subsequent series of 26 unrelated Danish patients, a phenotypic diagnosis of clinical idiopathic haemochromatosis was made before blood samples were taken for HFE genotyping. The total series consisted of 58 patients (40 men and 18 women) with a median age of 60 yrs (range 18-74). HFE genotyping was performed by the polymerase chain reaction (PCR) technique. Among the patients, 55 of 58 (94.8%) were C282Y/C282Y homozygous. One 63-year-old woman (1.7%) was compound C282Y/H63D heterozygous. Two women (3.4%), aged 42 and 43 yrs were negative for both the C282Y and the H63D mutation. In the Danish population, homozygosity for the C282Y mutation appears to be the prevailing cause of clinically overt genetic haemochromatosis. This finding has implications both for the evaluation of patients with iron overload disorders and for the strategy in future population screening surveys.

Potency and stability of frozen urokinase solutions in syringes.

PubMed

Dedrick, Stephen C; Ramirez-Rico, José

2004-08-01

The stability and potency of frozen urokinase solutions in syringes were studied. To determine the stability and potency of compounded urokinase dilutions after multiple freeze-thaw cycles, a total of 160 syringes containing five urokinase concentrations (2,500, 5,000, 7,500, 12,500, and 25,000 IU/mL) were prepared. For each of the five concentrations tested, two syringes per concentration were reserved for baseline testing. The remaining 150 syringes were frozen at -30 degrees C. After 7 days, half of the syringes (group 1) were thawed at room temperature, tested, and left at room temperature for 12 hours before refreezing. The other half of the syringes (group 2) were kept frozen for 30 days. Thirty days after initial compounding, all syringes were thawed, and the samples' urokinase potency, pH, and physical appearance were evaluated. Syringes were visually inspected for color, clarity, and precipitation. Descriptive statistics were computed for each concentration group and testing day. The compounded dilutions were stable under each experimental condition, with no physical deterioration or loss of in vitro potency after two freeze-thaw cycles. The reduced waste associated with the ability to refreeze unused urokinase could substantially lower the cost of procedures such as thrombolysis after intraventricular hemorrhage and catheter clearance by as much as 95%. Dilutions of urokinase 2,500-25,000 IU/mL were stable in single-use syringes after being frozen for 7 days, thawed, and refrozen for another 23 days.

Efficacy of freezing, frozen storage and edible antimicrobial coatings used in combination for control of Listeria monocytogenes on roasted turkey stored at chiller temperatures.

PubMed

Jiang, Zheng; Neetoo, Hudaa; Chen, Haiqiang

2011-10-01

The presence and growth of Listeria monocytogenes on ready-to-eat (RTE) turkey is an important food safety issue. The antilisterial efficacy of four polysaccharide-based edible coatings (starch, chitosan, alginate and pectin) incorporating sodium lactate (SL) and sodium diacetate (SD) as well as commercial preparations Opti.Form PD4, NovaGARD™ CB1, Protect-M and Guardian™ NR100 were compared against L. monocytogenes on roasted turkey. Pectin coating treatments incorporating SL/SD, Opti.Form PD4 with or without Protect-M, and NovaGARD™ CB1 displayed higher antimicrobial efficacy against. L. monocytogenes than the other antimicrobials and coating materials. In the second phase of the study, it was investigated whether frozen storage could enhance the antilisterial effectiveness of pectin coating treatments on chilled roasted turkey. Inoculated roasted turkey samples coated with pectin-based treatments were frozen for up to 4 weeks and subsequently stored at 4 °C for 8 weeks. Frozen storage significantly enhanced the antilisterial activity of various coating treatments; with selected treatments reducing the L. monocytogenes populations by as much as 1.1 log CFU/cm(2) during the subsequent 8-week chilled storage. This study demonstrates that pectin-based antimicrobial edible coatings hold promise in enhancing the safety of RTE poultry products and frozen storage has the potential to enhance their effectiveness. Copyright © 2011 Elsevier Ltd. All rights reserved.

Rheological properties of ice cream mixes and frozen ice creams containing fat and fat replacers.

PubMed

Adapa, S; Dingeldein, H; Schmidt, K A; Herald, T J

2000-10-01

Ice cream mixes and frozen ice creams at milk fat levels of 12%, 8%, 6%, 6% plus a protein-based fat replacer, and 6% plus a carbohydrate-based fat replacer were evaluated for viscoelastic properties by dynamic testing with sinusoidal oscillatory tests at various frequencies. The storage modulus (G'), loss modulus (G"), and tan delta (G"/G') were calculated for all the treatments to determine changes in the viscous and elastic properties of the mixes and frozen ice creams due to fat content. In ice cream mixes, G' and G" exhibited a strong frequency dependence. The G" was higher than G' throughout the frequency range (1 to 8 Hz) examined, without any crossover, except for the 12% mix. Elastic properties of the ice cream mixes decreased as fat content decreased. Tan delta values indicated that fat replacers did not enhance the elastic properties of the ice cream mixes. In all frozen ice creams, G' and G" again showed a frequency dependence throughout the range tested (0.5 to 10 Hz). The amount of fat in ice creams and the degree of fat destabilization affected the elasticity in the frozen product. Even though the ice creams did not have significant elastic properties, when compared as a group the samples with higher fat content had higher elastic properties. The addition of protein-based and carbohydrate-based fat replacers did not enhance the elastic properties of the ice creams but did increase the viscous properties.

Metals and trace elements in giant garter snakes (Thamnophis gigas) from the Sacramento Valley, California, USA

USGS Publications Warehouse

Wylie, G.D.; Hothem, R.L.; Bergen, D.R.; Martin, L.L.; Taylor, R.J.; Brussee, B.E.

2009-01-01

The giant garter snake (GGS; Thamnophis gigas) is a federally listed threatened species endemic to wetlands of the Central Valley of California. Habitat destruction has been the main factor in the decline of GGS populations, but the effects of contaminants on this species are unknown. To contribute to the recovery of these snakes, the U.S. Geological Survey (USGS) began studies of the life history and habitat use of GGSs in 1995. During a series of investigations conducted from 1995 to the present, specimens of dead GGSs were opportunistically collected from the Colusa National Wildlife Refuge (CNWR), the Natomas Basin, and other sites in northern California. Whole snakes were stored frozen for potential future analysis. As funding became available, we analyzed tissues of 23 GGSs to determine the concentrations of total mercury (Hg) and other trace elements in livers and concentrations of Hg in brains and tail clips. Mercury concentrations (??g/g, wet weight) ranged from 0.08 to 1.64 in livers, 0.01 to 0.18 in brains, and 0.02 to 0.32 in tail clips. In livers, geometric mean concentrations (??g/g, dry weight) of arsenic (25.7) and chromium (1.02) were higher than most values from studies of other snakes. Mercury concentrations in tail clips were positively correlated with concentrations in livers and brains, with the most significant correlations occurring at the Natomas Basin and when Natomas and CNWR were combined. Results indicate the value of using tail clips as a nonlethal bioindicator of contaminant concentrations. ?? 2008 US Government.

Effects of adding different levels of Glutamine to modified Beltsville extender on the survival of frozen rooster semen.

PubMed

Khiabani, Aytak Bakhshayesh; Moghaddam, Gholamali; Kia, Hossein Daghigh

2017-09-01

The aim of the present study was to investigate the effects of l-glutamine on the quality of frozen-thawed rooster semen. Semen samples were collected from eight mature roosters (Ross 308). After initial semen assessments, samples of adequate quality were mixed together and diluted with modified Beltsville extender without l-glutamine (control) and supplemented with 2.5, 5, and 7.5mM l-glutamine. Semen straws were subjected to cryopreservation and evaluated twice at 15-day intervals. After thawing, sperm viability, total and progressive sperm motilities were measured by Eosin-Nigrosine and Computer-Aided Sperm Analysis (CASA), respectively. The results showed that sperm functions decreased on day 30 compared to day 15. The extender supplemented with 5mM glutamine improved (p<0.05) sperm viability, total and progressive sperm motilities compared to other treatments and the control group. The best level of glutamine appeared to be 2.5mM, as it provided the highest sperm membrane integrity and the lowest level of abnormalities. The results of this study suggest that the addition of glutamine to the diluent improves semen quality and using glutamine allows rooster sperm to be frozen for longer. Copyright © 2017 Elsevier B.V. All rights reserved.

Cryogenic preservation of semen from the Aleutian Canada goose (Branta canadensis leucopareia)

USGS Publications Warehouse

Gee, G.F.; Sexton, T.J.

1990-01-01

Aleutian Canada geese (Branta canadensis leucopareia) were inseminated with frozen-thawed semen containing 6% or 7% dimethylsulfoxide (DMSO) resulting in 32 fertile eggs and 17 goslings; with 7% DMSO, 19 of 31 eggs were fertile. Beltsville Poultry Semen Extender (BPSE), adjusted to 270 ? 30 mOs and 7.5 ? 0.4 pH, was used to dilute semen samples and the DMSO before cryopreservation. About half of the live spermatozoa in the fresh semen (92.9 ? 2.5% live cells, laboratory studies; 87.3 ? 7.3%, insemination trials) survived the freeze-thaw process (46.7 ? 7.8%, laboratory; 33.3 ? 17.8%, insemination trials). Samples of frozen-thawed semen contained a greater percentage of bent spermatozoa (27.1 ? 8.4% of live cells) than fresh semen (14.4 ? 3.0% of live cells). Fecal- and urate-contaminated semen (a common problem when collecting goose semen) reduced the sperm motility score from 3.2 ? 0.6 to 2.7? 0.7 and number of live spermatozoa in frozen-thawed semen from 49 ? 9% to 24 ?18%. Other variables examined that had less of an effect on semen quality included semen extenders, semen holding temperature, dilution and equilibration, relationship between hour of semen collection and level of semen contamination, and the relationship between season and sperm concentration.

Protein crowding in solution, frozen and freeze-dried states: small-angle neutron and X-ray scattering study of lysozyme/sorbitol/water systems

NASA Astrophysics Data System (ADS)

Krueger, Susan; Khodadadi, Sheila; Clark, Nicholas; McAuley, Arnold; Cristiglio, Viviana; Theyencheri, Narayanan; Curtis, Joseph; Shalaev, Evgenyi

2015-03-01

For effective preservation, proteins are often stored as frozen solutions or in glassy states using a freeze-drying process. However, aggregation is often observed after freeze-thaw or reconstitution of freeze-dried powder and the stability of the protein is no longer assured. In this study, small-angle neutron and X-ray scattering (SANS and SAXS) have been used to investigate changes in protein-protein interaction distances of a model protein/cryoprotectant system of lysozyme/sorbitol/water, under representative pharmaceutical processing conditions. The results demonstrate the utility of SAXS and SANS methods to monitor protein crowding at different stages of freezing and drying. The SANS measurements of solution samples showed at least one protein interaction peak corresponding to an interaction distance of ~ 90 Å. In the frozen state, two protein interaction peaks were observed by SANS with corresponding interaction distances at 40 Å as well as 90 Å. On the other hand, both SAXS and SANS data for freeze-dried samples showed three peaks, suggesting interaction distances ranging from ~ 15 Å to 170 Å. Possible interpretations of these interaction peaks will be discussed, as well as the role of sorbitol as a cryoprotectant during the freezing and drying process.

Methods to decrease blood loss during liver resection: a network meta-analysis.

PubMed

Moggia, Elisabetta; Rouse, Benjamin; Simillis, Constantinos; Li, Tianjing; Vaughan, Jessica; Davidson, Brian R; Gurusamy, Kurinchi Selvan

2016-10-31

Liver resection is a major surgery with significant mortality and morbidity. Specialists have tested various methods in attempts to limit blood loss, transfusion requirements, and morbidity during elective liver resection. These methods include different approaches (anterior versus conventional approach), use of autologous blood donation, cardiopulmonary interventions such as hypoventilation, low central venous pressure, different methods of parenchymal transection, different methods of management of the raw surface of the liver, different methods of vascular occlusion, and different pharmacological interventions. A surgeon typically uses only one of the methods from each of these seven categories. The optimal method to decrease blood loss and transfusion requirements in people undergoing liver resection is unknown. To assess the effects of different interventions for decreasing blood loss and blood transfusion requirements during elective liver resection. We searched the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, Embase, and Science Citation Index Expanded to September 2015 to identify randomised clinical trials. We also searched trial registers and handsearched the references lists of identified trials. We included only randomised clinical trials (irrespective of language, blinding, or publication status) comparing different methods of decreasing blood loss and blood transfusion requirements in people undergoing liver resection. Two review authors independently identified trials and collected data. We assessed the risk of bias using Cochrane domains. We conducted a Bayesian network meta-analysis using the Markov chain Monte Carlo method in WinBUGS 1.4, following the guidelines of the National Institute for Health and Care Excellence Decision Support Unit guidance documents. We calculated the odds ratios (OR) with 95% credible intervals (CrI) for the binary outcomes, mean differences (MD) with 95% CrI for continuous outcomes, and rate ratios with 95% CrI for count outcomes, using a fixed-effect model or random-effects model according to model-fit. We assessed the evidence with GRADE. We identified 67 randomised clinical trials involving a total of 6197 participants. All the trials were at high risk of bias. A total of 5771 participants from 64 trials provided data for one or more outcomes included in this review. There was no evidence of differences in most of the comparisons, and where there was, these differences were in single trials, mostly of small sample size. We summarise only the evidence that was available in more than one trial below. Of the primary outcomes, the only one with evidence of a difference from more than one trial under the pair-wise comparison was in the number of adverse events (complications), which was higher with radiofrequency dissecting sealer than with the clamp-crush method (rate ratio 1.85, 95% CrI 1.07 to 3.26; 250 participants; 3 studies; very low-quality evidence). Among the secondary outcomes, the only differences we found from more than one trial under the pair-wise comparison were the following: blood transfusion (proportion) was higher in the low central venous pressure group than in the acute normovolemic haemodilution plus low central venous pressure group (OR 3.19, 95% CrI 1.56 to 6.95; 208 participants; 2 studies; low-quality evidence); blood transfusion quantity (red blood cells) was lower in the fibrin sealant group than in the control (MD -0.53 units, 95% CrI -1.00 to -0.07; 122 participants; 2; very low-quality evidence); blood transfusion quantity (fresh frozen plasma) was higher in the oxidised cellulose group than in the fibrin sealant group (MD 0.53 units, 95% CrI 0.36 to 0.71; 80 participants; 2 studies; very low-quality evidence); blood loss (MD -0.34 L, 95% CrI -0.46 to -0.22; 237 participants; 4 studies; very low-quality evidence), total hospital stay (MD -2.42 days, 95% CrI -3.91 to -0.94; 197 participants; 3 studies; very low-quality evidence), and operating time (MD -15.32 minutes, 95% CrI -29.03 to -1.69; 192 participants; 4 studies; very low-quality evidence) were lower with low central venous pressure than with control. For the other comparisons, the evidence for difference was either based on single small trials or there was no evidence of differences. None of the trials reported health-related quality of life or time needed to return to work. Paucity of data meant that we could not assess transitivity assumptions and inconsistency for most analyses. When direct and indirect comparisons were available, network meta-analysis provided additional effect estimates for comparisons where there were no direct comparisons. However, the paucity of data decreases the confidence in the results of the network meta-analysis. Low-quality evidence suggests that liver resection using a radiofrequency dissecting sealer may be associated with more adverse events than with the clamp-crush method. Low-quality evidence also suggests that the proportion of people requiring a blood transfusion is higher with low central venous pressure than with acute normovolemic haemodilution plus low central venous pressure; very low-quality evidence suggests that blood transfusion quantity (red blood cells) was lower with fibrin sealant than control; blood transfusion quantity (fresh frozen plasma) was higher with oxidised cellulose than with fibrin sealant; and blood loss, total hospital stay, and operating time were lower with low central venous pressure than with control. There is no evidence to suggest that using special equipment for liver resection is of any benefit in decreasing the mortality, morbidity, or blood transfusion requirements (very low-quality evidence). Radiofrequency dissecting sealer should not be used outside the clinical trial setting since there is low-quality evidence for increased harm without any evidence of benefits. In addition, it should be noted that the sample size was small and the credible intervals were wide, and we cannot rule out considerable benefit or harm with a specific method of liver resection.

Distribution of cadmium in leg muscle and liver of game birds from Serbia

NASA Astrophysics Data System (ADS)

Nikolić, D.; Đinović-Stojanović, J.; Stefanović, S.; Radičević, T.; Trbović, D.; Spirić, D.; Janković, S.

2017-09-01

The aim of this study was to present the distribution of cadmium (Cd) levels in leg muscle and liver of game birds. Samples (n=464) of: pheasants (n=182), mallards (n=25), Eurasian jay (n=7), partridges (n=5), woodcocks (n=8) and common quail (n=5) were collected during regular hunting seasons within the Serbian National Residue Monitoring Program from 2013 to 2016. Analysis of Cd was performed by ICP-MS. In all liver samples, Cd levels were above the limit of detection (LOD=0.001 mg/kg) while in 66.4% of muscle samples, Cd was detected. Statistical analysis showed significant differences between Cd levels in leg muscle and liver of woodcocks and others game birds. The highest mean Cd level was observed in muscle samples of woodcocks (0.042 mg/kg). The lowest mean Cd levels in liver were observed in common quails (0.130 mg/kg) and mallards (0.160 mg/kg) while the highest levels were measured in woodcocks (1.247 mg/kg) and pheasants (0.262 mg/kg). During four years of the Serbian National Residue Monitoring Program, leg muscle samples of woodcocks (n=3), liver samples of pheasants (n=23), woodcocks (n=6) and mallards (n=3) exceeded the maximum residue limit (MRL).

Determination of 17alpha-ethynylestradiol, carbamazepine, diazepam, simvastatin, and oxybenzone in fish livers.

PubMed

Kwon, Jeong-Wook; Armbrust, Kevin L; Vidal-Dorsch, Doris; Bay, Steven M

2009-01-01

A method using liquid chromatography/tandem mass spectrometry (LC/MS/MS) was developed for the determination of 17alpha-ethynylestradiol in fish liver; a second method using LC/MS was developed for the determination of carbamazepine, diazepam, simvastatin, and oxybenzone in fish liver. The fish liver samples were extracted and cleaned up by using liquid-liquid extraction and solid-phase extraction before the extracts were analyzed by LC/MS or LC/MS/MS with electrospray negative and positive ionization. Recoveries of the 5 target compounds from spiked catfish liver ranged from 72 +/- 2 to 100 +/- 3%. Limits of quantification for the 5 compounds were between 4.2 and 12.3 ng/g (wet weight). Ten turbot (Pleuronichthys verticalis) liver samples were analyzed; levels of 17alpha-ethynylestradiol, carbamazepine, simvastatin, and oxybenzone were below the detection limits. Diazepam was detected in all 10 fish liver samples at concentrations ranging from 23 to 110 ng/g (wet weight).

Comparison and optimization of detection methods for noroviruses in frozen strawberries containing different amounts of RT-PCR inhibitors.

PubMed

Bartsch, Christina; Szabo, Kathrin; Dinh-Thanh, Mai; Schrader, Christina; Trojnar, Eva; Johne, Reimar

2016-12-01

Frozen berries have been repeatedly identified as vehicles for norovirus (NoV) transmission causing large gastroenteritis outbreaks. However, virus detection in berries is often hampered by the presence of RT-PCR-inhibiting substances. Here, several virus extraction methods for subsequent real-time RT-PCR-based NoV-RNA detection in strawberries were compared and optimized. NoV recovery rates (RRs) between 0.21 ± 0.13% and 10.29 ± 6.03% were found when five different artificially contaminated strawberry batches were analyzed by the ISO/TS15216-2 method indicating the presence of different amounts of RT-PCR inhibitors. A comparison of five different virus extraction methods using artificially contaminated strawberries containing high amounts of RT-PCR inhibitors revealed the best NoV RRs for the ISO/TS15216 method. Further improvement of NoV RRs from 2.83 ± 2.92% to 15.28 ± 9.73% was achieved by the additional use of Sephacryl(®)-based columns for RNA purification. Testing of 22 frozen strawberry samples from a batch involved in a gastroenteritis outbreak resulted in 5 vs. 13 NoV GI-positive and in 9 vs. 20 NoV GII-positive samples using the original ISO/TS15216 method vs. the extended protocol, respectively. It can be concluded that the inclusion of an additional RNA purification step can increase NoV detection by the ISO/TS15216-2 method in frozen berries containing high amounts of RT-PCR inhibitors. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Management of risk of microbial cross-contamination from uncooked frozen hamburgers by alcohol-based hand sanitizer.

PubMed

Schaffner, Donald W; Schaffner, Kristin M

2007-01-01

This research was undertaken to determine the effectiveness of an alcohol-based hand sanitizer on hands contaminated with a nonpathogen surrogate for Escherichia coli O157:H7, where the source of the contamination was frozen hamburger patties. A nonpathogenic nalidixic acid-resistant food-grade strain of Enterobacter aerogenes was used to inoculate frozen hamburger patties composed of 76% lean beef and 24% fat. Thirty-two individuals participated to produce the data used in this study. Each participant handled nine patties at least three times, a sample for microbiological analysis was collected from the surface of one hand, the participant sanitized both hands, and a sample was collected from the other hand. Burger handling created perceptible and visible food debris on the hands of most participants. Computer simulations also were used to perform a variety of risk calculations. The average reduction in bacteria from the use of sanitizer on hands contaminated by frozen burgers containing E. aerogenes was 2.6 +/- 0.7 log CFU per hand. An experiment designed to simultaneously test the effect of sanitizer on E. aerogenes and E. coli O157:H7 also revealed no significant difference in sanitizer effectiveness against the two organisms. The results of the real-world risk estimation calculations (using the actual prevalence and concentration of E. coli O157:H7 in ground beef) predict that once in 1 million trials, a single pathogen cell will be transferred to a single lettuce piece. The effectiveness of this sanitizer intervention was similar to that for hand washing and glove use previously reported. The person-to-person microbial reduction variability from sanitizer use is similar to published data for glove use and was less variable than published data on hand washing effectiveness.

[Study of the reineta protein modifications (Brama australis), put under freezing and storage to -18 degrees C and -30 degrees C].

PubMed

Abugoch, Lilian; Quitral, Vilma; Larraín, M Angélica; Vinagre, Julia; Kriukov, Andrei; Chávez, Gloria

2006-12-01

The objective of the present work was to study functional and thermal properties of reineta (Brama australis) frozen meat, analysed by water retention capacity (WRC), gel forming capacity (GFC), texture, emulsifying capacity and differential scanning calorimetry (DSC). For this study, reineta fillets were obtained and extracted by the same conditions, and cutted, packaged, frozen and stored at -18 degrees C and -30 degrees C for 7 months. The results obtained, showed that there were no signifficant differences in the responses to thermal treatment for all the specimens. For samples frozen at -18 degrees C and -30 degrees C, the protein contents were 23.5 + 0.0 and 25.4 + 1.0%, respectively. The WRC values were 0.45 + 0.1 and 1.59 +/- 0.0 g water/g protein, respectively. The gel forming capacity was only present in the fresh samples, whereas the frozen stored ones only form protein aggregates. The emulsifying capacity was between 960 and 1400 g oil / g protein, and the storage time increased this value. The miosin denaturation temperature (Td) and denaturation enthalpy (?H), obtained by DSC, fluctuated between 39.2 +/- 0.5 to 44.8 +/- 0.8 degrees C and 1.12 +/- 0.3 to 0.52 +/- 0.2 J/g, respectively. The actina values were between 71.0 +/- 0.6 to 75.3 +/- 0.5 degrees C and between 0.5 +/- 0.1 to 0.7 +/- 0.1 J/g. Cooperativity decreased as the storage time increased. This is showing a certain degree of protein displacement. The values found by thermal analyses showed a direct relationship with the functional properties, both decreasing with storage time.

Histological staining methods preparatory to laser capture microdissection significantly affect the integrity of the cellular RNA.

PubMed

Wang, Hongyang; Owens, James D; Shih, Joanna H; Li, Ming-Chung; Bonner, Robert F; Mushinski, J Frederic

2006-04-27

Gene expression profiling by microarray analysis of cells enriched by laser capture microdissection (LCM) faces several technical challenges. Frozen sections yield higher quality RNA than paraffin-imbedded sections, but even with frozen sections, the staining methods used for histological identification of cells of interest could still damage the mRNA in the cells. To study the contribution of staining methods to degradation of results from gene expression profiling of LCM samples, we subjected pellets of the mouse plasma cell tumor cell line TEPC 1165 to direct RNA extraction and to parallel frozen sectioning for LCM and subsequent RNA extraction. We used microarray hybridization analysis to compare gene expression profiles of RNA from cell pellets with gene expression profiles of RNA from frozen sections that had been stained with hematoxylin and eosin (H&E), Nissl Stain (NS), and for immunofluorescence (IF) as well as with the plasma cell-revealing methyl green pyronin (MGP) stain. All RNAs were amplified with two rounds of T7-based in vitro transcription and analyzed by two-color expression analysis on 10-K cDNA microarrays. The MGP-stained samples showed the least introduction of mRNA loss, followed by H&E and immunofluorescence. Nissl staining was significantly more detrimental to gene expression profiles, presumably owing to an aqueous step in which RNA may have been damaged by endogenous or exogenous RNAases. RNA damage can occur during the staining steps preparatory to laser capture microdissection, with the consequence of loss of representation of certain genes in microarray hybridization analysis. Inclusion of RNAase inhibitor in aqueous staining solutions appears to be important in protecting RNA from loss of gene transcripts.

Histological staining methods preparatory to laser capture microdissection significantly affect the integrity of the cellular RNA

PubMed Central

Wang, Hongyang; Owens, James D; Shih, Joanna H; Li, Ming-Chung; Bonner, Robert F; Mushinski, J Frederic

2006-01-01

Background Gene expression profiling by microarray analysis of cells enriched by laser capture microdissection (LCM) faces several technical challenges. Frozen sections yield higher quality RNA than paraffin-imbedded sections, but even with frozen sections, the staining methods used for histological identification of cells of interest could still damage the mRNA in the cells. To study the contribution of staining methods to degradation of results from gene expression profiling of LCM samples, we subjected pellets of the mouse plasma cell tumor cell line TEPC 1165 to direct RNA extraction and to parallel frozen sectioning for LCM and subsequent RNA extraction. We used microarray hybridization analysis to compare gene expression profiles of RNA from cell pellets with gene expression profiles of RNA from frozen sections that had been stained with hematoxylin and eosin (H&E), Nissl Stain (NS), and for immunofluorescence (IF) as well as with the plasma cell-revealing methyl green pyronin (MGP) stain. All RNAs were amplified with two rounds of T7-based in vitro transcription and analyzed by two-color expression analysis on 10-K cDNA microarrays. Results The MGP-stained samples showed the least introduction of mRNA loss, followed by H&E and immunofluorescence. Nissl staining was significantly more detrimental to gene expression profiles, presumably owing to an aqueous step in which RNA may have been damaged by endogenous or exogenous RNAases. Conclusion RNA damage can occur during the staining steps preparatory to laser capture microdissection, with the consequence of loss of representation of certain genes in microarray hybridization analysis. Inclusion of RNAase inhibitor in aqueous staining solutions appears to be important in protecting RNA from loss of gene transcripts. PMID:16643667

Gerst and Swanson perform blood draw in Columbus module

NASA Image and Video Library

2014-06-04

Astronaut Alexander Gerst,Expedition 40 flight engineer (background),and Expedition 40 Commander Steve Swanson are photographed performing blood sample collection in the Columbus module as part of HRF Generic Frozen Blood Collection Operations.

Detection of hepatitis C virus RNA using ligation-dependent polymerase chain reaction in formalin-fixed, paraffin-embedded liver tissues.

PubMed Central

Park, Y. N.; Abe, K.; Li, H.; Hsuih, T.; Thung, S. N.; Zhang, D. Y.

1996-01-01

Reverse transcription polymerase chain reaction (RT-PCR) has been used to detect hepatitis C virus (HCV) sequences in liver tissue. However, RT-PCR has a variable detection sensitivity, especially on routinely processed formalin-fixed, paraffin-embedded (FFPE) specimens. RNA-RNA and RNA-protein cross-links formed during formalin fixation is the major limiting factor preventing reverse trans criptase from extending the primers. To overcome this problem, we applied the ligation-dependent PCR (LD-PCR) for the detection of HCV RNA in FFPE liver tissue. This method uses two capture probes for RNA isolation and two hemiprobes for the subsequent PCR. Despite cross-links, the capture probes and the hemiprobes are able to form hybrids with HCV RNAs released from the FFPE tissue. The hybrids are isolated through binding of the capture probes to paramagnetic beads. The hemiprobes are then ligated by a T4 DNA ligase to form a full probe that serves as a template for the Taq DNA polymerase. A total of 22 FFPE liver specimens, 21 with hepatocellular carcinoma (HCC) and 1 with biliary cirrhosis secondary to bile duct atresia were selected for this study, of which 13 patients were HCV seropositive and 9 seronegative. HCV RNA was detectable by ID-PCR from all 13 HCV-seropositive HCCs and from 5 of 8 HCV-seronegative HCCs but not from the HCV-seronegative liver with biliary atresia. By contrast, RT-PCR detected HCV sequences in only 5 of the HCV-sero-positive and in 1 of the HCV-seronegative HCCs. To resolve the discordance between the LD-PCR and RT-PCR results, RT-PCR was performed on frozen liver tissue of the discrepant specimens, which confirmed the LD-PCR positive results. In conclusion, LD-PCR is a more sensitive method than RT-PCR for the detection of HCV sequences in routinely processed liver tissues. A high rate of HCV infection (86%) is found in HCC specimens, indicating a previously underestimated role of HCV in HCC pathogenesis. Images Figure 2 PMID:8909238

Factors associated with blood transfusion in donor hepatectomy: results from 2344 donors at a large single center.

PubMed

Choi, Seong-Soo; Cho, Seong-Sik; Kim, Sung-Hoon; Jun, In-Gu; Hwang, Gyu-Sam; Kim, Young-Kug

2013-12-15

The safety of healthy living donors undergoing hepatic resection for living-donor liver transplantation is of paramount concern. Although blood transfusions have been associated with morbidity and mortality after hepatectomy, there is limited information about the risk factors associated with blood transfusion in living liver donors. We retrospectively analyzed 2344 donors who underwent a hepatectomy for living-donor liver transplantation. Logistic regression analysis was performed to determine blood transfusion predictors in living-donor hepatectomy. Of these donors, 48 (2.0%) and 97 (4.1%) were transfused with packed red blood cell (PRBC) and fresh-frozen plasma (FFP), respectively. The amount of PRBC and FFP administered to donors transfused with blood products were 1.9±0.8 and 3.7±2.5 units, respectively. In multivariate logistic regression analysis, a low preoperative hemoglobin level was found to be an independent predictor of PRBC transfusion in donor hepatectomy (odds ratio=0.585; 95% confidence interval=0.451-0.758; P<0.001). A high graft-to-donor weight ratio predicted an FFP transfusion in donor hepatectomy (odds ratio=2.997; 95% confidence interval=1.226-7.327; P=0.016). These results indicate that, in donor hepatectomy, the preoperative hemoglobin value and graft-to-donor weight ratio can provide useful information on the probability of PRBC and FFP transfusion, respectively.

Leptospira spp. infection in sheep herds in southeast Brazil

PubMed Central

2014-01-01

Background With the aim of studying Leptospira spp. infection in sheep herds, blood samples and respective kidney and liver fragments were collected from 100 animals from twenty different properties during slaughter at a meat company in the Sorocaba region, São Paulo state, southeast Brazil. The microscopic agglutination test (MAT) was performed with 29 strains of Leptospira spp. To identify the agent in the liver and kidney, 100 samples of each tissue were submitted to culture in Fletcher medium and analyzed by the polymerase chain reaction (PCR) for Leptospira spp. Results MAT detected 23 samples serologically positive for one or more Leptospira spp. serovars and significantly more for Autumnalis. Eight (4%) samples were positive in culture (four kidneys and four livers), corresponding to five animals with positive serology (one animal simultaneously positive for both kidney and liver) and two negatives. PCR detected Leptospira spp. in 14 samples (seven kidneys and seven livers) corresponding to 12 positive animals (two animals simultaneously positive for kidney and liver), of which ten were serologically positive and two negative. Conclusions PCR was faster, more practical and more sensitive than culture for detecting leptospires. The results reinforce the importance of sheep in the epidemiological context of leptospirosis. PMID:24822059

Investigating the influence of standard staining procedures on the copper distribution and concentration in Wilson's disease liver samples by laser ablation-inductively coupled plasma-mass spectrometry.

PubMed

Hachmöller, Oliver; Aichler, Michaela; Schwamborn, Kristina; Lutz, Lisa; Werner, Martin; Sperling, Michael; Walch, Axel; Karst, Uwe

2017-12-01

The influence of rhodanine and haematoxylin and eosin (HE) staining on the copper distribution and concentration in liver needle biopsy samples originating from patients with Wilson's disease (WD), a rare autosomal recessive inherited disorder of the copper metabolism, is investigated. In contemporary diagnostic of WD, rhodanine staining is used for histopathology, since rhodanine and copper are forming a red to orange-red complex, which can be recognized in the liver tissue using a microscope. In this paper, a laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) method is applied for the analysis of eight different WD liver samples. Apart from a spatially resolved elemental detection as qualitative information, this LA-ICP-MS method offers also quantitative information by external calibration with matrix-matched gelatine standards. The sample set of this work included an unstained and a rhodanine stained section of each WD liver sample. While unstained sections of WD liver samples showed very distinct structures of the copper distribution with high copper concentrations, rhodanine stained sections revealed a blurred copper distribution with significant decreased concentrations in a range from 20 to more than 90%. This implies a copper removal from the liver tissue by complexation during the rhodanine staining. In contrast to this, a further HE stained sample of one WD liver sample did not show a significant decrease in the copper concentration and influence on the copper distribution in comparison to the unstained section. Therefore, HE staining can be combined with the analysis by means of LA-ICP-MS in two successive steps from one thin section of a biopsy specimen. This allows further information to be gained on the elemental distribution by LA-ICP-MS additional to results obtained by histological staining. Copyright © 2017 Elsevier GmbH. All rights reserved.

Distribution of (/sup 14/C)acrylamide in male and pregnant Swiss-Webster mice studied by whole-body autoradiography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marlowe, C.; Clark, M.J.; Mast, R.W.

1986-12-01

Male and 13.5- and 17.5-day pregnant Swiss-Webster mice were administered 120 mg/kg (2,3-14C)acrylamide orally. The male mice were frozen 0.33, 1, 3, 9, 24, 72, and 216 hr later, and the pregnant mice at each gestational period were frozen at 3 and 24 hr. Whole-body autoradiographs from the male mice at early time intervals revealed accumulation of radioactivity in the contents of the gastrointestinal tract, liver, pancreas, testis, brain and gallbladder, and epithelia of oral cavity, esophagus, and bronchi. The distribution appears to be similar in the male and pregnant mice. Absorption from the stomach was virtually complete by 3more » hr; renal and hepatic elimination was essentially complete at 24 hr. Radioactivity in the male reproductive tract appeared in the parenchyma of the testis at 1 hr, moved to the seminiferous tubules and head of the epididymis at 9 hr, and by 9 days remained only in the tail of the epididymis and the crypts of the epithelium of the glans penis. This movement parallels that of spermatids. The 13.5-day fetuses were uniformly labeled except for a slightly increased uptake in fetal brain. The distribution of radioactivity in the 17.5-day fetal tissues resembled that in maternal tissues; the remarkable exception was an intense accumulation in fetal skin. This study indicates that acrylamide is efficiently absorbed from the stomach and eliminated by the liver, kidney, and possibly the pancreas. A previously unrecognized affinity of acrylamide or a metabolic product was demonstrated for fetal skin in late gestation and for adult epithelia of oral cavity, esophagus, forestomach, and bronchi. Also, acrylamide or a metabolite appears to bind to spermatids at a specific stage near maturation.« less

Acetate supplementation attenuates lipopolysaccharide-induced neuroinflammation.

PubMed

Reisenauer, Chris J; Bhatt, Dhaval P; Mitteness, Dane J; Slanczka, Evan R; Gienger, Heidi M; Watt, John A; Rosenberger, Thad A

2011-04-01

Glyceryl triacetate (GTA), a compound effective at increasing circulating and tissue levels of acetate was used to treat rats subjected to a continual 28 day intra-ventricular infusion of bacterial lipopolysaccharide (LPS). This model produces a neuroinflammatory injury characterized by global neuroglial activation and a decrease in choline acetyltransferase immunoreactivity in the basal forebrain. During the LPS infusion, rats were given a daily treatment of either water or GTA at a dose of 6 g/kg by oral gavage. In parallel experiments, free-CoA and acetyl-CoA levels were measured in microwave fixed brains and flash frozen heart, liver, kidney and muscle following a single oral dose of GTA. We found that a single oral dose of GTA significantly increased plasma acetate levels by 15 min and remained elevated for up to 4 h. At 30 min the acetyl-CoA levels in microwave-fixed brain and flash frozen heart and liver were increased at least 2.2-fold. The concentrations of brain acetyl-CoA was significantly increased between 30 and 45 min following treatment and remained elevated for up to 4 h. The concentration of free-CoA in brain was significantly decreased compared to controls at 240 min. Immunohistochemical and morphological analysis demonstrated that a daily treatment with GTA significantly reduced the percentage of reactive glial fibrillary acidic protein-positive astrocytes and activated CD11b-positive microglia by 40-50% in rats subjected to LPS-induced neuroinflammation. Further, in rats subjected to neuroinflammation, GTA significantly increased the number of choline acetyltransferase (ChAT)-positive cells by 40% in the basal forebrain compared to untreated controls. These data suggest that acetate supplementation increases intermediary short chain acetyl-CoA metabolism and that treatment is potentially anti-inflammatory and neuroprotective with regards to attenuating neuroglial activation and increasing ChAT immunoreactivity in this model. © 2011 The Authors. Journal of Neurochemistry © 2011 International Society for Neurochemistry.

Acetate supplementation attenuates lipopolysaccharide-induced neuroinflammation

PubMed Central

Reisenauer, Chris J.; Bhatt, Dhaval P.; Mitteness, Dane J.; Slanczka, Evan R.; Gienger, Heidi M.; Watt, John A.; Rosenberger, Thad A.

2011-01-01

Glyceryl triacetate (GTA), a compound effective at increasing circulating and tissue levels of acetate was used to treat rats subjected to a continual 28 day intra-ventricular infusion of bacterial lipopolysaccharide (LPS). This model produces a neuroinflammatory injury characterized by global neuroglial activation and a decrease in choline acetyltransferase immunoreactivity in the basal forebrain. During the LPS infusion, rats were given a daily treatment of either water or GTA at a dose of 6g/kg by oral gavage. In parallel experiments free-CoA and acetyl-CoA levels were measured in microwave fixed brains and flash frozen heart, liver, kidney and muscle following a single oral dose of GTA. We found that a single oral dose of GTA significantly increased plasma acetate levels by 15 min and remained elevated for up to 4 hr. At 30 min the acetyl-CoA levels in microwave-fixed brain and flash frozen heart and liver were increased at least 2.2-fold. The concentrations of brain acetyl-CoA was significantly increased between 30 and 45 min following treatment and remained elevated for up to 4 hr. The concentration of free-CoA in brain was significantly decreased compared to controls at 240 min. Immunohistochemical and morphological analysis demonstrated that a daily treatment with GTA significantly reduced the percentage of reactive GFAP-positive astrocytes and activated CD11b-positive microglia by 40–50% in rats subjected to LPS-induced neuroinflammation. Further, in rats subjected to neuroinflammation, GTA significantly increased the number of ChAT-positive cells by 40% in the basal forebrain compared to untreated controls. These data suggest that acetate supplementation increases intermediary short chain acetyl-CoA metabolism and that treatment is potentially anti-inflammatory and neuroprotective with regards to attenuating neuroglial activation and increasing ChAT immunoreactivity in this model. PMID:21272004

Occurrence and characterization of Salmonella from chicken nuggets, strips, and pelleted broiler feed.

PubMed

Bucher, O; Holley, R A; Ahmed, R; Tabor, H; Nadon, C; Ng, L K; D'Aoust, J Y

2007-10-01

Raw, frozen chicken nuggets and strips have been identified as a significant risk factor in contracting foodborne salmonellosis. Cases of salmonellosis as a result of consuming partly cooked chicken nuggets may be due in part to Salmonella strains originating in broiler feed. This study was undertaken to determine the occurrence and characterize the strains of Salmonella contaminating chicken nuggets, strips, and pelleted feeds, in an attempt to demonstrate whether the same Salmonella strains present in broiler feed could be isolated from raw, frozen chicken nuggets and strips available for human consumption. Salmonellae were recovered using the Health Canada MFHPB-20 method for the isolation and identification of Salmonella from foods. Strains were characterized by serotyping, phage typing, antimicrobial resistance typing (R-typing), and by pulsed-field gel electrophoresis (PFGE). Salmonellae were isolated from 25-g samples in 27% (n=92) of nugget and strip samples, 95% (n=20) of chicken nugget meat samples, and from 9% (n=111) of pelleted feed samples. Salmonella Heidelberg, Salmonella Enteritidis, and Salmonella Orion were the most commonly isolated serovars from chicken nuggets and strips, nugget and strip meat, and pelleted broiler feeds, respectively. Salmonella Enteritidis phage type (PT) 13a with PFGE pattern SENXAI.0006 and R-type sensitive as well as Salmonella Enteritidis PT13a with PFGE pattern SENXAI.0068 and R-type sensitive were isolated from pelleted feed, and chicken nugget and strip meat in two separate instances. Data showed that Salmonella strains isolated from broiler feed were indistinguishable from strains isolated from packaged raw, frozen chicken nuggets and strips. However, results did not rule out the possibility that breeding stock or contamination during processing may have contributed to chicken meat contamination by Salmonella.

Outbreak of hepatitis A in the USA associated with frozen pomegranate arils imported from Turkey: an epidemiological case study.

PubMed

Collier, Melissa G; Khudyakov, Yury E; Selvage, David; Adams-Cameron, Meg; Epson, Erin; Cronquist, Alicia; Jervis, Rachel H; Lamba, Katherine; Kimura, Akiko C; Sowadsky, Rick; Hassan, Rashida; Park, Sarah Y; Garza, Eric; Elliott, Aleisha J; Rotstein, David S; Beal, Jennifer; Kuntz, Thomas; Lance, Susan E; Dreisch, Rebecca; Wise, Matthew E; Nelson, Noele P; Suryaprasad, Anil; Drobeniuc, Jan; Holmberg, Scott D; Xu, Fujie

2014-10-01

In May, 2013, an outbreak of symptomatic hepatitis A virus infections occurred in the USA. Federal, state, and local public health officials investigated the cause of the outbreak and instituted actions to control its spread. We investigated the source of the outbreak and assessed the public health measures used. We interviewed patients, obtained their shopping information, and did genetic analysis of hepatitis A virus recovered from patients' serum and stool samples. We tested products for the virus and traced supply chains. Of 165 patients identified from ten states, 69 (42%) were admitted to hospital, two developed fulminant hepatitis, and one needed a liver transplant; none died. Illness onset occurred from March 31 to Aug 12, 2013. The median age of patients was 47 years (IQR 35-58) and 91 (55%) were women. 153 patients (93%) reported consuming product B from retailer A. 40 patients (24%) had product B in their freezers, and 113 (68%) bought it according to data from retailer A. Hepatitis A virus genotype IB, uncommon in the Americas, was recovered from specimens from 117 people with hepatitis A virus illness. Pomegranate arils that were imported from Turkey--where genotype IB is common--were identified in product B. No hepatitis A virus was detected in product B. Imported frozen pomegranate arils were identified as the vehicle early in the investigation by combining epidemiology--with data from several sources--genetic analysis of patient samples, and product tracing. Product B was removed from store shelves, the public were warned not to eat product B, product recalls took place, and postexposure prophylaxis with both hepatitis A virus vaccine and immunoglobulin was provided. Our findings show that modern public health actions can help rapidly detect and control hepatitis A virus illness caused by imported food. Our findings show that postexposure prophylaxis can successfully prevent hepatitis A illness when a specific product is identified. Imported food products combined with waning immunity in some adult populations might make this type of intervention necessary in the future. US Centers for Disease Control and Prevention, US Food and Drug Administration, and US state and local public health departments. Copyright © 2014 Elsevier Ltd. All rights reserved.

NS3 protease resistance-associated substitutions in liver tissue and plasma samples from patients infected by hepatitis C virus genotype 1A or 1B.

PubMed

Morsica, Giulia; Andolina, Andrea; Merli, Marco; Messina, Emanuela; Hasson, Hamid; Lazzarin, Adriano; Uberti-Foppa, Caterina; Bagaglio, Sabrina

2017-08-01

The presence of naturally occurring resistance-associated substitutions (RASs) in the HCV-protease domain has been poorly investigated in the liver, the main site of HCV replication. We evaluated the natural resistance of the virus to NS3 protease inhibitors in liver tissue and plasma samples taken from HCV-infected patients. RASs were investigated by means of viral population sequencing in liver tissue samples from 18 HCV-infected patients harbouring genotype 1a or genotype 1b; plasma samples from 12 of these patients were also available for virological investigation. A discordant genotype was found in two of the 12 patients (16.6%) who provided samples from both compartments. Sequence analysis of the NS3 protease domain showed the presence of RASs in four of the 18 liver tissue samples (22.2%), two of which showed cross-resistance to protease inhibitors in clinical use or phase 2-3 trials. The analysis of the 12 paired tissues and plasma samples excluded the presence of RASs in the plasma compartment. The dominance of discordant genotypes in the paired liver and plasma samples of some HCV-infected patients suggests mixed infection possibly leading to the selective advantage of different genotype in the two compartments. The presence of RASs at intra-hepatic level is not uncommon and may lead to the early emergence of cross-resistant strains.

Red fox (Vulpes vulpes Linnaeus, 1758) as biological indicator for environmental pollution in Hungary.

PubMed

Heltai, Miklós; Markov, Georgi

2012-10-01

Our aim were to establish the metal (Cu, Ni, Zn, Co, Cd, and Pb) levels of red fox liver and the kidney samples (n = 10) deriving from central part of Hungary and compare the results with other countries' data. According to our results the concentrations of residues of the targeted elements (mg/kg dry weight) in liver and kidney samples were, respectively in liver: Cu: 21.418, Zn: 156.928, Ni: 2.079, Co: 1.611, Pb: 1.678 and Cd: 0.499; and kidney samples: Cu: 9.236; Zn: 87.159; Ni: 2.514; Co: 2.455; Pb: 2.63 and Cd: 0.818. Pb levels of Hungarian red fox liver samples significantly exceed the values of Italian specimens' samples, whilst the same element's concentrations of Hungarian red fox kidney samples were higher than the results published in Germany.

The effects of freezing and thawing rates on tenderness, sensory quality, and retail display of beef subprimals.

PubMed

Hergenreder, J E; Hosch, J J; Varnold, K A; Haack, A L; Senaratne, L S; Pokharel, S; Beauchamp, C; Lobaugh, B; Calkins, C R

2013-01-01

The objective of this study was to evaluate processing methods for frozen beef subprimals; the effects of freezing and thawing rates on tenderness, sensory properties, and retail display were evaluated. There were 6 treatments: fresh, never frozen 14 d wet aged (14D); fresh, never frozen 21 d wet aged (21D); blast frozen-fast thawed (BF); blast frozen-slow thawed (BS); conventionally frozen-fast thawed (CF); and conventionally frozen-slow thawed (CS). All frozen beef subprimals were aged for 14 d before freezing. Three beef subprimal cuts, rib eye roll (n=90), strip loin (n=90), and top sirloin butt (n=90), were used with 3 replications of 5 samples per treatment per week (total of 9 wk, n=270). Blast freezing occurred by placing spacers between the boxes of meat on pallets at -28°C with high air velocity for 3 to 5 d. Conventional freezing occurred with boxes of meat stacked on pallets and placed in a -28°C freezer with minimal air movement for at least 10 d. Fast thawing of subprimals (to an internal temperature of -1°C to 1°C) occurred by immersion in a circulating water bath (<12°C) for 21 h, and slow thawing of subprimals occurred over a 2-wk period by placing individual subprimals on tables at 0°C. Steaks (2.5 cm thick) were cut from the longissimus thoracis (LT), longissimus lumborum (LL), and gluteus medius (GM) for Warner-Bratzler shear force (WBS), trained sensory evaluation, and retail display. For LL and GM beef steaks, frozen treatments were equal or lower in WBS values to 14D and 21D beef steaks. No differences were detected in WBS among the treatments applied to GM beef steaks (P=0.08). There were no differences in sensory tenderness among the LL, LT, and GM (P>0.05). All LL and LT beef steaks had approximately 4 d to 40% discoloration, and all GM steaks had over 3 d to 40% discoloration. Steaks from the LL and LT began to discolor at about 3 d, and the GM began to discolor after 1 d. For all beef subprimals, purge loss during storage and thawing was significantly greater for the slow-thawed subprimals (P<0.01), and all fast-thawed subprimals were equal or superior to 14D and 21D (P<0.01) in storage and thawing purge. During retail display, the greatest purge loss occurred in fast-thawed treatments (P<0.01). Overall, freezing rate did not affect purge loss, and neither freezing nor thawing rates had significant meaningful effects on WBS, and sensory properties were comparable with fresh, never-frozen subprimals.

Influence of experimental conditions on data variability in the liver comet assay.

PubMed

Guérard, M; Marchand, C; Plappert-Helbig, U

2014-03-01

The in vivo comet assay has increasingly been used for regulatory genotoxicity testing in recent years. While it has been demonstrated that the experimental execution of the assay, for example, electrophoresis or scoring, can have a strong impact on the results; little is known on how initial steps, that is, from tissue sampling during necropsy up to slide preparation, can influence the comet assay results. Therefore, we investigated which of the multitude of steps in processing the liver for the comet assay are most critical. All together eight parameters were assessed by using liver samples of untreated animals. In addition, two of those parameters (temperature and storage time of liver before embedding into agarose) were further investigated in animals given a single oral dose of ethyl methanesulfonate at dose levels of 50, 100, and 200 mg/kg, 3 hr prior to necropsy. The results showed that sample cooling emerged as the predominant influence factor, whereas variations in other elements of the procedure (e.g., size of the liver piece sampled, time needed to process the liver tissue post-mortem, agarose temperature, or time of lysis) seem to be of little relevance. Storing of liver samples of up to 6 hr under cooled conditions did not cause an increase in tail intensity. In contrast, storing the tissue at room temperature, resulted in a considerable time-dependent increase in comet parameters. Copyright © 2013 Wiley Periodicals, Inc.

A multicolor flow cytometric assay for measurement of platelet-derived microparticles.

PubMed

Mobarrez, Fariborz; Antovic, Jovan; Egberg, Nils; Hansson, Mona; Jörneskog, Gun; Hultenby, Kjell; Wallén, Håkan

2010-03-01

Flow cytometry (FCM) is the most commonly used method for detection of platelet-derived microparticles (PDMPs), but it is poorly standardized and mainly used for "bedside" analyses in fresh samples. If PDMPs could be analyzed in previously frozen samples it would increase the usefulness of the method. However, cell membrane fragments from contaminating cells created during freezing/thawing may cause artifacts and disturb measurements. PDMPs were labeled with monoclonal antibodies directed against CD42a and CD62P, or CD42a and CD142. The PDMP gate was determined using forward scatter (FSC) and CD42a expression. The mean fluorescence intensities (MFIs) of CD62P or CD142 positive particles were translated into MESF -values (Molecules of Equivalent Soluble Fluorochrome) using a standard curve. FITC-labeled phalloidin (which binds to intracellular actin) was used to detect destroyed cells/cell fragments. Phalloidin-positive particles were significantly more common in supernatants of frozen/thawed platelet rich and platelet poor plasma samples compared with supernatants of platelet free plasma. High-speed centrifugation was then used to obtain PDMP samples with low contamination of cell fragments. Electron microscopy showed that these samples contained numerous round stained particles with cellular membranes of a size of 100-700 nm. Reproducibility experiments using plasma samples from healthy individuals showed that the coefficients of variation (CVs) of MESF values of CD62P and CD142 (both intra- and interassay) were <10%, and the variation between two cytometers in two different laboratories was <5%. We also found that PDMP expression of CD142 (i.e. tissue factor [TF]) and CD62P (i.e P-selectin) was around two times higher in samples from type 1-diabetes patients compared with those from healthy controls (p<0.001). The use of MESF values to quantify PDMP expression of P-selectin and TF yields reproducible data and enables comparison of data between laboratories. If high-speed centrifugation is performed, contamination of cell fragments is low in frozen/thawed samples. (c) 2009 Elsevier Ltd. All rights reserved.

Subpopulations of bovine T lymphocytes collected during foot-and-mouth disease virus infection are affected by freezing, but are subsequently stable in frozen samples

USDA-ARS?s Scientific Manuscript database

Immunophenotyping of peripheral-blood lymphocytes by flow cytometry is an important tool for infectious disease research. In many live-animal experiments and other longitudinal studies, the processing, prompt staining, and analysis of fresh samples is a logistical challenge and daily variation can c...

Effect of Processing Delay and Storage Conditions on Urine Albumin-to-Creatinine Ratio.

PubMed

Herrington, William; Illingworth, Nicola; Staplin, Natalie; Kumar, Aishwarya; Storey, Ben; Hrusecka, Renata; Judge, Parminder; Mahmood, Maria; Parish, Sarah; Landray, Martin; Haynes, Richard; Baigent, Colin; Hill, Michael; Clark, Sarah

2016-10-07

Because there is substantial biologic intraindividual variation in albumin excretion, randomized trials of albuminuria-reducing therapies may need multiple urine samples to estimate daily urinary albumin excretion. Mailing spot urine samples could offer a convenient and cost-effective method to collect multiple samples, but urine albumin-to-creatinine ratio stability in samples stored at ambient temperatures for several days is unknown. Patients with kidney disease provided fresh urine samples in two tubes (with and without boric acid preservative). Reference aliquots from each participant were analyzed immediately, whereas remaining aliquots were subject to different handling/storage conditions before analysis, including delayed processing for up to 7 days at three different storage temperatures (4°C, 18°C, and 30°C), multiple freeze-thaw cycles, and long-term frozen storage at -80°C, -40°C, and -20°C. We calculated the mean percentage change in urine albumin-to-creatinine ratio for each condition, and we considered samples stable if the 95% confidence interval was within a ±5% threshold. Ninety-three patients provided samples with detectable albuminuria in the reference aliquot. Median (interquartile range) urine albumin-to-creatinine ratio was 87 (20-499) mg/g. The inclusion of preservative had minimal effect on fresh urine albumin-to-creatinine ratio measurements but reduced the changes in albumin and creatinine in samples subject to processing delay and storage conditions. The urine albumin-to-creatinine ratio was stable for 7 days in samples containing preservative at 4°C and 18°C and 2 days when stored at 30°C. It was also stable in samples with preservative after three freeze-thaw cycles and in frozen storage for 6 months at -80°C or -40°C but not at -20°C. Mailed urine samples collected with preservative and received within 7 days if ambient temperature is ≤18°C, or within 2 days if the temperature is higher but does not exceed 30°C, are suitable for the measurement of urine albumin-to-creatinine ratio in randomized trials. Preserved samples frozen to -40°C or -80°C for 6 months before analysis also seem suitable. Copyright © 2016 by the American Society of Nephrology.

Effect of Processing Delay and Storage Conditions on Urine Albumin-to-Creatinine Ratio

PubMed Central

Illingworth, Nicola; Staplin, Natalie; Kumar, Aishwarya; Storey, Ben; Hrusecka, Renata; Judge, Parminder; Mahmood, Maria; Parish, Sarah; Landray, Martin; Haynes, Richard; Baigent, Colin; Hill, Michael; Clark, Sarah

2016-01-01

Background and objectives Because there is substantial biologic intraindividual variation in albumin excretion, randomized trials of albuminuria-reducing therapies may need multiple urine samples to estimate daily urinary albumin excretion. Mailing spot urine samples could offer a convenient and cost-effective method to collect multiple samples, but urine albumin-to-creatinine ratio stability in samples stored at ambient temperatures for several days is unknown. Design, setting, participants, & measurements Patients with kidney disease provided fresh urine samples in two tubes (with and without boric acid preservative). Reference aliquots from each participant were analyzed immediately, whereas remaining aliquots were subject to different handling/storage conditions before analysis, including delayed processing for up to 7 days at three different storage temperatures (4°C, 18°C, and 30°C), multiple freeze-thaw cycles, and long–term frozen storage at −80°C, −40°C, and −20°C. We calculated the mean percentage change in urine albumin-to-creatinine ratio for each condition, and we considered samples stable if the 95% confidence interval was within a ±5% threshold. Results Ninety-three patients provided samples with detectable albuminuria in the reference aliquot. Median (interquartile range) urine albumin-to-creatinine ratio was 87 (20–499) mg/g. The inclusion of preservative had minimal effect on fresh urine albumin-to-creatinine ratio measurements but reduced the changes in albumin and creatinine in samples subject to processing delay and storage conditions. The urine albumin-to-creatinine ratio was stable for 7 days in samples containing preservative at 4°C and 18°C and 2 days when stored at 30°C. It was also stable in samples with preservative after three freeze-thaw cycles and in frozen storage for 6 months at −80°C or −40°C but not at −20°C. Conclusions Mailed urine samples collected with preservative and received within 7 days if ambient temperature is ≤18°C, or within 2 days if the temperature is higher but does not exceed 30°C, are suitable for the measurement of urine albumin-to-creatinine ratio in randomized trials. Preserved samples frozen to −40°C or −80°C for 6 months before analysis also seem suitable. PMID:27654930

The forensiX evidence collection tube and its impact on DNA preservation and recovery.

PubMed

Garvin, Alex M; Holzinger, Ralf; Berner, Florian; Krebs, Walter; Hostettler, Bernhard; Lardi, Elges; Hertli, Christian; Quartermaine, Roy; Stamm, Christoph

2013-01-01

Biological samples are vulnerable to degradation from the time they are collected until they are analysed at the laboratory. Biological contaminants, such as bacteria, fungi, and enzymes, as well as environmental factors, such as sunlight, heat, and humidity, can increase the rate of DNA degradation. Currently, DNA samples are normally dried or frozen to limit their degradation prior to their arrival at the laboratory. In this study, the effect of the sample drying rate on DNA preservation was investigated, as well as a comparison between drying and freezing methods. The drying performances of two commercially available DNA collection tools (swab and drying tube) with different drying rates were evaluated. The swabs were used to collect human saliva, placed into the drying tubes, and stored in a controlled environment at 25°C and 60% relative humidity, or frozen at -20°C, for 2 weeks. Swabs that were stored in fast sample drying tubes yielded 95% recoverable DNA, whereas swabs stored in tubes with slower sample drying rates yielded only 12% recoverable DNA; saliva stored in a microtube at -20°C was used as a control. Thus, DNA sampling tools that offer rapid drying can significantly improve the preservation of DNA collected on a swab, increasing the quantity of DNA available for subsequent analysis.

The DNA isolation method has effect on allele drop-out and on the results of fluorescent PCR and DNA fragment analysis.

PubMed

Nagy, Bálint; Bán, Zoltán; Papp, Zoltán

2005-10-01

The quality and the quantity of isolated DNA have an effect on PCR amplifications. The authors studied three DNA isolation protocols (resin binding method using fresh and frozen amniotic fluid samples, and silica adsorption method using fresh samples) on the quantity and on the quality of the isolated DNA. Amniotic fluid samples were obtained from 20 pregnant women. The isolated DNA concentrations were determined by real-time fluorimeter using SYBRGreen I method. Each sample was studied for the presence of 8 STR markers. The authors compared the number of the detected alleles, electrophoretograms and peak areas. There was a significant difference between the concentration of the obtained DNA and in the peak areas between the three isolation protocols. The numbers of detected alleles were different, we observed the most allele drop outs in the resin type DNA isolation protocol from the fresh sample (detected allele numbers 182), followed by resin binding protocol from the frozen samples (detected allele number 243) and by the silica adsorption method (detected allele number 264). The authors demonstrated that the DNA isolation method has an effect on the quantity and quality of the isolated DNA, and on further PCR amplifications.

Effects of Holder pasteurization on the protein profile of human milk.

PubMed

Peila, Chiara; Coscia, Alessandra; Bertino, Enrico; Cavaletto, Maria; Spertino, Stefano; Icardi, Sara; Tortone, Claudia; Visser, Gerard H A; Gazzolo, Diego

2016-04-07

The most widespread method for the treatment of donor milk is the Holder pasteurization (HoP). The available literature data show that HoP may cause degradation of some bioactive components. The aim of this study was to determine the effect of HoP on the protein profile of human milk (HM) using a GeLC-MS method, a proteomic approach and a promising technique able to offer a qualitative HM protein profile. HM samples were collected by standardized methods from 20 mothers carrying both preterm and term newborns. A aliquot of each sample was immediately frozen at -80 °C, whilst another one was Holder pasteurized and then frozen. All samples were then analyzed by GeLC-MS. The protein bands of interest were excised from the gel, digested with trypsin and identified by nano-HPLC-MS/MS analysis. The protein profile before and after HoP showed qualitative differences only in 6 samples out of 20, while in the remaining 14 no detectable differences were found. The differences interested only colostrums and transitional milk samples and regarded the decrease of the electrophoretic bands corresponding to alpha and beta-casein, tenascin, lactoferrin and immunoglobulin. In the majority of samples, HoP did not cause any modification, thereby preserving the biological activity of HM proteins.

7 CFR 58.349 - Frozen cream.

Code of Federal Regulations, 2010 CFR

2010-01-01

... tested in accordance with §§ 58.336 and 58.337. Samples for analysis should be taken prior to freezing of...: Copper content, not more than 0.3 ppm; iron content not more than 1.0 ppm. Supplemental Specifications...

High density FTA plates serve as efficient long-term sample storage for HLA genotyping.

PubMed

Lange, V; Arndt, K; Schwarzelt, C; Boehme, I; Giani, A S; Schmidt, A H; Ehninger, G; Wassmuth, R

2014-02-01

Storage of dried blood spots (DBS) on high-density FTA(®) plates could constitute an appealing alternative to frozen storage. However, it remains controversial whether DBS are suitable for high-resolution sequencing of human leukocyte antigen (HLA) alleles. Therefore, we extracted DNA from DBS that had been stored for up to 4 years, using six different methods. We identified those extraction methods that recovered sufficient high-quality DNA for reliable high-resolution HLA sequencing. Further, we confirmed that frozen whole blood samples that had been stored for several years can be transferred to filter paper without compromising HLA genotyping upon extraction. Concluding, DNA derived from high-density FTA(®) plates is suitable for high-resolution HLA sequencing, provided that appropriate extraction protocols are employed. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Quantification of C4d deposition and hepatitis C virus RNA in tissue in cases of graft rejection and hepatitis C recurrence after liver transplantation

PubMed Central

Song, Alice Tung Wan; de Mello, Evandro Sobroza; Alves, Venâncio Avancini Ferreira; Cavalheiro, Norma de Paula; Melo, Carlos Eduardo; Bonazzi, Patricia Rodrigues; Tengan, Fatima Mitiko; Freire, Maristela Pinheiro; Barone, Antonio Alci; D'Albuquerque, Luiz Augusto Carneiro; Abdala, Edson

2015-01-01

Histology is the gold standard for diagnosing acute rejection and hepatitis C recurrence after liver transplantation. However, differential diagnosis between the two can be difficult. We evaluated the role of C4d staining and quantification of hepatitis C virus (HCV) RNA levels in liver tissue. This was a retrospective study of 98 liver biopsy samples divided into four groups by histological diagnosis: acute rejection in patients undergoing liver transplant for hepatitis C (RejHCV+), HCV recurrence in patients undergoing liver transplant for hepatitis C (HCVTx+), acute rejection in patients undergoing liver transplant for reasons other than hepatitis C and chronic hepatitis C not transplanted (HCVTx-). All samples were submitted for immunohistochemical staining for C4d and HCV RNA quantification. Immunoexpression of C4d was observed in the portal vessels and was highest in the HCVTx- group. There was no difference in C4d expression between the RejHCV+ and HCVTx+ groups. However, tissue HCV RNA levels were higher in the HCVTx+ group samples than in the RejHCV+ group samples. Additionally, there was a significant correlation between tissue and serum levels of HCV RNA. The quantification of HCV RNA in liver tissue might prove to be an efficient diagnostic test for the recurrence of HCV infection. PMID:25742264

Application of RT-PCR in formalin-fixed and paraffin-embedded lung cancer tissues.

PubMed

Zhang, Fan; Wang, Zhuo-min; Liu, Hong-yu; Bai, Yun; Wei, Sen; Li, Ying; Wang, Min; Chen, Jun; Zhou, Qing-hua

2010-01-01

To analyze gene expression in formalin-fixed, paraffin-embedded lung cancer tissues using modified method. Total RNA from frozen tissues was extracted using TRIZOL reagent. RNA was extracted from formalin-fixed, paraffin-embedded tissues by digestion with proteinase K before the acid-phenol:chloroform extraction and carrier precipitation. We modified this method by using a higher concentration of proteinase K and a longer digestion time, optimized to 16 hours. RT-PCR and real-time RT-PCR were used to check reproducibility and the concordance between frozen and paraffin-embedded samples. The results showed that the RNA extracted from the paraffin-embedded lung tissues had high quality with the most fragment length between 28S and 18S bands (about 1000 to 2000 bases). The housekeeping gene GUSB exhibited low variation of expression in frozen and paraffin-embedded lung tissues, whereas PGK1 had the lowest variation in lymphoma tissues. Furthermore, real-time PCR analysis of the expression of known prognostic genes in non-small cell lung carcinoma (NSCLC) demonstrated an extremely high correlation (r>0.880) between the paired frozen and formalin-fixed, paraffin-embedded specimens. This improved method of RNA extraction is suitable for real-time quantitative RT-PCR, and may be used for global gene expression profiling of paraffin-embedded tissues.

Effect of freezing and canning on the content of selected vitamins and pigments in seeds of two grass pea (Lathyrus sativus L.) cultivars at the not fully mature stage.

PubMed

Korus, Anna; Lisiewska, Zofia; Kmiecik, Waldemar

2002-08-01

Seeds of the grass pea (Lathyrus sativus L.) cultivars Derek and Krab, with a dry matter content of about 33%, were used for freezing and for canning. The content of vitamins C, B1, and B2 and of carotenoids, beta-carotene, and chlorophylls was determined in raw and blanched material, in frozen products after 6-month storage before and after cooking to consumption consistency, and in canned products after 6-month storage. In comparison with the cultivar Krab, raw seeds of Derek contained 45% more vitamin C, 14% more total chlorophylls, 13% less thiamine (vitamin B1), and 7% less riboflavin (vitamin B2). The level of carotenoids was similar. Blanching of seeds led to a statistically significant decrease only in the content of vitamin C. Freezing and frozen storage significantly lowered the level of vitamin C and chlorophylls. The cooking of frozen seeds and the production of canned products and their storage resulted in a statistically verified reduction in the content of components analysed in all the samples. Greater losses were found in products prepared from seeds of the cv. Krab. After cooking, frozen seeds contained more of all the analysed components than the canned products.

Effect of Novel Quick Freezing Techniques Combined with Different Thawing Processes on Beef Quality

PubMed Central

Yoo, Seon-Mi; Han, Gui-Jung

2014-01-01

This study investigated the effect of various freezing and thawing techniques on the quality of beef. Meat samples were frozen using natural convection freezing (NF), individual quick freezing (IQF), or cryogenic freezing (CF) techniques, followed by natural convection thawing (NCT) or running water thawing (RT). The meat was frozen until the core temperature reached -12℃ and then stored at -24℃, followed by thawing until the temperature reached 5℃. Quality parameters, such as the pH, water binding properties, CIE color, shear force, and microstructure of the beef were elucidated. Although the freezing and thawing combinations did not cause remarkable changes in the quality parameters, rapid freezing, in the order of CF, IQF, and NF, was found to minimize the quality deterioration. In the case of thawing methods, NCT was better than RT and the meat quality was influence on the thawing temperature rather than the thawing rate. Although the microstructure of the frozen beef exhibited an excessive loss of integrity after the freezing and thawing, it did not cause any remarkable change in the beef quality. Taken together, these results demonstrate that CF and NCT form the best combination for beef processing; however, IQF and NCT may have practical applications in the frozen food industry. PMID:26761674

Intra-Hepatic Depletion of Mucosal-Associated Invariant T Cells in Hepatitis C Virus-Induced Liver Inflammation.

PubMed

Bolte, Fabian J; O'Keefe, Ashley C; Webb, Lauren M; Serti, Elisavet; Rivera, Elenita; Liang, T Jake; Ghany, Marc; Rehermann, Barbara

2017-11-01

Chronic hepatitis affects phenotypes of innate and adaptive immune cells. Mucosal-associated invariant T (MAIT) cells are enriched in the liver as compared with the blood, respond to intra-hepatic cytokines, and (via the semi-invariant T-cell receptor) to bacteria translocated from the gut. Little is known about the role of MAIT cells in livers of patients with chronic hepatitis C virus (HCV) infection and their fate after antiviral therapy. We collected blood samples from 42 patients with chronic HCV infection who achieved a sustained virologic response after 12 weeks of treatment with sofosbuvir and velpatasvir. Mononuclear cells were isolated from blood before treatment, at weeks 4 and 12 during treatment, and 24 weeks after the end of treatment. Liver biopsies were collected from 37 of the patients prior to and at week 4 of treatment. Mononuclear cells from 56 blood donors and 10 livers that were not suitable for transplantation were used as controls. Liver samples were assessed histologically for inflammation and fibrosis. Mononuclear cells from liver and blood were studied by flow cytometry and analyzed for responses to cytokine and bacterial stimulation. The frequency of MAIT cells among T cells was significantly lower in blood and liver samples of patients with HCV infection than of controls (median, 1.31% vs 2.32% for blood samples, P = .0048; and median, 4.34% vs 13.40% for liver samples, P = .001). There was an inverse correlation between the frequency of MAIT cells in the liver and histologically determined levels of liver inflammation (r = -.5437, P = .0006) and fibrosis (r = -.5829, P = .0002). MAIT cells from the liver had higher levels of activation and cytotoxicity than MAIT cells from blood (P < .0001). Production of interferon gamma by MAIT cells was dependent on monocyte-derived interleukin 18, and was reduced in patients with HCV infection in response to T-cell receptor-mediated but not cytokine-mediated stimulation, as compared with controls. Anti-viral therapy rapidly decreased liver inflammation and MAIT cell activation and cytotoxicity, and increased the MAIT cell frequency among intra-hepatic but not blood T cells. The MAIT cell response to T-cell receptor-mediated stimulation did not change during the 12 weeks of antiviral therapy. In analyses of paired blood and liver samples from patients with chronic HCV infection before, during, and after antiviral therapy with sofosbuvir and velpatasvir, we found that intrahepatic MAIT cells are activated by monocyte-derived cytokines and depleted in HCV-induced liver inflammation. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Microstructure and physical changes in the Mexican cooked lamb meat barbacoa made with chilled and frozen meat.

PubMed

Estrada-Solís, Joaquín; Figueroa-Rodríguez, Katia A; Figueroa-Sandoval, Benjamín; Hernández-Rosas, Francisco; Hernández-Cazares, Aleida S

2016-08-01

Longissimus dorci (LD) samples of different origin (imported and domestic) with pre-treatments (imported meat stored at -18°C for 6months, domestic meat stored at -18°C for 10days, and domestic meat stored at 4°C for 24h) were cooked as barbacoa and frozen using two treatments (air blast and liquid immersion) and then evaluated after 30days of storage. The results showed that the origin and pre-treatment of meat affected L*, a*, instrumental texture and microstructure; that the storage time affected pH, aw, b* and microstructure; and that the freezing treatments did not affect the meat. Overall, the frozen cooked lamb dish barbacoa could present some problems at the conservation stage due to an increase in pH, aw and changes in microstructure; however, the physical traits (color and texture) remained mostly unchanged and depended more on the quality of the raw meat. Copyright © 2016 Elsevier Ltd. All rights reserved.

Gravity-induced anomalies in interphase spacing reported for binary eutectics.

PubMed

Smith, Reginald W

2002-10-01

It has been reasoned that desirable microstructural refinement in binary eutectics could result from freezing in reduced-gravity. It is recognized that the interphase spacing in a binary eutectic is controlled by solute transport and that, on Earth, buoyancy-driven convection may enhance this. Hence, it has been presumed that the interphase spacing ought to decrease when a eutectic alloy is frozen under conditions of much-reduced gravity, where such buoyancy effects would be largely absent. The result of such speculation has been that many workers have frozen various eutectics under reduced gravity and have reported that, although some eutectics became finer, others showed no change, and some even became coarser. This reported varied behavior will be reviewed in the light of long term studies by the author at Queen's University, including recent microgravity studies in which samples of two eutectic alloy systems, MnBi-Bi and MnSb-Sb, were frozen under very stable conditions and showed no change in interphase spacing.

The Effect of Edible Coating Enriched With Kaffir Lime Leaf Essential Oil (Citrus hystrix DC) on Beef Sausage Quality During Frozen Storage (-18°±2°C)

NASA Astrophysics Data System (ADS)

Utami, R.; Kawiji; Khasanah, L. U.; Solikhah, R.

2018-03-01

The aim of this study was to determine the effect of edible coating enriched with kaffir lime (Citrus hystrix DC) leaves essential oil at various concentration on beef sausage quality during frozen storage (-18°±2°C). The concentration of kaffir lime leaves essential oil enriched in edible coating were varied at 0%; 0.2%; 1.4%. Microbiological, physical and chemical characteristics (TPC, color, TBA, TVB, and pH) were investigated at 0, 1, 2, 3, and 4 months of storage. The result showed that edible coating with the addition of kaffir lime leaves essential oils decreased the microbial growth, TVB value, and TBA value of beef sausage. The color and pH of samples can be stabilized during storage. The selected kaffir lime leaves essential oil concentrations based on microbial, physical, and chemical characteristics of beef sausages during frozen storage at -18°C was 0.2%.

Effects of processing and storage conditions of cocoyam strips on the quality of fries.

PubMed

Oguntowo, Oyindamola; Obadina, Adewale O; Sobukola, Olajide P; Adegunwa, Mojisola O

2016-11-01

The effects of blanching time and temperature on the sensory and textural properties of frozen cocoyam strips were studied for cocoyam varieties. The most preferred variety after sensory evaluation was blanched at 90°C for 5 min, reproduced, and frozen at a temperature of -18°C for storage studies over a period of 12 weeks with Irish potato as control. Sensory evaluation and instrumental texture analysis of frozen fried samples were conducted at 3 weeks intervals for 12 weeks. Sensory evaluation during storage showed no significant difference ( P  < 0.05) in taste, aroma, and mouth feel attributes between control and cocoyam fries. The sensory score for taste, sogginess, and mouth feel increased while those for aroma and color decreased in comparison with the control fries over storage. The texture increased during storage and for control fries. There was a significant negative correlation between sogginess, hardness, and dry matter, respectively.

Paritaprevir and Ritonavir Liver Concentrations in Rats as Assessed by Different Liver Sampling Techniques

PubMed Central

Venuto, Charles S.; Markatou, Marianthi; Woolwine-Cunningham, Yvonne; Furlage, Rosemary; Ocque, Andrew J.; DiFrancesco, Robin; Dumas, Emily O.; Wallace, Paul K.; Morse, Gene D.

2017-01-01

ABSTRACT The liver is crucial to pharmacology, yet substantial knowledge gaps exist in the understanding of its basic pharmacologic processes. An improved understanding for humans requires reliable and reproducible liver sampling methods. We compared liver concentrations of paritaprevir and ritonavir in rats by using samples collected by fine-needle aspiration (FNA), core needle biopsy (CNB), and surgical resection. Thirteen Sprague-Dawley rats were evaluated, nine of which received paritaprevir/ritonavir at 30/20 mg/kg of body weight by oral gavage daily for 4 or 5 days. Drug concentrations were measured using liquid chromatography-tandem mass spectrometry on samples collected via FNA (21G needle) with 1, 3, or 5 passes (FNA1, FNA3, and FNA5); via CNB (16G needle); and via surgical resection. Drug concentrations in plasma were also assessed. Analyses included noncompartmental pharmacokinetic analysis and use of Bland-Altman techniques. All liver tissue samples had higher paritaprevir and ritonavir concentrations than those in plasma. Resected samples, considered the benchmark measure, resulted in estimations of the highest values for the pharmacokinetic parameters of exposure (maximum concentration of drug in serum [Cmax] and area under the concentration-time curve from 0 to 24 h [AUC0–24]) for paritaprevir and ritonavir. Bland-Altman analyses showed that the best agreement occurred between tissue resection and CNB, with 15% bias, followed by FNA3 and FNA5, with 18% bias, and FNA1 and FNA3, with a 22% bias for paritaprevir. Paritaprevir and ritonavir are highly concentrated in rat liver. Further research is needed to validate FNA sampling for humans, with the possible derivation and application of correction factors for drug concentration measurements. PMID:28264852

Paritaprevir and Ritonavir Liver Concentrations in Rats as Assessed by Different Liver Sampling Techniques.

PubMed

Venuto, Charles S; Markatou, Marianthi; Woolwine-Cunningham, Yvonne; Furlage, Rosemary; Ocque, Andrew J; DiFrancesco, Robin; Dumas, Emily O; Wallace, Paul K; Morse, Gene D; Talal, Andrew H

2017-05-01

The liver is crucial to pharmacology, yet substantial knowledge gaps exist in the understanding of its basic pharmacologic processes. An improved understanding for humans requires reliable and reproducible liver sampling methods. We compared liver concentrations of paritaprevir and ritonavir in rats by using samples collected by fine-needle aspiration (FNA), core needle biopsy (CNB), and surgical resection. Thirteen Sprague-Dawley rats were evaluated, nine of which received paritaprevir/ritonavir at 30/20 mg/kg of body weight by oral gavage daily for 4 or 5 days. Drug concentrations were measured using liquid chromatography-tandem mass spectrometry on samples collected via FNA (21G needle) with 1, 3, or 5 passes (FNA 1 , FNA 3 , and FNA 5 ); via CNB (16G needle); and via surgical resection. Drug concentrations in plasma were also assessed. Analyses included noncompartmental pharmacokinetic analysis and use of Bland-Altman techniques. All liver tissue samples had higher paritaprevir and ritonavir concentrations than those in plasma. Resected samples, considered the benchmark measure, resulted in estimations of the highest values for the pharmacokinetic parameters of exposure (maximum concentration of drug in serum [ C max ] and area under the concentration-time curve from 0 to 24 h [AUC 0-24 ]) for paritaprevir and ritonavir. Bland-Altman analyses showed that the best agreement occurred between tissue resection and CNB, with 15% bias, followed by FNA 3 and FNA 5 , with 18% bias, and FNA 1 and FNA 3 , with a 22% bias for paritaprevir. Paritaprevir and ritonavir are highly concentrated in rat liver. Further research is needed to validate FNA sampling for humans, with the possible derivation and application of correction factors for drug concentration measurements. Copyright © 2017 American Society for Microbiology.

Comparison of Fecal Collection Methods for Microbiota Studies in Bangladesh

PubMed Central

Chen, Jun; Kibriya, Muhammad G.; Chen, Yu; Islam, Tariqul; Eunes, Mahbubul; Ahmed, Alauddin; Naher, Jabun; Rahman, Anisur; Amir, Amnon; Shi, Jianxin; Abnet, Christian C.; Nelson, Heidi; Knight, Rob; Chia, Nicholas; Ahsan, Habibul; Sinha, Rashmi

2017-01-01

ABSTRACT To our knowledge, fecal microbiota collection methods have not been evaluated in low- and middle-income countries. Therefore, we evaluated five different fecal sample collection methods for technical reproducibility, stability, and accuracy within the Health Effects of Arsenic Longitudinal Study (HEALS) in Bangladesh. Fifty participants from the HEALS provided fecal samples in the clinic which were aliquoted into no solution, 95% ethanol, RNAlater, postdevelopment fecal occult blood test (FOBT) cards, and fecal immunochemical test (FIT) tubes. Half of the aliquots were frozen immediately at −80°C (day 0) and the remaining samples were left at ambient temperature for 96 h and then frozen (day 4). Intraclass correlation coefficients (ICC) were calculated for the relative abundances of the top three phyla, for two alpha diversity measures, and for four beta diversity measures. The duplicate samples had relatively high ICCs for technical reproducibility at day 0 and day 4 (range, 0.79 to 0.99). The FOBT card and samples preserved in RNAlater and 95% ethanol had the highest ICCs for stability over 4 days. The FIT tube had lower stability measures overall. In comparison to the “gold standard” method using immediately frozen fecal samples with no solution, the ICCs for many of the microbial metrics were low, but the rank order appeared to be preserved as seen by the Spearman correlation. The FOBT cards, 95% ethanol, and RNAlater were effective fecal preservatives. These fecal collection methods are optimal for future cohort studies, particularly in low- and middle-income countries. IMPORTANCE The collection of fecal samples in prospective cohort studies is essential to provide the opportunity to study the effect of the human microbiota on numerous health conditions. However, these collection methods have not been adequately tested in low- and middle-income countries. We present estimates of technical reproducibility, stability at ambient temperature for 4 days, and accuracy comparing a “gold standard” for fecal samples in no solution, 95% ethanol, RNAlater, postdevelopment fecal occult blood test cards, and fecal immunochemical test tubes in a study conducted in Bangladesh. Fecal occult blood test cards and fecal samples stored in 95% ethanol or RNAlater adequately preserve fecal samples in this setting. Therefore, new studies in low- and middle-income countries should include collection of fecal samples using fecal occult blood test cards, 95% ethanol, or RNAlater for prospective cohort studies. PMID:28258145

Human serum cholinesterase from liver pathological samples exhibit highly elevated aryl acylamidase activity.

PubMed

Boopathy, Rathanam; Rajesh, Ramanna Valmiki; Darvesh, Sultan; Layer, Paul G

2007-05-01

Although aspartate aminotransferase (AST) and gamma-glutamyltransferase (gamma GT) enzymes are widely used as markers for liver disorders, the ubiquitous enzyme butyrylcholinesterase (BChE), synthesized in liver is also used as marker in the assessment of liver pathophysiology. This BChE enzyme in addition to its esterase activity has yet another enzymatic function designated as aryl acylamidase (AAA) activity. It is determined in in vitro based on the hydrolysis of the synthetic substrate o-nitroacetanilide. In the present study, human serum cholinesterase (BChE) activity was studied with respect to its AAA activity on the BChE protein (AAA(BChE)) in patients with liver disorders. AST and gamma GT values were taken into account in this study as known markers for liver disorders. Blood samples were grouped into 3 based on esterase activity associated with BChE protein. They are normal, low, and very low BChE activity but with markedly increased AST and gamma GT levels. These samples were tested for their respective AAA function. Association of AAA with BChE from samples was proved using BChE monoclonal antibody precipitation experiment. The absolute levels of AAA were increased as BChE activity decreased while deviating from normal samples and such deviation was directly proportional to the severity of the liver disorder. Differences between these groups became prominent after determining the ratios of AAA(BChE) to BChE activities. Samples showing very high AAA(BChE) to BChE ratio were also showing high to very high gamma GT values. These findings establish AAA(BChE) as an independently regulated enzymatic activity on BChE especially in liver disorders. Moreover, since neither the low esterase activity of BChE by itself nor increased levels of AST/gamma GT are sufficient pathological indicators, this pilot study merits replication with large sample numbers.

Changes in expression of cellular oncogenes and endogenous retrovirus-like sequences during hepatocarcinogenesis induced by a peroxisome proliferator.

PubMed Central

Hsieh, L. L.; Shinozuka, H.; Weinstein, I. B.

1991-01-01

Previous studies have demonstrated that BR-931, a hepatic peroxisome proliferator, can induce liver tumours in mice and rats. Since alterations in gene expression may play a critical role in multistage hepatocarcinogenesis, the present studies examined the expression of the c-myc, c-H-ras, epidermal growth factor (EGF) receptor and ODC (ornithine decarboxylase) genes, as well as endogenous retrovirus-like sequences, in F344 rat liver during the first 8 weeks of feeding a 0.16% Br931 diet and in liver tumours induced by chronic feeding of this diet. Northern blot analysis of poly A + liver RNA samples showed an increase in the level of RNAs homologous to rat leukaemia virus (RaLV) but no significant change in the level of 30S-retrovirus related RNAs in the liver RNA samples obtained from rats during the first 8 weeks of feeding the diet containing BR931. An increase in the levels of c-myc, c-H-ras and ODC transcripts was also seen in the liver RNA samples from the treated rats. Of particular interest was a decrease in the abundance of EGF receptor transcripts in the liver RNA samples from rats fed the BR931 diet. Increased levels of RaLV, c-myc, and ODC RNAs were also seen in the tumours induced by BR931, but this was not the case for 30S and c-H-ras. The liver tumour samples also showed a decrease in EGF receptor RNA. These changes in cellular levels of specific RNAs resemble, in several respect, those we previously described in rodent liver during regeneration and tumour promotion, and also those seen in rodent hepatomas induced by other agents. Therefore, they may reflect a common profile of gene expression relevant to liver proliferation and carcinogenesis. Images Figure 1 Figure 2 PMID:1931600

Tumoural specimens for forensic purposes: comparison of genetic alterations in frozen and formalin-fixed paraffin-embedded tissues.

PubMed

Ananian, Viviana; Tozzo, Pamela; Ponzano, Elena; Nitti, Donato; Rodriguez, Daniele; Caenazzo, Luciana

2011-05-01

In certain circumstances, tumour tissue specimens are the only DNA resource available for forensic DNA analysis. However, cancer tissues can show microsatellite instability and loss of heterozygosity which, if concerning the short tandem repeats (STRs) used in the forensic field, can cause misinterpretation of the results. Moreover, though formalin-fixed paraffin-embedded tissues (FFPET) represent a large resource for these analyses, the quality of the DNA obtained from this kind of specimen can be an important limit. In this study, we evaluated the use of tumoural tissue as biological material for the determination of genetic profiles in the forensic field, highlighting which STR polymorphisms are more susceptible to tumour genetic alterations and which of the analysed tumours show a higher genetic variability. The analyses were conducted on samples of the same tissues conserved in different storage conditions, to compare genetic profiles obtained by frozen tissues and formalin-fixed paraffin-embedded tissues. The importance of this study is due to the large number of specimens analysed (122), the large number of polymorphisms analysed for each specimen (39), and the possibility to compare, many years after storage, the same tissue frozen and formalin-fixed paraffin-embedded. In the comparison between the genetic profiles of frozen tumour tissues and FFPET, the same genetic alterations have been reported in both kinds of specimens. However, FFPET showed new alterations. We conclude that the use of FFPET requires greater attention than frozen tissues in the results interpretation and great care in both pre-extraction and extraction processes.

Effects of semen preservation on boar spermatozoa head membranes.

PubMed

Buhr, M M; Canvin, A T; Bailey, J L

1989-08-01

Head plasma membranes were isolated from the sperm-rich fraction of boar semen and from sperm-rich semen that had been subjected to three commercial preservation processes: Extended for fresh insemination (extended), prepared for freezing but not frozen (cooled), and stored frozen for 3-5 weeks (frozen-thawed). Fluorescence polarization was used to determine fluidity of the membranes of all samples for 160 min at 25 degrees C and also for membranes from the sperm-rich and extended semen during cooling and reheating (25 to 5 to 40 degrees C, 0.4 degrees C/min). Head plasma membranes from extended semen were initially more fluid than from other sources (P less than 0.05). Fluidity of head membranes from all sources decreased at 25 degrees C, but the rate of decrease was significantly lower for membranes from cooled and lower again for membranes from frozen-thawed semen. Cooling to 5 degrees C reduced the rate of fluidity change for plasma membranes from the sperm-rich fraction, while heating over 30 degrees C caused a significantly greater decrease. The presence of Ca++ (10 mM) lowered the fluidity of the head plasma membranes from sperm-rich and extended semen over time at 25 degrees C but did not affect the membranes from the cooled or frozen-thawed semen. The change in head plasma membrane fluidity at 25 degrees C may reflect the dynamic nature of spermatozoa membranes prior to fertilization. Extenders, preservation processes and temperature changes have a strong influence on head plasma membrane fluidity and therefore the molecular organization of this membrane.

Pre-selection by double layer density gradient centrifugation improves the fertilising capacity of frozen-thawed, capacitated stallion sperm.

PubMed

Morató, Roser; Soares, Juleide M De Souza; Orero, Guifré; Mogas, Teresa; Miró, Jordi

2013-06-01

The effect of combining double layer density gradient centrifugation (DL-DGC) with different capacitation treatments on the fertilising capacity of frozen-thawed stallion sperm was examined via a heterologous assay involving in vitro-matured, zona pellucida-free bovine oocytes. In a first experiment, aliquots of frozen-thawed stallion sperm were subjected to one of five capacitation treatments without DL-DGC - ionomycin at 1.0μM, 0.1μM, 0.05μM or 0.01μM, or caffeine at 200μg/mL. The fertilising capacity of the semen was then assessed at 18h by staining the above oocytes with 4,6-diamidino-2-phenylindole (DAPI) and examining for sperm penetration, the number of penetrated spermatozoa per oocyte, and male pronucleus formation. In a second experiment, aliquots of frozen-thawed stallion sperm were subjected to DL-DGC selection - or not - and then further subjected to the two best capacitation treatments (0.1μM and 0.05μM ionomycin). The fertilising capacity of the semen was then determined as above. The DL-DGC/capacitated sperm samples showed the highest mean penetration rates: 24.16% following capacitation with 0.1μM ionomycin, and 12.21% following capacitation with 0.05μM ionomycin. The capacitated but non-DL-DGC-selected sperm returned significantly lower values: 6.26% and 7.02% for the same ionomycin treatments respectively. These findings suggest that combining DL-DGC selection with ionomycin capacitation improves the fertilising capacity of frozen-thawed stallion sperm. Copyright © 2013 Elsevier B.V. All rights reserved.

Development of methods for cryopreservation of rooster sperm from the endangered breed "Gallina Valenciana de Chulilla" using low glycerol concentrations.

PubMed

Blanch, E; Tomás, C; Casares, L; Gómez, E A; Sansano, S; Giménez, I; Mocé, E

2014-06-01

Glycerol (11%; v:v) is the cryoprotectant most often used for the cryopreservation of rooster sperm. However, chicken breeds differ in the resistance of their sperm to the cryopreservation process and endangered or local breeds usually present low fertilizing ability when conventional sperm cryopreservation protocols are used. The objective of this study was to optimize the protocol for the cryopreservation of the sperm from the endangered breed "Gallina Valenciana de Chulilla". For this purpose, 10 pools of semen from 43 roosters of this breed were cryopreserved using 8%, 7%, 6%, or 4% glycerol, and the sperm quality was determined immediately after thawing and in the insemination doses. Lohmann Brown Classic laying hens (n = 40) were used for the insemination trials. The sperm quality after cryopreservation progressively decreased as the glycerol concentration was reduced (P < 0.01); samples frozen using 4% glycerol exhibited the lowest quality (38% total motile sperm and 49% live sperm), and samples frozen using 8% glycerol exhibited the highest quality (67% total motile sperm and 66% live sperm). These differences were also observed after the glycerol was removed (P < 0.01). However, the sperm fertilizing ability was similar for all the treatments (23%-30% fertilized eggs), and increased as the glycerol concentration decreased. In conclusion, semen from roosters frozen using 4% glycerol exhibited lower sperm quality but similar fertilizing ability compared with samples processed using higher glycerol concentrations. These results may provide useful information for developing cryopreservation protocols for other breeds. Copyright © 2014 Elsevier Inc. All rights reserved.

Determination of toxic elements (mercury, cadmium, lead, tin and arsenic) in fish and shellfish samples. Risk assessment for the consumers.

PubMed

Olmedo, P; Pla, A; Hernández, A F; Barbier, F; Ayouni, L; Gil, F

2013-09-01

Although fish intake has potential health benefits, the presence of metal contamination in seafood has raised public health concerns. In this study, levels of mercury, cadmium, lead, tin and arsenic have been determined in fresh, canned and frozen fish and shellfish products and compared with the maximum levels currently in force. In a further step, potential human health risks for the consumers were assessed. A total of 485 samples of the 43 most frequently consumed fish and shellfish species in Andalusia (Southern Spain) were analyzed for their toxic elements content. High mercury concentrations were found in some predatory species (blue shark, cat shark, swordfish and tuna), although they were below the regulatory maximum levels. In the case of cadmium, bivalve mollusks such as canned clams and mussels presented higher concentrations than fish, but almost none of the samples analyzed exceeded the maximum levels. Lead concentrations were almost negligible with the exception of frozen common sole, which showed median levels above the legal limit. Tin levels in canned products were far below the maximum regulatory limit, indicating that no significant tin was transferred from the can. Arsenic concentrations were higher in crustaceans such as fresh and frozen shrimps. The risk assessment performed indicated that fish and shellfish products were safe for the average consumer, although a potential risk cannot be dismissed for regular or excessive consumers of particular fish species, such as tuna, swordfish, blue shark and cat shark (for mercury) and common sole (for lead). Copyright © 2013 Elsevier Ltd. All rights reserved.

EFFECT ON PERFUSION VALUES OF SAMPLING INTERVAL OF CT PERFUSION ACQUISITIONS IN NEUROENDOCRINE LIVER METASTASES AND NORMAL LIVER

PubMed Central

Ng, Chaan S.; Hobbs, Brian P.; Wei, Wei; Anderson, Ella F.; Herron, Delise H.; Yao, James C.; Chandler, Adam G.

2014-01-01

Objective To assess the effects of sampling interval (SI) of CT perfusion acquisitions on CT perfusion values in normal liver and liver metastases from neuroendocrine tumors. Methods CT perfusion in 16 patients with neuroendocrine liver metastases were analyzed by distributed parameter modeling to yield tissue blood flow, blood volume, mean transit time, permeability, and hepatic arterial fraction, for tumor and normal liver. CT perfusion values for the reference sampling interval of 0.5s (SI0.5) were compared with those of SI datasets of 1s, 2s, 3s and 4s, using mixed-effects model analyses. Results Increases in SI beyond 1s were associated with significant and increasing departures of CT perfusion parameters from reference values at SI0.5 (p≤0.0009). CT perfusion values deviated from reference with increasing uncertainty with increasing SIs. Findings for normal liver were concordant. Conclusion Increasing SIs beyond 1s yield significantly different CT perfusion parameter values compared to reference values at SI0.5. PMID:25626401

The role of intraoperative narrow-band imaging in transoral laser microsurgery for early and moderately advanced glottic cancer.

PubMed

Klimza, Hanna; Jackowska, Joanna; Piazza, Cesare; Banaszewski, Jacek; Wierzbicka, Malgorzata

2018-03-01

Trans-oral laser microsurgery is an established technique for the treatment of early and moderately advanced laryngeal cancer. The authors intend to test the usefulness of narrow-band imaging in the intraoperative assessment of the larynx mucosa in terms of specifying surgical margins. Forty-four consecutive T1-T2 glottic cancers treated with trans-oral laser microsurgery Type I-VI cordectomy were presented. Suspected areas (90 samples/44 patients) were biopsied under the guidance of narrow-band imaging and white light and sent for frozen section. Our study revealed that 75 of 90 (83.3%) white light and narrow-band imaging-guided samples were histopathologically positive: 30 (40%) were confirmed as carcinoma in situ or invasive carcinoma and 45 (60%) as moderate to severe dysplasia. In 6 patients mucosa was suspected only in narrow-band imaging, with no suspicion under white light. Thus, in these 6 patients 18/90 (20%) samples were taken. In 5/6 patients 16/18 (88.8%) samples were positive in frozen section: in 6/18 (33.3%) carcinoma (2 patients), 10/18 (66.6%) severe dysplasia was confirmed (3 patients). In 1 patient 2/18 (11.1%) samples were negative in frozen section. Presented analysis showed, that sensitivity, specificity and accuracy of white light was 79.5%, 20% and 71.1% respectively, while narrow-band imaging was 100%, 0.0% and 85.7%, respectively. The intraoperative use of narrow-band imaging proved to be valuable in the visualization of suspect areas of the mucosa. Narrow-band imaging confirms the suspicions undertaken in white light and importantly, it showed microlesions beyond the scope of white light. Copyright © 2018 Associação Brasileira de Otorrinolaringologia e Cirurgia Cérvico-Facial. Published by Elsevier Editora Ltda. All rights reserved.

Liver lead concentrations in raptors in New Jersey, USA, 2008-2010.

PubMed

Stansley, William; Murphy, Lisa A

2011-08-01

Lead exposure in New Jersey raptors was assessed by analyzing liver samples from carcasses obtained from wildlife rehabilitators. Samples were collected from 221 individuals representing 13 species. Concentrations were within the range of normal background exposure in 12 species. One red-tailed hawk had a liver lead concentration consistent with clinical poisoning (7.4 μg/g wet weight), which represents an incidence of 1% (1/104) in that species and 0.5% (1/221) in the overall sample. A second red-tailed hawk had a liver lead concentration consistent with subclinical exposure (2.1 μg/g wet weight). The combined incidence of elevated exposure (subclinical exposure + clinical poisoning) was 2% (2/104) in red-tailed hawks and 1% (2/221) in the overall sample.

Fertility disturbances of dimethylacetamide and glycerol in rooster sperm diluents: Discrimination among effects produced pre and post freezing-thawing process.

PubMed

Abouelezz, F M K; Sayed, M A M; Santiago-Moreno, J

2017-09-01

With avian sperm cryopreservation protocols, the most widely used cryoprotectants (CPAs) are the glycerol (GLY; in gradual freezing: in-straw freezing method), and the dimethylacetamide (DMA; in pellets by plunging into liquid nitrogen: in-pellet rapid freezing method). Use of both methods results in a small portion of thawed live sperm with lesser fertilizing ability compared with the semen samples immediately after collection. This study was conducted to assess the pre-freezing damage occurring to the sperm due to the interaction with the cryoprotectants (CPAs) GLY (8%) and DMA (5%), as well as the post-freezing damage resulting from both freezing methods Data for each treatment, in fresh and frozen-thawed samples, were compared for sperm motility, fertilizing capacity and sperm-egg penetration holes/germinal disc (SP holes/GD). Hens (n=50) were artificially inseminated (10 hens/treatment) six times with 3day intervals between inseminations. The treatment of fresh sperm with DMA led to a reduction (P<0.05) in the count of SP holes/GD (21.4) and the fertility rate (66.7%). The addition and elimination of GLY in fresh samples resulted in a lesser (P<0.05) number of SP holes/GD (11.8) and the fertility rate (i.e., 50.0%). The number of SP-holes/GD was least in frozen-thawed samples using both DMA and GLY (14.2 and 9.2, respectively). The fertility rate when using semen frozen with DMA in- pellets was greater (P<0.05) than with use of semen that had been frozen using GLY in straws (46.4% compared with 31.3%). The reduction in fertility compared with the control when semen was cryopreserved using GLY was 64.1%; the GLY addition and elimination was responsible for two thirds of this reduction. The reduction in fertility when using semen cryopreserved with DMA was 46.7%; half of the reduction was attributed to the treatment with DMA. In conclusion, the mechanical damage attributed to the process for reducing GLY concentrations was more harmful to sperm fertilizing capacity than the toxicity of DMA and freeze/thaw process. For both freezing methods, the amount of sperm cryo-damage was similar, when the damage attributed to the CPA addition and elimination process was excluded. Copyright © 2017 Elsevier B.V. All rights reserved.

Cryopreservation of sperm of red abalone (Haliotis rufescens)

USGS Publications Warehouse

Salinas-Flores, L.; Paniagua-Chavez, C. G.; Jenkins, J.A.; Tiersch, T.R.

2005-01-01

Abalone culture, a developing industry in Baja California, Mexico, would benefit from genetic improvement and controlled breeding. The use of cryopreserved sperm would allow germplasm availability, and this study was designed to develop sperm cryopreservation protocols for red abalone Haliotis rufescens. The acute toxic effects of the cryoprotectants dimethyl sulfoxide (DMSO), propylene glycol (PG), and glycerol (GLY) were assessed after suspending sperm in different concentrations, whereby cryoprotectant treatments of 10% DMSO and 10% GLY equilibrated for 10 min yielded the highest range of motile sperm in preliminary freezing trials and were used for cryopreservation studies. To determine effective cooling rates, three freezing chambers were tested. Replicate samples of sperm from 4 males were placed in 0.5-mL French straws and frozen using a commercial freezing chamber (CFC) used for bull sperm, a programmable rate chamber (PRC), and a manually controlled styrofoam chamber (MCC). For the CFC, the cooling rate was 16??C/min, from 4??C to -140??C. For the PRC and MCC, it was 1??C/min, from -20??C to -30??C. The samples were held at -30??C for 5 min before being plunged into liquid nitrogen (-196??C) for storage, and each sample was thawed in a water bath at 45??C for 8 s. The quality of thawed sperm was determined by estimating percent motility, evaluating membrane integrity using a dual-staining technique and flow cytometry, and estimating fertilization rate. Statistical analyses were performed using 2-way ANOVA where chamber and treatment were the independent variables. Sperm quality parameters were independent. For motilities, a significant interaction was noted between the cryoprotective treatment and the chamber type, whereby motilities for DMSO and GLY were higher (P = 0.0055) using MCC. Membrane integrities were significantly lower after using the PRC than the CFC or the MCC (P = 0.0167). The highest post-thaw motility (48 ?? 7%) was found using sperm suspended in 10% glycerol and frozen in the MCC. The highest percent of intact membranes (56 ?? 11%) was for sperm suspended in 10% glycerol and frozen in the CFC. The highest fertilization rate (29 ?? 10%) was with samples frozen with 10% glycerol in the CFC. The use of cryopreserved sperm from red abalone provides an alternative breeding option for culture and the protocols delineated are the first developed for this species.

Comparison of pinniped and cetacean prey tissue lipids with lipids of their elasmobranch predator.

PubMed

Davidson, Bruce; Cliff, Geremy

2014-01-01

The great white shark is known to include pinnipeds and cetaceans in its diet. Both groups of marine mammals deposit thick blubber layers around their bodies. Elasmobranchs do not produce adipose tissue, but rather store lipid in their livers, thus a great white predating on a marine mammal will deposit the lipids in its liver until required. Samples from great white liver and muscle, Cape fur seal, Indian Ocean bottlenose dolphin and common dolphin liver, muscle and blubber were analyzed for their lipid and fatty acid profiles. The great white liver and marine mammal blubber samples showed a considerable degree of homogeneity, but there were significant differences when comparing between the muscle samples. Blubber from all three marine mammal species was calculated to provide greater than 95% of lipid intake for the great white shark from the tissues analyzed. Sampling of prey blubber may give a good indication of the lipids provided to the shark predator.

Psychrophiles and astrobiology: microbial life of frozen worlds

NASA Astrophysics Data System (ADS)

Pikuta, Elena V.; Hoover, Richard B.

2003-01-01

Most bodies of our Solar System are "Frozen Worlds" where the prevailing surface temperature remains at or below freezing. On Earth there are vast permanently frozen regions of permafrost, polar ice sheets, and glaciers and the deep oceans and deep-sea marine sediments have remained at 2 - 4°C for eons. Psychrophilic and psychrotrophic microbiota that inhabit these regimes provide analogs for microbial life that might inhabit ice sheets and permafrost of Mars, comets, or the ice/water interfaces or sediments deep beneath the icy crusts of Europa, Callisto, or Ganymede. Cryopreserved micro-organisms can remain viable (in a deep anabiotic state) for millions of years frozen in permafrost and ice. Psychrophilic and psychrotrophic (cold-loving) microbes can carry out metabolic processes in water films and brine, acidic, or alkaline chanels in permafrost or ice at temperatures far below 0°C. These microbes of the cryosphere help define the thermal and temporal limits of life on Earth and may provide clues to where and how to search for evidence of life elsewhere in the Cosmos. Astrobiologists at the NASA Marshall Space Flight Center have collected microbial extremophiles from the Pleistocene ice wedges and frozen thermokarst ponds from the Fox Permafrost Tunnel of Alaska. Microbes have also been isolated from samples of Magellanic Penguin guano from Patagonia; deep-sea marine muds near hydrothermal vents; snow and permafrost from Siberia, and deep ice cores, ice-bubble and cryoconite rocks of the Central Antarctic Ice Sheet. These samples have yielded microbial extremophiles representing a wide variety of anaerobic bacteria and archaea. These microbes have been isolated, cultured, characterized and analyzed by phylogenetic and genomic methods. Images were obtained by Phase Contrast, Environmental, Field Emission Scanning and Transmission Electron Microscopes to study the ultra-microstructure and elemental distribution in the composition of these micro-organisms. We consider the Astrobiological significance of the Fox Tunnel with its rich assemblage of frozen microbes as proxy for developing techniques that may help optimize the search for evidence of life in the permafrost of Mars. We provide images of a novel anaerobic, heterotrophic, psychrotrophic bacterium (str.FTR1) isolated in pure culture from the Fox Tunnel. We also describe novel psychrotrophs isolated from guano of the Magellanic penguin (Spheniscus magellanicus) from the southern tip of Patagonia. These strains PmagG1 and PPP2) represent new species and genera of anaerobic microbes that grow at very low temperatures. The lowest limit for growth without morphological changes of str.PmagG1 is -4°C.

Automated cell disruption is a reliable and effective method of isolating RNA from fresh snap-frozen normal and malignant oral mucosa samples.

PubMed

Van der Vorst, Sébastien; Dekairelle, Anne-France; Irenge, Léonid; Hamoir, Marc; Robert, Annie; Gala, Jean-Luc

2009-01-01

This study compared automated vs. manual tissue grinding in terms of RNA yield obtained from oral mucosa biopsies. A total of 20 patients undergoing uvulectomy for sleep-related disorders and 10 patients undergoing biopsy for head and neck squamous cell carcinoma were enrolled in the study. Samples were collected, snap-frozen in liquid nitrogen, and divided into two parts of similar weight. Sample grinding was performed on one sample from each pair, either manually or using an automated cell disruptor. The performance and efficacy of each homogenization approach was compared in terms of total RNA yield (spectrophotometry, fluorometry), mRNA quantity [densitometry of specific TP53 amplicons and TP53 quantitative reverse-transcribed real-time PCR (qRT-PCR)], and mRNA quality (functional analysis of separated alleles in yeast). Although spectrophotometry and fluorometry results were comparable for both homogenization methods, TP53 expression values obtained by amplicon densitometry and qRT-PCR were significantly and consistently better after automated homogenization (p<0.005) for both uvula and tumor samples. Functional analysis of separated alleles in yeast results was better with the automated technique for tumor samples. Automated tissue homogenization appears to be a versatile, quick, and reliable method of cell disruption and is especially useful in the case of small malignant samples, which show unreliable results when processed by manual homogenization.

Effect of Repeated Freezing and Thawing on 18 Clinical Chemistry Analytes in Rat Serum

PubMed Central

Kale, Vijay P; Patel, Sweta G; Gunjal, Prashant S; Wakchaure, Santosh U; Sundar, Rajesh S; Ranvir, Ramchandra K; Jain, Mukul R

2012-01-01

In a preclinical research laboratory, using serum samples that have been frozen and thawed repeatedly is sometimes unavoidable when needing to confirm previous results or perform additional analysis. Here we determined the effects of multiple cycles of refrigeration or freezing and thawing of rat serum at 3 temperature conditions for different storage times on clinical chemistry analytes. Serum samples obtained from adult Wistar rats were stored at 2 to 8 °C and −10 to −20 °C for as long as 72 h and at −70 °C for as long as 30 d. At different time points (24, 48, and 72 h for samples stored at 2 to 8 °C or −10 to −20 °C and 1, 7, and 30 d for samples stored at −70 °C), the samples were brought to room temperature, analyzed, and then stored again at the designated temperature. The results obtained after each storage cycle were compared with those obtained from the initial analysis of fresh samples. Of the 18 serum analytes evaluated, 14 were stable without significant changes, even after 3 freeze–thaw cycles at the tested temperature ranges. Results from this study will help researchers working with rat serum to interpret the biochemical data obtained from serum samples that have been frozen and thawed repeatedly. PMID:23043814

Transjugular Liver Biopsy

PubMed Central

Behrens, George; Ferral, Hector

2012-01-01

Liver biopsy is considered the gold standard for the evaluation of acute and chronic liver disorders. Transjugular liver biopsy (TJLB) was described by Dotter in 1964 and clinically performed for the first time by Hanafee in 1967. TJLB consists of obtaining liver tissue through a rigid cannula introduced into one of the hepatic veins, typically using jugular venous access. The quality of the TJLB specimens has improved so much that the samples obtained by this method are comparable with those obtained with the percutaneous technique. TJLB is indicated for patients with coagulopathy, ascites, peliosis hepatis, morbid obesity, liver transplant, or in patients undergoing a transjugular intrahepatic portosystemic shunt procedure. The technical success rate for a TJLB procedure ranges from 87 to 97%. Sample fragmentation has been reported in 14 to 25% of the TJLB samples. The complication rates are low and range between 1.3% and 6.5%. The purpose of this article is to provide a review of the fundamental aspects of the TJLB procedure, including technique, indications, contraindications, results, and complications. PMID:23729981

Unfolding epidemiological stories: how the WHO made frozen blood into a flexible resource for the future.

PubMed

Radin, Joanna

2014-09-01

In the decades after World War II, the World Health Organization (WHO) played an important role in managing the process of stabilizing collections of variable blood samples as a fundamentally unstable, protean, and unfolding biomedical resource. In this system, known and as yet unknown constituents of blood were positioned as relevant to the work of multiple constituencies including human population geneticists, physical anthropologists, and immunologists. To facilitate serving these and other constituencies, it was crucial to standardize practices of collecting and preserving samples of blood from globally distributed human populations. The WHO achieved this by linking its administrative infrastructure-comprised of expert advisory groups and technical reports-to key laboratories, which served as sites for demonstrating and also for disseminating standards for working with variable blood samples. The practices that were articulated in making blood samples into a flexible resource contributes to emerging histories of global health that highlight the centrality of new institutions, like the WHO, new forms of expertise, like population genetics and serological epidemiology, and new kinds of research materials, like frozen blood. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cryopreservation of human sperm: efficacy and use of a new nitrogen-free controlled rate freezer versus liquid nitrogen vapour freezing.

PubMed

Creemers, E; Nijs, M; Vanheusden, E; Ombelet, W

2011-12-01

Preservation of spermatozoa is an important aspect of assisted reproductive medicine. The aim of this study was to investigate the efficacy and use of a recently developed liquid nitrogen and cryogen-free controlled rate freezer and this compared with the classical liquid nitrogen vapour freezing method for the cryopreservation of human spermatozoa. Ten patients entering the IVF programme donated semen samples for the study. Samples were analysed according to the World Health Organization guidelines. No significant difference in total sperm motility after freeze-thawing between the new technique and classical technique was demonstrated. The advantage of the new freezing technique is that it uses no liquid nitrogen during the freezing process, hence being safer to use and clean room compatible. Investment costs are higher for the apparatus but running costs are only 1% in comparison with classical liquid nitrogen freezing. In conclusion, post-thaw motility of samples frozen with the classical liquid nitrogen vapour technique was comparable with samples frozen with the new nitrogen-free freezing technique. This latter technique can thus be a very useful asset to the sperm cryopreservation laboratory. © 2011 Blackwell Verlag GmbH.

A Comparison of RNA-Seq Results from Paired Formalin-Fixed Paraffin-Embedded and Fresh-Frozen Glioblastoma Tissue Samples

PubMed Central

Esteve-Codina, Anna; Arpi, Oriol; Martinez-García, Maria; Pineda, Estela; Mallo, Mar; Gut, Marta; Carrato, Cristina; Rovira, Anna; Lopez, Raquel; Tortosa, Avelina; Dabad, Marc; Del Barco, Sonia; Heath, Simon; Bagué, Silvia; Ribalta, Teresa; Alameda, Francesc; de la Iglesia, Nuria

2017-01-01

The molecular classification of glioblastoma (GBM) based on gene expression might better explain outcome and response to treatment than clinical factors. Whole transcriptome sequencing using next-generation sequencing platforms is rapidly becoming accepted as a tool for measuring gene expression for both research and clinical use. Fresh frozen (FF) tissue specimens of GBM are difficult to obtain since tumor tissue obtained at surgery is often scarce and necrotic and diagnosis is prioritized over freezing. After diagnosis, leftover tissue is usually stored as formalin-fixed paraffin-embedded (FFPE) tissue. However, RNA from FFPE tissues is usually degraded, which could hamper gene expression analysis. We compared RNA-Seq data obtained from matched pairs of FF and FFPE GBM specimens. Only three FFPE out of eleven FFPE-FF matched samples yielded informative results. Several quality-control measurements showed that RNA from FFPE samples was highly degraded but maintained transcriptomic similarities to RNA from FF samples. Certain issues regarding mutation analysis and subtype prediction were detected. Nevertheless, our results suggest that RNA-Seq of FFPE GBM specimens provides reliable gene expression data that can be used in molecular studies of GBM if the RNA is sufficiently preserved. PMID:28122052

Study on the contribution of cryosphere to runoff in the cold alpine basin: A case study of Hulugou River Basin in the Qilian Mountains

NASA Astrophysics Data System (ADS)

Zongxing, Li; Qi, Feng; Wei, Liu; Tingting, Wang; Aifang, Cheng; Yan, Gao; Xiaoyan, Guo; Yanhui, Pan; Jianguo, Li; Rui, Guo; Bing, Jia

2014-11-01

Global warming would inevitably lead to the increased glacier-snow meltwater and mountainous discharge. Taking an example the Hulugou River Basin in the Qilian Mountains, this study confirmed the contribution of cryosphere to runoff by means of the isotope hydrograph separation. The hydro-geochemistry and the isotope geochemistry suggested that both the meltwater and rainwater infiltrated into the subsurface and fed into the river runoff of the Hulugou River Basin in the form of springs. The isotopic composition of river water and underground water was close to the Local Meteoric Water Line, and the δ18O and δD ranged among precipitation, glacier-snow meltwater and frozen soil meltwater. The results indicated that 68% of the recharge of the Hulugou River water was the precipitation, thereinto, glacier-snow meltwater and frozen soil meltwater contributing 11% and 21%, respectively. For tributary-1, precipitation accounted for 77% of the total stream runoff, with frozen soil meltwater accounting for 17%, and glacier-snow meltwater only supplied 6%. During the sampling period, the contribution of surface runoff from precipitation was 44% to tributary-2, and glacier-snow meltwater had contributed 42%; only 14% from frozen soil meltwater. For tributary-3, precipitation accounted for 63% of the total runoff, and other 37% originated from the frozen soil meltwater. According to the latest observational data, the glacier-snow meltwater has accounted for 11.36% of the total runoff in the stream outlet, in which the calculation has been verified by hydrograph separation. It is obvious that the contribution of cryosphere has accounted for 1/3 of the outlet runoff in the Hulugou River Basin, which has been an important part of river sources. This study demonstrated that the alpine regions of western China, especially those basins with glaciers, snow and frozen soil, have played a crucial role in regional water resource provision under global warming.

Quantification of liver fibrosis via second harmonic imaging of the Glisson's capsule from liver surface.

PubMed

Xu, Shuoyu; Kang, Chiang Huen; Gou, Xiaoli; Peng, Qiwen; Yan, Jie; Zhuo, Shuangmu; Cheng, Chee Leong; He, Yuting; Kang, Yuzhan; Xia, Wuzheng; So, Peter T C; Welsch, Roy; Rajapakse, Jagath C; Yu, Hanry

2016-04-01

Liver surface is covered by a collagenous layer called the Glisson's capsule. The structure of the Glisson's capsule is barely seen in the biopsy samples for histology assessment, thus the changes of the collagen network from the Glisson's capsule during the liver disease progression are not well studied. In this report, we investigated whether non-linear optical imaging of the Glisson's capsule at liver surface would yield sufficient information to allow quantitative staging of liver fibrosis. In contrast to conventional tissue sections whereby tissues are cut perpendicular to the liver surface and interior information from the liver biopsy samples were used, we have established a capsule index based on significant parameters extracted from the second harmonic generation (SHG) microscopy images of capsule collagen from anterior surface of rat livers. Thioacetamide (TAA) induced liver fibrosis animal models was used in this study. The capsule index is capable of differentiating different fibrosis stages, with area under receiver operating characteristics curve (AUC) up to 0.91, making it possible to quantitatively stage liver fibrosis via liver surface imaging potentially with endomicroscopy. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High and variable copper status identified among dairy herds in the Waikato region by concentrations of Cu in liver sourced from biopsies and cull cows.

PubMed

Grace, N D; Knowles, S O; Hittmann, A R

2010-06-01

To document the Cu supplementation practices on dairy farms in the Waikato region, determine the Cu status of those herds, and compare the suitability of liver samples sourced from biopsies and cull cows for assessing Cu status. During spring 2008, concentrations of Cu, Mo and S were determined from pasture samples from 24 dairy farms. Feeding regimens, herd size, milksolids production, soil type, fertiliser policy and Cu supplementation practices were recorded for each property. Based on these data, 10 monitor farms were selected to represent a range of Cu intakes for herds, from 5 to 12 mg Cu/kg dry matter (DM). On each monitor farm 12 healthy lactating cows were selected for liver biopsy and collection of blood samples during the following autumn. Around the same time, livers were collected from 12 cull cows per farm when they were slaughtered, and samples of pasture were again collected from each farm. Concentrations of Cu were measured in all tissue samples. Concentrations of Cu in pasture tended to be higher (mean 10.4 vs 8.2 mg/kg DM) in the autumn than spring, while concentrations of Mo were lower in the autumn (mean 0.35 vs 1.07 mg/kg DM). Most of the 24 farms used Cu supplementation in some form. Mean concentrations of Cu in liver for herds ranged from 640 (SD 544) to 2,560 (SD 474) micromol/kg fresh tissue in biopsies, and 520 (SD 235) to 2,610 (SD 945) micromol/kg in liver from cull cows. Mean concentrations of Cu in serum ranged from 7.9 to 13.4 micromol/L. The variability in concentrations of Cu for each farm was greater for liver (CV 50%) than serum (CV 21%). For individual cows, concentrations of Cu in liver, obtained by biopsy, and serum were not correlated. The concentration of Cu in liver of dairy cows reflected widely differing dietary intakes of Cu between herds, although levels indicated an adequate Cu status on all farms in this study. Use of either biopsy samples or livers from cull cows were indicative of the Cu status of the herd. Wide variation in observed concentrations of Cu in liver indicated that at least 12 cows per herd should be sampled. On farms with intensive, long-term Cu supplementation programmes there is a risk of chronic Cu toxicity in some animals. Thus, the Cu status of dairy herds should be determined, and monitored, before making any recommendations regarding supplementation.

Inhibition of lipid oxidation in frozen farmed ovate pompano (Trachinotus ovatus L.) fillets stored at -18 °C by chitosan coating incorporated with citric acid or licorice extract.

PubMed

Qiu, Xujian; Chen, Shengjun; Liu, Guangming; Lin, Hong

2016-08-01

Lipid oxidation can occur in fish fillets during long-term frozen storage and cause quality and nutrition loss, which is a major concern in the seafood industry. Our previous study showed that chitosan combined with citric acid or licorice extract can have a preserving effect on fresh fish fillets stored at 4 °C. It is of interest to further study their antioxidant effects on fish fillets during frozen storage. Chitosan, chitosan and citric acid, chitosan and licorice extract can inhibit primary and secondary lipid oxidation, as indicated by lower peroxide value (PV) and thiobarbituric acid reactive substances (TBARS) values compared to the control samples. In addition, drip loss was decreased in the treatment samples. Both citric acid and licorice extract enhanced the antioxidant effects of chitosan. Among all the three treatments, chitosan and licorice extract showed the best antioxidant effects, reducing both PV and TBARS significantly at the end of storage. The combination of chitosan and citric acid or licorice extract showed significant antioxidant effects on ovate pompano fillets at -18 °C during 6 months of storage. They could be applied as natural antioxidant preservatives for use in seafood products or other meat products. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Preoperative Thromboelastometry as a Predictor of Transfusion Requirements during Adult Living Donor Liver Transplantation.

PubMed

Fayed, Nirmeen; Mourad, Wessam; Yassen, Khaled; Görlinger, Klaus

2015-03-01

The ability to predict transfusion requirements may improve perioperative bleeding management as an integral part of a patient blood management program. Therefore, the aim of our study was to evaluate preoperative thromboelastometry as a predictor of transfusion requirements for adult living donor liver transplant recipients. The correlation between preoperative thromboelastometry variables in 100 adult living donor liver transplant recipients and intraoperative blood transfusion requirements was examined by univariate and multivariate linear regression analysis. Thresholds of thromboelastometric parameters for prediction of packed red blood cells (PRBCs), fresh frozen plasma (FFP), platelets, and cryoprecipitate transfusion requirements were determined with receiver operating characteristics analysis. The attending anesthetists were blinded to the preoperative thromboelastometric analysis. However, a thromboelastometry-guided transfusion algorithm with predefined trigger values was used intraoperatively. The transfusion triggers in this algorithm did not change during the study period. Univariate analysis confirmed significant correlations between PRBCs, FFP, platelets or cryoprecipitate transfusion requirements and most thromboelastometric variables. Backward stepwise logistic regression indicated that EXTEM coagulation time (CT), maximum clot firmness (MCF) and INTEM CT, clot formation time (CFT) and MCF are independent predictors for PRBC transfusion. EXTEM CT, CFT and FIBTEM MCF are independent predictors for FFP transfusion. Only EXTEM and INTEM MCF were independent predictors of platelet transfusion. EXTEM CFT and MCF, INTEM CT, CFT and MCF as well as FIBTEM MCF are independent predictors for cryoprecipitate transfusion. Thromboelastometry-based regression equation accounted for 63% of PRBC, 83% of FFP, 61% of cryoprecipitate, and 44% of platelet transfusion requirements. Preoperative thromboelastometric analysis is helpful to predict transfusion requirements in adult living donor liver transplant recipients. This may allow for better preparation and less cross-matching prior to surgery. The findings of our study need to be re-validated in a second prospective patient population.

21 CFR 160.110 - Frozen eggs.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 2 2010-04-01 2010-04-01 false Frozen eggs. 160.110 Section 160.110 Food and... CONSUMPTION EGGS AND EGG PRODUCTS Requirements for Specific Standardized Eggs and Egg Products § 160.110 Frozen eggs. (a) Frozen eggs, frozen whole eggs, frozen mixed eggs is the food prepared by freezing...

21 CFR 160.110 - Frozen eggs.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 2 2011-04-01 2011-04-01 false Frozen eggs. 160.110 Section 160.110 Food and... CONSUMPTION EGGS AND EGG PRODUCTS Requirements for Specific Standardized Eggs and Egg Products § 160.110 Frozen eggs. (a) Frozen eggs, frozen whole eggs, frozen mixed eggs is the food prepared by freezing...

Isolation of circulating tumor cells from pancreatic cancer by automated filtration

PubMed Central

Brychta, Nora; Drosch, Michael; Driemel, Christiane; Fischer, Johannes C.; Neves, Rui P.; Esposito, Irene; Knoefel, Wolfram; Möhlendick, Birte; Hille, Claudia; Stresemann, Antje; Krahn, Thomas; Kassack, Matthias U.; Stoecklein, Nikolas H.; von Ahsen, Oliver

2017-01-01

It is now widely recognized that the isolation of circulating tumor cells based on cell surface markers might be hindered by variability in their protein expression. Especially in pancreatic cancer, isolation based only on EpCAM expression has produced very diverse results. Methods that are independent of surface markers and therefore independent of phenotypical changes in the circulating cells might increase CTC recovery also in pancreatic cancer. We compared an EpCAM-dependent (IsoFlux) and a size-dependent (automated Siemens Healthineers filtration device) isolation method for the enrichment of pancreatic cancer CTCs. The recovery rate of the filtration based approach is dramatically superior to the EpCAM-dependent approach especially for cells with low EpCAM-expression (filtration: 52%, EpCAM-dependent: 1%). As storage and shipment of clinical samples is important for centralized analyses, we also evaluated the use of frozen diagnostic leukapheresis (DLA) as source for isolating CTCs and subsequent genetic analysis such as KRAS mutation detection analysis. Using frozen DLA samples of pancreatic cancer patients we detected CTCs in 42% of the samples by automated filtration. PMID:29156783

Isolation of circulating tumor cells from pancreatic cancer by automated filtration.

PubMed

Brychta, Nora; Drosch, Michael; Driemel, Christiane; Fischer, Johannes C; Neves, Rui P; Esposito, Irene; Knoefel, Wolfram; Möhlendick, Birte; Hille, Claudia; Stresemann, Antje; Krahn, Thomas; Kassack, Matthias U; Stoecklein, Nikolas H; von Ahsen, Oliver

2017-10-17

It is now widely recognized that the isolation of circulating tumor cells based on cell surface markers might be hindered by variability in their protein expression. Especially in pancreatic cancer, isolation based only on EpCAM expression has produced very diverse results. Methods that are independent of surface markers and therefore independent of phenotypical changes in the circulating cells might increase CTC recovery also in pancreatic cancer. We compared an EpCAM-dependent (IsoFlux) and a size-dependent (automated Siemens Healthineers filtration device) isolation method for the enrichment of pancreatic cancer CTCs. The recovery rate of the filtration based approach is dramatically superior to the EpCAM-dependent approach especially for cells with low EpCAM-expression (filtration: 52%, EpCAM-dependent: 1%). As storage and shipment of clinical samples is important for centralized analyses, we also evaluated the use of frozen diagnostic leukapheresis (DLA) as source for isolating CTCs and subsequent genetic analysis such as KRAS mutation detection analysis. Using frozen DLA samples of pancreatic cancer patients we detected CTCs in 42% of the samples by automated filtration.

Microbiological and nutritional quality of the goat meat by-product "sarapatel".

PubMed

Brasil, Luciana; Queiroz, Angela; Silva, Josevan; Bezerra, Taliana; Arcanjo, Narciza; Magnani, Marciane; Souza, Evandro; Madruga, Marta

2014-01-16

Goat "sarapatel" is a product made from blood and viscera. For the first time, the microbiological and nutritional quality of "sarapatel" samples (n=48) sold under different conditions (in street markets, butcher shops, and supermarkets under refrigeration, frozen or at room temperature) was evaluated. Goat "sarapatel" is a nutritive food, with each 100 g providing, on average, 72 g of moisture, 2 g of ash, 18 g of protein, 9 g of lipids, 2 g of carbohydrates, 282 mg of cholesterol, and high amounts of unsaturated fatty acids and essential amino acids. The analysis of the "sarapatel" samples shows that none of them contain Salmonella spp. or L. monocytogenes. High counts (>104) of total coliforms, thermotolerant coliforms, and sulfite-reducing Clostridium were detected, and coagulase-positive Staphylococcus was found in 31.25% of samples. The storage conditions evaluated (refrigeration, frozen or at room temperature) did not affect the physicochemical quality of the "sarapatel"; however, the unsatisfactory microbiological quality indicates that it is necessary to improve the health-sanitary aspects of the processing and sale of this product.

Solid state nuclear magnetic resonance with magic-angle spinning and dynamic nuclear polarization below 25 K

PubMed Central

Thurber, Kent R.; Potapov, Alexey; Yau, Wai-Ming; Tycko, Robert

2012-01-01

We describe an apparatus for solid state nuclear magnetic resonance (NMR) with dynamic nuclear polarization (DNP) and magic-angle spinning (MAS) at 20–25 K and 9.4 Tesla. The MAS NMR probe uses helium to cool the sample space and nitrogen gas for MAS drive and bearings, as described earlier (Thurber et al., J. Magn. Reson. 2008) [1], but also includes a corrugated waveguide for transmission of microwaves from below the probe to the sample. With a 30 mW circularly polarized microwave source at 264 GHz, MAS at 6.8 kHz, and 21 K sample temperature, greater than 25-fold enhancements of cross-polarized 13C NMR signals are observed in spectra of frozen glycerol/water solutions containing the triradical dopant DOTOPA-TEMPO when microwaves are applied. As demonstrations, we present DNP-enhanced one-dimensional and two-dimensional 13C MAS NMR spectra of frozen solutions of uniformly 13C-labeled L-alanine and melittin, a 26-residue helical peptide that we have synthesized with four uniformly 13C-labeled amino acids. PMID:23238592

The influence of normal and high ultimate muscle pH on the microbiology and colour stability of previously frozen black wildebeest (Connochaetes gnou) meat.

PubMed

Shange, Nompumelelo; Makasi, Thandeka N; Gouws, Pieter A; Hoffman, Louwrens C

2018-01-01

Changes in pH, colour and microbiological counts were investigated in previously frozen Biceps femoris (BF) muscles from black wildebeest. Samples were stored under vacuum at refrigerated conditions (4.2±0.8°C) for 12days. Seven BF muscles had a high pH (DFD) (pH≥6) and five had a normal pH (pH<6). Overtime the pH of DFD did not significantly change whilst that of normal pH meat decreased. Browning under anaerobic storage conditions was seen, more for normal meat than DFD meat. Initial total viable counts, lactic acid bacteria and coliform counts from samples with normal pH, were significantly higher than counts from the DFD samples. However, overtime DFD meat showed a faster increase for all microorganisms tested compared to normal pH meat. Overall, this study revealed that DFD meat can have a shorter shelf-life than normal pH meat stored at 4.2±0.8°C. Copyright © 2017 Elsevier Ltd. All rights reserved.

Solid state nuclear magnetic resonance with magic-angle spinning and dynamic nuclear polarization below 25 K.

PubMed

Thurber, Kent R; Potapov, Alexey; Yau, Wai-Ming; Tycko, Robert

2013-01-01

We describe an apparatus for solid state nuclear magnetic resonance (NMR) with dynamic nuclear polarization (DNP) and magic-angle spinning (MAS) at 20-25 K and 9.4 Tesla. The MAS NMR probe uses helium to cool the sample space and nitrogen gas for MAS drive and bearings, as described earlier, but also includes a corrugated waveguide for transmission of microwaves from below the probe to the sample. With a 30 mW circularly polarized microwave source at 264 GHz, MAS at 6.8 kHz, and 21 K sample temperature, greater than 25-fold enhancements of cross-polarized (13)C NMR signals are observed in spectra of frozen glycerol/water solutions containing the triradical dopant DOTOPA-TEMPO when microwaves are applied. As demonstrations, we present DNP-enhanced one-dimensional and two-dimensional (13)C MAS NMR spectra of frozen solutions of uniformly (13)C-labeled l-alanine and melittin, a 26-residue helical peptide that we have synthesized with four uniformly (13)C-labeled amino acids. Published by Elsevier Inc.

MGMT promoter methylation in Peruvian patients with glioblastoma

PubMed Central

Belmar-Lopez, Carolina; Castaneda, Carlos A; Castillo, Miluska; García-Corrochano, Pamela; Orrego, Enrique; Meléndez, Barbara; Casavilca, Sandro; Flores, Claudio; Orrego, Enrique

2018-01-01

Purpose O6-methylguanine–DNA methyltransferase (MGMT) promoter methylation predicts the outcome and response to alkylating chemotherapy in glioblastoma. The aim of this study is to evaluate the prevalence of MGMT methylation in Peruvian glioblastoma cases. Patients and methods We evaluated retrospectively 50 cases of resected glioblastoma during the period 2008–2013 at Instituto Nacional de Enfermedades Neoplasicas in Peru. Samples consisted of paraffin embedded and frozen tumour tissue. MGMT-promoter methylation status and the expression level of MGMT gene were evaluated by methylation-specific PCR and real-time PCR, respectively. Results Unmethylated, methylated and partially methylated statuses were found in 54%, 20% and 26% of paraffin-embedded samples, respectively. Methylation status was confirmed in the Virgen de la Salud Hospital and frozen samples. There was an association between the status of MGMT-promoter methylation and the level of gene expression (p = 0.001). Methylation was associated with increased progression-free survival (p = 0.002) and overall survival (OS) (p < 0.001). Conclusion MGMT-promoter methylation frequency in Peruvian glioblastoma is similar to that reported in other populations and the detection test has been standardised. PMID:29515653

Cold-stage microscopy system for fast-frozen liquids.

PubMed

Talmon, Y; Davis, H T; Scriven, L E; Thomas, E L

1979-06-01

The least artifact-laden fixation technique for examining colloidal suspensions, microemulsions, and other microstructured liquids in the electron microscope appears to be thermal fixation, i.e., ultrafast freezing of the liquid specimen. For rapid-enough cooling and for observation in TEM/STEM a thin sample is needed. The need is met by trapping a thin layer ( approximately 100 nm) of liquid between two polyimide films ( approximately 40 nm thickness) mounted on copper grids and immersing the resulting sandwich in liquid nitrogen at its melting point. For liquids containing water, polyimides films are used since this polymer is far less susceptible to the electron beam damage observed for the commonly used polymer films such as Formvar and collodion in contact with ice. Transfer of the frozen sample into the microscope column without deleterious frost deposition and warming is accomplished with a new transfer module for the cooling stage of the JEOL JEM-100CX microscope, which makes a true cold stage out of a device originally intended for cooling specimens inside the column. Sample results obtained with the new fast-freeze, cold-stage microscopy system are given.

Characterization of HIFU ablation using DNA fragmentation labeling as apoptosis stain

NASA Astrophysics Data System (ADS)

Anquez, Jeremie; Corréas, Jean-Michel; Pau, Bernard; Lacoste, François; Yon, Sylvain

2012-11-01

The goal of this work was to compare modalities to precisely quantify the extent of thermally induced lesions: gross pathology vs. histopathology vs. devascularization. Liver areas of 14 rabbits were targeted with HIFU and RF ablations in an acute study. Contrast enhanced computorized tomography (CE-CT) scan images were acquired two hours after HIFU and RF treatment to obtain the devascularized volumes of the livers. The animals were then euthanized and deep frozen. The livers were sliced and each slice was photographed and stacked yielding a volume of gross pathology. The volume VGP of the HIFU lesions were derived. The area AGP of the lesions were computed on a particular slice. The lesions were segmented as hypo intense (devascularized) regions on CE-CT images and their volumes VC were computed. The ratios VC/VGP were computed for all the HIFU lesions on all the 14 subjects with a mean value of 1.2. Histology was performed on the livers using Hematoxyline Eosine Staining (HES) and DNA Fragmentation labeling (TUNEL® technology) which characterizes apoptosis. Apoptotic regions of area AT were segmented on the images stained by TUNEL®. No necrosis was identified on the HES data. While TUNEL® did not mark the cores of the RF lesions as apoptotic, the periphery of HIFU and RF lesions was always recognized with TUNEL® as apoptotic. The ratio AGP/AT was computed. The mean value was 0.95 and 0.25 for HIFU and RF lesions respectively. These findings show that the devascularized territory seen on CE-CT scan coincide with the coagulated territories seen with gross pathology. Those actually correspond to cells in apoptosis. It is confirmed that HES stain does not show necrosis 2 hours after thermal ablation. TUNEL® technology for DNA fragmentation labeling appears as a useful marker for thermally induced acute lesions in the liver.

Metabolic profiling of ob/ob mouse fatty liver using HR-MAS 1H-NMR combined with gene expression analysis reveals alterations in betaine metabolism and the transsulfuration pathway.

PubMed

Gogiashvili, Mikheil; Edlund, Karolina; Gianmoena, Kathrin; Marchan, Rosemarie; Brik, Alexander; Andersson, Jan T; Lambert, Jörg; Madjar, Katrin; Hellwig, Birte; Rahnenführer, Jörg; Hengstler, Jan G; Hergenröder, Roland; Cadenas, Cristina

2017-02-01

Metabolic perturbations resulting from excessive hepatic fat accumulation are poorly understood. Thus, in this study, leptin-deficient ob/ob mice, a mouse model of fatty liver disease, were used to investigate metabolic alterations in more detail. Metabolites were quantified in intact liver tissues of ob/ob (n = 8) and control (n = 8) mice using high-resolution magic angle spinning (HR-MAS) 1 H-NMR. In addition, after demonstrating that HR-MAS 1 H-NMR does not affect RNA integrity, transcriptional changes were measured by quantitative real-time PCR on RNA extracted from the same specimens after HR-MAS 1 H-NMR measurements. Importantly, the gene expression changes obtained agreed with those observed by Affymetrix microarray analysis performed on RNA isolated directly from fresh-frozen tissue. In total, 40 metabolites could be assigned in the spectra and subsequently quantified. Quantification of lactate was also possible after applying a lactate-editing pulse sequence that suppresses the lipid signal, which superimposes the lactate methyl resonance at 1.3 ppm. Significant differences were detected for creatinine, glutamate, glycine, glycolate, trimethylamine-N-oxide, dimethylglycine, ADP, AMP, betaine, phenylalanine, and uridine. Furthermore, alterations in one-carbon metabolism, supported by both metabolic and transcriptional changes, were observed. These included reduced demethylation of betaine to dimethylglycine and the reduced expression of genes coding for transsulfuration pathway enzymes, which appears to preserve methionine levels, but may limit glutathione synthesis. Overall, the combined approach is advantageous as it identifies changes not only at the single gene or metabolite level but also deregulated pathways, thus providing critical insight into changes accompanying fatty liver disease. Graphical abstract A Evaluation of RNA integrity before and after HR-MAS 1 H-NMR of intact mouse liver tissue. B Metabolite concentrations and gene expression levels assessed in ob/ob (steatotic) and ob/+ (control) mice using HR-MAS 1 H-NMR and qRT-PCR, respectively.

RAB GTPASES ASSOCIATE WITH ISOLATED LIPID DROPLETS (LDS) AND SHOW ALTERED CONTENT AFTER ETHANOL ADMINISTRATION: POTENTIAL ROLE IN ALCOHOL-IMPAIRED LD METABOLISM

PubMed Central

Rasineni, Karuna; McVicker, Benita L.; Tuma, Dean J.; McNiven, Mark A.; Casey, Carol A.

2013-01-01

Background Alcoholic liver disease is manifested by the presence of fatty liver, primarily due to accumulation of hepatocellular lipid droplets (LDs). The presence of membrane-trafficking proteins (e.g. Rab GTPases) with LDs indicates that LDs may be involved in trafficking pathways known to be altered in ethanol damaged hepatocytes. Since these Rab GTPases are crucial regulators of protein trafficking, we examined the effect ethanol administration has on hepatic Rab protein content and association with LDs. Methods Male Wistar rats were pair-fed Lieber-DeCarli diets for 5 to 8 weeks. Whole liver and isolated LD fractions were analyzed. Identification of LDs and associated Rab proteins was performed in frozen liver or paraffin-embedded sections followed by immunohistochemical analysis. Results Lipid accumulation was characterized by larger LD vacuoles and increased total triglyceride content in ethanol-fed rats. Rabs 1, 2, 3d, 5, 7 and 18 were analyzed in post-nuclear supernatant (PNS) as well as LDs. All of the Rabs were found in the PNS, and Rabs 1, 2, 5 and 7 did not show alcohol-altered content, while Rab 3d content was reduced by over 80%, and Rab 18 also showed ethanol-induced reduction in content. Rab 3d was not found to associate with LDs, while all other Rabs were found in the LD fractions, and several showed an ethanol-related decrease (Rabs 2, 5, 7, 18). Immunohistochemical analysis revealed the enhanced content of a LD-associated protein, perilipin 2 (PLIN2) that was paralleled with an associated decrease of Rab 18 in ethanol-fed rat sections. Conclusion Chronic ethanol feeding was associated with increased PLIN2 and altered Rab GTPase content in enriched LD fractions. Although mechanisms driving these changes are not established, further studies on intracellular protein trafficking and LD biology after alcohol administration will likely contribute to our understanding of fatty liver disease. PMID:24117505

Successful Total En Bloc Spondylectomy of T7 Vertebra for Hepatocellular Carcinoma Metastasis After Living Donor Liver Transplantation.

PubMed

Kimura, Hiroaki; Fujibayashi, Shunsuke; Shimizu, Takayoshi; Otsuki, Bungo; Murakami, Hideki; Kaido, Toshimi; Uemoto, Shinji; Matsuda, Shuichi

2015-08-15

Case report. We report a patient who was successfully treated with total en bloc spondylectomy (TES) for T7 metastasis after living donor liver transplantation for hepatocellular carcinoma (HCC). Spinal metastasis from HCC has a poor prognosis. There are only a few studies on surgical outcomes of spinal metastasis from HCC. Because of the high surgical morbidity and short life expectancy in patients with HCC with spinal metastasis, TES is not considered in these patients, although several studies have reported satisfactory results for TES for some types of metastatic spinal tumors. Liver transplantation (LT) is the curative treatment option for early HCC. However, the recurrence of HCC is a possible problem after LT, although no reports on surgery for spinal metastasis following LT for HCC have been published. We report on the first case of a patient who was successfully treated with TES for T7 metastasis after living donor LT for HCC. The patient was a 65-year-old man, who had undergone living donor LT for HCC 2 years before. His main symptom was progressive gait disturbance because of the spinal cord compression by the tumor at T7. Radiology and pathology examinations revealed a solitary metastasis at T7 with neither recurrence in the liver nor metastasis in the other organs. We performed TES using a pedicle screw system and a mesh cage filled with frozen autografts. After surgery, the patient showed clear improvement in neurological symptoms. At 3 months after surgery, a T4 metastasis was detected with magnetic resonance imaging, and the patient was treated with heavy ion radiotherapy. He could walk without a cane and there was no evidence of recurrence at 1.5 years after surgery. Solitary spinal metastasis of HCC may become an indication for TES if liver function improves after LT. 5.

Photometric properties of Mars soils analogs

USGS Publications Warehouse

Pommerol, A.; Thomas, N.; Jost, B.; Beck, P.; Okubo, C.; McEwen, A.S.

2013-01-01

We have measured the bidirectional reflectance of analogs of dry, wet, and frozen Martian soils over a wide range of phase angles in the visible spectral range. All samples were produced from two geologic samples: the standard JSC Mars-1 soil simulant and Hawaiian basaltic sand. In a first step, experiments were conducted with the dry samples to investigate the effects of surface texture. Comparisons with results independently obtained by different teams with similar samples showed a satisfying reproducibility of the photometric measurements as well as a noticeable influence of surface textures resulting from different sample preparation procedures. In a second step, water was introduced to produce wet and frozen samples and their photometry investigated. Optical microscope images of the samples provided information about their microtexture. Liquid water, even in relatively low amount, resulted in the disappearance of the backscattering peak and the appearance of a forward-scattering peak whose intensity increases with the amount of water. Specular reflections only appeared when water was present in an amount large enough to allow water to form a film at the surface of the sample. Icy samples showed a wide variability of photometric properties depending on the physical properties of the water ice. We discuss the implications of these measurements in terms of the expected photometric behavior of the Martian surface, from equatorial to circum-polar regions. In particular, we propose some simple photometric criteria to improve the identification of wet and/or icy soils from multiple observations under different geometries.

A Method of Visualizing Three-Dimensional Distribution of Yeast in Bread Dough

NASA Astrophysics Data System (ADS)

Maeda, Tatsurou; Do, Gab-Soo; Sugiyama, Junichi; Oguchi, Kosei; Shiraga, Seizaburou; Ueda, Mitsuyoshi; Takeya, Koji; Endo, Shigeru

A novel technique was developed to monitor the change in three-dimensional (3D) distribution of yeast in frozen bread dough samples in accordance with the progress of mixing process. Application of a surface engineering technology allowed the identification of yeast in bread dough by bonding EGFP (Enhanced Green Fluorescent Protein) to the surface of yeast cells. The fluorescent yeast (a biomarker) was recognized as bright spots at the wavelength of 520 nm. A Micro-Slicer Image Processing System (MSIPS) with a fluorescence microscope was utilized to acquire cross-sectional images of frozen dough samples sliced at intervals of 1 μm. A set of successive two-dimensional images was reconstructed to analyze 3D distribution of yeast. Samples were taken from each of four normal mixing stages (i.e., pick up, clean up, development, and final stages) and also from over mixing stage. In the pick up stage yeast distribution was uneven with local areas of dense yeast. As the mixing progressed from clean up to final stages, the yeast became more evenly distributed throughout the dough sample. However, the uniformity in yeast distribution was lost in the over mixing stage possibly due to the breakdown of gluten structure within the dough sample.

Visualization and quantification of three-dimensional distribution of yeast in bread dough.

PubMed

Maeda, Tatsuro; DO, Gab-Soo; Sugiyama, Junichi; Araki, Tetsuya; Tsuta, Mizuki; Shiraga, Seizaburo; Ueda, Mitsuyoshi; Yamada, Masaharu; Takeya, Koji; Sagara, Yasuyuki

2009-07-01

A three-dimensional (3-D) bio-imaging technique was developed for visualizing and quantifying the 3-D distribution of yeast in frozen bread dough samples in accordance with the progress of the mixing process of the samples, applying cell-surface engineering to the surfaces of the yeast cells. The fluorescent yeast was recognized as bright spots at the wavelength of 520 nm. Frozen dough samples were sliced at intervals of 1 microm by an micro-slicer image processing system (MSIPS) equipped with a fluorescence microscope for acquiring cross-sectional images of the samples. A set of successive two-dimensional images was reconstructed to analyze the 3-D distribution of the yeast. The average shortest distance between centroids of enhanced green fluorescent protein (EGFP) yeasts was 10.7 microm at the pick-up stage, 9.7 microm at the clean-up stage, 9.0 microm at the final stage, and 10.2 microm at the over-mixing stage. The results indicated that the distribution of the yeast cells was the most uniform in the dough of white bread at the final stage, while the heterogeneous distribution at the over-mixing stage was possibly due to the destruction of the gluten network structure within the samples.

Amplifying Dynamic Nuclear Polarization of Frozen Solutions by Incorporating Dielectric Particles

PubMed Central

2014-01-01

There is currently great interest in understanding the limits on NMR signal enhancements provided by dynamic nuclear polarization (DNP), and in particular if the theoretical maximum enhancements can be achieved. We show that over a 2-fold improvement in cross-effect DNP enhancements can be achieved in MAS experiments on frozen solutions by simply incorporating solid particles into the sample. At 9.4 T and ∼105 K, enhancements up to εH = 515 are obtained in this way, corresponding to 78% of the theoretical maximum. We also underline that degassing of the sample is important to achieve highest enhancements. We link the amplification effect to the dielectric properties of the solid material, which probably gives rise to scattering, diffraction, and amplification of the microwave field in the sample. This is substantiated by simulations of microwave propagation. A reduction in sample heating at a given microwave power also likely occurs due to reduced dielectric loss. Simulations indicate that the microwave field (and thus the DNP enhancement) is inhomogeneous in the sample, and we deduce that in these experiments between 5 and 10% of the solution actually yields the theoretical maximum signal enhancement of 658. The effect is demonstrated for a variety of particles added to both aqueous and organic biradical solutions. PMID:25285480

Terminologie alimentaire (Food Terminology).

ERIC Educational Resources Information Center

Pelletier, Jean-Francois

1980-01-01

Translations and descriptions are given in French for a number of English food terms: convenience foods, fast foods, fast foods industry, fast foods restaurant, frozen foods, deep frozen foods, fast frozen foods, quick frozen foods, dry frozen foods. (MSE)

Mueller matrix microscope: a quantitative tool to facilitate detections and fibrosis scorings of liver cirrhosis and cancer tissues.

PubMed

Wang, Ye; He, Honghui; Chang, Jintao; He, Chao; Liu, Shaoxiong; Li, Migao; Zeng, Nan; Wu, Jian; Ma, Hui

2016-07-01

Today the increasing cancer incidence rate is becoming one of the biggest threats to human health.Among all types of cancers, liver cancer ranks in the top five in both frequency and mortality rate all over the world. During the development of liver cancer, fibrosis often evolves as part of a healing process in response to liver damage, resulting in cirrhosis of liver tissues. In a previous study, we applied the Mueller matrix microscope to pathological liver tissue samples and found that both the Mueller matrix polar decomposition (MMPD) and Mueller matrix transformation (MMT) parameters are closely related to the fibrous microstructures. In this paper,we take this one step further to quantitatively facilitate the fibrosis detections and scorings of pathological liver tissue samples in different stages from cirrhosis to cancer using the Mueller matrix microscope. The experimental results of MMPD and MMT parameters for the fibrotic liver tissue samples in different stages are measured and analyzed. We also conduct Monte Carlo simulations based on the sphere birefringence model to examine in detail the influence of structural changes in different fibrosis stages on the imaging parameters. Both the experimental and simulated results indicate that the polarized light microscope and transformed Mueller matrix parameter scan provide additional quantitative information helpful for fibrosis detections and scorings of liver cirrhosis and cancers. Therefore, the polarized light microscope and transformed Mueller matrix parameters have a good application prospect in liver cancer diagnosis.

Mueller matrix microscope: a quantitative tool to facilitate detections and fibrosis scorings of liver cirrhosis and cancer tissues

NASA Astrophysics Data System (ADS)

Wang, Ye; He, Honghui; Chang, Jintao; He, Chao; Liu, Shaoxiong; Li, Migao; Zeng, Nan; Wu, Jian; Ma, Hui

2016-07-01

Today the increasing cancer incidence rate is becoming one of the biggest threats to human health. Among all types of cancers, liver cancer ranks in the top five in both frequency and mortality rate all over the world. During the development of liver cancer, fibrosis often evolves as part of a healing process in response to liver damage, resulting in cirrhosis of liver tissues. In a previous study, we applied the Mueller matrix microscope to pathological liver tissue samples and found that both the Mueller matrix polar decomposition (MMPD) and Mueller matrix transformation (MMT) parameters are closely related to the fibrous microstructures. In this paper, we take this one step further to quantitatively facilitate the fibrosis detections and scorings of pathological liver tissue samples in different stages from cirrhosis to cancer using the Mueller matrix microscope. The experimental results of MMPD and MMT parameters for the fibrotic liver tissue samples in different stages are measured and analyzed. We also conduct Monte Carlo simulations based on the sphere birefringence model to examine in detail the influence of structural changes in different fibrosis stages on the imaging parameters. Both the experimental and simulated results indicate that the polarized light microscope and transformed Mueller matrix parameters can provide additional quantitative information helpful for fibrosis detections and scorings of liver cirrhosis and cancers. Therefore, the polarized light microscope and transformed Mueller matrix parameters have a good application prospect in liver cancer diagnosis.

Frozen shoulder and the Big Five personality traits.

PubMed

Debeer, Philippe; Franssens, Fien; Roosen, Isabelle; Dankaerts, Wim; Claes, Laurence

2014-02-01

In the past, several studies have suggested the existence of a "periarthritic personality" in patients with frozen shoulder. We conducted a study to determine differences in personality traits in patients with primary and secondary frozen shoulders. We prospectively evaluated 118 patients (84 women and 34 men; mean age, 53.8 years; SD 7.56) with a frozen shoulder. Of these patients, 48 had an idiopathic frozen shoulder and 70 had a secondary frozen shoulder. Personality traits were determined by the NEO Five-Factor Inventory (NEO-FFI) scale. This questionnaire measures the 5 major personality traits and is based on the norms determined in a neutral test situation for 2415 controls. Compared with healthy controls, no differences in personality traits were found in patients with primary and secondary frozen shoulder, except for Conscientiousness and Extraversion, for which patients with secondary frozen shoulder scored significantly higher than healthy controls. Patients with primary frozen shoulder scored significantly higher on Openness to Experience than did patients with secondary frozen shoulder; on the other 4 Big Five personality traits, no significant differences were found between patients with primary and secondary frozen shoulder. More specifically, patients with idiopathic frozen shoulder did not score higher on the trait Neuroticism as would be expected from previous publications. Our study results do not indicate that patients with an idiopathic frozen shoulder have a specific personality compared with healthy controls. Only a few differences were found in personality traits when the entire frozen shoulder group was compared with healthy controls and between patients with primary and secondary frozen shoulders. The results of this study suggest that these differences are not sufficient to speak about a specific "frozen shoulder personality." Copyright © 2014 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Mosby, Inc. All rights reserved.

The liver tissue bank and clinical database in China.

PubMed

Yang, Yuan; Liu, Yi-Min; Wei, Ming-Yue; Wu, Yi-Fei; Gao, Jun-Hui; Liu, Lei; Zhou, Wei-Ping; Wang, Hong-Yang; Wu, Meng-Chao

2010-12-01

To develop a standardized and well-rounded material available for hepatology research, the National Liver Tissue Bank (NLTB) Project began in 2008 in China to make well-characterized and optimally preserved liver tumor tissue and clinical database. From Dec 2008 to Jun 2010, over 3000 individuals have been enrolled as liver tumor donors to the NLTB, including 2317 cases of newly diagnosed hepatocellular carcinoma (HCC) and about 1000 cases of diagnosed benign or malignant liver tumors. The clinical database and sample store can be managed easily and correctly with the data management platform used. We believe that the high-quality samples with detailed information database will become the cornerstone of hepatology research especially in studies exploring the diagnosis and new treatments for HCC and other liver diseases.

The Eccentric Behavior of Nearly Frozen Orbits

NASA Technical Reports Server (NTRS)

Sweetser, Theodore H.; Vincent, Mark A.

2013-01-01

Frozen orbits are orbits which have only short-period changes in their mean eccentricity and argument of periapse, so that they basically keep a fixed orientation within their plane of motion. Nearly frozen orbits are those whose eccentricity and argument of periapse have values close to those of a frozen orbit. We call them "nearly" frozen because their eccentricity vector (a vector whose polar coordinates are eccentricity and argument of periapse) will stay within a bounded distance from the frozen orbit eccentricity vector, circulating around it over time. For highly inclined orbits around the Earth, this distance is effectively constant over time. Furthermore, frozen orbit eccentricity values are low enough that these orbits are essentially eccentric (i.e., off center) circles, so that nearly frozen orbits around Earth are bounded above and below by frozen orbits.

Rational approach to transfusion in liver transplantation.

PubMed

Saner, Fuat H; Abeysundara, Lasitha; Hartmann, Matthias; Mallett, Susan V

2018-03-01

For over 50 years patients with liver cirrhosis were considered to be at markedly increased risk of bleeding. This dogma was seemingly supported by abnormalities in standard laboratory tests (SLTs), such as the prothrombin time, that were interpreted as indicating a bleeding diathesis. However, publications from the last decade have revealed SLTs to be poor predictors of bleeding and it is now understood that stable patients with cirrhosis have a rebalanced haemostatic system and preserved thrombin generation. Viscoelastic tests (VETs), such as ROTEM® or TEG™ allow dynamic assessment of the entire coagulation process and provide a better illustration of the interactions between pro- and anticoagulants as well as platelets. Despite their documented success in reducing transfusion rates in liver transplantation more than 30 years ago, the adoption of VETs has been met with some resistance and has only recently gained significant momentum. Bleeding risk should be assessed in every patient undergoing invasive intervention and must consider markers of disease severity, underlying coagulation incompetence, anaemia and surgical factors. The recognition that bleeding in this patient cohort is predominantly linked to mechanistic factors such as portal hypertension, rather than primary coagulopathy, has led to a paradigm shift in their perioperative management. Cognizant of their detrimental effect, the use of large volumes of fresh frozen plasma to correct derangements in SLTs has given way to more refined haemostatic management with specific factor concentrates guided by VETs, coupled with measures to minimize portal venous pressure and meticulous surgical hemostasis.

Optimization of the elution buffer and concentration method for detecting hepatitis E virus in swine liver using a nested reverse transcription-polymerase chain reaction and real-time reverse transcription-polymerase chain reaction.

PubMed

Son, Na Ry; Seo, Dong Joo; Lee, Min Hwa; Seo, Sheungwoo; Wang, Xiaoyu; Lee, Bog-Hieu; Lee, Jeong-Su; Joo, In-Sun; Hwang, In-Gyun; Choi, Changsun

2014-09-01

The aim of this study was to develop an optimal technique for detecting hepatitis E virus (HEV) in swine livers. Here, three elution buffers and two concentration methods were compared with respect to enhancing recovery of HEV from swine liver samples. Real-time reverse transcription-polymerase chain reaction (RT-PCR) and nested RT-PCR were performed to detect HEV RNA. When phosphate-buffered saline (PBS, pH 7.4) was used to concentrate HEV in swine liver samples using ultrafiltration, real-time RT-PCR detected HEV in 6 of the 26 samples. When threonine buffer was used to concentrate HEV using polyethylene glycol (PEG) precipitation and ultrafiltration, real-time RT-PCR detected HEV in 1 and 3 of the 26 samples, respectively. When glycine buffer was used to concentrate HEV using ultrafiltration and PEG precipitation, real-time RT-PCR detected HEV in 1 and 3 samples of the 26 samples, respectively. When nested RT-PCR was used to detect HEV, all samples tested negative regardless of the type of elution buffer or concentration method used. Therefore, the combination of real-time RT-PCR and ultrafiltration with PBS buffer was the most sensitive and reliable method for detecting HEV in swine livers. Copyright © 2014 Elsevier B.V. All rights reserved.

Effect of DNA extraction and sample preservation method on rumen bacterial population.

PubMed

Fliegerova, Katerina; Tapio, Ilma; Bonin, Aurelie; Mrazek, Jakub; Callegari, Maria Luisa; Bani, Paolo; Bayat, Alireza; Vilkki, Johanna; Kopečný, Jan; Shingfield, Kevin J; Boyer, Frederic; Coissac, Eric; Taberlet, Pierre; Wallace, R John

2014-10-01

The comparison of the bacterial profile of intracellular (iDNA) and extracellular DNA (eDNA) isolated from cow rumen content stored under different conditions was conducted. The influence of rumen fluid treatment (cheesecloth squeezed, centrifuged, filtered), storage temperature (RT, -80 °C) and cryoprotectants (PBS-glycerol, ethanol) on quality and quantity parameters of extracted DNA was evaluated by bacterial DGGE analysis, real-time PCR quantification and metabarcoding approach using high-throughput sequencing. Samples clustered according to the type of extracted DNA due to considerable differences between iDNA and eDNA bacterial profiles, while storage temperature and cryoprotectants additives had little effect on sample clustering. The numbers of Firmicutes and Bacteroidetes were lower (P < 0.01) in eDNA samples. The qPCR indicated significantly higher amount of Firmicutes in iDNA sample frozen with glycerol (P < 0.01). Deep sequencing analysis of iDNA samples revealed the prevalence of Bacteroidetes and similarity of samples frozen with and without cryoprotectants, which differed from sample stored with ethanol at room temperature. Centrifugation and consequent filtration of rumen fluid subjected to the eDNA isolation procedure considerably changed the ratio of molecular operational taxonomic units (MOTUs) of Bacteroidetes and Firmicutes. Intracellular DNA extraction using bead-beating method from cheesecloth sieved rumen content mixed with PBS-glycerol and stored at -80 °C was found as the optimal method to study ruminal bacterial profile. Copyright © 2013 Elsevier Ltd. All rights reserved.

Effects of Chilling and Partial Freezing on Rigor Mortis Changes of Bighead Carp (Aristichthys nobilis) Fillets: Cathepsin Activity, Protein Degradation and Microstructure of Myofibrils.

PubMed

Lu, Han; Liu, Xiaochang; Zhang, Yuemei; Wang, Hang; Luo, Yongkang

2015-12-01

To investigate the effects of chilling and partial freezing on rigor mortis changes in bighead carp (Aristichthys nobilis), pH, cathepsin B, cathepsin B+L activities, SDS-PAGE of sarcoplasmic and myofibrillar proteins, texture, and changes in microstructure of fillets at 4 °C and -3 °C were determined at 0, 2, 4, 8, 12, 24, 48, and 72 h after slaughter. The results indicated that pH of fillets (6.50 to 6.80) was appropriate for cathepsin function during the rigor mortis. For fillets that were chilled and partially frozen, the cathepsin activity in lysosome increased consistently during the first 12 h, followed by a decrease from the 12 to 24 h, which paralleled an increase in activity in heavy mitochondria, myofibrils and sarcoplasm. There was no significant difference in cathepsin activity in lysosomes between fillets at 4 °C and -3 °C (P > 0.05). Partially frozen fillets had greater cathepsin activity in heavy mitochondria than chilled samples from the 48 to 72 h. In addition, partially frozen fillets showed higher cathepsin activity in sarcoplasm and lower cathepsin activity in myofibrils compared with chilled fillets. Correspondingly, we observed degradation of α-actinin (105 kDa) by cathepsin L in chilled fillets and degradation of creatine kinase (41 kDa) by cathepsin B in partially frozen fillets during the rigor mortis. The decline of hardness for both fillets might be attributed to the accumulation of cathepsin in myofibrils from the 8 to 24 h. The lower cathepsin activity in myofibrils for fillets that were partially frozen might induce a more intact cytoskeletal structure than fillets that were chilled. © 2015 Institute of Food Technologists®

Detection of viable Salmonella in ice cream by TaqMan real-time polymerase chain reaction assay combining propidium monoazide.

PubMed

Wang, Yuexia; Yang, Ming; Liu, Shuchun; Chen, Wanyi; Suo, Biao

2015-09-01

Real-time polymerase chain reaction (PCR) allows rapid detection of Salmonella in frozen dairy products, but it might cause a false positive detection result because it might amplify DNA from dead target cells as well. In this study, Salmonella-free frozen ice cream was initially inoculated with heat-killed Salmonella Typhimurium cells and stored at -18°C. Bacterial DNA extracted from the sample was amplified using TaqMan probe-based real-time PCR targeting the invA gene. Our results indicated that DNA from the dead cells remained stable in frozen ice cream for at least 20 days, and could produce fluorescence signal for real-time PCR as well. To overcome this limitation, propidium monoazide (PMA) was combined with real-time PCR. PMA treatment can effectively prevent PCR amplification from heat-killed Salmonella cells in frozen ice cream. The PMA real-time PCR assay can selectively detect viable Salmonella at as low as 10 3  CFU/mL. Combining 18 hours of pre-enrichment with the assay allows for the detection of viable Salmonella at 10 0  CFU/mL and avoiding the false-positive result of dead cells. The PMA real-time PCR assay provides an alternative specifically for detection of viable Salmonella in ice cream. However, when the PMA real-time PCR assay was evaluated in ice cream subjected to frozen storage, it obviously underestimated the contamination situation of viable Salmonella, which might lead to a false negative result. According to this result, the use of enrichment prior to PMA real-time PCR analysis remains as the more appropriate approach. Copyright © 2015. Published by Elsevier B.V.

Studies on the quantitative autoradiography. III. Quantitative comparison of a novel tissue-mold measurement technique "paste-mold method," to the semiquantitative whole body autoradiography (WBA), using the same animals.

PubMed

Motoji, N; Hamai, Y; Niikura, Y; Shigematsu, A

1995-01-01

A novel preparation technique, so called "Paste Mold," was devised for organ and tissue distribution studies. This is the most powerful by joining with autoradioluminography (ARLG), which was established and validated recently in the working group of Forum '93 of Japanese Society for study of xenobiotics. A small piece (10-50 mg) of each organ or tissue was available for measuring its radioactive concentration and it was sampled from the remains of frozen carcass used for macroautoradiography (MARG). The solubilization of the frozen pieces was performed with mixing a suitable volume of gelatine and strong alkaline solution prior to mild heating kept at 40 degrees C for a few hours. After that, the tissue paste was molded in template pattern to form the small plates. The molded plates were contacted with Imaging plate (IP) for recording their radioactive concentration. The recorded IP was processed by BAS2000. The molded plate was formed in thickness of 200 microns, so called infinit thickness against soft beta rays, and therefore the resulting relative intensities, represented by (PSL-BG)/S values, indicated practically responsible ratio of the radioactive concentration in organs and tissues, without any calibulation for beta-self absorption coefficiency. On the other hand, the left half body of the frozen carcass was used for making whole body autoradiography (WBA) before the Paste-Mold preparation. Comparison was performed for difference in (PSL-BG)/S values of organs and tissues between frozen and dried sections. A good concordance in relative intensities, (PSL-BG)/S by the Paste-Mold preparation was given with those by the frozen sections rather than dried sections.(ABSTRACT TRUNCATED AT 250 WORDS)