In-vivo immunofluorescence confocal microscopy of herpes simplex virus type 1 keratitis

NASA Astrophysics Data System (ADS)

Kaufman, Stephen C.; Laird, Jeffery A.; Beuerman, Roger W.

1996-05-01

The white-light confocal microscope offers an in vivo, cellular-level resolution view of the cornea. This instrument has proven to be a valuable research and diagnostic tool for the study of infectious keratitis. In this study, we investigate the direct visualization of herpes simplex virus type 1 (HSV-1)-infected corneal epithelium, with in vivo confocal microscopy, using HSV-1 immunofluorescent antibodies. New Zealand white rabbits were infected with McKrae strain of HSV-1 in one eye; the other eye of each rabbit was used as an uninfected control. Four days later, the rabbits were anesthetized and a cellulose sponge was applied to each cornea, and a drop of direct HSV fluorescein-tagged antibody was placed on each sponge every 3 to 5 minutes for 1 hour. Fluorescence confocal microscopy was then performed. The HSV-infected corneas showed broad regions of hyperfluorescent epithelial cells. The uninfected corneas revealed no background fluorescence. Thus, using the confocal microscope with a fluorescent cube, we were able to visualize HSV-infected corneal epithelial cells tagged with a direct fluorescent antibody. This process may prove to be a useful clinical tool for the in vivo diagnosis of HSV keratitis.

Flow cytometric immunofluorescence of rat anterior pituitary cells

NASA Technical Reports Server (NTRS)

Hatfield, J. Michael; Hymer, W. C.

1985-01-01

A flow cytometric immunofluorescence technique was developed for the quantification of growth hormone, prolactin, and luteinizing hormone producing cells. The procedure is based on indirect-immunofluorescence of intracellular hormone using an EPICS V cell sorter and can objectively count 50,000 cells in about 3 minutes. It can be used to study the dynamics of pituitary cell populations under various physiological and pharmacological conditions.

Performance analysis of automated evaluation of Crithidia luciliae-based indirect immunofluorescence tests in a routine setting - strengths and weaknesses.

PubMed

Hormann, Wymke; Hahn, Melanie; Gerlach, Stefan; Hochstrate, Nicola; Affeldt, Kai; Giesen, Joyce; Fechner, Kai; Damoiseaux, Jan G M C

2017-11-27

Antibodies directed against dsDNA are a highly specific diagnostic marker for the presence of systemic lupus erythematosus and of particular importance in its diagnosis. To assess anti-dsDNA antibodies, the Crithidia luciliae-based indirect immunofluorescence test (CLIFT) is one of the assays considered to be the best choice. To overcome the drawback of subjective result interpretation that inheres indirect immunofluorescence assays in general, automated systems have been introduced into the market during the last years. Among these systems is the EUROPattern Suite, an advanced automated fluorescence microscope equipped with different software packages, capable of automated pattern interpretation and result suggestion for ANA, ANCA and CLIFT analysis. We analyzed the performance of the EUROPattern Suite with its automated fluorescence interpretation for CLIFT in a routine setting, reflecting the everyday life of a diagnostic laboratory. Three hundred and twelve consecutive samples were collected, sent to the Central Diagnostic Laboratory of the Maastricht University Medical Centre with a request for anti-dsDNA analysis over a period of 7 months. Agreement between EUROPattern assay analysis and the visual read was 93.3%. Sensitivity and specificity were 94.1% and 93.2%, respectively. The EUROPattern Suite performed reliably and greatly supported result interpretation. Automated image acquisition is readily performed and automated image classification gives a reliable recommendation for assay evaluation to the operator. The EUROPattern Suite optimizes workflow and contributes to standardization between different operators or laboratories.

Accuracy of Immunofluorescence in the Diagnosis of Primary Ciliary Dyskinesia

PubMed Central

Frost, Emily; Dixon, Mellisa; Ollosson, Sarah; Kilpin, Kate; Patel, Mitali; Scully, Juliet; Rogers, Andrew V.; Mitchison, Hannah M.; Bush, Andrew; Hogg, Claire

2017-01-01

Rationale: The standard approach to diagnosis of primary ciliary dyskinesia (PCD) in the United Kingdom consists of assessing ciliary function by high-speed microscopy and ultrastructure by election microscopy, but equipment and expertise is not widely available internationally. The identification of biallelic disease-causing mutations is also diagnostic, but many disease-causing genes are unknown, and testing is not widely available outside the United States. Fluorescent antibodies to ciliary proteins are used to validate research genetic studies, but diagnostic utility in this disease has not been systematically evaluated. Objectives: To determine utility of a panel of six fluorescent labeled antibodies as a diagnostic tool for PCD. Methods: The study used immunofluorescent labeling of nasal brushings from a discovery cohort of 35 patients diagnosed with PCD by ciliary ultrastructure, and a diagnostic accuracy cohort of 386 patients referred with symptoms suggestive of disease. The results were compared with diagnostic outcome. Measurements and Main Results: Immunofluorescence correctly identified mislocalized or absent staining in 100% of the discovery cohort. In the diagnostic cohort immunofluorescence successfully identified 22 of 25 patients with PCD and normal staining in all 252 in whom PCD was considered highly unlikely. In addition, immunofluorescence provided a result in 55% (39) of cases that were previously inconclusive. Immunofluorescence results were available within 14 days, costing $187 per sample compared with electron microscopy (27 days; cost $1,452). Conclusions: Immunofluorescence is a highly specific diagnostic test for PCD, and it improves the speed and availability of diagnostic testing. However, sensitivity is limited and immunofluorescence is not suitable as a stand-alone test. PMID:28199173

The kinetics-based enzyme-linked immunosorbent assay for coronavirus antibodies in cats: calibration to the indirect immunofluorescence assay and computerized standardization of results through normalization to control values.

PubMed

Barlough, J E; Jacobson, R H; Downing, D R; Lynch, T J; Scott, F W

1987-01-01

The computer-assisted, kinetics-based enzyme-linked immunosorbent assay for coronavirus antibodies in cats was calibrated to the conventional indirect immunofluorescence assay by linear regression analysis and computerized interpolation (generation of "immunofluorescence assay-equivalent" titers). Procedures were developed for normalization and standardization of kinetics-based enzyme-linked immunosorbent assay results through incorporation of five different control sera of predetermined ("expected") titer in daily runs. When used with such sera and with computer assistance, the kinetics-based enzyme-linked immunosorbent assay minimized both within-run and between-run variability while allowing also for efficient data reduction and statistical analysis and reporting of results.

Choline acetyltransferase-like immunofluorescence in epidermis of human skin.

PubMed

Johansson, O; Wang, L

1993-01-01

Using the indirect immunofluorescence approach the occurrence of choline acetyltransferase-like immunoreactivity in epidermis, except stratum basale, of human skin is described. Immunoreactive cells were also found in hair follicles, sweat gland ducts and sebaceous glands.

Immunofluorescence of spirochetes with serum from swine recovered from swine dysentery using an indirect fluorescent antibody test.

PubMed Central

Lee, C H; Olson, L D

1976-01-01

Using an indirect fluorescent antibody test, immunofluorescence of large spirochetes was observed with serum from swine that had recovered from swine dysentery. The spirochetes were obtained from scrapings of the colonic mucosa on the first day of diarrhea which was the time when the spirochete population was observed to be the highest. Of 29 exposed nonmedicated swine which developed and recovered from a diarrhea characteristic of swine dysentery 27 had antispirochete serum titers which ranged from 1:2 to 1:16. None of the 50 nonexposes swine developed a titer. Of 19 swine with a serum titer and reexposed with infective swine dysentery inoculum, 18 did not develop a diarrhea and were presumed to be immune. Considering these findings it is possible that this test could be used to detect antispirochete antibody in unknown swine serum. Images Fig. 1. PMID:793698

Detection of Prosthetic Hip Infection at Revision Arthroplasty by Immunofluorescence Microscopy and PCR Amplification of the Bacterial 16S rRNA Gene

PubMed Central

Tunney, Michael M.; Patrick, Sheila; Curran, Martin D.; Ramage, Gordon; Hanna, Donna; Nixon, James R.; Gorman, Sean P.; Davis, Richard I.; Anderson, Neil

1999-01-01

In this study the detection rates of bacterial infection of hip prostheses by culture and nonculture methods were compared for 120 patients with total hip revision surgery. By use of strict anaerobic bacteriological practice during the processing of samples and without enrichment, the incidence of infection by culture of material dislodged from retrieved prostheses after ultrasonication (sonicate) was 22%. Bacteria were observed by immunofluorescence microscopy in 63% of sonicate samples with a monoclonal antibody specific for Propionibacterium acnes and polyclonal antiserum specific for Staphylococcus spp. The bacteria were present either as single cells or in aggregates of up to 300 bacterial cells. These aggregates were not observed without sonication to dislodge the biofilm. Bacteria were observed in all of the culture-positive samples, and in some cases in which only one type of bacterium was identified by culture, both coccoid and coryneform bacteria were observed by immunofluorescence microscopy. Bacteria from skin-flake contamination were readily distinguishable from infecting bacteria by immunofluorescence microscopy. Examination of skin scrapings did not reveal large aggregates of bacteria but did reveal skin cells. These were not observed in the sonicates. Bacterial DNA was detected in 72% of sonicate samples by PCR amplification of a region of the bacterial 16S rRNA gene with universal primers. All of the culture-positive samples were also positive for bacterial DNA. Evidence of high-level infiltration either of neutrophils or of lymphocytes or macrophages into associated tissue was observed in 73% of patients. Our results indicate that the incidence of prosthetic joint infection is grossly underestimated by current culture detection methods. It is therefore imperative that current clinical practice with regard to the detection and subsequent treatment of prosthetic joint infection be reassessed in the light of these results. PMID:10488193

The kinetics-based enzyme-linked immunosorbent assay for coronavirus antibodies in cats: calibration to the indirect immunofluorescence assay and computerized standardization of results through normalization to control values.

PubMed Central

Barlough, J E; Jacobson, R H; Downing, D R; Lynch, T J; Scott, F W

1987-01-01

The computer-assisted, kinetics-based enzyme-linked immunosorbent assay for coronavirus antibodies in cats was calibrated to the conventional indirect immunofluorescence assay by linear regression analysis and computerized interpolation (generation of "immunofluorescence assay-equivalent" titers). Procedures were developed for normalization and standardization of kinetics-based enzyme-linked immunosorbent assay results through incorporation of five different control sera of predetermined ("expected") titer in daily runs. When used with such sera and with computer assistance, the kinetics-based enzyme-linked immunosorbent assay minimized both within-run and between-run variability while allowing also for efficient data reduction and statistical analysis and reporting of results. PMID:3032390

Comparison between Indirect Immunofluorescence Assay and Shell Vial Culture for Detection of Mumps Virus from Clinical Samples

PubMed Central

Reina, Jordi; Ballesteros, Francisca; Ruiz de Gopegui, Enrique; Munar, Maria; Mari, Margarita

2003-01-01

We report a prospective comparison of the efficacies of an indirect immunofluorescence assay (IFA) and shell vial culture (SVC) of throat swab and urine samples from patients with mumps. Throat swab samples were used for the IFA; the urine samples and throat swabs were inoculated into vials of Vero cells. We studied 62 patients by using 62 throat swabs and 50 urine samples (50 patients with both samples). Sixty (96.7%) throat samples were positive in the SVC, and 61 (98.3%) were positive in the IFA. For the 50 patients from whom both samples were available, the IFA was positive in 50 (100%) cases, the urine sample was positive in 49 (98%) cases, and the throat swab was positive in 48 (96%) cases (P > 0.05). This comparison of throat swabs and urine samples has shown that the two clinical samples are similar in efficacy. PMID:14605158

Immunofluorescence Analysis of Endogenous and Exogenous Centromere-kinetochore Proteins

PubMed Central

Niikura, Yohei; Kitagawa, Katsumi

2016-01-01

"Centromeres" and "kinetochores" refer to the site where chromosomes associate with the spindle during cell division. Direct visualization of centromere-kinetochore proteins during the cell cycle remains a fundamental tool in investigating the mechanism(s) of these proteins. Advanced imaging methods in fluorescence microscopy provide remarkable resolution of centromere-kinetochore components and allow direct observation of specific molecular components of the centromeres and kinetochores. In addition, methods of indirect immunofluorescent (IIF) staining using specific antibodies are crucial to these observations. However, despite numerous reports about IIF protocols, few discussed in detail problems of specific centromere-kinetochore proteins.1-4 Here we report optimized protocols to stain endogenous centromere-kinetochore proteins in human cells by using paraformaldehyde fixation and IIF staining. Furthermore, we report protocols to detect Flag-tagged exogenous CENP-A proteins in human cells subjected to acetone or methanol fixation. These methods are useful in detecting and quantifying endogenous centromere-kinetochore proteins and Flag-tagged CENP-A proteins, including those in human cells. PMID:26967065

Development of confocal immunofluorescence FRET microscopy to Investigate eNOS and GSNOR localization and interaction in pulmonary endothelial cells

NASA Astrophysics Data System (ADS)

Rehman, Shagufta; Brown-Steinke, Kathleen; Palmer, Lisa; Periasamy, Ammasi

2015-03-01

Confocal FRET microscopy is a widely used technique for studying protein-protein interactions in live or fixed cells. Endothelial nitric oxide synthase (eNOS) and S-nitrosoglutathione reductase (GSNOR) are enzymes involved in regulating the bioavailability of S-nitrosothiols (SNOs) in the pulmonary endothelium and have roles in the development of pulmonary arterial hypertension. Labeling of endogenous proteins to better understand a disease process can be challenging. We have used immunofluorescence to detect endogenous eNOS and GSNOR in primary pulmonary endothelial cells to co-localize these proteins as well as to study their interaction by FRET. The challenge has been in selecting the right immunofluorescence labeling condition, right antibody, the right blocking reagent, the right FRET pair and eliminating cross-reactivity of secondary antibodies. We have used Alexa488 and Alexa568 as a FRET pair. After a series of optimizations, the data from Confocal Laser Scanning Microscopy (CLSM) demonstrate co-localization of eNOS and GSNOR in the perinuclear region of the pulmonary endothelial cell primarily within the cis-Golgi with lower levels of co-localization seen within the trans-Golgi. FRET studies demonstrate, for the first time, interaction between eNOS and GSNOR in both murine and bovine pulmonary endothelial cells. Further characterization of eNOSGSNOR interaction and the subcellular location of this interaction will provide mechanistic insight into the importance of S-nitrosothiol signaling in pulmonary biology, physiology and pathology.

Immunofluorescent localization of Staphylococcos aureus antigen in acute bacterial endocarditis nephritis.

PubMed

Yum, M; Wheat, L J; Maxwell, D; Edwards, J L

1978-11-01

A 75-year-old man with Staphylococcus aureus endocarditis in whom acute diffuse proliferative glomerulonephritis developed is described. The light- and electron-microscopic changes of the glomeruli in this case were identical to those of acute poststreptococcal glomerulonephritis. Immunofluorescence revealed deposition of immunoglobulins and complement in the glomeruli. In addition, bacterial antigenic material was demonstrated in the glomeruli by indirect immunofluorescence. These observations further support the hypothesis of an immune-complex pathogenesis in this form of glomerulonephritis.

In situ detection of the activation of Rac1 and RalA small GTPases in mouse adipocytes by immunofluorescent microscopy following in vivo and ex vivo insulin stimulation.

PubMed

Takenaka, Nobuyuki; Nihata, Yuma; Ueda, Sho; Satoh, Takaya

2017-11-01

Rac1 has been implicated in insulin-dependent glucose uptake by mechanisms involving plasma membrane translocation of the glucose transporter GLUT4 in skeletal muscle. Although the uptake of glucose is also stimulated by insulin in adipose tissue, the role for Rac1 in adipocyte insulin signaling remains controversial. As a step to reveal the role for Rac1 in adipocytes, we aimed to establish immunofluorescent microscopy to detect the intracellular distribution of activated Rac1. The epitope-tagged Rac1-binding domain of a Rac1-specific target was utilized as a probe that specifically recognizes the activated form of Rac1. Rac1 activation in response to ex vivo and in vivo insulin stimulations in primary adipocyte culture and mouse white adipose tissue, respectively, was successfully observed by immunofluorescent microscopy. These Rac1 activations were mediated by phosphoinositide 3-kinase. Another small GTPase RalA has also been implicated in insulin-stimulated glucose uptake in skeletal muscle and adipose tissue. Similarly to Rac1, immunofluorescent microscopy using an activated RalA-specific polypeptide probe allowed us to detect intracellular distribution of insulin-activated RalA in adipocytes. These novel approaches to visualize the activation status of small GTPases in adipocytes will largely contribute to the understanding of signal transduction mechanisms particularly for insulin action. Copyright © 2017 Elsevier Inc. All rights reserved.

Microtubules in Plant Cells: Strategies and Methods for Immunofluorescence, Transmission Electron Microscopy and Live Cell Imaging

PubMed Central

Celler, Katherine; Fujita, Miki; Kawamura, Eiko; Ambrose, Chris; Herburger, Klaus; Wasteneys, Geoffrey O.

2016-01-01

Microtubules are required throughout plant development for a wide variety of processes, and different strategies have evolved to visualize and analyze them. This chapter provides specific methods that can be used to analyze microtubule organization and dynamic properties in plant systems and summarizes the advantages and limitations for each technique. We outline basic methods for preparing samples for immunofluorescence labelling, including an enzyme-based permeabilization method, and a freeze-shattering method, which generates microfractures in the cell wall to provide antibodies access to cells in cuticle-laden aerial organs such as leaves. We discuss current options for live cell imaging of MTs with fluorescently tagged proteins (FPs), and provide chemical fixation, high pressure freezing/freeze substitution, and post-fixation staining protocols for preserving MTs for transmission electron microscopy and tomography. PMID:26498784

Gemella sanguinis endocarditis with c-ANCA/anti-PR-3-associated immune complex necrotizing glomerulonephritis with a ‘full-house’ pattern on immunofluorescence microscopy

PubMed Central

Rousseau-Gagnon, Mathieu; Riopel, Julie; Desjardins, Anne; Garceau, Daniel; Agharazii, Mohsen; Desmeules, Simon

2013-01-01

A 67-year-old man was evaluated for haematuria, with a rising creatinine level from 88 to 906 µmol/L and positive c-anti-neutrophil cytoplasm antibody (ANCA)/anti-proteinase 3 (anti-PR3). A kidney biopsy revealed necrotizing glomerulonephritis with a ‘full-house’ pattern on immunofluorescence microscopy. Echocardiography and blood cultures growing Gemella sanguinis diagnosed endocarditis. Dialysis was required for a month. Three months later, following valve replacement, glucocorticoids and 2 months of antibiotic therapy, the creatinine level decreased to 62 µmol/L and c-ANCA/anti-PR3 disappeared. This first case of c-ANCA/anti-PR3 positive glomerulonephritis with a ‘full-house’ immunofluorescence pattern due to bacterial endocarditis underlines the importance of ruling out infection with ANCA positivity or kidney biopsy suggestive of lupus nephritis. PMID:26064489

Development of an indirect immunofluorescence technique for the diagnosis of toxoplasmosis in bottlenose dolphins.

PubMed

Bernal-Guadarrama, María José; Salichs, Joan; Almunia, Javier; García-Parraga, Daniel; Fernández-Gallardo, Nuhacet; Santana-Morales, María Ángeles; Pacheco, Víctor; Afonso-Lehmann, Raquel N; Déniz, Daniel; Lorenzo-Morales, Jacob; Valladares, Basilio; Martínez-Carretero, Enrique

2014-02-01

The diagnosis of toxoplasmosis is often complicated by the lack of specific clinical symptoms or postmortem features, in humans and other animals. The only diagnostic test described so far for the serological diagnosis of Toxoplasma gondii in marine mammals is the modified agglutination test (Dubey et al., Am J Vet Res 48(8):1239-1243, 1987). The development of more sensible and specific immunological techniques requires specific antibodies, which are currently unavailable in the scientific market. Indirect immunofluorescence (IIF) is one of the most widely used methods for the diagnosis of toxoplasmosis in humans (Auer et al., Parasitol Res 12:965-970, 2000). In order to develop and apply this technique to the bottlenose dolphin (Tursiops truncatus), immunoglobulins were firstly purified using ion-exchange chromatography. The purified immunoglobulins were then injected in New Zealand rabbits in order to obtain polyclonal antibodies. These antisera were validated by the IIF technique, using as controls serum samples of dolphins infected by Toxoplasma. The results were visualized using antirabbit IgG labeled with fluorescein. This newly developed and specific serological assay was then tested with the dolphin collection of Loro Parque, Tenerife, Spain (group I), and L'Oceanogràfic of Valencia, Spain (group II). The obtained results in this study showed that none of the dolphins from group 1 were infected by T. gondii and two animals were positive in group 2. Furthermore, we conclude that this study has produced antibodies with high specificity against dolphin immunoglobulins and an IIF method which may be used as immunological diagnostic tools, especially for the serological diagnosis of toxoplasmosis.

Pattern of glomerular diseases in Oman: a study based on light microscopy and immunofluorescence.

PubMed

Alwahaibi, Nasar Yousuf; Alhabsi, Taiseer Ahmed; Alrawahi, Samira Abdullah

2013-03-01

Light microscopy and immunofluorescence play an important part in the final diagnosis of renal biopsy. The aim of this study was to analyze the pattern of various glomerular diseases in Oman. A total of 424 renal biopsies were retrospectively analyzed at the Sultan Qaboos University Hospital between 1999 and 2010. Focal and segmental glomerulosclerosis (FSGS), minimal change disease (MCD), membranous glomerulopathy (MGN) and IgA nephropathy were the most common primary glomerular diseases encountered, accounting for 21.2%, 17%, 12.3% and 8.3%, respectively, of all cases. Lupus nephritis was the most common secondary glomerular disease and was the most prevalent among all biopsies, accounting for 30.4% of all biopsies. Amyloidosis was seen in only two cases. The presence of fluorescein isothiocyanatefibrin in all renal cases was low when compared with IgG, IgA, IgM, C3 and C1q markers. In conclusion, based on the findings of this study, lupus nephritis was the most common of all glomerular diseases and FSGS was the most common primary glomerular disease. The importance of fluorescein isothiocyanate-fibrin in the diagnosis of renal biopsy needs to be further investigated.

Accuracy of molecular diagnostics in pemphigus and bullous pemphigoid: comparison of commercial and modified mosaic indirect immunofluorescence tests as well as enzyme-linked immunosorbent assays.

PubMed

Gornowicz-Porowska, Justyna; Seraszek-Jaros, Agnieszka; Bowszyc-Dmochowska, Monika; Kaczmarek, Elżbieta; Pietkiewicz, Paweł; Bartkiewicz, Paweł; Dmochowski, Marian

2017-02-01

Pemphigus and bullous pemphigoid (BP) are identified by autoantibodies (abs) against desmoglein 1, 3 (DSG1/3) and BP180/BP230, respectively. A novel mosaic to indirect immunofluorescence (IIF) using purified BP180 recombinant proteins spotted on slide and transfected cells expressing BP230, DSG1, DSG3 is available. The commercial (IgG detection) and modified (IgG4 detection) mosaic for indirect immunofluorescence (IIFc - IIF commercial, IIFm - IIF modified) and IgG ELISAs were evaluated in pemphigus and bullous pemphigoid (BP) molecular diagnostics. To compare diagnostic accuracy of commercial (IgG detection) and modified (IgG4 detection) mosaic IIF assay and to examine the diagnostic value of ELISAs in relation to mosaic IIF in routine laboratory diagnostics of pemphigus and BP. Sera from 37 BP and 19 pemphigus patients were studied. Associations between tests were assessed using Fisher's exact test. There are associations between the positive/negative samples detected by IIFc with desmoglein1 (DSG1)/desmoglein3 (DSG3)/BP230 transfected cells and ELISAs and no association between anti-BP180 IgG detection by IIFc and ELISA. IIFm with DSG1 and DSG3 showed both 100% sensitivity and 100% and 78% specificity, respectively, and 100% and 83% positive predictive value in relation to IIFc. IIFm with BP230 had 87% specificity, 55% sensitivity, whereas IIFm with BP180 had a 100% sensitivity and 13% specificity in relation to IIFc. The IIFc with DSG1/DSG3/BP230 transfected cells, excluding BP180 spots, is an alternative method to ELISA in pemphigus/BP diagnostics. IgG4 antibodies, both pathogenically and diagnostically important, are inconsistently detectable with IIFm.

Comparison of diagnostic value of indirect immunofluorescence assay and desmoglein ELISA in the diagnosis of pemphigus.

PubMed

Marinović, Branka; Fabris, Zrinka; Lipozencić, Jasna; Stulhofer Buzina, Daska; Lakos Jukić, Ines

2010-01-01

Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are autoimmune blistering diseases characterized by intraepidermal separation as the result of autoantibodies directed to desmoglein 1 and desmoglein 3, adhesion molecules that have a pathogenic role in blister formation. Both PV and PF are diagnosed according to clinical picture, histopathologic, immunopathologic and molecular biologic features. In the present study, the value of indirect immunofluorescence (IIF) and enzyme linked immunosorbent assay (ELISA) for desmoglein 1 (Dsg 1) and desmoglein 3 (Dsg 3) at baseline visit was compared. The study was performed as a retrospective study that included 22 patients, 19 of them with PV and three with PF. Patient sera were tested with IIF and Dsg 1 and Dsg 3 ELISA. In the group of 19 PV patients, 12 patients had positive IIF, Dsg 3 and Dsg 1 ELISA; two had positive IIF and positive anti Dsg 3 but negative anti Dsg 1; three had negative IIF but positive both Dsg 1 and Dsg 3 antibodies; and two had negative IIF and Dsg 1 but positive Dsg 3 antibodies. In the group of PF patients, all three patients had positive IIF, positive Dsg 1 ELISA and negative Dsg 3 ELISA. Results of our study supported previous reports confirming Dsg 1 and Dsg 3 ELISA to be a sensitive and specific tool for the diagnosis of PV and PF.

ANAlyte: A modular image analysis tool for ANA testing with indirect immunofluorescence.

PubMed

Di Cataldo, Santa; Tonti, Simone; Bottino, Andrea; Ficarra, Elisa

2016-05-01

The automated analysis of indirect immunofluorescence images for Anti-Nuclear Autoantibody (ANA) testing is a fairly recent field that is receiving ever-growing interest from the research community. ANA testing leverages on the categorization of intensity level and fluorescent pattern of IIF images of HEp-2 cells to perform a differential diagnosis of important autoimmune diseases. Nevertheless, it suffers from tremendous lack of repeatability due to subjectivity in the visual interpretation of the images. The automatization of the analysis is seen as the only valid solution to this problem. Several works in literature address individual steps of the work-flow, nonetheless integrating such steps and assessing their effectiveness as a whole is still an open challenge. We present a modular tool, ANAlyte, able to characterize a IIF image in terms of fluorescent intensity level and fluorescent pattern without any user-interactions. For this purpose, ANAlyte integrates the following: (i) Intensity Classifier module, that categorizes the intensity level of the input slide based on multi-scale contrast assessment; (ii) Cell Segmenter module, that splits the input slide into individual HEp-2 cells; (iii) Pattern Classifier module, that determines the fluorescent pattern of the slide based on the pattern of the individual cells. To demonstrate the accuracy and robustness of our tool, we experimentally validated ANAlyte on two different public benchmarks of IIF HEp-2 images with rigorous leave-one-out cross-validation strategy. We obtained overall accuracy of fluorescent intensity and pattern classification respectively around 85% and above 90%. We assessed all results by comparisons with some of the most representative state of the art works. Unlike most of the other works in the recent literature, ANAlyte aims at the automatization of all the major steps of ANA image analysis. Results on public benchmarks demonstrate that the tool can characterize HEp-2 slides in terms of

Comparison of an indirect immunofluorescence assay and a modified sensitive immunoblot assay for the study of the autoantibody in pemphigus vulgaris.

PubMed

Mohimen, A; Ahmed, A R

1995-01-01

Some patients with pemphigus vulgaris (PV) have positive direct immunofluorescence (DIF) but are negative by indirect immunofluorescence (IIF). The purpose of this study was (1) to compare the sensitivity of an IIF assay with an immunoblot (IB) assay, (2) to compare the IIF and the IB assay in PV patients in whom the clinical picture and DIF were consistent, but the IIF was negative and (3) to compare the IIF and the IB assay in patients in clinical remission for 3 years or more. A comparison was made of the titers of PV autoantibody in the IIF assay using monkey esophagus as substrate and the modified sensitive IB assay using preabsorbed normal human skin lysate and COLO-16 lysate as a substrate in the three groups of patients. The sensitivity of the Western blot was enhanced by modifications in the extraction procedure of the lysate, by absorption of lysate with normal human serum and by the use of an enzygraphic web. In group 1, comprising 23 PV patients with active generalized disease, the titers of the autoantibody in the IB assay were 2-4-fold higher than in the IIF assay. This difference was highly significant (P = 0.0001). In group 2, comprising 10 patients with limited or minimal PV who were positive on DIF and negative on IIF, all the patients were positive in the IB assay. In group 3, comprising 9 patients clinically free of disease and off all therapy for at least 3 years and negative in IIF assay, all the patients were positive in the IB assay.(ABSTRACT TRUNCATED AT 250 WORDS)

Simple Elimination of Background Fluorescence in Formalin-Fixed Human Brain Tissue for Immunofluorescence Microscopy.

PubMed

Sun, Yulong; Ip, Philbert; Chakrabartty, Avijit

2017-09-03

Immunofluorescence is a common method used to visualize subcellular compartments and to determine the localization of specific proteins within a tissue sample. A great hindrance to the acquisition of high quality immunofluorescence images is endogenous autofluorescence of the tissue caused by aging pigments such as lipofuscin or by common sample preparation processes such as aldehyde fixation. This protocol describes how background fluorescence can be greatly reduced through photobleaching using white phosphor light emitting diode (LED) arrays prior to treatment with fluorescent probes. The broad-spectrum emission of white phosphor LEDs allow for bleaching of fluorophores across a range of emission peaks. The photobleaching apparatus can be constructed from off-the-shelf components at very low cost and offers an accessible alternative to commercially available chemical quenchers. A photobleaching pre-treatment of the tissue followed by conventional immunofluorescence staining generates images free of background autofluorescence. Compared to established chemical quenchers which reduced probe as well as background signals, photobleaching treatment had no effect on probe fluorescence intensity while it effectively reduced background and lipofuscin fluorescence. Although photobleaching requires more time for pre-treatment, higher intensity LED arrays may be used to reduce photobleaching time. This simple method can potentially be applied to a variety of tissues, particularly postmitotic tissues that accumulate lipofuscin such as the brain and cardiac or skeletal muscles.

X-ray microscopy of live biological micro-organisms

NASA Astrophysics Data System (ADS)

Raja Al-Ani, Ma'an Nassar

Real-time, compact x-ray microscopy has the potential to benefit many scientific fields, including microbiology, pharmacology, organic chemistry, and physics. Single frame x-ray micro-radiography, produced by a compact, solid-state laser plasma source, allows scientists to use x-ray emission for elemental analysis, and to observe biological specimens in their natural state. In this study, x-ray images of mouse kidney tissue, live bacteria, Pseudomonas aeruginosa and Burkholderia cepacia, and the bacteria's interaction with the antibiotic gentamicin, are examined using x-ray microscopy. For the purposes of comparing between confocal microscopy and x-ray microscopy, we introduced to our work the technique of gold labeling. Indirect immunofluorescence staining and immuno-gold labeling were applied on human lymphocytes and human tumor cells. Differential interference contrast microscopy (DIC) showed the lymphocyte body and nucleus, as did x-ray microscopy. However, the high resolution of x-ray microscopy allows us to differentiate between the gold particles bound to the antibodies and the free gold. A compact, tabletop Nd: glass laser is used in this study to produce x-rays from an Yttrium target. An atomic force microscope is used to scan the x-ray images from the developed photo-resist. The use of compact, tabletop laser plasma sources, in conjunction with x-ray microscopy, is a new technique that has great potential as a flexible, user-friendly scientific research tool.

Immunity to human cytomegalovirus measured and compared by complement fixation, indirect fluorescent-antibody, indirect hemagglutination, and enzyme-linked immunosorbent assays.

PubMed Central

Brandt, J A; Kettering, J D; Lewis, J E

1984-01-01

The complement fixation test is currently the test employed most frequently to determine the presence of antibody to human cytomegalovirus. Several other techniques have been adapted for this purpose. A comparison of cytomegalovirus antibody titers was made between the complement fixation test, a commercially available enzyme-linked immunosorbent assay, an indirect immunofluorescent technique, and a modified indirect hemagglutination test. Forty-three serum samples were tested for antibodies by each of the above procedures. The enzyme-linked immunosorbent, immunofluorescent, and indirect hemagglutination assays were in close agreement on all samples tested; the titers obtained with these methods were all equal to or greater than the complement fixation titer for 38 of the 41 samples (92.6%). Two samples were anticomplementary in the complement fixation test but gave readable results in the other tests. The complement fixation test was the least sensitive of the procedures examined. The commercial enzyme-linked immunosorbent assay system was the most practical method and offered the highest degree of sensitivity in detecting antibodies to cytomegalovirus. PMID:6321544

Multiplex RT-PCR and indirect immunofluorescence assays for detection and subtyping of human influenza virus in Tunisia.

PubMed

Ben M'hadheb, Manel; Harrabi, Myriam; Souii, Amira; Jrad-Battikh, Nadia; Gharbi, Jawhar

2015-03-01

Influenza viruses are negative stranded segmented RNA viruses belonging to Orthomyxoviridae family. They are classified into three types A, B, and C. Type A influenza viruses are classified into subtypes according to the antigenic characters of the surface glycoproteins: hemagglutinin (H) and neuraminidase (N). The aim of the present study is to develop a fast and reliable multiplex RT-PCR technique for detecting simultaneously the subtypes A/H1N1 and A/H3N2 of influenza virus. Our study included 398 patients (mean age 30.33 ± 19.92 years) with flu or flu-like syndromes, consulting physicians affiliated with collaborating teams. A multiplex RT-PCR detecting A/H1N1 and A/H3N2 influenza viruses and an examination by indirect immunofluorescence (IFI) were performed. In the optimized conditions, we diagnosed by IFI a viral infection in 90 patients (22.6 %): 85 cases of influenza type A, four cases of influenza type B, and only one case of coinfection with types A and B. An evaluation of the technique was performed on 19 clinical specimens positive in IFI, and we detected eight cases of A/H3N2, five cases of A/H1N1, one case of influenza virus type A which is not an H1N1 nor H3N2, and five negative cases. Multiplex RT-PCR is a sensitive technique allowing an effective and fast diagnosis of respiratory infections caused by influenza viruses in which the optimization often collides with problems of sensibility.

The detection of Giardia muris and Giardia lamblia cysts by immunofluorescence in animal tissues and fecal samples subjected to cycles of freezing and thawing.

PubMed

Erlandsen, S L; Sherlock, L A; Bemrick, W J

1990-04-01

The effects of freezing and thawing on the detection of selected Giardia spp. cysts were investigated using immunofluorescence, bright field microscopy, and low voltage scanning electron microscopy (SEM). Giardia muris cysts were obtained from either animal carcasses, fecal pellets, or isolated cyst preparations, whereas Giardia lamblia cysts were isolated from fecal samples. These samples were stained using an immunofluorescence technique after 1-3 freezing (-16 C) and thawing (20 C) cycles. Cysts were detected successfully by immunofluorescence in all samples. However, in those samples subjected to freeze-thawing, the cyst walls often became distorted and then were not detectable by bright field microscopy. Low voltage SEM demonstrated that the filaments in the distorted cyst wall underwent rearrangements of interfilament spacing. Quantitation of cyst recovery after freezing and thawing demonstrated that a substantial loss occurred after 1 cycle of alternating temperature when low concentrations of cysts were used, but not with high concentrations of cysts. Cyst recovery, after 3 freezing and thawing cycles, was dramatically lowered irrespective of the initial cyst concentration. These results demonstrated that immunofluorescence was an effective technique for the detection of Giardia spp. cysts in frozen samples and would suggest that freezing and thawing of fecal samples could prevent the detection of cysts when only bright field microscopy was employed.

3D multiplexed immunoplasmonics microscopy

NASA Astrophysics Data System (ADS)

Bergeron, Éric; Patskovsky, Sergiy; Rioux, David; Meunier, Michel

2016-07-01

Selective labelling, identification and spatial distribution of cell surface biomarkers can provide important clinical information, such as distinction between healthy and diseased cells, evolution of a disease and selection of the optimal patient-specific treatment. Immunofluorescence is the gold standard for efficient detection of biomarkers expressed by cells. However, antibodies (Abs) conjugated to fluorescent dyes remain limited by their photobleaching, high sensitivity to the environment, low light intensity, and wide absorption and emission spectra. Immunoplasmonics is a novel microscopy method based on the visualization of Abs-functionalized plasmonic nanoparticles (fNPs) targeting cell surface biomarkers. Tunable fNPs should provide higher multiplexing capacity than immunofluorescence since NPs are photostable over time, strongly scatter light at their plasmon peak wavelengths and can be easily functionalized. In this article, we experimentally demonstrate accurate multiplexed detection based on the immunoplasmonics approach. First, we achieve the selective labelling of three targeted cell surface biomarkers (cluster of differentiation 44 (CD44), epidermal growth factor receptor (EGFR) and voltage-gated K+ channel subunit KV1.1) on human cancer CD44+ EGFR+ KV1.1+ MDA-MB-231 cells and reference CD44- EGFR- KV1.1+ 661W cells. The labelling efficiency with three stable specific immunoplasmonics labels (functionalized silver nanospheres (CD44-AgNSs), gold (Au) NSs (EGFR-AuNSs) and Au nanorods (KV1.1-AuNRs)) detected by reflected light microscopy (RLM) is similar to the one with immunofluorescence. Second, we introduce an improved method for 3D localization and spectral identification of fNPs based on fast z-scanning by RLM with three spectral filters corresponding to the plasmon peak wavelengths of the immunoplasmonics labels in the cellular environment (500 nm for 80 nm AgNSs, 580 nm for 100 nm AuNSs and 700 nm for 40 nm × 92 nm AuNRs). Third, the developed

Decoupling indirect topographic cross-talk in band excitation piezoresponse force microscopy imaging and spectroscopy

DOE PAGES

Mazet, Lucie; Jesse, Stephen; Niu, Gang; ...

2016-06-20

Here, all scanning probe microscopies are subjected to topographic cross-talk, meaning the topography-related contrast in functional images. Here, we investigate the signatures of indirect topographic cross-talk in piezoresponse force microscopy (PFM) imaging and spectroscopy and its decoupling using band excitation (BE) method in ferroelectric BaTiO 3 deposited on the Si substrates with free standing nanopillars of diameter 50 nm. Comparison between the single-frequency PFM and BE-PFM results shows that the measured signal can be significantly distorted by topography-induced shifts in the contact resonance frequency and cantilever transfer function. However, with proper correction, such shifts do not affect PFM imaging andmore » hysteresis loop measurements. This suggests the necessity of an advanced approach, such as BE-PFM, for detection of intrinsic sample piezoresponse on the topographically non-uniform surfaces.« less

Indirect Immunofluorescence Assay for the Simultaneous Detection of Antibodies against Clinically Important Old and New World Hantaviruses

PubMed Central

Lederer, Sabine; Lattwein, Erik; Hanke, Merle; Sonnenberg, Karen; Stoecker, Winfried; Lundkvist, Åke; Vaheri, Antti; Vapalahti, Olli; Chan, Paul K. S.; Feldmann, Heinz; Dick, Daryl; Schmidt-Chanasit, Jonas; Padula, Paula; Vial, Pablo A.; Panculescu-Gatej, Raluca; Ceianu, Cornelia; Heyman, Paul; Avšič-Županc, Tatjana; Niedrig, Matthias

2013-01-01

In order to detect serum antibodies against clinically important Old and New World hantaviruses simultaneously, multiparametric indirect immunofluorescence assays (IFAs) based on biochip mosaics were developed. Each of the mosaic substrates consisted of cells infected with one of the virus types Hantaan (HTNV), Puumala (PUUV), Seoul (SEOV), Saaremaa (SAAV), Dobrava (DOBV), Sin Nombre (SNV) or Andes (ANDV). For assay evaluation, serum IgG and IgM antibodies were analyzed using 184 laboratory-confirmed hantavirus-positive sera collected at six diagnostic centers from patients actively or previously infected with the following hantavirus serotypes: PUUV (Finland, n = 97); SEOV (China, n = 5); DOBV (Romania, n = 7); SNV (Canada, n = 23); ANDV (Argentina and Chile, n = 52). The control panel comprised 89 sera from healthy blood donors. According to the reference tests, all 184 patient samples were seropositive for hantavirus-specific IgG (n = 177; 96%) and/or IgM (n = 131; 72%), while all control samples were tested negative. In the multiparametric IFA applied in this study, 183 (99%) of the patient sera were IgG and 131 (71%) IgM positive (accordance with the reference tests: IgG, 96%; IgM, 93%). Overall IFA sensitivity for combined IgG and IgM analysis amounted to 100% for all serotypes, except for SNV (96%). Of the 89 control sera, 2 (2%) showed IgG reactivity against the HTNV substrate, but not against any other hantavirus. Due to the high cross-reactivity of hantaviral nucleocapsid proteins, endpoint titrations were conducted, allowing serotype determination in >90% of PUUV- and ANDV-infected patients. Thus, multiparametric IFA enables highly sensitive and specific serological diagnosis of hantavirus infections and can be used to differentiate PUUV and ANDV infection from infections with Murinae-borne hantaviruses (e.g. DOBV and SEOV). PMID:23593524

Yaba Tumor Poxvirus Synthesis In Vitro I. Cytopathological, Histochemical, and Immunofluorescent Studies

PubMed Central

Yohn, David S.; Haendiges, Victoria A.; Grace, James T.

1966-01-01

Yohn, David S. (Roswell Park Memorial Institute, Buffalo, N.Y.), Victoria A. Haendiges, and James T. Grace, Jr. Yaba tumor poxvirus synthesis in vitro. I. Cytopathological, histochemical, and immunofluorescent studies. J. Bacteriol. 91:1977–1985. 1966.—Yaba tumor poxvirus synthesis in BSC-1 cell culture was followed sequentially with light microscopy, immunofluorescent microscopy, and various histochemical stains. The first evidence of infection was detected at 24 hr when nucleoli became hypertrophic, reflecting enhanced ribonucleic acid (RNA) synthesis. At 36 hr, deoxyribonucleic acid synthesis was detected in the cytoplasm. This was immediately followed by or associated with antigen synthesis at paranuclear sites and enhanced RNA synthesis in the cytoplasm. Cytoplasmic inclusions were readily apparent at 4 days in initially infected cells. Contiguous spread of virus was judged to have occurred around the third day of infection. The time required to complete the synthetic cycle from time of infection to production of infectious progeny was estimated to be of the order of 60 hr. This cycle is 6 to 10 times longer than for vaccinia virus. By light microscopy, cytopathic effects (CPE) were detectable in 5 days in heavily infected cultures. With 100 units or less of infectious virus, CPE was not readily detected until the 10th to 14th day. At this time, focal areas of infection classified either as microtumors or microplaques were present. Secondary foci appeared during the 4th week of incubation. Images PMID:4287080

[Evaluation of a Computer-Aided Microscope System and Its Anti-Nuclear Antibody Test Kit for Indirect Immunofluorescence Assay].

PubMed

Hayashi, Nobuhide; Saegusa, Jun; Uto, Kenichi; Oyabu, Chinami; Saito, Toshiharu; Sato, Itsuko; Kawano, Seiji; Kumagai, Shunichi

2016-02-01

Antinuclear antibody (ANA) testing is indispensable for diagnosing and understanding clinical conditions of autoimmune diseases. The indirect immunofluorescence assay (IFA) is the gold standard for ANA screening, and it can detect more than 100 different antibodies, such as anti-PCNA as well as anti-cytoplasmic antibodies. However, complicated procedures of conventional IFA and visual interpretation require highly skilled laboratory staff. This study evaluates the capability, characteristics, and applicability of the recently developed ANA detection system (EUROPattern Cosmic IFA System, EPA) using HEp20-10 cells and the automated pattern recognition microscope. Findings using EPA and conventional methods were compared in 282 sera obtained from connective tissue disease patients and 250 sera from healthy individuals. The concordance of the positivity rate, antibody titer (within +/- 1 tube difference), and the accurate recognition rate of ANA patterns between the automated EPA method and the microscopic judgement of the EPA image by eye was 98.9, 97.4, and 55.3%, respectively. The EPA method showed concordance of the positivity rate as high as 93.3% and concordance of the antibody titer as high as 94.0% (within +/- 1 titer) compared with the conventional method. Regarding the four typical patterns of ANA (homogeneous, speckled, nucleolar, and centromere), large differences between the EPA and conventional methods were not observed, and the rate of concordance between the final EPA result and the conventional method was from 94.1 to 100%. The positivity rate of ANA using the EPA and conventional methods showed marked agreement among the six connective tissue diseases (SLE, MCTD, SSc, PM/DM, and SS) and healthy individuals. Although the EPA system is not considered a complete system and laboratory staff should verify the results, it is a useful system for routine ANA analysis because it contributes to ANA standardization and an efficient workflow.

DropArray™, a wall-less 96-well plate for uptake and immunofluorescence microscopy, confirms CD22 recycles.

PubMed

Ingle, Gladys S; Scales, Suzie J

2014-03-01

CD22 is a cell surface glycoprotein restricted to normal and malignant B-cells and is the target of several anti-CD22 antibody-based cancer therapies. For therapeutic antibody-payload conjugates, it is important to understand the subcellular trafficking of anti-CD22 antibodies to optimize antibody and/or linker-drug properties to maximize antitumor efficacy. It is agreed that anti-CD22 antibodies rapidly internalize, but controversial whether they recycle or are degraded in lysosomes, and it is unclear if trafficking is antibody or cell-type dependent. No studies examined anti-CD22 trafficking to either pathway in B-cells over time by dual immunofluorescence microscopy, likely partly because multiple samples of suspension cells are tedious to stain. We overcame this by using DropArray™, a novel wall-less 96-well plate technology allowing rapid simultaneous staining of suspension or adherent cells in small (10-20 μL) volumes. We examined the time-course of trafficking of five different anti-CD22 antibodies in eight B-cell lines representing four B-cell cancer types and show that in all cases antibodies internalize within 5 min and recycle, with only small amounts eventually trafficking to lysosomes. CD22 also localizes to recycling endosomes at steady state in the absence of antibody. Our data may help explain the differential efficacies of anti-CD22 antibodies conjugated to different therapeutic payloads. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Diagnosis of equine monocytic ehrlichiosis (Potomac horse fever) by indirect immunofluorescence.

PubMed

Ristic, M; Holland, C J; Dawson, J E; Sessions, J; Palmer, J

1986-07-01

The recent establishment of a system for the continuous in vitro propagation of Ehrlichia risticii, the causative agent of equine monocytic ehrlichiosis (EME; synonym, Potomac horse fever), has facilitated the development of an indirect fluorescent antibody test for the diagnosis of this disease under laboratory and field conditions. The field diagnostic application of the test has aided in the recognition of the disease in 16 states of the United States and in 1 province of Canada. A limited epidemiologic study conducted between January and September 1985, in an area where the disease is known to be enzootic, revealed that conversion from seronegative to seropositive status is not always accompanied by clinical manifestations of the disease. Confirmatory findings in experimentally inoculated horses suggest the existence of clinically undetectable infections.

Autofluorescence and non-specific immunofluorescent labeling in frozen bovine intestinal tissue sections: Solutions for multi-color immunofluorescence experiments

USDA-ARS?s Scientific Manuscript database

Autofluorescence and non-specific immunofluorescent labeling are common challenges associated with immunofluorescence experiments. Autofluorescence typically demonstrates a broad emission spectrum, increasing the potential for overlap with experiments that utilize multiple fluorophores. During immun...

An Indirect Immunofluorescence Method Facilitates Detection of Thrombospondin Type 1 Domain–Containing 7A–Specific Antibodies in Membranous Nephropathy

PubMed Central

Hoxha, Elion; Beck, Laurence H.; Wiech, Thorsten; Tomas, Nicola M.; Probst, Christian; Mindorf, Swantje; Meyer-Schwesinger, Catherine; Zahner, Gunther; Stahl, Phillip R.; Schöpper, Ruth; Panzer, Ulf; Harendza, Sigrid; Helmchen, Udo; Salant, David J.

2017-01-01

Thrombospondin type 1 domain–containing 7A (THSD7A) is a target antigen identified in adult membranous nephropathy (MN) along with the major antigen phospholipase A2 receptor 1 (PLA2R1). The prevalence of THSD7A-Ab–positive patients is unknown, and it is unclear whether the clinical presentation differs between patients positive for PLA2R1-Ab or THSD7A-Ab. We screened serum samples of 1276 patients with MN from three different cohorts for the presence of THSD7A-Ab by Western blot analysis and a newly developed indirect immunofluorescence test (IFT). Compared with Western blot analysis, the IFT had a 92% sensitivity and a 100% specificity. The prevalence of THSD7A-associated MN in a prospective cohort of 345 patients with MN was 2.6%, and most were women. In this cohort, the percentage of patients with THSD7A-associated MN and malignant disease significantly exceeded that of patients with PLA2R1-associated MN and malignant disease. In all cohorts, we identified 40 patients with THSD7A-associated MN, eight of whom developed a malignancy within a median time of 3 months from diagnosis of MN. In one patient with THSD7A-associated MN and metastases of an endometrial carcinoma, immunohistochemistry showed THSD7A expression on the metastatic cells and within follicular dendritic cells of the metastasis–infiltrated lymph node. We conclude that the IFT allows sensitive and specific measurement of circulating THSD7A-Ab in patients with MN. Patients with THSD7A-associated MN differ in their clinical characteristics from patients with PLA2R1-associated MN, and more intensive screening for the presence of malignancies may be warranted in those with THSD7A-associated MN. PMID:27436855

Comparison of growth inhibition and immunofluorescence tests in serotyping clinical isolates of Ureaplasma urealyticum.

PubMed Central

Piot, P

1977-01-01

Typestrains of the eight serotypes of Ureaplasma urealyticum and 39 genital isolates were typed by the growth inhibition (GI) and indirect immunofluorescence (IF) tests. The GI test proved to be very specific and simple, demanding large volumes of serum. It was less sensitive than the IF test, which produced more cross-reactions, was economical in serum, and able to detect mixed infections. Of the 39 isolates, 27 were serotyped by GI and 34 by IF. Mixed cultures occurred in as many as 36% of the isolates. PMID:326348

Comparison of sensitivity and specificity of 4 methods for detection of Giardia duodenalis in feces: immunofluorescence and PCR are superior to microscopy of concentrated iodine-stained samples.

PubMed

Gotfred-Rasmussen, Helle; Lund, Marianne; Enemark, Heidi L; Erlandsen, Mogens; Petersen, Eskild

2016-03-01

For decades, microscopy of feces after formol-ethylacetate (FEA) concentration and iodine staining has been the routine test for intestinal protozoa. Lately, polymerase chain reaction or fluorescence-labeled parasite-specific antibodies have been introduced, but their place in everyday routine diagnostics has not yet been established. We compared FEA and salt-sugar flotation (SSF) concentration followed by microscopy of iodine-stained concentrate and immunofluorescence assay (IFA) and real-time polymerase chain reaction (qPCR) for detection of Giardia duodenalis in human feces. The median number of Giardia cysts found by FEA in 19 Giardia-positive samples was 50 cysts per gram (CPG), by SSF 350 CPG, by IFA 76,700 CPG, and by qPCR 316,000 CPG. We next tested 455 consecutive samples for presence of Giardia cysts. Using IFA as reference, qPCR had a sensitivity of 91%, specificity of 95.1%, a false-positive rate of 50%, a false-negative rate of 0.48%, a positive predictive value of 50%, and a negative predictive value of 99.5%. In conclusion, qPCR and IFA were significantly more sensitive than microscopy of iodine-stained concentrates using either FEA or SSF. We suggest, when using qPCR, that positive samples are verified by IFA to prevent false-positive results. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Characterizing DNA Repair Processes at Transient and Long-lasting Double-strand DNA Breaks by Immunofluorescence Microscopy.

PubMed

Murthy, Vaibhav; Dacus, Dalton; Gamez, Monica; Hu, Changkun; Wendel, Sebastian O; Snow, Jazmine; Kahn, Andrew; Walterhouse, Stephen H; Wallace, Nicholas A

2018-06-08

The repair of double-stranded breaks (DSBs) in DNA is a highly coordinated process, necessitating the formation and resolution of multi-protein repair complexes. This process is regulated by a myriad of proteins that promote the association and disassociation of proteins to these lesions. Thanks in large part to the ability to perform functional screens of a vast library of proteins, there is a greater appreciation of the genes necessary for the double-strand DNA break repair. Often knockout or chemical inhibitor screens identify proteins involved in repair processes by using increased toxicity as a marker for a protein that is required for DSB repair. Although useful for identifying novel cellular proteins involved in maintaining genome fidelity, functional analysis requires the determination of whether the protein of interest promotes localization, formation, or resolution of repair complexes. The accumulation of repair proteins can be readily detected as distinct nuclear foci by immunofluorescence microscopy. Thus, association and disassociation of these proteins at sites of DNA damage can be accessed by observing these nuclear foci at representative intervals after the induction of double-strand DNA breaks. This approach can also identify mis-localized repair factor proteins, if repair defects do not simultaneously occur with incomplete delays in repair. In this scenario, long-lasting double-strand DNA breaks can be engineered by expressing a rare cutting endonuclease (e.g., I-SceI) in cells where the recognition site for the said enzyme has been integrated into the cellular genome. The resulting lesion is particularly hard to resolve as faithful repair will reintroduce the enzyme's recognition site, prompting another round of cleavage. As a result, differences in the kinetics of repair are eliminated. If repair complexes are not formed, localization has been impeded. This protocol describes the methodology necessary to identify changes in repair kinetics as

Immunofluorescence findings in rapid whitening of scalp hair.

PubMed

Guin, J D; Kumar, V; Petersen, B H

1981-09-01

Rapid whitening of scalp hair developed during a three-month period along with a diffuse, subtotal alopecia in a patient. Immunofluorescence microscopy of biopsy material showed prominent deposits of IgG and IgM in a granular pattern in the epithelium of the lower portions of hair follicles. Some return of the color and amount of scalp hair occurred within a year, but occasional bouts of hair loss continued to occur. It is theorized that the rapid graying was caused by a selective loss of pigmented hair, which was perhaps caused by an immunologic mechanism. Some of the findings suggest that the cause of this patient's loss of hair color may be different from those of patients who have been previously described as having rapid whitening of scalp hair because of alopecia areata or vitiligo.

SNOM and AFM microscopy techniques to study the effect of non-ionizing radiation on the morphological and biochemical properties of human keratinocytes cell line (HaCaT).

PubMed

Rieti, S; Manni, V; Lisi, A; Giuliani, L; Sacco, D; D'Emilia, E; Cricenti, A; Generosi, R; Luce, M; Grimaldi, S

2004-01-01

In this study we have employed atomic force microscopy (AFM) and scanning near-field optical microscopy (SNOM) techniques to study the effect of the interaction between human keratinocytes (HaCaT) and electromagnetic fields at low frequency. HaCaT cells were exposed to a sinusoidal magnetic field at a density of 50 Hz, 1 mT. AFM analysis revealed modification in shape and morphology in exposed cells with an increase in the areas of adhesion between cells. This latter finding was confirmed by SNOM indirect immunofluorescence analysis performed with a fluorescent antibody against the adhesion marker beta4 integrin, which revealed an increase of beta4 integrin segregation in the cell membrane of 50-Hz exposed cells, suggesting that a higher percentage of these cells shows a modified pattern of this adhesion marker.

Diagnosis of herpes simplex virus infection by immunofluorescence.

PubMed Central

Taber, L H; Brasier, F; Couch, R B; Greenberg, S B; Jones, D; Knight, V

1976-01-01

The utility of the indirect immunofluorescent antibody (IFA) technique for diagnosis of herpes simplex virus (HSV) infection was examined by testing specimens for this agent from 31 patients with encephalitis or meningitis, 17 with conjunctivitis, 19 with genital disease, and 1 with genital disease and meningitis. Brain biopsy tissue from four patients with encephalitis was positive by IFA and virus culture for HSV. Leukocytes in cerebrospinal fluid from these four patients and one with HSV meningitis were also positive by IFA, but virus isolation attempts on the fluid were all negative. Conjunctival scrapings from two patients with conjunctivitis were positive for HSV by both IFA and virus culture. Eleven of 12 culture-positive lesions of herpes progenitalis were positive by IFA, and 1 dark field-positive syphilitic chancre was also positive for HSV by both IFA and culture. Evidence for specificity of the results was provided by internal controls in each test and negative results from patients with other diagnoses. Thus, the IFA technique constituted a rapid, sensitive, and specific diagnostic method for the diagnosis of HSV infections. PMID:178689

Double-label immunofluorescence method for simultaneous detection of adenovirus and herpes simplex virus from the eye.

PubMed

Walpita, P; Darougar, S

1989-07-01

The development and application of a double-label immunofluorescence method which has the potential to screen for single or dual infections from any site, in single shell vial cultures, is described. In this study, a total of 1,141 ocular specimens were inoculated in shell vials, centrifuged at 15,000 X g for 1 h, incubated at 37 degrees C for 48 h, and fixed in methanol at room temperature for 15 min. The virus inclusions were detected by staining with a double-label indirect immunofluorescence procedure using mixtures of appropriate first antibodies, followed by fluorescein- and rhodamine-conjugated second antibodies. Each specimen was also inoculated in parallel by the conventional virus isolation method. The sensitivity and specificity of the double-label shell vial procedure were comparable to those with the conventional method, and the former test took only 48 h to complete. The test offers a rapid and simple single-vial procedure which allows for individual or simultaneous detection of multiple pathogens. It results in savings in time and cost over the conventional virus isolation method and other shell vial procedures.

Quantum dot immunocytochemical localization of somatostatin in somatostatinoma by Widefield Epifluorescence, super-resolution light, and immunoelectron microscopy.

PubMed

Killingsworth, Murray C; Lai, Ken; Wu, Xiaojuan; Yong, Jim L C; Lee, C Soon

2012-11-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy.

Quantum Dot Immunocytochemical Localization of Somatostatin in Somatostatinoma by Widefield Epifluorescence, Super-resolution Light, and Immunoelectron Microscopy

PubMed Central

Lai, Ken; Wu, Xiaojuan; Yong, Jim L. C.; Lee, C. Soon

2012-01-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy. PMID:22899862

Immunofluorescent studies on human spermatozoa. III. Immunoglobulin classes of human spermatozoal antibodies

PubMed Central

Hansen, K. Brogaard

1972-01-01

Antibodies in human sera against four different antigens of human spermatozoa discovered by means of an indirect two-layer immunofluorescence technique (IFT) were characterized by determination of the class of immunoglobulins to which they belonged. A three-layer IFT using monospecific antisera against human IgG, IgA or IgM as the second layer was employed together with fractionation of sera on Sephadex G-200 or DEAE-cellulose followed by testing of the concentrated pools in a two-layer IFT. The study revealed that antibodies against the antigen in the front part of the acrosome were primarily IgM and those against the antigen in the tail primarily IgG. Antibodies against antigens in the equatorial segment and the postnuclear cap showed a varying predominance of these two immunoglobulins. Spermatozoal antibody as IgA was found only in small amounts. PMID:4558409

Kidney lesions in Rocky Mountain spotted fever: a light-, immunofluorescence-, and electron-microscopic study.

PubMed Central

Bradford, W. D.; Croker, B. P.; Tisher, C. C.

1979-01-01

The essential pathologic lesion in Rocky Mountain spotted fever (RMSF) is a vasculitis that may involve the kidneys as well as the heart, brain, skin, and subcutaneous tissues. Histopathologic information concerning the response of the kidneys in RMSF is rather limited, however. In this study renal tissue from 17 children who died of RMSF was examined by light, electron, and immunofluorescence microscopy. A lymphocytic or mixed inflammation, or both, involving vessels and interstitium of the kidney was found in all patients. In addition, 10 patients had histologic evidence of acute tubular necrosis, and another 3 had glomerular lesions consisting of focal segmental tuft necrosis or increased cellularity secondary to neutophilic infiltration, or both. Immunofluorescence- and electron-microscopic studies failed to demonstrate immune-complex deposition within glomeruli, a finding that suggests that immunoglobulin and classic immune complexes were not involved in the pathogenesis of the renal lesions at the time of death. These findings suggest the possibility that the pathogenesis of the renal lesion in RMSF may be due to a direct action of the organism (Rickettsia rickettsii) on the vessel wall. Images Figure 2 Figure 1 PMID:525676

Characterization of myosin heavy chain and its gene in Amoeba proteus.

PubMed

Oh, S W; Jeon, K W

1998-01-01

Monoclonal antibodies against the myosin heavy chain of Amoeba proteus were obtained and used to localize myosin inside amoebae and to clone cDNAs encoding myosin. Myosin was found throughout the amoeba cytoplasm but was more concentrated in the ectoplasmic regions as determined by indirect immunofluorescence microscopy. In symbiont-bearing xD amoebae, myosin was also found on the symbiosome membranes, as checked by indirect immunofluorescence microscopy and by immunoelectron microscopy. The open reading frame of a cloned myosin cDNA contained 6,414 nucleotides, coding for a polypeptide of 2,138 amino acids. While the amino-acid sequence of the globular head region of amoeba's myosin had a high degree of similarity with that of myosins from various organisms, the tail region building a coiled-coil structure did not show a significant sequence similarity. There appeared to be at least three different isoforms of myosins in amoebae, with closely related amino acids in the globular head region.

Improved Serial Sectioning Techniques for Correlative Light-Electron Microscopy Mapping of Human Langerhans Islets

PubMed Central

Saitoh, Sei; Ohno, Nobuhiko; Saitoh, Yurika; Terada, Nobuo; Shimo, Satoshi; Aida, Kaoru; Fujii, Hideki; Kobayashi, Tetsuro; Ohno, Shinichi

2018-01-01

Combined analysis of immunostaining for various biological molecules coupled with investigations of ultrastructural features of individual cells is a powerful approach for studies of cellular functions in normal and pathological conditions. However, weak antigenicity of tissues fixed by conventional methods poses a problem for immunoassays. This study introduces a method of correlative light and electron microscopy imaging of the same endocrine cells of compact and diffuse islets from human pancreatic tissue specimens. The method utilizes serial sections obtained from Epon-embedded specimens fixed with glutaraldehyde and osmium tetroxide. Double-immunofluorescence staining of thick Epon sections for endocrine hormones (insulin and glucagon) and regenerating islet-derived gene 1 α (REG1α) was performed following the removal of Epoxy resin with sodium ethoxide, antigen retrieval by autoclaving, and de-osmification treatment with hydrogen peroxide. The immunofluorescence images of endocrine cells were superimposed with the electron microscopy images of the same cells obtained from serial ultrathin sections. Immunofluorescence images showed well-preserved secretory granules in endocrine cells, whereas electron microscopy observations demonstrated corresponding secretory granules and intracellular organelles in the same cells. In conclusion, the correlative imaging approach developed by us may be useful for examining ultrastructural features in combination with immunolocalisation of endocrine hormones in the same human pancreatic islets. PMID:29622846

Immunofluorescence Staining Protocols for Major Autophagy Proteins Including LC3, P62, and ULK1 in Mammalian Cells in Response to Normoxia and Hypoxia.

PubMed

Li, Wen; Li, Shupeng; Li, Yifang; Lin, Xiaoying; Hu, Yongquan; Meng, Tian; Wu, Baojin; He, Rongrong; Feng, Du

2018-03-27

Immunofluorescence is an invaluable technique widely used in cell biology. This technique allows visualization of the subcellular distribution of different target proteins or organelles, by specific recognition of the antibody to the endogenous protein itself or to its antigen via the epitope. This technique can be used on tissue sections, cultured cells, or individual cells. Meanwhile, immunofluorescence can also be used in combination with non-antibody fluorescent staining, such as DAPI or fluorescent fusion proteins, e.g., GFP or YFP, etc.Autophagy is a catabolic pathway in which dysfunctional organelles and cellular components are degraded via lysosomes. During this process, cytoplasmic LC3 translocates to autophagosomal membranes. Therefore, cells undergoing autophagy can be identified by visualizing fluorescently labeled LC3 or other autophagy markers. Immunofluorescence is an important part of autophagy detection methods even if observation of the formation of autophagosome by transmission electron microscopy has become a gold standard for characterizing autophagy.By observing the immunofluorescence staining of some key autophagy proteins, we can intuitively evaluate the levels of autophagy in samples. Herein, this protocol describes the predominant method used for the research of autophagy, which mainly focuses on the immunofluorescence staining of cellular LC3, P62, and ULK1 in response to normoxia and hypoxia, by presenting the detailed materials required and methodology.

Comparison of direct and indirect methods of measuring airborne chrysotile fibre concentration.

PubMed

Eypert-Blaison, Celine; Veissiere, Sylvie; Rastoix, Olivier; Kauffer, Edmond

2010-01-01

Transmission electron microscopy observations most frequently form a basis for estimating asbestos fibre concentration in the environment and in buildings with asbestos-containing materials. Sampled fibres can be transferred to microscope grids by applying either a direct [ISO (1995) Draft International ISO/DIS 10312. Ambient air. Determination of asbestos fibres. Direct transfer transmission electron microscopy procedure. Geneva, Switzerland: International Standardization Organization] or an indirect [AFNOR (1996) Détermination de la concentration en fibres d'amiante par microscopie électronique à transmission-Méthode indirecte. Cedex, France: AFNOR, p. 42; ISO (1997) Draft International ISO/DIS 13794. Ambient air. Determination of asbestos fibres. Indirect-transfer transmission electron microscopy procedure. Geneva, Switzerland: International Standardization Organization] method. In the latter case, ISO Standard 13794 recommends filtering calcination residues either on a polycarbonate (PC) filter (PC indirect method) or on a cellulose ester (CE) membrane (CE indirect method). The PC indirect method requires that fibres deposited on a PC filter be covered by a carbon layer, whereas in the CE indirect method, the CE membrane has to be directly processed using a method described in ISO Standard 10312. The purpose of this study was to compare results obtained using, on the one hand, direct preparation methods and, on the other hand, PC indirect or CE indirect methods, for counting asbestos fibres deposited on filters as a result of liquid filtration or air sampling. In direct method-based preparation, we observed that an etching time of 6-14 min does not affect the measured densities, except for fibres <1 microm deposited by liquid filtration. Moreover, in all cases, the direct method gives higher densities than the PC indirect method because of possible fibre disappearance when using the carbon evaporator implemented in the PC indirect method. The CE membrane

Immunofluorescence detection of pea protein in meat products.

PubMed

Petrášová, Michaela; Pospiech, Matej; Tremlová, Bohuslava; Javůrková, Zdeňka

2016-08-01

In this study we developed an immunofluorescence method to detect pea protein in meat products. Pea protein has a high nutritional value but in sensitive individuals it may be responsible for causing allergic reactions. We produced model meat products with various additions of pea protein and flour; the detection limit (LOD) of the method for pea flour was 0.5% addition, and for pea protein it was 0.001% addition. The repeatabilities and reproducibilities for samples both positive and negative for pea protein were all 100%. In a blind test with model products and commercial samples, there was no statistically significant difference (p > 0.05) between the declared concentrations of pea protein and flour and the immunofluorescence method results. Sensitivity was 1.06 and specificity was 1.00. These results show that the immunofluorescence method is suitable for the detection of pea protein in meat products.

Optimization of Single- and Dual-Color Immunofluorescence Protocols for Formalin-Fixed, Paraffin-Embedded Archival Tissues.

PubMed

Kajimura, Junko; Ito, Reiko; Manley, Nancy R; Hale, Laura P

2016-02-01

Performance of immunofluorescence staining on archival formalin-fixed paraffin-embedded human tissues is generally not considered to be feasible, primarily due to problems with tissue quality and autofluorescence. We report the development and application of procedures that allowed for the study of a unique archive of thymus tissues derived from autopsies of individuals exposed to atomic bomb radiation in Hiroshima, Japan in 1945. Multiple independent treatments were used to minimize autofluorescence and maximize fluorescent antibody signals. Treatments with NH3/EtOH and Sudan Black B were particularly useful in decreasing autofluorescent moieties present in the tissue. Deconvolution microscopy was used to further enhance the signal-to-noise ratios. Together, these techniques provide high-quality single- and dual-color fluorescent images with low background and high contrast from paraffin blocks of thymus tissue that were prepared up to 60 years ago. The resulting high-quality images allow the application of a variety of image analyses to thymus tissues that previously were not accessible. Whereas the procedures presented remain to be tested for other tissue types and archival conditions, the approach described may facilitate greater utilization of older paraffin block archives for modern immunofluorescence studies. © 2016 The Histochemical Society.

Quick Fluorescent In Situ Hybridization Protocol for Xist RNA Combined with Immunofluorescence of Histone Modification in X-chromosome Inactivation

PubMed Central

Yamada, Norishige; Ogawa, Akiyo; Ogawa, Yuya

2014-01-01

Combining RNA fluorescent in situ hybridization (FISH) with immunofluorescence (immuno-FISH) creates a technique that can be employed at the single cell level to detect the spatial dynamics of RNA localization with simultaneous insight into the localization of proteins, epigenetic modifications and other details which can be highlighted by immunofluorescence. X-chromosome inactivation is a paradigm for long non-coding RNA (lncRNA)-mediated gene silencing. X-inactive specific transcript (Xist) lncRNA accumulation (called an Xist cloud) on one of the two X-chromosomes in mammalian females is a critical step to initiate X-chromosome inactivation. Xist RNA directly or indirectly interacts with various chromatin-modifying enzymes and introduces distinct epigenetic landscapes to the inactive X-chromosome (Xi). One known epigenetic hallmark of the Xi is the Histone H3 trimethyl-lysine 27 (H3K27me3) modification. Here, we describe a simple and quick immuno-FISH protocol for detecting Xist RNA using RNA FISH with multiple oligonucleotide probes coupled with immunofluorescence of H3K27me3 to examine the localization of Xist RNA and associated epigenetic modifications. Using oligonucleotide probes results in a shorter incubation time and more sensitive detection of Xist RNA compared to in vitro transcribed RNA probes (riboprobes). This protocol provides a powerful tool for understanding the dynamics of lncRNAs and its associated epigenetic modification, chromatin structure, nuclear organization and transcriptional regulation. PMID:25489864

Sudan Black B masks Mycobacterium avium subspecies paratuberculosis immunofluorescent antibody labeling

USDA-ARS?s Scientific Manuscript database

Autofluorescence and non-specific immunofluorescent labeling are common challenges associated with immunofluorescence experiments. Autofluorescence typically demonstrates a broad emission spectrum, increasing the potential for overlap with experiments that utilize multiple fluorophores. During immun...

Immunofluorescent detection in the ovary of host antibodies against a secretory ferritin injected into female Haemaphysalis longicornis ticks.

PubMed

Galay, Remil Linggatong; Matsuo, Tomohide; Hernandez, Emmanuel Pacia; Talactac, Melbourne Rio; Kusakisako, Kodai; Umemiya-Shirafuji, Rika; Mochizuki, Masami; Fujisaki, Kozo; Tanaka, Tetsuya

2018-04-01

Due to the continuous threat of ticks and tick-borne diseases to human and animal health worldwide, and the drawbacks of chemical acaricide application, many researchers are exploring vaccination as an alternative tick control method. Earlier studies have shown that host antibodies can circulate in the ticks, but it has not been confirmed whether these antibodies can be passed on to the eggs. We previously reported that ticks infesting rabbits immunized with a recombinant secretory ferritin of Haemaphysalis longicornis (HlFER2) had reduced egg production and hatching. Here we attempted to detect the presence of antibodies against HlFER2 in the ovary and eggs of female ticks through immunofluorescent visualization. Purified anti-HlFER2 antibodies or rabbit IgG for control was directly injected to engorged female H. longicornis. Ovaries and eggs after oviposition were collected and prepared for an indirect immunofluorescent antibody test. Positive fluorescence was detected in ovaries one day post-injection of anti-HlFER2 antibodies. Through silencing of Hlfer2 gene, we also determined whether the injected antibodies can specifically bind to native HlFER2. Immunofluorescence was observed in the oocytes of dsLuciferase control ticks injected with anti-HlFER2 antibodies, but not in the oocytes of Hlfer2-silenced ticks also injected with anti-HlFER2 antibodies. Our current findings suggest that host antibodies can be passed on to the oocytes, which is significant in formulating a vaccine that can disrupt tick reproduction. Copyright © 2017 Elsevier B.V. All rights reserved.

Interpretation of ANA Indirect Immunofluorescence Test Outside the Darkroom Using NOVA View Compared to Manual Microscopy

PubMed Central

Copple, Susan S.; Jaskowski, Troy D.; Giles, Rashelle; Hill, Harry R.

2014-01-01

Objective. To evaluate NOVA View with focus on reading archived images versus microscope based manual interpretation of ANA HEp-2 slides by an experienced, certified medical technologist. Methods. 369 well defined sera from: 44 rheumatoid arthritis, 50 systemic lupus erythematosus, 35 scleroderma, 19 Sjögren's syndrome, and 10 polymyositis patients as well as 99 healthy controls were examined. In addition, 12 defined sera from the Centers for Disease Control and 100 random patient sera sent to ARUP Laboratories for ANA HEp-2 IIF testing were included. Samples were read using the archived images on NOVA View and compared to results obtained from manual reading. Results. At a 1 : 40/1 : 80 dilution the resulting comparison demonstrated 94.8%/92.9% positive, 97.4%/97.4% negative, and 96.5%/96.2% total agreements between manual IIF and NOVA View archived images. Agreement of identifiable patterns between methods was 97%, with PCNA and mixed patterns undetermined. Conclusion. Excellent agreements were obtained between reading archived images on NOVA View and manually on a fluorescent microscope. In addition, workflow benefits were observed which need to be analyzed in future studies. PMID:24741573

Magneto immunofluorescence assay for diagnosis of celiac disease.

PubMed

Kergaravat, Silvina V; Beltramino, Luis; Garnero, Nidia; Trotta, Liliana; Wagener, Marta; Fabiano, Silvia N; Pividori, Maria Isabel; Hernandez, Silvia R

2013-10-10

A magneto immunofluorescence assay for the detection of anti-transglutaminase antibodies (ATG2) in celiac disease was developed. The ATG2 were recognized by transglutaminase enzyme immobilized on the magnetic beads and then the immunological reaction was revealed by antibodies labeled with peroxidase. The fluorescent response of the enzymatic reaction with o-phenylenediamine and H2O2 as substrates was correlated with anti-transglutaminase titer, showing EC50 and LOD values of 1:11,600 and 1:74,500 of antibody titers, respectively. A total number of 29 sera samples from clinically confirmed cases of celiac disease and 19 negative control samples were tested by the novel magneto immunofluorescence assay. The data were submitted to the receiver-operating characteristic plot (ROC) analysis which indicated that 8.1 U was the most effective cut-off value to discriminate correctly between celiac and non-celiac patients. The immunofluorescence assay exhibited a sensitivity of 96.6%, a specificity of 89.5% and an efficiency 93.8% compared with the commercial optical ELISA kit. Copyright © 2013 Elsevier B.V. All rights reserved.

Open-source do-it-yourself multi-color fluorescence smartphone microscopy

PubMed Central

Sung, Yulung; Campa, Fernando; Shih, Wei-Chuan

2017-01-01

Fluorescence microscopy is an important technique for cellular and microbiological investigations. Translating this technique onto a smartphone can enable particularly powerful applications such as on-site analysis, on-demand monitoring, and point-of-care diagnostics. Current fluorescence smartphone microscope setups require precise illumination and imaging alignment which altogether limit its broad adoption. We report a multi-color fluorescence smartphone microscope with a single contact lens-like add-on lens and slide-launched total-internal-reflection guided illumination for three common tasks in investigative fluorescence microscopy: autofluorescence, fluorescent stains, and immunofluorescence. The open-source, simple and cost-effective design has the potential for do-it-yourself fluorescence smartphone microscopy. PMID:29188104

High-resolution imaging by scanning electron microscopy of semithin sections in correlation with light microscopy.

PubMed

Koga, Daisuke; Kusumi, Satoshi; Shodo, Ryusuke; Dan, Yukari; Ushiki, Tatsuo

2015-12-01

In this study, we introduce scanning electron microscopy (SEM) of semithin resin sections. In this technique, semithin sections were adhered on glass slides, stained with both uranyl acetate and lead citrate, and observed with a backscattered electron detector at a low accelerating voltage. As the specimens are stained in the same manner as conventional transmission electron microscopy (TEM), the contrast of SEM images of semithin sections was similar to TEM images of ultrathin sections. Using this technique, wide areas of semithin sections were also observed by SEM, without the obstruction of grids, which was inevitable for traditional TEM. This study also applied semithin section SEM to correlative light and electron microscopy. Correlative immunofluorescence microscopy and immune-SEM were performed in semithin sections of LR white resin-embedded specimens using a FluoroNanogold-labeled secondary antibody. Because LR white resin is hydrophilic and electron stable, this resin is suitable for immunostaining and SEM observation. Using correlative microscopy, the precise localization of the primary antibody was demonstrated by fluorescence microscopy and SEM. This method has great potential for studies examining the precise localization of molecules, including Golgi- and ER-associated proteins, in correlation with LM and SEM. © The Author 2015. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Immunofluorescence Staining — EDRN Public Portal

Cancer.gov

Direct immunofluorescence method is used to detect the deposit of immunoglobulins, complement components, fibrinogen, etc. in tissues. This technique is usually performed on frozen sections. The primary antibody is conjugated to fluorescein binds directly with the antigen and can be detected by the fluorescent tag using a fluorescent microscope.

In Situ Detection of Anaplasma spp. by DNA Target-Primed Rolling-Circle Amplification of a Padlock Probe and Intracellular Colocalization with Immunofluorescently Labeled Host Cell von Willebrand Factor ▿

PubMed Central

Wamsley, Heather L.; Barbet, Anthony F.

2008-01-01

Endothelial cell culture and preliminary immunofluorescent staining of Anaplasma-infected tissues suggest that endothelial cells may be an in vivo nidus of mammalian infection. To investigate endothelial cells and other potentially cryptic sites of Anaplasma sp. infection in mammalian tissues, a sensitive and specific isothermal in situ technique to detect localized Anaplasma gene sequences by using rolling-circle amplification of circularizable, linear, oligonucleotide probes (padlock probes) was developed. Cytospin preparations of uninfected or Anaplasma-infected cell cultures were examined using this technique. Via fluorescence microscopy, the technique described here, and a combination of differential interference contrast microscopy and von Willebrand factor immunofluorescence, Anaplasma phagocytophilum and Anaplasma marginale were successfully localized in situ within intact cultured mammalian cells. This work represents the first application of this in situ method for the detection of a microorganism and forms the foundation for future applications of this technique to detect, localize, and analyze Anaplasma nucleotide sequences in the tissues of infected mammalian and arthropod hosts and in cell cultures. PMID:18495855

Differential serological testing by simultaneous indirect immunofluorescent antibody test in canine leishmaniosis and ehrlichiosis.

PubMed

Guillén Llera, J L; López García, M L; Martín Reinoso, E; De Vivar González, R

2002-11-11

A mixed indirect fluorescence antibody test (IFAT), based on cultured promastigotes Leishmania infantum and formol-inactivated suspension of cells infected with the bacteria Ehrlichia canis, was applied to make a differential diagnosis between canine ehrlichiosis and leishmaniosis. A titre greater than 80 was considered positive for antibodies to E. canis and suggestive of antibodies to L. infantum. Positive sera were titrated subsequently by serial dilutions to confirm antibodies positive to Leishmania and establishing the antibody titre of both pathogens. Fluorescence was absent with negative control sera and background staining was minimal. No serological cross-reactions between positive sera for L. infantum or E. canis were detected. Results obtained by mixed IFAT did not differ when the same serum IFAT standard was compared. The test showed equivalent sensitivity (100%). The specifities were 100% for L. infantum and 98.5% for E. canis. The equivalence in sensitivity was confirmed by calculating the correlation coefficient between IFAT standards and mixed IFAT (r>or=0.99 for both pathogens). The results of our investigations demonstrated that mixed IFAT is a specific means of establishing serological differential diagnosis of canine leishmaniosis and ehrlichiosis.

AMP deaminase histochemical activity and immunofluorescent isozyme localization in rat skeletal muscle

NASA Technical Reports Server (NTRS)

Thompson, J. L.; Sabina, R. L.; Ogasawara, N.; Riley, D. A.

1992-01-01

The cellular distribution of AMP deaminase (AMPda) isozymes was documented for rat soleus and plantaris muscles, utilizing immunofluorescence microscopy and immunoprecipitation methods. AMPda is a ubiquitous enzyme existing as three distinct isozymes, A, B and C, which were initially purified from skeletal muscle, liver (and kidney), and heart, respectively. AMPda-A is primarily concentrated subsarcolemmally and intermyofibrillarly within muscle cells, while isozymes B and C are concentrated within non-myofiber elements of muscle tissue. AMPda-B is principally associated with connective tissues surrounding neural elements and the muscle spindle capsule, and AMPda-C is predominantly associated with circulatory elements, such as arterial and venous walls, capillary endothelium, and red blood cells. These specific localizations, combined with documented differences in kinetic properties, suggest multiple functional roles for the AMPda isozymes or temporal segregation of similar AMPda functions. Linkage of the AMPda substrate with adenosine production pathways at the AMP level and the localization of isozyme-C in vascular tissue suggest a regulatory role in the microcirculation.

Difference of EGCg adhesion on cell surface between Staphylococcus aureus and Escherichia coli visualized by electron microscopy after novel indirect staining with cerium chloride.

PubMed

Nakayama, Motokazu; Shigemune, Naofumi; Tsugukuni, Takashi; Tokuda, Hajime; Miyamoto, Takahisa

2011-07-01

We developed a novel method using indirect staining with cerium chloride for visualization of the catechin derivative epigallocatechin gallate (EGCg) on the surface of particles, i.e., polystyrene beads and bacterial cells, by electron microscopy. The staining method is based on the fact that in an alkaline environment, EGCg produces hydrogen peroxide, and then hydrogen peroxide reacts with cerium, resulting in a cerium hydroperoxide precipitate. This precipitate subsequently reacts with EGCg to produce larger deposits. The amount of precipitate is proportional to the amount of EGCg. Highly EGCg-sensitive Staphylococcus aureus and EGCg-resistant Escherichia coli were treated with EGCg under various pH conditions. Transmission electron microscopy observation showed that the amount of deposits on S. aureus increased with an increase in EGCg concentration. After treating bacterial cells with 0.5mg/mL EGCg (pH 6.0), attachment of EGCg was significantly lower to E. coli than to S. aureus. This is the first report that shows differences in affinity of EGCg to the cell surfaces of Gram-positive and -negative bacteria by electron microscopy. Copyright © 2011 Elsevier B.V. All rights reserved.

Autoradiography and Immunofluorescence Combined for Autecological Study of Single Cell Activity with Nitrobacter as a Model System1

PubMed Central

Fliermans, C. B.; Schmidt, E. L.

1975-01-01

Specific detection of a particular bacterium by immunofluorescence was combined with estimation of its metabolic activity by autoradiography. The nitrifying bacteria Nitrobacter agilis and N. winogradskyi were used as a model system. Nitrobacter were incubated with NaH14CO3 and 14CO2 prior to study. The same preparations made for autoradiograms were stained with fluorescent antibodies specific for the Nitrobacter species. Examination by epifluorescence and transmitted dark-field microscopy revealed Nitrobacter cells with and without associated silver grains. Direct detection and simultaneous evaluation of metabolic activity of Nitrobacter was demonstrated in pure cultures, in a simple mixed culture, and in a natural soil. Images PMID:1103733

Local viscoelastic response of direct and indirect dental restorative composites measured by AFM.

PubMed

Grattarola, Laura; Derchi, Giacomo; Diaspro, Alberto; Gambaro, Carla; Salerno, Marco

2018-06-08

We investigated the viscoelastic response of direct and indirect dental restorative composites by the novel technique of AM-FM atomic force microscopy. We selected four composites for direct restorations (Adonis, Optifil, EPH, CME) and three composites for indirect restorations (Gradia, Estenia, Signum). Scanning electron microscopy with micro-analysis was also used to support the results. The mean storage modulus of all composites was in the range of 10.2-15.2 GPa. EPH was the stiffest (p<0.05 vs. all other composites but Adonis and Estenia), while no significant difference was observed between direct and indirect group (p≥0.05). For the loss tangent, Gradia had the highest value (~0.3), different (p<0.05) from Optifil (~0.01) and EPH (~0.04) despite the large coefficient of variation (24%), and the direct composites showed higher loss tangent (p<0.01) than the indirect composites. All composites exhibited minor contrast at the edge of fillers, showing that these are pre-polymerized, as confirmed by EDS.

Cell-based quantification of biomarkers from an ultra-fast microfluidic immunofluorescent staining: application to human breast cancer cell lines

NASA Astrophysics Data System (ADS)

Migliozzi, D.; Nguyen, H. T.; Gijs, M. A. M.

2018-02-01

Immunohistochemistry (IHC) is one of the main techniques currently used in the clinics for biomarker characterization. It consists in colorimetric labeling with specific antibodies followed by microscopy analysis. The results are then used for diagnosis and therapeutic targeting. Well-known drawbacks of such protocols are their limited accuracy and precision, which prevent the clinicians from having quantitative and robust IHC results. With our work, we combined rapid microfluidic immunofluorescent staining with efficient image-based cell segmentation and signal quantification to increase the robustness of both experimental and analytical protocols. The experimental protocol is very simple and based on fast-fluidic-exchange in a microfluidic chamber created on top of the formalin-fixed-paraffin-embedded (FFPE) slide by clamping it a silicon chip with a polydimethyl siloxane (PDMS) sealing ring. The image-processing protocol is based on enhancement and subsequent thresholding of the local contrast of the obtained fluorescence image. As a case study, given that the human epidermal growth factor receptor 2 (HER2) protein is often used as a biomarker for breast cancer, we applied our method to HER2+ and HER2- cell lines. We report very fast (5 minutes) immunofluorescence staining of both HER2 and cytokeratin (a marker used to define the tumor region) on FFPE slides. The image-processing program can segment cells correctly and give a cell-based quantitative immunofluorescent signal. With this method, we found a reproducible well-defined separation for the HER2-to-cytokeratin ratio for positive and negative control samples.

Confocal Laser Scanning Microscopy and Ultrastructural Study of VGLUT2 Thalamic Input to Striatal Projection Neurons in Rats

PubMed Central

Lei, Wanlong; Deng, Yunping; Liu, Bingbing; Mu, Shuhua; Guley, Natalie M.; Wong, Ting; Reiner, Anton

2014-01-01

We examined thalamic input to striatum in rats using immunolabeling for the vesicular glutamate transporter (VGLUT2). Double immunofluorescence viewed with confocal laser scanning microscopy (CLSM) revealed that VGLUT2+ terminals are distinct from VGLUT1+ terminals. CLSM of Phaseolus vulgaris-leucoagglutinin (PHAL)-labeled cortical or thalamic terminals revealed that VGLUT2 is rare in corticostriatal terminals but nearly always present in thalamostriatal terminals. Electron microscopy revealed that VGLUT2+ terminals made up 39.4% of excitatory terminals in striatum (with VGLUT1+ corticostriatal terminals constituting the rest), and 66.8% of VGLUT2+ terminals synapsed on spines and the remainder on dendrites. VGLUT2+ axo-spinous terminals had a mean diameter of 0.624 lm, while VGLUT2+ axodendritic terminals a mean diameter of 0.698 µm. In tissue in which we simultaneously immunolabeled thalamostriatal terminals for VGLUT2 and striatal neurons for D1 (with about half of spines immunolabeled for D1), 54.6% of VGLUT2+ terminals targeted D1+ spines (i.e., direct pathway striatal neurons), and 37.3% of D1+ spines received VGLUT2+ synaptic contacts. By contrast, 45.4% of VGLUT2+ terminals targeted D1-negative spines (i.e., indirect pathway striatal neurons), and only 25.8% of D1-negative spines received VGLUT2+ synaptic contacts. Similarly, among VGLUT2+ axodendritic synaptic terminals, 59.1% contacted D1+ dendrites, and 40.9% contacted D1-negative dendrites. VGLUT2+ terminals on D1+ spines and dendrites tended to be slightly smaller than those on D1-negative spines and dendrites. Thus, thala-mostriatal terminals contact both direct and indirect pathway striatal neurons, with a slight preference for direct. These results are consistent with physiological studies indicating slightly different effects of thalamic input on the two types of striatal projection neurons. PMID:23047588

Confocal laser scanning microscopy and ultrastructural study of VGLUT2 thalamic input to striatal projection neurons in rats.

PubMed

Lei, Wanlong; Deng, Yunping; Liu, Bingbing; Mu, Shuhua; Guley, Natalie M; Wong, Ting; Reiner, Anton

2013-04-15

We examined thalamic input to striatum in rats using immunolabeling for the vesicular glutamate transporter (VGLUT2). Double immunofluorescence viewed with confocal laser scanning microscopy (CLSM) revealed that VGLUT2+ terminals are distinct from VGLUT1+ terminals. CLSM of Phaseolus vulgaris-leucoagglutinin (PHAL)-labeled cortical or thalamic terminals revealed that VGLUT2 is rare in corticostriatal terminals but nearly always present in thalamostriatal terminals. Electron microscopy revealed that VGLUT2+ terminals made up 39.4% of excitatory terminals in striatum (with VGLUT1+ corticostriatal terminals constituting the rest), and 66.8% of VGLUT2+ terminals synapsed on spines and the remainder on dendrites. VGLUT2+ axospinous terminals had a mean diameter of 0.624 μm, while VGLUT2+ axodendritic terminals a mean diameter of 0.698 μm. In tissue in which we simultaneously immunolabeled thalamostriatal terminals for VGLUT2 and striatal neurons for D1 (with about half of spines immunolabeled for D1), 54.6% of VGLUT2+ terminals targeted D1+ spines (i.e., direct pathway striatal neurons), and 37.3% of D1+ spines received VGLUT2+ synaptic contacts. By contrast, 45.4% of VGLUT2+ terminals targeted D1-negative spines (i.e., indirect pathway striatal neurons), and only 25.8% of D1-negative spines received VGLUT2+ synaptic contacts. Similarly, among VGLUT2+ axodendritic synaptic terminals, 59.1% contacted D1+ dendrites, and 40.9% contacted D1-negative dendrites. VGLUT2+ terminals on D1+ spines and dendrites tended to be slightly smaller than those on D1-negative spines and dendrites. Thus, thalamostriatal terminals contact both direct and indirect pathway striatal neurons, with a slight preference for direct. These results are consistent with physiological studies indicating slightly different effects of thalamic input on the two types of striatal projection neurons. Copyright © 2012 Wiley Periodicals, Inc.

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Mycobacterium tuberculosis immunofluorescent reagents. 866.3370 Section 866.3370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents...

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Mycobacterium tuberculosis immunofluorescent reagents. 866.3370 Section 866.3370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents...

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Mycobacterium tuberculosis immunofluorescent reagents. 866.3370 Section 866.3370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents...

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Mycobacterium tuberculosis immunofluorescent reagents. 866.3370 Section 866.3370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents...

21 CFR 866.3370 - Mycobacterium tuberculosis immunofluorescent reagents.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Mycobacterium tuberculosis immunofluorescent reagents. 866.3370 Section 866.3370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents...

Easy performance of 6-color confocal immunofluorescence with 4-laser line microscopes.

PubMed

Eissing, Nathalie; Heger, Lukas; Baranska, Anna; Cesnjevar, Robert; Büttner-Herold, Maike; Söder, Stephan; Hartmann, Arndt; Heidkamp, Gordon F; Dudziak, Diana

2014-09-01

Confocal laser scanning microscopy is an advanced technique for imaging tissue samples in vitro and in vivo at high optical resolution. The development of new fluorochrome variants do not only make it possible to perform multicolor flow cytometry of single cells, but in combination with high resolution laser scanning systems also to investigate the distribution of cells in lymphoid tissues by confocal immunofluorescence analyses, thus allowing the distinction of various cell populations directly in the tissue. Here, we provide a protocol for the visualization of at least six differently fluorochrome-labeled antibodies at the same time using a conventional confocal laser scanning microscope with four laser lines (405 nm, 488 nm, 555 nm, and 639 nm laser wavelength) in both murine and human tissue samples. We further demonstrate that compensation correction algorithms are not necessary to reduce spillover of fluorochromes into other channels when the used fluorochromes are combined according to their specific emission bands and the varying Stokes shift for co-excited fluorochromes with the same laser line. Copyright © 2014 Elsevier B.V. All rights reserved.

Immunofluorescence of bovine virus diarrhea viral antigen in white blood cells from experimentally infected immunocompetent calves.

PubMed Central

Bezek, D M; Baker, J C; Kaneene, J B

1988-01-01

A study to evaluate the detection of bovine virus diarrhea viral antigen using immunofluorescence testing of white blood cells was conducted. Five colostrum-deprived calves were inoculated intravenously with a cytopathic strain of the virus. Lymphocyte and buffy coat smears were prepared daily for direct immunofluorescent staining for detection of antigen. Lymphocytes were separated from heparinized blood using a Ficoll density procedure. Buffy coat smears were prepared from centrifuged blood samples collected using ethylenediaminetetraacetic acid as an anticoagulant. Bovine viral diarrhea virus antigen was detected by immunofluorescence between 3 and 11 days postinfection in lymphocyte smears and 3 to 12 days postinfection in buffy coat smears. Isolation of virus from both lymphocytes and buffy coat preparations correlated with detection of immunofluorescence. Serum neutralizing antibody to bovine virus diarrhea virus was detected on day 10 postinfection. Buffy coat smears were as sensitive as lymphocyte smears for the detection of antigen by immunofluorescence. It appeared that immunofluorescent staining of white blood cells was an effective method of detecting bovine virus diarrhea viral antigen. PMID:2836047

21 CFR 866.3460 - Rabiesvirus immuno-fluorescent reagents.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Rabiesvirus immuno-fluorescent reagents. 866.3460 Section 866.3460 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3460 Rabiesvirus...

21 CFR 866.3460 - Rabiesvirus immuno-fluorescent reagents.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Rabiesvirus immuno-fluorescent reagents. 866.3460 Section 866.3460 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3460 Rabiesvirus...

21 CFR 866.3460 - Rabiesvirus immuno-fluorescent reagents.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Rabiesvirus immuno-fluorescent reagents. 866.3460 Section 866.3460 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3460 Rabiesvirus...

21 CFR 866.3460 - Rabiesvirus immuno-fluorescent reagents.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Rabiesvirus immuno-fluorescent reagents. 866.3460 Section 866.3460 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3460 Rabiesvirus...

21 CFR 866.3460 - Rabiesvirus immuno-fluorescent reagents.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Rabiesvirus immuno-fluorescent reagents. 866.3460 Section 866.3460 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3460 Rabiesvirus...

Confocal fluorescence microscopy in a murine model of microdissection testicular sperm extraction to improve sperm retrieval.

PubMed

Smith, Ryan P; Lowe, Greg J; Kavoussi, Parviz K; Steers, William D; Costabile, Raymond A; Herr, John C; Shetty, Jagathpala; Lysiak, Jeffrey J

2012-05-01

Microdissection testicular sperm extraction markedly improves the sperm retrieval rates in men with nonobstructive azoospermia. However, localizing sperm foci can be time-consuming and it is not always successful. Fiberoptic confocal fluorescent microscopy offers the advantage of rapid in vivo detection of fluorescently labeled sperm in the seminiferous tubules. After establishing the feasibility of fiberoptic confocal fluorescent microscopy to identify antibody labeled sperm in vivo C57/B6 mice underwent intraperitoneal injection of busulfan to induce azoospermia. During spermatogenesis reestablishment at approximately 16 weeks the mice were anesthetized and the testes were delivered through a low midline incision. Fluorescein isothiocyanate labeled antibody to intra-acrosomal protein Hs-14 was injected retrograde into a single murine rete testis. The testes were imaged in vivo with fiberoptic confocal fluorescent microscopy and sperm foci were detected. The respective seminiferous tubules were excised and squash prepared for immunofluorescence microscopy. Sperm foci were identified in the testis injected with fluorescently tagged antibody by in vivo fiberoptic confocal fluorescence microscopy. The contralateral control testis of each mouse showed no specific signal. Immunofluorescence microscopy of the excised tubules provided morphological confirmation of the presence of labeled sperm with an absence in controls. Findings were consistent in the feasibility portion of the study and in the busulfan model of nonobstructive azoospermia. Fiberoptic confocal fluorescent microscopy was feasible during microdissection testicular sperm extraction in an azoospermic mouse model to identify fluorescently labeled sperm in vivo. Translation to the clinical setting could decrease operative time and improve the sperm harvest rate. Copyright © 2012 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

[Laboratory diagnosis of genital herpes--direct immunofluorescence method].

PubMed

Majewska, Anna; Romejko-Wolniewicz, Ewa; Zareba-Szczudlik, Julia; Kilijańczyk, Marek; Gajewska, Małgorzata; Młynarczyk, Grazyna

2013-07-01

Aim of the study was to determine clinical usefulness of direct immunofluorescence method in the laboratory diagnosis of genital herpes in women. Overall 187 anogenital swabs were collected from 120 women. Using a dacron-tipped applicator 83 swabs were collected from women suspected of genital herpes and 104 from patients with no signs of genital infection. All samples were tested using cell culture (Vero cell line) and then direct immunofluorescence method (DIF) for the identification of antigens of herpes simplex viruses: HSV-1 and HSV-2. Characteristic cytopathic effect (CPE), indicative of alphaherpesvirus infection, was observed in 43.4% of cultures with clinical specimens collected from women with suspected genital herpes and in 29.8% of cultures of clinical specimens taken from patients with no clinical symptoms of genital herpes. Herpes simplex viruses were determined in 73 samples by direct immunofluorescence method after amplification of the virus in cell culture. The DIF test confirmed the diagnosis based on the microscopic CPE observation in 85%. In 15% of samples (taken from pregnant women without clinical signs of infection) we reported positive immunofluorescence in the absence of CPE. The frequency of antigen detection was statistically significantly higher in samples that were positive by culture study (chi-square test with Yates's correction, p < 0.01). This method proved to be highly sensitive (97%) in women with clinically suspected infection. High negative predictive value (99%) proves the clinical utility of the DIF in these group of patients. In asymptomatic infections, viral antigens were detected most frequently in the swabs from the cervical canal, and in cases of suspected genital herpes in swabs taken from the vestibule of the vagina and the vulva. However, there was no statistically significant difference in the frequency of detection of Herpes Simplex Virus antigens in specimens from different parts of the genital tract in both groups

Automated Analysis of siRNA Screens of Virus Infected Cells Based on Immunofluorescence Microscopy

NASA Astrophysics Data System (ADS)

Matula, Petr; Kumar, Anil; Wörz, Ilka; Harder, Nathalie; Erfle, Holger; Bartenschlager, Ralf; Eils, Roland; Rohr, Karl

We present an image analysis approach as part of a high-throughput microscopy screening system based on cell arrays for the identification of genes involved in Hepatitis C and Dengue virus replication. Our approach comprises: cell nucleus segmentation, quantification of virus replication level in cells, localization of regions with transfected cells, cell classification by infection status, and quality assessment of an experiment. The approach is fully automatic and has been successfully applied to a large number of cell array images from screening experiments. The experimental results show a good agreement with the expected behavior of positive as well as negative controls and encourage the application to screens from further high-throughput experiments.

Advanced Diagnostic Techniques in Autoimmune Bullous Diseases

PubMed Central

Jindal, Anuradha; Rao, Raghavendra; Bhogal, Balbir S

2017-01-01

Autoimmune blistering diseases are diverse group of conditions characterized by blisters in the skin with or without mucosal lesions. There may be great degree of clinical and histopathological overlap; hence, advanced immunological tests may be necessary for more precise diagnosis of these conditions. Direct immunofluorescence microscopy is the gold standard tests to demonstrate the tissue-bound antibodies and should be done in all cases. Magnitude of antibody level in patient’ serum can be assessed by indirect immunofluorescence and enzyme linked immunosorbent assay. In this article we have reviewed the various techniques that are available in the diagnosis of autoimmune blistering diseases. PMID:28584369

Variations of attractors and wavelet spectra of the immunofluorescence distributions for women in the pregnant period

NASA Astrophysics Data System (ADS)

Galich, Nikolay E.

2008-07-01

Communication contains the description of the immunology data treatment. New nonlinear methods of immunofluorescence statistical analysis of peripheral blood neutrophils have been developed. We used technology of respiratory burst reaction of DNA fluorescence in the neutrophils cells nuclei due to oxidative activity. The histograms of photon count statistics the radiant neutrophils populations' in flow cytometry experiments are considered. Distributions of the fluorescence flashes frequency as functions of the fluorescence intensity are analyzed. Statistic peculiarities of histograms set for women in the pregnant period allow dividing all histograms on the three classes. The classification is based on three different types of smoothing and long-range scale averaged immunofluorescence distributions, their bifurcation and wavelet spectra. Heterogeneity peculiarities of long-range scale immunofluorescence distributions and peculiarities of wavelet spectra allow dividing all histograms on three groups. First histograms group belongs to healthy donors. Two other groups belong to donors with autoimmune and inflammatory diseases. Some of the illnesses are not diagnosed by standards biochemical methods. Medical standards and statistical data of the immunofluorescence histograms for identifications of health and illnesses are interconnected. Peculiarities of immunofluorescence for women in pregnant period are classified. Health or illness criteria are connected with statistics features of immunofluorescence histograms. Neutrophils populations' fluorescence presents the sensitive clear indicator of health status.

OUR EXPERIENCE WITH IMMUNOFLUORESCENCE (RELATING TO 250 SERA STUDIED AT THE NANCY DERMATOLOGICAL CLINIC)

DTIC Science & Technology

The principle of immunofluorescence, discovered by Coons in 1942, was applied to the diagnosis of syphilis by Deacon, Falcon and Harris in 1957...technique and to determine its value (Dauget, Fribourg-Blanc, Thivolet). The research reported here, relating to 250 sera, seems to confirm the great importance of the immunofluorescence test (IF) in the diagnosis of syphilis .

Application of the Indirect Immunoperoxidase Stain Technique to the Flagella of Azospirillum brasilense

PubMed Central

Hall, Patrick G.; Krieg, Noel R.

1984-01-01

An indirect immunoperoxidase stain was used to demonstrate by electron microscopy that an antigenic difference exists between the polar flagellum and the lateral flagella of Azospirillum brasilense ATCC 29145. Images PMID:16346482

Nucleocytoplasmic shuttling: the ins and outs of quantitative imaging.

PubMed

Molenaar, Chris; Weeks, Kate L

2018-05-17

Nucleocytoplasmic protein shuttling is integral to the transmission of signals between the nucleus and the cytoplasm. The nuclear/cytoplasmic distribution of proteins of interest can be determined via fluorescence microscopy, following labelling of the target protein with fluorophore-conjugated antibodies (immunofluorescence) or by tagging the target protein with an autofluorescent protein, such as green fluorescent protein (GFP). The latter enables live cell imaging, a powerful approach that precludes many of the artefacts associated with indirect immunofluorescence in fixed cells. In this review, we discuss important considerations for the design and implementation of fluorescence microscopy experiments to quantify the nuclear/cytoplasmic distribution of a protein of interest. We summarise the pros and cons of detecting endogenous proteins in fixed cells by immunofluorescence and ectopically-expressed fluorescent fusion proteins in living cells. We discuss the suitability of widefield fluorescence microscopy and of 2D, 3D and 4D imaging by confocal microscopy for different applications, and describe two different methods for quantifying the nuclear/cytoplasmic distribution of a protein of interest from the fluorescent signal. Finally, we discuss the importance of eliminating sources of bias and subjectivity during image acquisition and post-imaging analyses. This is critical for the accurate and reliable quantification of nucleocytoplasmic shuttling. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Bayesian Validation of the Indirect Immunofluorescence Assay and Its Superiority to the Enzyme-Linked Immunosorbent Assay and the Complement Fixation Test for Detecting Antibodies against Coxiella burnetii in Goat Serum.

PubMed

Muleme, Michael; Stenos, John; Vincent, Gemma; Campbell, Angus; Graves, Stephen; Warner, Simone; Devlin, Joanne M; Nguyen, Chelsea; Stevenson, Mark A; Wilks, Colin R; Firestone, Simon M

2016-06-01

Although many studies have reported the indirect immunofluorescence assay (IFA) to be more sensitive in detection of antibodies to Coxiella burnetii than the complement fixation test (CFT), the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the assay have not been previously established for use in ruminants. This study aimed to validate the IFA by describing the optimization, selection of cutoff titers, repeatability, and reliability as well as the DSe and DSp of the assay. Bayesian latent class analysis was used to estimate diagnostic specifications in comparison with the CFT and the enzyme-linked immunosorbent assay (ELISA). The optimal cutoff dilution for screening for IgG and IgM antibodies in goat serum using the IFA was estimated to be 1:160. The IFA had good repeatability (>96.9% for IgG, >78.0% for IgM), and there was almost perfect agreement (Cohen's kappa > 0.80 for IgG) between the readings reported by two technicians for samples tested for IgG antibodies. The IFA had a higher DSe (94.8%; 95% confidence interval [CI], 80.3, 99.6) for the detection of IgG antibodies against C. burnetii than the ELISA (70.1%; 95% CI, 52.7, 91.0) and the CFT (29.8%; 95% CI, 17.0, 44.8). All three tests were highly specific for goat IgG antibodies. The IFA also had a higher DSe (88.8%; 95% CI, 58.2, 99.5) for detection of IgM antibodies than the ELISA (71.7%; 95% CI, 46.3, 92.8). These results underscore the better suitability of the IFA than of the CFT and ELISA for detection of IgG and IgM antibodies in goat serum and possibly in serum from other ruminants. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Bayesian Validation of the Indirect Immunofluorescence Assay and Its Superiority to the Enzyme-Linked Immunosorbent Assay and the Complement Fixation Test for Detecting Antibodies against Coxiella burnetii in Goat Serum

PubMed Central

Stenos, John; Vincent, Gemma; Campbell, Angus; Graves, Stephen; Warner, Simone; Devlin, Joanne M.; Nguyen, Chelsea; Stevenson, Mark A.; Wilks, Colin R.; Firestone, Simon M.

2016-01-01

Although many studies have reported the indirect immunofluorescence assay (IFA) to be more sensitive in detection of antibodies to Coxiella burnetii than the complement fixation test (CFT), the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the assay have not been previously established for use in ruminants. This study aimed to validate the IFA by describing the optimization, selection of cutoff titers, repeatability, and reliability as well as the DSe and DSp of the assay. Bayesian latent class analysis was used to estimate diagnostic specifications in comparison with the CFT and the enzyme-linked immunosorbent assay (ELISA). The optimal cutoff dilution for screening for IgG and IgM antibodies in goat serum using the IFA was estimated to be 1:160. The IFA had good repeatability (>96.9% for IgG, >78.0% for IgM), and there was almost perfect agreement (Cohen's kappa > 0.80 for IgG) between the readings reported by two technicians for samples tested for IgG antibodies. The IFA had a higher DSe (94.8%; 95% confidence interval [CI], 80.3, 99.6) for the detection of IgG antibodies against C. burnetii than the ELISA (70.1%; 95% CI, 52.7, 91.0) and the CFT (29.8%; 95% CI, 17.0, 44.8). All three tests were highly specific for goat IgG antibodies. The IFA also had a higher DSe (88.8%; 95% CI, 58.2, 99.5) for detection of IgM antibodies than the ELISA (71.7%; 95% CI, 46.3, 92.8). These results underscore the better suitability of the IFA than of the CFT and ELISA for detection of IgG and IgM antibodies in goat serum and possibly in serum from other ruminants. PMID:27122484

Nano-fEM: protein localization using photo-activated localization microscopy and electron microscopy.

PubMed

Watanabe, Shigeki; Richards, Jackson; Hollopeter, Gunther; Hobson, Robert J; Davis, Wayne M; Jorgensen, Erik M

2012-12-03

Mapping the distribution of proteins is essential for understanding the function of proteins in a cell. Fluorescence microscopy is extensively used for protein localization, but subcellular context is often absent in fluorescence images. Immuno-electron microscopy, on the other hand, can localize proteins, but the technique is limited by a lack of compatible antibodies, poor preservation of morphology and because most antigens are not exposed to the specimen surface. Correlative approaches can acquire the fluorescence image from a whole cell first, either from immuno-fluorescence or genetically tagged proteins. The sample is then fixed and embedded for electron microscopy, and the images are correlated (1-3). However, the low-resolution fluorescence image and the lack of fiducial markers preclude the precise localization of proteins. Alternatively, fluorescence imaging can be done after preserving the specimen in plastic. In this approach, the block is sectioned, and fluorescence images and electron micrographs of the same section are correlated (4-7). However, the diffraction limit of light in the correlated image obscures the locations of individual molecules, and the fluorescence often extends beyond the boundary of the cell. Nano-resolution fluorescence electron microscopy (nano-fEM) is designed to localize proteins at nano-scale by imaging the same sections using photo-activated localization microscopy (PALM) and electron microscopy. PALM overcomes the diffraction limit by imaging individual fluorescent proteins and subsequently mapping the centroid of each fluorescent spot (8-10). We outline the nano-fEM technique in five steps. First, the sample is fixed and embedded using conditions that preserve the fluorescence of tagged proteins. Second, the resin blocks are sectioned into ultrathin segments (70-80 nm) that are mounted on a cover glass. Third, fluorescence is imaged in these sections using the Zeiss PALM microscope. Fourth, electron dense structures are

Fibrinogen Demonstration in Oral Lichen Planus: An Immunofluorescence Study on Archival Tissues.

PubMed

Shirol, Pallavi D; Naik, Veena; Kale, Alka

2015-10-01

Lichen planus is a premalignant condition with minimal diagnostic aids. This study is an attempt to use paraffin embedded sections of lichen planus with immunofluorescein stain and to evaluate the immunofluorescent sections to establish pattern of fibrinogen deposition. Thirty-five paraffin embedded sections of old and new cases of oral lichen planus (study group) and five normal oral mucosa (control group) were chosen. Two sections of each (H & E) case were taken, one was stained with hematoxylin and eosin and another with fluorescein isothiocynate conjugate (FITC) polyclonal rabbit antibody against fibrinogen. Fluorescent findings were examined with a fluorescent microscope. A high statistical significant correlation was found in respect to fluorescence positivity, intensity of fluorescence and distribution of fluorescence each with p < 0.0001 and fluorescence at blood vessel walls (p = 0.0003). This study suggested that paraffin embedded sections can be successfully used in direct immunofluorescence staining in routine set up where only formalin fixed tissues are received. Paraffin embedded sections can be successfully used in direct immunofluorescence staining when only formalin fixed tissues are received.

Expression and analysis of exogenous proteins in epidermal cells.

PubMed

Dagnino, Lina; Ho, Ernest; Chang, Wing Y

2010-01-01

In this chapter we review protocols for transient transfection of primary keratinocytes. The ability to transfect primary epidermal cells regardless of their differentiation status allows the biochemical and molecular characterization of multiple proteins. We review methods to analyze exogenous protein abundance in transfected keratinocytes by immunoblot and immunoprecipitation. We also present protocols to determine the subcellular distribution of these proteins by indirect immunofluorescence microscopy approaches.

Familial primary cutaneous amyloidosis. Clinical, genetic, and immunofluorescent studies.

PubMed

Vasily, D B; Bhatia, S G; Uhlin, S R

1978-08-01

Familial primary cutaneous amyloidosis, a rare, autosomal dominant genodermatosis, affected 16 of 46 family members of German descent. Previous case reports involved families of Russian, Spanish, or Chinese descent. The finding of IgG, IgM, C3 in the amyloid deposits confirms recent reports of immunofluorescent dermal amyloid deposits.

Delay, change and bifurcation of the immunofluorescence distribution attractors in health statuses diagnostics and in medical treatment

NASA Astrophysics Data System (ADS)

Galich, Nikolay E.; Filatov, Michael V.

2008-07-01

Communication contains the description of the immunology experiments and the experimental data treatment. New nonlinear methods of immunofluorescence statistical analysis of peripheral blood neutrophils have been developed. We used technology of respiratory burst reaction of DNA fluorescence in the neutrophils cells nuclei due to oxidative activity. The histograms of photon count statistics the radiant neutrophils populations' in flow cytometry experiments are considered. Distributions of the fluorescence flashes frequency as functions of the fluorescence intensity are analyzed. Statistic peculiarities of histograms set for healthy and unhealthy donors allow dividing all histograms on the three classes. The classification is based on three different types of smoothing and long-range scale averaged immunofluorescence distributions and their bifurcation. Heterogeneity peculiarities of long-range scale immunofluorescence distributions allow dividing all histograms on three groups. First histograms group belongs to healthy donors. Two other groups belong to donors with autoimmune and inflammatory diseases. Some of the illnesses are not diagnosed by standards biochemical methods. Medical standards and statistical data of the immunofluorescence histograms for identifications of health and illnesses are interconnected. Possibilities and alterations of immunofluorescence statistics in registration, diagnostics and monitoring of different diseases in various medical treatments have been demonstrated. Health or illness criteria are connected with statistics features of immunofluorescence histograms. Neutrophils populations' fluorescence presents the sensitive clear indicator of health status.

Chemical reactivation of fluorescein isothiocyanate immunofluorescence-labeled resin-embedded samples

NASA Astrophysics Data System (ADS)

Li, Longhui; Rao, Gong; Lv, Xiaohua; Chen, Ruixi; Cheng, Xiaofeng; Wang, Xiaojun; Zeng, Shaoqun; Liu, Xiuli

2018-02-01

Resin embedding is widely used and facilitates microscopic imaging of biological tissues. In contrast, quenching of fluorescence during embedding process hinders the application of resin embedding for imaging of fluorescence-labeled samples. For samples expressing fluorescent proteins, it has been demonstrated that the weakened fluorescence could be recovered by reactivating the fluorophore with alkaline buffer. We extended this idea to immunofluorescence-labeling technology. We showed that the fluorescence of pH-sensitive fluorescein isothiocyanate (FITC) was quenched after resin embedding but reactivated after treating by alkaline buffer. We observed 138.5% fluorescence preservation ratio of reactivated state, sixfold compared with the quenched state in embedding resin, which indicated its application for fluorescence imaging of high signal-to-background ratio. Furthermore, we analyzed the chemical reactivation mechanism of FITC fluorophore. This work would show a way for high-resolution imaging of immunofluorescence-labeled samples embedded in resin.

Pathological Gleason prediction through gland ring morphometry in immunofluorescent prostate cancer images

NASA Astrophysics Data System (ADS)

Scott, Richard; Khan, Faisal M.; Zeineh, Jack; Donovan, Michael; Fernandez, Gerardo

2016-03-01

The Gleason score is the most common architectural and morphological assessment of prostate cancer severity and prognosis. There have been numerous quantitative techniques developed to approximate and duplicate the Gleason scoring system. Most of these approaches have been developed in standard H and E brightfield microscopy. Immunofluorescence (IF) image analysis of tissue pathology has recently been proven to be extremely valuable and robust in developing prognostic assessments of disease, particularly in prostate cancer. There have been significant advances in the literature in quantitative biomarker expression as well as characterization of glandular architectures in discrete gland rings. In this work we leverage a new method of segmenting gland rings in IF images for predicting the pathological Gleason; both the clinical and the image specific grade, which may not necessarily be the same. We combine these measures with nuclear specific characteristics as assessed by the MST algorithm. Our individual features correlate well univariately with the Gleason grades, and in a multivariate setting have an accuracy of 85% in predicting the Gleason grade. Additionally, these features correlate strongly with clinical progression outcomes (CI of 0.89), significantly outperforming the clinical Gleason grades (CI of 0.78). This work presents the first assessment of morphological gland unit features from IF images for predicting the Gleason grade.

Immunofluorescence localization of dissociation supernatant and extracellular matrix components in Lytechinus pictus sectioned embryos. M.S. Thesis

NASA Technical Reports Server (NTRS)

Garciaflack, Ana Leticia

1988-01-01

Indirect immunofluorescence was used to localize specific extracellular components in embryos of the sea urchin Lytechinus pictus. Hyalin and S2 (a group of components found in the disaggregation supernatant from Strongylocentrotus purpuratus blastulae) were uniformly present at all stages (unfertilized up to 32 hr) except hyalin could not be detected at the 12 hour early blastula stage. Laminin was found in 16 cell, 32 cell, 6 hour, 18 hour, 24 hour, and 32 hour stages, with especially bright fluorescence at 18 hours. Collagen I was present at all stages (freshly fertilized up to 32 hour) except little was detected at 12 hours. Fibronectin was uniformly present in blastocoelar fibers stained with anto-collagen I and anti-fibronectin. These results were compared with those for S. purpuratus to produce an overview of the localization of specific extracellular matrix components during development of two species of sea urchins. The results set the stage for future studies that will examine the function of these components at the various developmental stages.

Methylation and in vivo expression of the surface-exposed Leptospira interrogans outer-membrane protein OmpL32.

PubMed

Eshghi, Azad; Pinne, Marija; Haake, David A; Zuerner, Richard L; Frank, Ami; Cameron, Caroline E

2012-03-01

Recent studies have revealed that bacterial protein methylation is a widespread post-translational modification that is required for virulence in selected pathogenic bacteria. In particular, altered methylation of outer-membrane proteins has been shown to modulate the effectiveness of the host immune response. In this study, 2D gel electrophoresis combined with MALDI-TOF MS identified a Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 protein, corresponding to ORF LIC11848, which undergoes extensive and differential methylation of glutamic acid residues. Immunofluorescence microscopy implicated LIC11848 as a surface-exposed outer-membrane protein, prompting the designation OmpL32. Indirect immunofluorescence microscopy of golden Syrian hamster liver and kidney sections revealed expression of OmpL32 during colonization of these organs. Identification of methylated surface-exposed outer-membrane proteins, such as OmpL32, provides a foundation for delineating the role of this post-translational modification in leptospiral virulence.

GP50 as a promising early diagnostic antigen for Taenia multiceps infection in goats by indirect ELISA.

PubMed

Huang, Xing; Xu, Jing; Wang, Yu; Guo, Cheng; Chen, Lin; Gu, Xiaobin; Lai, Weimin; Peng, Xuerong; Yang, Guangyou

2016-12-01

Coenurosis is caused by coenurus, the metacestode of Taenia multiceps, which mainly parasitizes the brain and spinal cord of cattle, sheep and goats. To date, no widely-approved methods are available to identify early coenurus infection. In this study, we identified a full-length cDNA that encodes GP50 (TmGP50) from the transcriptome of T. multiceps, and then cloned and expressed in E. coli. The native proteins in adult stage and coenurus were located via immunofluorescence assays, while the potential of recombinant TmGP50 protein (rTmGP50) for indirect ELISA-based serodiagnostics was assessed using native goat sera. In addition, we orally infected 20 goats with mature T. multiceps eggs. Praziquantel (10%) was given to 10 of the goats 45 days post-infection (p.i.). Blood samples were collected for 17 weeks p.i. from the 20 goats and anti-rTmGP50 antibodies were evaluated using the indirect ELISA established here. The TmGP50 contains an 897 bp open reading frame, in which signal sequence resides in 1 ~ 48 sites and mature polypeptide consists of 282 amino acid residues. Immunofluorescence staining showed that native TmGP50 was localized to the microthrix and parenchymatous zone of the adult parasite and coenurus, and the coenurus cystic wall. The indirect ELISA based on rTmGP50 exhibited a sensitivity of 95.0% and a specificity of 92.6% when detecting GP50 antibodies in sera of naturally infected goats and sheep. In goats experimentally infected with T. multiceps, anti-TmGP50 antibody was detectable from 2 to 17 weeks p.i. in the control group, while the antibody fell below the cut-off value about 3 weeks after praziquantel treatment. Our results indicate that recombinant TmGP50 is a suitable early diagnostic antigen for coenurus infection in goats.

Visualization of early influenza A virus trafficking in human dendritic cells using STED microscopy.

PubMed

Baharom, Faezzah; Thomas, Oliver S; Lepzien, Rico; Mellman, Ira; Chalouni, Cécile; Smed-Sörensen, Anna

2017-01-01

Influenza A viruses (IAV) primarily target respiratory epithelial cells, but can also replicate in immune cells, including human dendritic cells (DCs). Super-resolution microscopy provides a novel method of visualizing viral trafficking by overcoming the resolution limit imposed by conventional light microscopy, without the laborious sample preparation of electron microscopy. Using three-color Stimulated Emission Depletion (STED) microscopy, we visualized input IAV nucleoprotein (NP), early and late endosomal compartments (EEA1 and LAMP1 respectively), and HLA-DR (DC membrane/cytosol) by immunofluorescence in human DCs. Surface bound IAV were internalized within 5 min of infection. The association of virus particles with early endosomes peaked at 5 min when 50% of NP+ signals were also EEA1+. Peak association with late endosomes occurred at 15 min when 60% of NP+ signals were LAMP1+. At 30 min of infection, the majority of NP signals were in the nucleus. Our findings illustrate that early IAV trafficking in human DCs proceeds via the classical endocytic pathway.

Comparison of Immunofluorescence and Desmoglein Enzyme-linked Immunosorbent Assay in the Diagnosis of Pemphigus: A Prospective, Cross-sectional Study in a Tertiary Care Hospital

PubMed Central

Ravi, Deepthi; Prabhu, S Smitha; Rao, Raghavendra; Balachandran, C; Bairy, Indira

2017-01-01

Background: Pemphigus is an acquired immunobullous disorder in which antibodies are directed against epidermal cadherins. Despite the commercial availability and less cost of enzyme-linked immunosorbent assays (ELISAs) to detect antidesmoglein 1 (Dsg1) and anti-Dsg3, immunofluorescence is still widely used for confirmation of diagnosis. Aims: (1) To compare the usefulness of indirect immunofluorescence (IIF) and ELISA tests in the diagnosis of pemphigus. (2) To find the clinical correlation between the tests and severity of the disease. Materials and Methods: Sixty-one patients (27 women and 34 men, age distribution from 20 to 75) were clinically diagnosed as pemphigus (pemphigus foliaceus - 11, pemphigus vulgaris - 50) and were recruited for the study. IIF and Dsg ELISA were performed and the findings were compared with each other and with the pemphigus area activity score. Data were entered in SPSS and were analyzed using Kruskal–Wallis test. Results: There was a moderate positive correlation between the cutaneous score and Dsg1 titer, and mucosal score and Dsg3 titer. The titer of IIF showed statistically significant positive correlation with the cutaneous score but not the mucosal score. Dsg ELISA showed higher sensitivity (90.2%) than IIF (75.4%) in the diagnosis of pemphigus. Conclusions: Dsg ELISA is a more sensitive method than IIF and shows more correlation with the disease severity. PMID:28400637

Nonlinear Focal Modulation Microscopy.

PubMed

Zhao, Guangyuan; Zheng, Cheng; Kuang, Cuifang; Zhou, Renjie; Kabir, Mohammad M; Toussaint, Kimani C; Wang, Wensheng; Xu, Liang; Li, Haifeng; Xiu, Peng; Liu, Xu

2018-05-11

We demonstrate nonlinear focal modulation microscopy (NFOMM) to achieve superresolution imaging. Traditional approaches to superresolution that utilize point scanning often rely on spatially reducing the size of the emission pattern by directly narrowing (e.g., through minimizing the detection pinhole in Airyscan, Zeiss) or indirectly peeling its outer profiles [e.g., through depleting the outer emission region in stimulated emission depletion (STED) microscopy]. We show that an alternative conceptualization that focuses on maximizing the optical system's frequency shifting ability offers advantages in further improving resolution while reducing system complexity. In NFOMM, a spatial light modulator and a suitably intense laser illumination are used to implement nonlinear focal-field modulation to achieve a transverse spatial resolution of ∼60  nm (∼λ/10). We show that NFOMM is comparable with STED microscopy and suitable for fundamental biology studies, as evidenced in imaging nuclear pore complexes, tubulin and vimentin in Vero cells. Since NFOMM is readily implemented as an add-on module to a laser-scanning microscope, we anticipate wide utility of this new imaging technique.

Nonlinear Focal Modulation Microscopy

NASA Astrophysics Data System (ADS)

Zhao, Guangyuan; Zheng, Cheng; Kuang, Cuifang; Zhou, Renjie; Kabir, Mohammad M.; Toussaint, Kimani C.; Wang, Wensheng; Xu, Liang; Li, Haifeng; Xiu, Peng; Liu, Xu

2018-05-01

We demonstrate nonlinear focal modulation microscopy (NFOMM) to achieve superresolution imaging. Traditional approaches to superresolution that utilize point scanning often rely on spatially reducing the size of the emission pattern by directly narrowing (e.g., through minimizing the detection pinhole in Airyscan, Zeiss) or indirectly peeling its outer profiles [e.g., through depleting the outer emission region in stimulated emission depletion (STED) microscopy]. We show that an alternative conceptualization that focuses on maximizing the optical system's frequency shifting ability offers advantages in further improving resolution while reducing system complexity. In NFOMM, a spatial light modulator and a suitably intense laser illumination are used to implement nonlinear focal-field modulation to achieve a transverse spatial resolution of ˜60 nm (˜λ /10 ). We show that NFOMM is comparable with STED microscopy and suitable for fundamental biology studies, as evidenced in imaging nuclear pore complexes, tubulin and vimentin in Vero cells. Since NFOMM is readily implemented as an add-on module to a laser-scanning microscope, we anticipate wide utility of this new imaging technique.

Correlation between serological and immunofluorescence results in the investigation of brucellosis in veterinary surgeons.

PubMed Central

Henderson, R J; Hill, D M; Vickers, A A; Edwards, J M; Tillett, H

1976-01-01

Four serological tests and three immunofluorescence tests for IgG, IgM, and IgA were compared for value in the investigation of brucellosis in veterinary surgeons. No one serological test stood out over the others, and the immunofluorescence tests did not appear to have advantages over the serological tests. If a laboratory is limited in time and resources then the saline agglutination or the complement fixation test would be reasonably satisfactory. The 2-mercaptoethanol test and the antihuman globulin (Coombs' test) have no advantages over the other two and could be dropped. Immunofluorescence tests are not recommended for routine testing of brucellosis sera. The results and these recommendations apply to the 'vet' sera tested; it is reasonable to suppose that what applies to 'vet' sera will also apply to sera of those who work with or are in repeated contact with cattle and who will have had previous experience of brucella antigen, that is, dairy farmers, herdsmen, or slaughter house employees. PMID:765361

Development and characterization of monoclonal antibody to the lymphocystis disease virus of Japanese flounder Paralichthys olivaceus isolated from China.

PubMed

Cheng, Shunfeng; Zhan, Wenbin; Xing, Jing; Sheng, Xiuzhen

2006-08-01

Lymphocystis disease virus (LCDV) can infect, both naturally and experimentally, about 100 different teleost fish species. In this study, LCDV was purified using differential and gradient centrifugation from skin tumours of Japanese flounder Paralichthys olivaceus. A panel of five monoclonal antibodies (Mabs) against LCDV were produced by immunization of Balb/c mice with purified virus preparations. Analysed by the indirect enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), Western blot and immunogold electron microscopy (IEM), they showed specificity for LCDV. Immunofluorescent studies showed that the specific fluorescence signals appeared at the peripheral zone of hypertrophied cells cytoplasm where was the cytoplasmic inclusion bodies location and many of them formed ribbon-shaped. Western blot analysis demonstrated that two Mabs 1D7 and 2B6 reacted specifically to a single protein with an approximately molecular weight of 116kDa, Mab 3G3 reacted with two LCDV proteins at molecular mass of approximately 116 and 90kDa. Immunogold transmission electron-microscopy provided visualized evidence that the epitopes recognized by these Mabs were located on the outer surface of virions. The Mabs characterized should prove useful for developing LCDV diagnostic assays and for studying the biology of infection and pathogenesis of disease.

Evaluation of criteria for the postmortem diagnosis of mycoplasmal pneumonia of swine.

PubMed Central

Armstrong, C H; Scheidt, A B; Thacker, H L; Runnels, L J; Freeman, M J

1984-01-01

Ten swine from each of five herds believed to be affected with mycoplasmal pneumonia of swine and ten swine from each of five herds believed to be mycoplasmal pneumonia-free were selected for postmortem study. Lungs from the 100 swine were examined; grossly and microscopically for lesions typical of mycoplasmal pneumonia of swine and culturally and by an indirect immunofluorescent procedure for the presence of Mycoplasma hyopneumoniae. Nineteen of the lungs had both gross and microscopic lesions typical of mycoplasmal pneumonia of swine and 13 (68%) of these were infected, i.e. were culturally and/or indirect immunofluorescent positive. Absence of gross lesions did not prove freedom from mycoplasmal pneumonia, 14 of 73 (19%) grossly normal lungs were found to be infected with M. hyopneumoniae. Comparison of the indirect immunofluorescent and cultural examination, as methods of diagnosing mycoplasma pneumonia, revealed that neither procedure alone was reliable in the case of negative results. Ten lungs were indirect immunofluorescent negative and culturally positive and seven were culturally negative and indirect immunofluorescent positive (11 lungs were positive by both procedures). It was concluded that a definitive diagnosis of mycoplasmal pneumonia of swine requires that M. hyopneumoniae be visualized in indirect immunofluorescent stained lung sections or that it be recovered culturally. PMID:6434167

A history of urine microscopy.

PubMed

Cameron, J Stewart

2015-11-01

The naked-eye appearance of the urine must have been studied by shamans and healers since the Stone Age, and an elaborate interpretation of so-called Uroscopy began around 600 AD as a form of divination. A 1000 years later, the first primitive monocular and compound microscopes appeared in the Netherlands, and along with many other objects and liquids, urine was studied from around 1680 onwards as the enlightenment evolved. However, the crude early instruments did not permit fine study because of chromatic and linear/spherical blurring. Only after complex multi-glass lenses which avoided these problems had been made and used in the 1820s in London by Lister, and in Paris by Chevalier and Amici, could urinary microscopy become a practical, clinically useful tool in the 1830s. Clinical urinary microscopy was pioneered by Rayer and his pupils in Paris (especially Vigla), in the late 1830s, and spread to UK and Germany in the 1840s, with detailed descriptions and interpretations of cells and formed elements of the urinary sediment by Nasse, Henle, Robinson and Golding Bird. Classes were held, most notably by Donné in Paris. After another 50 years, optical microscopy had reached its apogee, with magnifications of over 1000 times obtainable free of aberration, using immersion techniques. Atlases of the urinary sediment were published in all major European countries and in the US. Polarised light and phase contrast was used also after 1900 to study urine, and by the early 20th century, photomicroscopy (pioneered by Donné and Daguerre 50 years previously, but then ignored) became usual for teaching and recording. In the 1940s electron microscopy began, followed by detection of specific proteins and cells using immunofluorescent antibodies. All this had been using handheld methodology. Around 1980, machine-assisted observations began, and have dominated progress since.

Localization of extracellular matrix components in developing mouse salivary glands by confocal microscopy

NASA Technical Reports Server (NTRS)

Hardman, P.; Spooner, B. S.

1992-01-01

The importance of the extracellular matrix (ECM) in epithelial-mesenchymal interactions in developing organisms is well established. Proteoglycans and interstitial collagens are required for the growth, morphogenesis, and differentiation of epithelial organs and the distribution of these molecules has been described. However, much less is known about other ECM macromolecules in developing epithelial organs. We used confocal microscopy to examine the distribution of laminin, heparan sulfate (BM-1) proteoglycan, fibronectin, and collagen types I, IV, and V, in mouse embryonic salivary glands. Organ rudiments were isolated from gestational day 13 mouse embryos and cultured for 24, 48, or 72 hours. Whole mounts were stained by indirect immunofluorescence and then examined using a Zeiss Laser Scan Microscope. We found that each ECM component examined had a distinct distribution and that the distribution of some molecules varied with culture time. Laminin was mainly restricted to the basement membrane. BM-1 proteoglycan was concentrated in the basement membrane and also formed a fine network throughout the mesenchyme. Type IV collagen was mainly located in the basement membrane of the epithelium, but it was also present throughout the mesenchyme. Type V collagen was distributed throughout the mesenchyme at 24 hours, but at 48 hours was principally located in the basement membrane. Type I collagen was distributed throughout the mesenchyme at all culture times, and accumulated in the clefts and particularly at the epithelial-mesenchymal interface as time in culture increased. Fibronectin was observed throughout the mesenchyme at all times.

Detection of antibody to Purpureocillium lilacinum by immunofluorescent assay and flow cytometry in serum of infected C57BL/6 mice.

PubMed

de Sequeira, Danielly C M; Peixoto, Mariana L P; De Luca, Paula M; Oliveira-Ferreira, Joseli; Antas, Paulo R Z; Borba, Cintia M

2013-10-31

Purpureocillium lilacinum is an emerging pathogenic fungus that can cause different clinical manifestations ranging from cutaneous and sub-cutaneous infections to severe oculomycosis. In this study, using both conventional indirect immunofluorescence and non-conventional flow cytometry approaches, IgG antibodies were readily detected in both C57BL/6 immunocompetent and immunosuppressed mice after i.v. infection with P. lilacinum. The humoral immune response was specific, since virtually no antibodies were detected in the serum of control mice. Flow cytometry assays also showed both quantitative and qualitative differences in total IgG and its isotypes in sera of immunocompetent and immunosupressed infected mice. Although a good starting point, it is clear that the effectiveness of serological assays for P. lilacinum hyalohyphomycosis identification in clinical studies still requires further standardization. Upon further validation in humans, these techniques have the potential to be suitable to detect P. lilacinum infection in patients, thereby avoiding current laborious and time-consuming culture techniques. © 2013.

Detection of anti-nuclear antibody by immunofluorescence assay and enzyme immunoassay in childhood systemic lupus erythematosus: experience from Bangladesh.

PubMed

Dipti, Tanjeem Rabika; Azam, Mohammad Shaiful; Sattar, Mohammad Humayun; Rahman, Shahana Akhter

2012-02-01

Systemic lupus erythematosus (SLE) is a multisystem, chronic but often episodic, autoimmune disease that is characterized by the presence of antinuclear antibodies (ANA). The criteria set by American College of Rheumatology are widely used for diagnosis of SLE. Elevation of ANA titer is the most sensitive of the ACR criteria. There are different methods for detection of ANA. Indirect immunofluorescence (ANA-IFA) and enzyme immunoassay (ANA-EIA) are commonly used methods. The sensitivity of ANA-IFA using HEp-2 cell substrate is 90-100% in systemic rheumatic diseases. In Bangladesh most of the laboratories use ANA-EIA for detection of ANA. As the sensitivity of ANA-EIA is lower than ANA-IFA it might be that we are missing many cases of ANA positivity in childhood SLE cases. To detect ANA by immunofluorescence assay using HEp-2 cell substrate and enzyme immunoassay in childhood SLE and to compare the diagnostic performance of these methods. This is a cross-sectional analytical study. A total of 40 patients were enrolled. Among them 20 were childhood SLE cases. Another 20 patients of childhood rheumatic diseases other than SLE were taken as the disease control group. In childhood SLE cases, 100% were ANA-positive by IFA and 55% were ANA positive by EIA. The sensitivity of ANA-IFA was 100%. In contrast, sensitivity of ANA-EIA was 55%.   ANA-IFA is superior to ANA-EIA for detection of ANA in childhood SLE patients. ANA-IFA should be the primary screening test for children with clinical features suggestive of SLE. © 2011 The Authors. International Journal of Rheumatic Diseases © 2011 Asia Pacific League of Associations for Rheumatology and Blackwell Publishing Asia Pty Ltd.

Applying fluorescence microscopy to the investigation of the behavior of foodborne pathogens on produce

NASA Astrophysics Data System (ADS)

Brandl, Maria T.

2009-05-01

In the past decade, the development of new tools to better visualize microbes at the cellular scale has spurred a renaissance in the application of microscopy to the study of bacteria in their natural environment. This renewed interest in microscopy may be largely attributable to the advent of the confocal laser scanning microscope (CLSM) and to the discovery of the green fluorescent protein. This article provides information about the use of fluorescence microscopy combined with fluorescent labels such as GFP, DsRed, and DNA stains, with immunofluorescence, and with digital image analysis, to examine the behavior of bacteria and other microbes on plant surfaces. Some of the advantages and pitfalls of these methods will be described using practical examples derived from studies of the ecology of foodborne pathogens, namely Salmonella enterica and E. coli O157:H7, on fresh fruit and vegetables. Confocal microscopy has been a powerful approach to uncover some of the factors involved in the association of produce with epidemics caused by these human pathogens and their interaction with other microbes in their nonhost environment.

Quantitative Immunofluorescence Analysis of Nucleolus-Associated Chromatin.

PubMed

Dillinger, Stefan; Németh, Attila

2016-01-01

The nuclear distribution of eu- and heterochromatin is nonrandom, heterogeneous, and dynamic, which is mirrored by specific spatiotemporal arrangements of histone posttranslational modifications (PTMs). Here we describe a semiautomated method for the analysis of histone PTM localization patterns within the mammalian nucleus using confocal laser scanning microscope images of fixed, immunofluorescence stained cells as data source. The ImageJ-based process includes the segmentation of the nucleus, furthermore measurements of total fluorescence intensities, the heterogeneity of the staining, and the frequency of the brightest pixels in the region of interest (ROI). In the presented image analysis pipeline, the perinucleolar chromatin is selected as primary ROI, and the nuclear periphery as secondary ROI.

Limbal explants from cryopreserved cadaver human corneas. Immunofluorescence and light microscopy of epithelial cells growing in culture.

PubMed

Bratanov, M; Neronov, A; Nikolova, E

2009-01-01

The aim of the present study was to determine whether human cadaver corneas, that were subject to cryopreservation, would be a source of migrating epithelial cells in vitro and what kind of morphological features these cells possess. Limbal explant culture was used for expanding the epithelial cells. Non-quantitative light microscopical examinations of the cultures within a period of 28 days were carried out. The phenotype of cultured cells, particularly of the presumed adult stem cell population, was examined by indirect fluorescent immunostaining using antibodies against corneal stem cell associated markers p63 and vimentin. The effectiveness of the freezing-thawing protocol was confirmed by cultivation of limbal explants taken from non-cryopreserved cadaver corneoscleral rims. The result clearly showed that limbal tissue, subjected to cryopreservation and long lasting (up to 12 months) storage in liquid nitrogen, retains the capacity to be source of migrating and proliferating epithelial cells in vitro including the presumed adult stem cells and transient amplifying cells.

Post-translational glutamylation and tyrosination in tubulin of tritrichomonads and the diplomonad Giardia intestinalis.

PubMed

Boggild, A K; Sundermann, C A; Estridge, B H

2002-01-01

Glutamylated and tyrosinated tubulin were localized in Giardia intestinalis and selected trichomonads of the Tritrichomonadinae subfamily, using specific monoclonal antibodies directed at each of the post-translational modifications. Analysis was carried out using indirect immunofluorescence microscopy. Although trichomonad tubulins remained unlabeled by anti-tyrosine tubulin (TUB-1A2), the presence of the glutamylation motif (GT 335) was confirmed and found to differ in distribution among tritrichomonads. Tritrichomonas muris was most heavily labeled with GT 335, while T. foetus was the least so. Like trichomonads, Giardia was unreactive to anti-tyrosine tubulin; however, the GT 335 antibody produced marked fluorescence in Giardia trophozoites. This study is the first to report immunofluorescent localization of tubulin glutamylation in Giardia and confirms previously reported mass spectrometry data.

Gliding Motility of Babesia bovis Merozoites Visualized by Time-Lapse Video Microscopy

PubMed Central

Asada, Masahito; Goto, Yasuyuki; Yahata, Kazuhide; Yokoyama, Naoaki; Kawai, Satoru; Inoue, Noboru; Kaneko, Osamu; Kawazu, Shin-ichiro

2012-01-01

Background Babesia bovis is an apicomplexan intraerythrocytic protozoan parasite that induces babesiosis in cattle after transmission by ticks. During specific stages of the apicomplexan parasite lifecycle, such as the sporozoites of Plasmodium falciparum and tachyzoites of Toxoplasma gondii, host cells are targeted for invasion using a unique, active process termed “gliding motility”. However, it is not thoroughly understood how the merozoites of B. bovis target and invade host red blood cells (RBCs), and gliding motility has so far not been observed in the parasite. Methodology/Principal Findings Gliding motility of B. bovis merozoites was revealed by time-lapse video microscopy. The recorded images revealed that the process included egress of the merozoites from the infected RBC, gliding motility, and subsequent invasion into new RBCs. The gliding motility of B. bovis merozoites was similar to the helical gliding of Toxoplasma tachyzoites. The trails left by the merozoites were detected by indirect immunofluorescence assay using antiserum against B. bovis merozoite surface antigen 1. Inhibition of gliding motility by actin filament polymerization or depolymerization indicated that the gliding motility was driven by actomyosin dependent process. In addition, we revealed the timing of breakdown of the parasitophorous vacuole. Time-lapse image analysis of membrane-stained bovine RBCs showed formation and breakdown of the parasitophorous vacuole within ten minutes of invasion. Conclusions/Significance This is the first report of the gliding motility of B. bovis. Since merozoites of Plasmodium parasites do not glide on a substrate, the gliding motility of B. bovis merozoites is a notable finding. PMID:22506073

[Fractal research of neurite growth in immunofluorescent images].

PubMed

Tang, Min; Wang, Huinan

2008-12-01

Fractal dimension has been widely used in medical images processing and analysis. The neurite growth of cultured dorsal root ganglion (DRG) was detected by fluorescent immunocytochemistry treated with nerve regeneration factor (0.1, 0.5, 2.0 mg/L). A novel method based on triangular prism surface area (TPSA) was introduced and adopted to calculate the fractal dimension of the two-dimensional immunofluorescent images. Experimental results demonstrate that this method is easy to understand and convenient to operate, and the quantititve results are concordant with the observational findings under microscope. This method can be guidelines for analyzing and deciding experimental results.

Use of a Granulocyte Immunofluorescence Assay Designed for Humans for Detection of Antineutrophil Cytoplasmic Antibodies in Dogs with Chronic Enteropathies.

PubMed

Florey, J; Viall, A; Streu, S; DiMuro, V; Riddle, A; Kirk, J; Perazzotti, L; Affeldt, K; Wagner, R; Vaden, S; Harris, T; Allenspach, K

2017-07-01

Perinuclear antineutrophil cytoplasmic antibodies (pANCA) previously have been shown to be serum markers in dogs with chronic enteropathies, with dogs that have food-responsive disease (FRD) having higher frequencies of seropositivity than dogs with steroid-responsive disease (SRD). The indirect immunofluorescence (IIF) assay used in previous publications is time-consuming to perform, with low interobserver agreement. We hypothesized that a commercially available granulocyte IIF assay designed for humans could be used to detect perinuclear antineutrophil cytoplasmic antibodies in dogs. Forty-four dogs with FRD, 20 dogs with SRD, 20 control dogs, and 38 soft-coated wheaten terrier (SCWT) or SCWT-cross dogs. A granulocyte assay designed for humans was used to detect pANCA, cANCA, and antinuclear antibodies (ANA), as well as antibodies against proteinase-3 protein (PR-3) and myeloperoxidase protein (MPO) in archived serum samples. Sensitivity of the granulocyte assay to predict FRD in dogs was 0.61 (95% confidence interval (CI), 0.45, 0.75), and specificity was 1.00 (95% CI, 0.91, 1.00). A significant association was identified between positive pANCA or cANCA result and diagnosis of FRD (P < 0.0001). Agreement between the two assays to detect ANCA in the same serum samples from SCWT with protein-losing enteropathy/protein-losing nephropathy (PLE/PLN) was substantial (kappa, 0.77; 95% CI, 0.53, 1.00). Eight ANCA-positive cases were positive for MPO or PR-3 antibodies. The granulocyte immunofluorescence assay used in our pilot study was easy and quick to perform. Agreement with the previously published method was good. Copyright © 2017 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

The importance of direct immunofluorescence in pemphigus herpetiformis diagnosis*

PubMed Central

de Faria, Paula Carolina Pessanha; Cruz, Camila Caberlon; Abulafia, Luna Azulay; Maceira, Juan Manuel Pineiro; Cassia, Flávia de Freire; Medeiros, Paula Mota

2017-01-01

Pemphigus herpetiformis is an autoimmune bullous disease, that combines clinical features of dermatitis herpetiformis and linear IgA bullous dermatosis and immunological characteristics of pemphigus, which makes this disease peculiar and this diagnosis rarely suspected in the first evaluation of the patient. The reported case is of a patient with clinically bullous disease similar to dermatitis herpetiformis, whose multiple biopsies were inconclusive, and only after direct immunofluorescence with a pemphigus pattern (intraepidermal intercellular pattern) the confirmation of the diagnosis was possible. PMID:29267475

Microcapsules on Streptococcus mutans serotypes by electron microscopy.

PubMed

Grenier, E M; Gray, R H; Loesche, W J; Eveland, W C

1977-02-01

Extracellular microcapsules have been demonstrated on cells of most serotypes of Streptococcus mutans by electron microscopy, using bacterial strains of the various serotypes and peroxidase labeled or unlabeled immune serum. A correlation was noted between the amount of capsular substance on the strains of S mutans examined and degree of antigenicity as expressed by the indirect fluorescent antibody (FA) title. A serotype d strain was shown to lose both antigenicity as determined by the FA reaction and capsular material as seen by electron microscopy with repeated in vitro passage. When 10% unheated rabbit serum was added to the medium, antigenicity and capsular material were restored.

Proof of the quantitative potential of immunofluorescence by mass spectrometry.

PubMed

Toki, Maria I; Cecchi, Fabiola; Hembrough, Todd; Syrigos, Konstantinos N; Rimm, David L

2017-03-01

Protein expression in formalin-fixed, paraffin-embedded patient tissue is routinely measured by Immunohistochemistry (IHC). However, IHC has been shown to be subject to variability in sensitivity, specificity and reproducibility, and is generally, at best, considered semi-quantitative. Mass spectrometry (MS) is considered by many to be the criterion standard for protein measurement, offering high sensitivity, specificity, and objective molecular quantification. Here, we seek to show that quantitative immunofluorescence (QIF) with standardization can achieve quantitative results comparable to MS. Epidermal growth factor receptor (EGFR) was measured by quantitative immunofluorescence in 15 cell lines with a wide range of EGFR expression, using different primary antibody concentrations, including the optimal signal-to-noise concentration after quantitative titration. QIF target measurement was then compared to the absolute EGFR concentration measured by Liquid Tissue-selected reaction monitoring mass spectrometry. The best agreement between the two assays was found when the EGFR primary antibody was used at the optimal signal-to-noise concentration, revealing a strong linear regression (R 2 =0.88). This demonstrates that quantitative optimization of titration by calculation of signal-to-noise ratio allows QIF to be standardized to MS and can therefore be used to assess absolute protein concentration in a linear and reproducible manner.

An inexpensive approach for bright-field and dark-field imaging by scanning transmission electron microscopy in scanning electron microscopy.

PubMed

Patel, Binay; Watanabe, Masashi

2014-02-01

Scanning transmission electron microscopy in scanning electron microscopy (STEM-in-SEM) is a convenient technique for soft materials characterization. Various specimen-holder geometries and detector arrangements have been used for bright-field (BF) STEM-in-SEM imaging. In this study, to further the characterization potential of STEM-IN-SEM, a new specimen holder has been developed to facilitate direct detection of BF signals and indirect detection of dark-field (DF) signals without the need for substantial instrument modification. DF imaging is conducted with the use of a gold (Au)-coated copper (Cu) plate attached to the specimen holder which directs highly scattered transmitted electrons to an off-axis yttrium-aluminum-garnet (YAG) detector. A hole in the copper plate allows for BF imaging with a transmission electron (TE) detector. The inclusion of an Au-coated Cu plate enhanced DF signal intensity. Experiments validating the acquisition of true DF signals revealed that atomic number (Z) contrast may be achieved for materials with large lattice spacing. However, materials with small lattice spacing still exhibit diffraction contrast effects in this approach. The calculated theoretical fine probe size is 1.8 nm. At 30 kV, in this indirect approach, DF spatial resolution is limited to 3.2 nm as confirmed experimentally.

Choice of Illumination System & Fluorophore for Multiplex Immunofluorescence on FFPE Tissue Sections

PubMed Central

Kishen, Ria E. B.; Kluth, David C.; Bellamy, Christopher O. C.

2016-01-01

The recent availability of novel dyes and alternative light sources to facilitate complex tissue immunofluorescence studies such as multiplex labelling has not been matched by reports critically evaluating the considerations and relative benefits of these new tools, particularly in combination. Product information is often limited to wavelengths used for older fluorophores (FITC, TRITC & corresponding Alexa dyes family). Consequently, novel agents such as Quantum dots are not widely appreciated or used, despite highly favourable properties including extremely bright emission, stability and potentially reduced tissue autofluorescence at the excitation wavelength. Using spectral analysis, we report here a detailed critical appraisal and comparative evaluation of different light sources and fluorophores in multiplex immunofluorescence of clinical biopsy sections. The comparison includes mercury light, metal halide and 3 different LED-based systems, using 7 Qdots (525, 565, 585, 605, 625, 705), Cy3 and Cy5. We discuss the considerations relevant to achieving the best combination of light source and fluorophore for accurate multiplex fluorescence quantitation. We highlight practical limitations and confounders to quantitation with filter-based approaches. PMID:27632367

Direct versus indirect ACL femoral attachment fibres and their implications on ACL graft placement.

PubMed

Moulton, Samuel G; Steineman, Brett D; Haut Donahue, Tammy L; Fontboté, Cristián A; Cram, Tyler R; LaPrade, Robert F

2017-01-01

To further elucidate the direct and indirect fibre insertion morphology within the human ACL femoral attachment using scanning electron microscopy and determine where in the footprint each fibre type predominates. The hypothesis was that direct fibre attachment would be found centrally in the insertion site, while indirect fibre attachment would be found posteriorly adjacent to the posterior articular cartilage. Ten cadaveric knees were dissected to preserve and isolate the entirety of the femoral insertion of the ACL. Specimens were then prepared and evaluated with scanning electron microscopy to determine insertional fibre morphology and location. The entirety of the fan-like projection of the ACL attachment site lay posterior to the lateral intercondylar ridge. In all specimens, a four-phase architecture, consistent with previous descriptions of direct fibres, was found in the centre of the femoral attachment site. The posterior margin of the ACL attachment attached directly adjacent to the posterior articular cartilage with some fibres coursing into it. The posterior portion of the ACL insertion had a two-phase insertion, consistent with previous descriptions of indirect fibres. The transition from the ligament fibres to bone had less interdigitations, and the interdigitations were significantly smaller (p < 0.001) compared to the transition in the direct fibre area. The interdigitations of the direct fibres were 387 ± 81 μm (range 282-515 μm) wide, while the interdigitations of indirect fibres measured 228 ± 75 μm (range 89-331 μm). The centre of the ACL femoral attachment consisted of a direct fibre structure, while the posterior portion had an indirect fibre structure. These results support previous animal studies reporting that the centre of the ACL femoral insertion was comprised of the strongest reported fibre type. Clinically, the femoral ACL reconstruction tunnel should be oriented to cover the entirety of the central direct ACL fibres

Effect of tooth brush abrasion and thermo-mechanical loading on direct and indirect veneer restorations.

PubMed

Rosentritt, Martin; Sawaljanow, Alexander; Behr, Michael; Kolbeck, Carola; Preis, Verena

2015-01-01

This study investigated toothbrush abrasion and in vitro aging on ceramic (indirect technique) and composite veneers (direct technique). Identical composite and individual human incisors were restored with industrially preformed composite veneers, indirectly produced ceramic veneers, and direct composite restorations. Surface roughness was determined before and after tooth brushing. A 5-year period of oral service was simulated by thermal cycling and mechanical loading (TCML). After TCML, all specimens were examined with microscopy and scanning electron microscopy. Specimens without failures during TCML were loaded until failure. analysis of variance; Bonferroni's post hoc analysis, Kaplan-Meier-Log Rank test (α = 0.05). Tooth brushing yielded a non-significant increase (p = 0.560) in roughness in all materials (industrial veneer, 0.12+/-0.07 μm, direct restoration, 0.18+/-0.14 μm, ceramic, 0.35+/-0.16 μm). No significant differences in roughness could be determined between the materials, neither before nor after testing (p < 0.001). After TCML of artificial teeth, direct and preformed composite veneers on composite teeth showed no failures or damages. Two ceramic veneers showed cracking in the labial area. After TCML of human teeth, transmission microscopy indicated a facial crack in a ceramic veneer and chipping in the cervical area of a preformed veneer. Two direct composite veneers lost retention. No significantly different survival rates were found between the three veneer groups. Fracture force on human teeth varied between 527.8+/-132.4 N (ceramic), 478.3+/-165.4 N (preformed composite), and 605.0+/-263.5 N (direct composite). All materials revealed comparable wear resistance. Indirect ceramic, direct restorative composite, and preformed composite veneers showed comparable failure rates and satisfying longevity. The results indicate similar longevity of the chosen materials for veneer restorations.

Increased serum anti-N-methyl-D-aspartate receptor antibody immunofluorescence in psychiatric patients with past catatonia

PubMed Central

Lin, Chin-Chuen; Hung, Yi-Yung; Tsai, Meng-Chang

2017-01-01

Objective Anti-N-methyl-D-aspartate receptor (NMDAR) antibody was thought to be the cause of anti-NMDAR encephalitis, with manifestations similar to catatonia and schizophrenia. Anti-NMDAR antibody in neuropsychiatric patients who had catatonia before were investigated in a follow-up evaluation. The intensity of antibody immunofluorescence was quantified and compared with healthy controls. Method Nineteen patients (eight males and eleven females) agreed to be followed-up. Thirteen had the diagnosis of schizophrenia, two had the diagnosis of major depressive disorder, two had bipolar disorder, one had postpartum depression, and one had herpes simplex encephalitis. No patient had catatonia during the follow-up. Nineteen sex-matched healthy controls were recruited. Results Using Mann-Whitney U test, patients had greater intensity of anti-NMDAR antibody immunofluorescence than the healthy controls (121,979 ± 86,526 vs. 47,692 ± 26,102, p = 0.003). No correlation was found between immunofluorescence intensity and catatonia scales or symptom severity scores. Neuropsychiatric patients with past catatonia showed greater anti-NMDAR antibody response than the healthy controls. Conclusion NMDAR dysfunction might play a role in the mechanism underlying catatonia. Further studies are needed to confirm this finding. PMID:29073246

Increased serum anti-N-methyl-D-aspartate receptor antibody immunofluorescence in psychiatric patients with past catatonia.

PubMed

Lin, Chin-Chuen; Hung, Yi-Yung; Tsai, Meng-Chang; Huang, Tiao-Lai

2017-01-01

Anti-N-methyl-D-aspartate receptor (NMDAR) antibody was thought to be the cause of anti-NMDAR encephalitis, with manifestations similar to catatonia and schizophrenia. Anti-NMDAR antibody in neuropsychiatric patients who had catatonia before were investigated in a follow-up evaluation. The intensity of antibody immunofluorescence was quantified and compared with healthy controls. Nineteen patients (eight males and eleven females) agreed to be followed-up. Thirteen had the diagnosis of schizophrenia, two had the diagnosis of major depressive disorder, two had bipolar disorder, one had postpartum depression, and one had herpes simplex encephalitis. No patient had catatonia during the follow-up. Nineteen sex-matched healthy controls were recruited. Using Mann-Whitney U test, patients had greater intensity of anti-NMDAR antibody immunofluorescence than the healthy controls (121,979 ± 86,526 vs. 47,692 ± 26,102, p = 0.003). No correlation was found between immunofluorescence intensity and catatonia scales or symptom severity scores. Neuropsychiatric patients with past catatonia showed greater anti-NMDAR antibody response than the healthy controls. NMDAR dysfunction might play a role in the mechanism underlying catatonia. Further studies are needed to confirm this finding.

Immunogold scanning electron microscopy can reveal the polysaccharide architecture of xylem cell walls

PubMed Central

Sun, Yuliang; Juzenas, Kevin

2017-01-01

Abstract Immunofluorescence microscopy (IFM) and immunogold transmission electron microscopy (TEM) are the two main techniques commonly used to detect polysaccharides in plant cell walls. Both are important in localizing cell wall polysaccharides, but both have major limitations, such as low resolution in IFM and restricted sample size for immunogold TEM. In this study, we have developed a robust technique that combines immunocytochemistry with scanning electron microscopy (SEM) to study cell wall polysaccharide architecture in xylem cells at high resolution over large areas of sample. Using multiple cell wall monoclonal antibodies (mAbs), this immunogold SEM technique reliably localized groups of hemicellulosic and pectic polysaccharides in the cell walls of five different xylem structures (vessel elements, fibers, axial and ray parenchyma cells, and tyloses). This demonstrates its important advantages over the other two methods for studying cell wall polysaccharide composition and distribution in these structures. In addition, it can show the three-dimensional distribution of a polysaccharide group in the vessel lateral wall and the polysaccharide components in the cell wall of developing tyloses. This technique, therefore, should be valuable for understanding the cell wall polysaccharide composition, architecture and functions of diverse cell types. PMID:28398585

BLIND TRIALS EVALUATING IN VITRO INFECTIVITY OF CRYPTOSPORIDIUM PARVUM OOCYSTS USING CELL CULTURE IMMUNOFLUORESCENCE

EPA Science Inventory

An optimized cell culture-immunofluorescence (IFA) procedure, using the HCT-8 cell line, was evaluated in 'blind' trials to determine the sensitivity and reproducibility for measuring infectivity of flow cytometry prepared inocula of C. parvum oocysts. In separate trials, suspens...

Quantitative comparison between full-spectrum and filter-based imaging in hyperspectral fluorescence microscopy

PubMed Central

GAO, L.; HAGEN, N.; TKACZYK, T.S.

2012-01-01

Summary We implement a filterless illumination scheme on a hyperspectral fluorescence microscope to achieve full-range spectral imaging. The microscope employs polarisation filtering, spatial filtering and spectral unmixing filtering to replace the role of traditional filters. Quantitative comparisons between full-spectrum and filter-based microscopy are provided in the context of signal dynamic range and accuracy of measured fluorophores’ emission spectra. To show potential applications, a five-colour cell immunofluorescence imaging experiment is theoretically simulated. Simulation results indicate that the use of proposed full-spectrum imaging technique may result in three times improvement in signal dynamic range compared to that can be achieved in the filter-based imaging. PMID:22356127

Electron and Fluorescence Microscopy of Extracellular Glucan and Aryl-Alcohol Oxidase during Wheat-Straw Degradation by Pleurotus eryngii

PubMed Central

Barrasa, J. M.; Gutiérrez, A.; Escaso, V.; Guillén, F.; Martínez, M. J.; Martínez, A. T.

1998-01-01

The ligninolytic fungus Pleurotus eryngii grown in liquid medium secreted extracellular polysaccharide (87% glucose) and the H2O2-producing enzyme aryl-alcohol oxidase (AAO). The production of both was stimulated by wheat-straw. Polyclonal antibodies against purified AAO were obtained, and a complex of glucanase and colloidal gold was prepared. With these tools, the localization of AAO and extracellular glucan in mycelium from liquid medium and straw degraded under solid-state fermentation conditions was investigated by transmission electron microscopy (TEM) and fluorescence microscopy. These studies revealed that P. eryngii produces a hyphal sheath consisting of a thin glucan layer. This sheath appeared to be involved in both mycelial adhesion to the straw cell wall during degradation and AAO immobilization on hyphal surfaces, with the latter evidenced by double labeling. AAO distribution during differential degradation of straw tissues was observed by immunofluorescence microscopy. Finally, TEM immunogold studies confirmed that AAO penetrates the plant cell wall during P. eryngii degradation of wheat straw. PMID:9435085

Indirection and computer security.

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

Berg, Michael J.

2011-09-01

The discipline of computer science is built on indirection. David Wheeler famously said, 'All problems in computer science can be solved by another layer of indirection. But that usually will create another problem'. We propose that every computer security vulnerability is yet another problem created by the indirections in system designs and that focusing on the indirections involved is a better way to design, evaluate, and compare security solutions. We are not proposing that indirection be avoided when solving problems, but that understanding the relationships between indirections and vulnerabilities is key to securing computer systems. Using this perspective, we analyzemore » common vulnerabilities that plague our computer systems, consider the effectiveness of currently available security solutions, and propose several new security solutions.« less

Analysis of the Cellular Stress Response During Ebola Virus Infection by Immunofluorescence.

PubMed

Nelson, Emily V; Schmidt, Kristina M

2017-01-01

In this chapter, the use of immunofluorescence analysis as a tool to examine stress granule (SG) formation in Ebola virus (EBOV)-infected cells is described. The following protocol focuses on the process of inducing and analyzing the cellular stress response, including treatment of cells with inducers and inhibitors of the SG formation, and also describes EBOV infection, DNA transfection, and the usage of different cell lines.

Using indirect covariance spectra to identify artifact responses in unsymmetrical indirect covariance calculated spectra.

PubMed

Martin, Gary E; Hilton, Bruce D; Blinov, Kirill A; Williams, Antony J

2008-02-01

Several groups of authors have reported studies in the areas of indirect and unsymmetrical indirect covariance NMR processing methods. Efforts have recently focused on the use of unsymmetrical indirect covariance processing methods to combine various discrete two-dimensional NMR spectra to afford the equivalent of the much less sensitive hyphenated 2D NMR experiments, for example indirect covariance (icv)-heteronuclear single quantum coherence (HSQC)-COSY and icv-HSQC-nuclear Overhauser effect spectroscopy (NOESY). Alternatively, unsymmetrical indirect covariance processing methods can be used to combine multiple heteronuclear 2D spectra to afford icv-13C-15N HSQC-HMBC correlation spectra. We now report the use of responses contained in indirect covariance processed HSQC spectra as a means for the identification of artifacts in both indirect covariance and unsymmetrical indirect covariance processed 2D NMR spectra. Copyright (c) 2007 John Wiley & Sons, Ltd.

Identification of Skeletal Muscle Satellite Cells by Immunofluorescence with Pax7 and Laminin Antibodies.

PubMed

Feng, Xuesong; Naz, Faiza; Juan, Aster H; Dell'Orso, Stefania; Sartorelli, Vittorio

2018-04-19

Immunofluorescence is an effective method that helps to identify different cell types on tissue sections. In order to study the desired cell population, antibodies for specific cell markers are applied on tissue sections. In adult skeletal muscle, satellite cells (SCs) are stem cells that contribute to muscle repair and regeneration. Therefore, it is important to visualize and trace the satellite cell population under different physiological conditions. In resting skeletal muscle, SCs reside between the basal lamina and myofiber plasma membrane. A commonly used marker for identifying SCs on the myofibers or in cell culture is the paired box protein Pax7. In this article, an optimized Pax7 immunofluorescence protocol on skeletal muscle sections is presented that minimizes non-specific staining and background. Another antibody that recognizes a protein (laminin) of the basal lamina was also added to help identify SCs. Similar protocols can also be used to perform double or triple labeling with Pax7 and antibodies for additional proteins of interest.

Autoantibodies against Leydig cells in patients after spermatic cord torsion.

PubMed Central

Zanchetta, R; Mastrogiacomo, I; Graziotti, P; Foresta, C; Betterle, C

1984-01-01

This study is aimed at searching for the presence of circulating antibodies against frozen sections of human testis, ovary and trophoblast in patients that had spermatic cord torsion. Sixty-eight sera samples were studied. Nine patients (13.2%) were positive for organ specific anti-testis autoantibodies. Six patients were positive for antibodies against Leydig cells: five were positive only with the indirect immunofluorescence technique of complement fixing (ITT/CF), the sixth patient was positive only with the indirect immunofluorescence technique (ITT). The other three patients were positive for antibodies against germ line cells: two patients were positive with both techniques, the third was positive only with indirect immunofluorescence technique. Eight of these patients were negative for antibodies against adrenal cortex while only one case was positive with indirect immunofluorescence technique both on adrenal cortex and Leydig cells. Human lyophilized testis absorbed the reactive antibodies against Leydig cells and germ line cells, while adrenal cortex and lyophilized testosterone were ineffective. This study shows the identification of a specific antibody against Leydig cells and germ line cells in patients after spermatic cord torsion. PMID:6362937

Detection of Pituitary Antibodies by Immunofluorescence: Approach and Results in Patients With Pituitary Diseases

PubMed Central

Ricciuti, Adriana; De Remigis, Alessandra; Landek-Salgado, Melissa A.; De Vincentiis, Ludovica; Guaraldi, Federica; Lupi, Isabella; Iwama, Shintaro; Wand, Gary S.; Salvatori, Roberto

2014-01-01

Context: Pituitary antibodies have been measured mainly to identify patients whose disease is caused or sustained by pituitary-specific autoimmunity. Although reported in over 100 publications, they have yielded variable results and are thus considered of limited clinical utility. Objectives: Our objectives were to analyze all publications reporting pituitary antibodies by immunofluorescence for detecting the major sources of variability, to experimentally test these sources and devise an optimized immunofluorescence protocol, and to assess prevalence and significance of pituitary antibodies in patients with pituitary diseases. Study Design and Outcome Measures: We first evaluated the effect of pituitary gland species, section fixation, autofluorescence quenching, blockade of unwanted antibody binding, and use of purified IgG on the performance of this antibody assay. We then measured cross-sectionally the prevalence of pituitary antibodies in 390 pituitary cases and 60 healthy controls, expressing results as present or absent and according to the (granular, diffuse, perinuclear, or mixed) staining pattern. Results: Human pituitary was the best substrate to detect pituitary antibodies and yielded an optimal signal-to-noise ratio when treated with Sudan black B to reduce autofluorescence. Pituitary antibodies were more common in cases (95 of 390, 24%) than controls (3 of 60, 5%, P = .001) but did not discriminate among pituitary diseases when reported dichotomously. However, when expressed according to their cytosolic staining, a granular pattern was highly predictive of pituitary autoimmunity (P < .0001). Conclusion: We report a comprehensive study of pituitary antibodies by immunofluorescence and provide a method and an interpretation scheme that should be useful for identifying and monitoring patients with pituitary autoimmunity. PMID:24606106

A dual-modality optical coherence tomography and selective plane illumination microscopy system for mouse embryonic imaging

NASA Astrophysics Data System (ADS)

Wu, Chen; Ran, Shihao; Le, Henry; Singh, Manmohan; Larina, Irina V.; Mayerich, David; Dickinson, Mary E.; Larin, Kirill V.

2017-02-01

Both optical coherence tomography (OCT) and selective plane illumination microscopy (SPIM) are frequently used in mouse embryonic research for high-resolution three-dimensional imaging. However, each of these imaging methods provide a unique and independent advantage: SPIM provides morpho-functional information through immunofluorescence and OCT provides a method for whole-embryo 3D imaging. In this study, we have combined rotational imaging OCT and SPIM into a single, dual-modality device to image E9.5 mouse embryos. The results demonstrate that the dual-modality setup is able to provide both anatomical and functional information simultaneously for more comprehensive tissue characterization.

ISOLATION AND DETECTION OF GIARDIA CYSTS FROM WATER USING DIRECT IMMUNOFLUORESCENCE.

USGS Publications Warehouse

Sorenson, Stephen K.; Riggs, John L.; Dileanis, Peter D.; Suk, Thomas J.

1986-01-01

A water-sampling apparatus used for the isolation and detection of Giardia cysts in water has been designed and tested. The sampling apparatus uses one of a variety of pumps or waterline pressure to move water through a filter. Two of the optional pumps are lightweight enough to make the apparatus portable and thus suitable for sampling in remote areas. This technique of sample processing produces good cyst recovery in much less time than is required with previously established methods. Giardia cysts are identified using direct immunofluorescence.

Rapid analysis and exploration of fluorescence microscopy images.

PubMed

Pavie, Benjamin; Rajaram, Satwik; Ouyang, Austin; Altschuler, Jason M; Steininger, Robert J; Wu, Lani F; Altschuler, Steven J

2014-03-19

Despite rapid advances in high-throughput microscopy, quantitative image-based assays still pose significant challenges. While a variety of specialized image analysis tools are available, most traditional image-analysis-based workflows have steep learning curves (for fine tuning of analysis parameters) and result in long turnaround times between imaging and analysis. In particular, cell segmentation, the process of identifying individual cells in an image, is a major bottleneck in this regard. Here we present an alternate, cell-segmentation-free workflow based on PhenoRipper, an open-source software platform designed for the rapid analysis and exploration of microscopy images. The pipeline presented here is optimized for immunofluorescence microscopy images of cell cultures and requires minimal user intervention. Within half an hour, PhenoRipper can analyze data from a typical 96-well experiment and generate image profiles. Users can then visually explore their data, perform quality control on their experiment, ensure response to perturbations and check reproducibility of replicates. This facilitates a rapid feedback cycle between analysis and experiment, which is crucial during assay optimization. This protocol is useful not just as a first pass analysis for quality control, but also may be used as an end-to-end solution, especially for screening. The workflow described here scales to large data sets such as those generated by high-throughput screens, and has been shown to group experimental conditions by phenotype accurately over a wide range of biological systems. The PhenoBrowser interface provides an intuitive framework to explore the phenotypic space and relate image properties to biological annotations. Taken together, the protocol described here will lower the barriers to adopting quantitative analysis of image based screens.

Anti-human neutrophil antigen-1a, -1b, and -2 antibodies in neonates and children with immune neutropenias analyzed by extracted granulocyte antigen immunofluorescence assay.

PubMed

Onodera, Rie; Kurita, Emi; Taniguchi, Kikuyo; Karakawa, Shuhei; Okada, Satoshi; Kihara, Hirotaka; Fujii, Teruhisa; Kobayashi, Masao

2017-11-01

Anti-human neutrophil antigen (HNA) antibodies have been implicated in the development of neonatal alloimmune neutropenia (NAN) and autoimmune neutropenia (AIN). There are many conventional assay methods that detect anti-HNA antibodies. However, a method to measure multiple samples and detect several anti-HNA antibodies simultaneously is needed. We developed a new method, the extracted granulocyte antigen immunofluorescence assay (EGIFA), to analyze anti-HNA-1a, -1b, and -2 antibodies in sera. The results obtained by EGIFA were evaluated in comparison with those from several standard assay methods. Anti-HNA antibodies in serum samples from nine familial cases with suspected NAN (n = 19) and children with suspected AIN (n = 88) were also measured by EGIFA. The evaluation of nine serum samples with anti-HNA antibodies suggested that EGIFA demonstrated equivalent specificity and superior sensitivity to monoclonal antibody-specific immobilization of granulocyte antigens and had comparable sensitivity to the granulocyte indirect immunofluorescence test. EGIFA successfully detected anti-HNA-1a or -1b antibodies in seven of nine familial cases with suspected NAN. EGIFA detected anti-HNA antibodies in 40.9% of children with suspected AIN. Among them, isolated anti-HNA-1a or -1b antibody was detected in 4.5 or 12.5% of children, respectively, and anti-HNA-2 antibody was identified in 3.4% of children. The 30.8% (16 of 52) of children negative for anti-HNA antibody by EGIFA were positive for anti-HLA antibody. EGIFA facilitated the measurement of anti-HNA-1a, -1b, and/or -2 antibodies in sera. The prompt measurement of anti-HNA antibodies will improve the diagnosis and clinical management of patients with suspected NAN or AIN. © 2017 AABB.

Release of Rhizobium spp. from Tropical Soils and Recovery for Immunofluorescence Enumeration

PubMed Central

Kingsley, Mark T.; Bohlool, B. Ben

1981-01-01

Limitations associated with immunofluorescence enumeration of bacteria in soil derive largely from the efficiency with which cells can be separated from soil particles and collected on membrane filters for staining. Many tropical soils fix added bacteria tightly, resulting in low recoveries. Eight soils, representative of three of the major soil orders found in the tropics (oxisols, vertisols, and inceptisols), were tested for recovery of added Rhizobium strains. All except one Hawaiian andept (Typic Eutrandept) yielded recoveries ranging from <1 to 13%. Recovery from the andept was 100%. In soil-sand mixtures, addition of only a small amount of soil caused a dramatic decrease in recovery of added rhizobia. Increasing the soil content of the mixture from 0% (10 g of sand) to 50% (5 g of soil-5 g of sand) reduced recoveries from >90 to <1%. Varying the ionic strength and pH of the extracting solution did not cause marked increases in recovery. Protein solutions, ethylenediaminetetraacetate, and NaHCO3, on the other hand, improved release of bacteria. We report a modification to the usual membrane filter immunofluorescence procedure which yielded consistently high and reproducible recovery (coefficient of variation, 30%) of rhizobia from several tropical soils. In the modified procedure, partially hydrolyzed gelatin, diluted in ammonium phosphate, was used to suspend the soil. This caused dispersion of the soil and release of the bacteria from soil flocs. The efficiency of recovery of Rhizobium spp. from several tropical and two temperate soils remained high as the content of these soils in soil-sand mixtures was increased from 0 to 100%. The modified membrane filter immunofluorescence procedure was used to follow the growth of a strain of chickpea (Cicer arietinum) Rhizobium in a sterilized oxisol. The results showed a close agreement with viable counts at different stages during the growth cycle. Diluent for the hydrolyzed gelatin also had a marked effect on

Release of Rhizobium spp. from Tropical Soils and Recovery for Immunofluorescence Enumeration.

PubMed

Kingsley, M T; Bohlool, B B

1981-08-01

Limitations associated with immunofluorescence enumeration of bacteria in soil derive largely from the efficiency with which cells can be separated from soil particles and collected on membrane filters for staining. Many tropical soils fix added bacteria tightly, resulting in low recoveries. Eight soils, representative of three of the major soil orders found in the tropics (oxisols, vertisols, and inceptisols), were tested for recovery of added Rhizobium strains. All except one Hawaiian andept (Typic Eutrandept) yielded recoveries ranging from <1 to 13%. Recovery from the andept was 100%. In soil-sand mixtures, addition of only a small amount of soil caused a dramatic decrease in recovery of added rhizobia. Increasing the soil content of the mixture from 0% (10 g of sand) to 50% (5 g of soil-5 g of sand) reduced recoveries from >90 to <1%. Varying the ionic strength and pH of the extracting solution did not cause marked increases in recovery. Protein solutions, ethylenediaminetetraacetate, and NaHCO(3), on the other hand, improved release of bacteria. We report a modification to the usual membrane filter immunofluorescence procedure which yielded consistently high and reproducible recovery (coefficient of variation, 30%) of rhizobia from several tropical soils. In the modified procedure, partially hydrolyzed gelatin, diluted in ammonium phosphate, was used to suspend the soil. This caused dispersion of the soil and release of the bacteria from soil flocs. The efficiency of recovery of Rhizobium spp. from several tropical and two temperate soils remained high as the content of these soils in soil-sand mixtures was increased from 0 to 100%. The modified membrane filter immunofluorescence procedure was used to follow the growth of a strain of chickpea (Cicer arietinum) Rhizobium in a sterilized oxisol. The results showed a close agreement with viable counts at different stages during the growth cycle. Diluent for the hydrolyzed gelatin also had a marked effect on

Localizing Proteins in Fixed Giardia lamblia and Live Cultured Mammalian Cells by Confocal Fluorescence Microscopy.

PubMed

Nyindodo-Ogari, Lilian; Schwartzbach, Steven D; Skalli, Omar; Estraño, Carlos E

2016-01-01

Confocal fluorescence microscopy and electron microscopy (EM) are complementary methods for studying the intracellular localization of proteins. Confocal fluorescence microscopy provides a rapid and technically simple method to identify the organelle in which a protein localizes but only EM can identify the suborganellular compartment in which that protein is present. Confocal fluorescence microscopy, however, can provide information not obtainable by EM but required to understand the dynamics and interactions of specific proteins. In addition, confocal fluorescence microscopy of cells transfected with a construct encoding a protein of interest fused to a fluorescent protein tag allows live cell studies of the subcellular localization of that protein and the monitoring in real time of its trafficking. Immunostaining methods for confocal fluorescence microscopy are also faster and less involved than those for EM allowing rapid optimization of the antibody dilution needed and a determination of whether protein antigenicity is maintained under fixation conditions used for EM immunogold labeling. This chapter details a method to determine by confocal fluorescence microscopy the intracellular localization of a protein by transfecting the organism of interest, in this case Giardia lamblia, with the cDNA encoding the protein of interest and then processing these organisms for double label immunofluorescence staining after chemical fixation. Also presented is a method to identify the organelle targeting information in the presequence of a precursor protein, in this case the presequence of the precursor to the Euglena light harvesting chlorophyll a/b binding protein of photosystem II precursor (pLHCPII), using live cell imaging of mammalian COS7 cells transiently transfected with a plasmid encoding a pLHCPII presequence fluorescent protein fusion and stained with organelle-specific fluorescent dyes.

Immunofluorescent staining of nuclear antigen in lymphoid cells transformed by Herpesvirus papio (HVP).

PubMed

Schmitz, H

1981-01-01

An improved fixation method for antigen detection in lymphoblastoid cells is described. Herpesvirus papio nuclear antigen (HUPNA) could be stained in several transformed lymphoid cell lines by anti-complement immunofluorescence (ACIF). Antibody to HUPNA was detected in many human sera containing antibodies to Epstein-Barr virus capsid and nuclear antigen (EBNA). Rheumatoid arthritis sera showed a high incidence of both anti-EBNA and anti-HUPNA antibodies.

The use of immunofluorescence techniques for the laboratory diagnosis of transmissible gastroenteritis of swine.

PubMed Central

Solorzano, R F; Morin, M; Morehouse, L G

1978-01-01

Over a four year period, 74 of 250 field outbreaks of enteric disease (30%) and 110 of 440 swine (25%) were positive for transmissible gastroenteritis by immunofluorescence procedures. Of 141 swine from herds positive for transmissible gastroenteritis 110 (78%) were positive by fluorescent antibody techniques. The fastest, easiest to perform and most effective procedure was the examination of frozen sections of the jejunum from acutely ill animals by the fluorescent antibody tissue section technique. Only two herds were found to be positive by the fluorescent antibody tissue culture technique which were negative by fluorescent antibody tissue section technique. A considerable number of outbreaks, 21 of 74 (28%), of transmissible gastroenteritis were detected by immunofluorescence in swine over two weeks of age. The majority of outbreaks of transmissible gastroenteritis, 50 of 74 (68%), occurred in Missouri during the months of January through April and 63 of 74 (85%) during the months of December through May. The recurrence of the disease in a number of counties over a four-year period suggest the possibility of endemic foci. PMID:217505

Definitive and differential diagnosis of desquamative gingivitis through direct immunofluorescence studies.

PubMed

Suresh, Lakshmanan; Neiders, Mirdza E

2012-10-01

Desquamative gingivitis (DG) is a common clinical manifestation of oral autoimmune vesiculobullous diseases (VBDs). Their polymorphous clinical presentations coupled with similar histologic features make diagnosis indistinguishable among the different VBDs. Direct immunofluorescence (IF) studies are valuable gold-standard diagnostic tests that allow for discrimination among the various VBDs that present with DG. There have been no recent detailed analyses done that have used conventional light microscopy and direct IF in diagnosis to document the clinical associations of DG with various autoimmune oral diseases. The aim of this study is to examine retrospectively a large cohort of patients with DG for associated diseases and to determine the utility of direct IF and conventional light microscopy in establishing a definitive diagnosis. During a 14-month period, our laboratory in Buffalo, New York, received 239 consecutive archival cases of gingival biopsy with a clinical diagnosis of DG. These specimens were submitted to establish or rule out a diagnosis of a direct IF-positive VBD. The demographic, clinical, and microscopic findings were tabulated using established inclusion and diagnostic criteria. Approximately half the number (48.1%) of biopsies received for direct IF studies were submitted by periodontists. Slightly more than half of the patients (53%) previously had biopsies submitted for both hematoxylin and eosin (H & E) and direct IF testing. There was a female predilection for all the diseases studied except for pemphigus and linear immunoglobulin A disease. Oral lichen planus was the most common disease presenting as DG, followed by pemphigoid. The clinical diagnosis of lichen planus correlated with the biopsy findings in 80% of the cases and with pemphigoid in 60%. Definitive diagnosis was rendered to ≈80% of the gingival biopsies submitted. Negative cases of direct IF presenting as DG had significant pathology, such as dysplasia and carcinoma, which

Cruel intentions on television and in real life: can viewing indirect aggression increase viewers' subsequent indirect aggression?

PubMed

Coyne, Sarah M; Archer, John; Eslea, Mike

2004-07-01

Numerous studies have shown that viewing violence in the media can influence an individual's subsequent aggression, but none have examined the effect of viewing indirect aggression. This study examines the immediate effect of viewing indirect and direct aggression on subsequent indirect aggression among 199 children ages 11 to 14 years. They were shown an indirect, direct, or no-aggression video and their subsequent indirect aggression was measured by negative evaluation of a confederate and responses to a vignette. Participants viewing indirect or direct aggression gave a more negative evaluation of and less money to a confederate than participants viewing no-aggression. Participants viewing indirect aggression gave less money to the confederate than those viewing direct aggression. Participants viewing indirect aggression gave more indirectly aggressive responses to an ambiguous situation and participants viewing direct aggression gave more directly aggressive responses. This study provides the first evidence that viewing indirect aggression in the media can have an immediate impact on subsequent aggression.

Development and application of an indirect enzyme-linked immunosorbent assay using recombinant truncated Cap protein for the diagnosis of porcine circovirus-like virus P1.

PubMed

Wen, Li-bin; Wen, Shi-fu; He, Kong-wang

2016-01-19

Porcine circovirus-like virus P1 is a newly discovered virus. To date, there has been no specific serological assay for use in the diagnosis of P1 infection. Because P1 has high homology to porcine circovirus type 2 (PCV2) at the nucleotide level, the C-terminal portion of the capsid protein (amino acids 73-114), a discriminative antigen, was expressed in a prokaryotic expression system. The recombinant product (rctCap), composed of three identical repeated domains, was shown to be strongly immunoreactive to P1-specific serum. This assay was validated by comparison with an indirect immunofluorescence assay (IFA). The diagnostic sensitivity and specificity of the rctCap enzyme-linked immunosorbent assay (ELISA) developed in this study are 93.6% and 98.3%, respectively, compared with the results from IFAs on 450 sera samples from pigs. The indirect ELISA that we developed with rctCap, the recombinant capsid fragment containing the 217-342 nt repeat domain, was sensitive, specific, and suitable for the large-scale detection of P1 infections in swine.

Neuromyelitis optica immunoglobulin G in Chinese patients detected by immunofluorescence assay on a monkey brain substrate.

PubMed

Long, Youming; Hu, Xueqiang; Peng, Fuhua; Lu, Zhengqi; Wang, Yuge; Yang, Yu; Qiu, Wei

2012-01-01

Serum neuromyelitis optica immunoglobulin G (NMO-IgG) is used as a biomarker to differentiate between neuromyelitis optica (NMO) and multiple sclerosis (MS). However, the original assay is expensive and complex and shows low sensitivity. Here, we investigated the potential of NMO-IgG detection using an indirect immunofluorescence (IIF) assay on monkey brains. NMO-IgG seroprevalence was determined in 168 samples by an IIF assay on a monkey brain substrate. The data were compared with those from a standard mouse brain IIF assay using McNemar and kappa tests. Thirty-one of 50 (62%) NMO patients, 7 of 18 (38.9%) longitudinally extensive transverse myelitis patients, 6 of 57 (10.5%) MS patients, and 5 of 10 (50%) optic neuritis patients were seropositive for NMO-IgG. None of the acute partial transverse myelitis patients (n = 3) or healthy controls (n = 20) was positive. Thus, the sensitivity of the test was 62% for the patients with clinically definite NMO. The specificity was 89.5%, considering the 57 MS patients as the control group. The modified IIF assay on monkey brains and the standard IIF assay based on mouse brains were not significantly different (McNemar test; p = 1.000). The two assays were concordant in 39 seropositive samples and 100 seronegative samples (kappa test; kappa = 0.592, p < 0.0001). Although the modified IIF monkey brain assay was no better than the standard mouse brain IIF assay, we affirmed that NMO-IgG is a sensitive and specific biomarker to differentiate between NMO and MS. Copyright © 2011 S. Karger AG, Basel.

Single-cell immunofluorescence assay for terminal transferase: human leukaemic and non-leukaemic cells.

PubMed

Okamura, S; Crane, F; Jamal, N; Messner, H A; Mak, T W

1980-02-01

The characteristics of a single-cell immunofluorescence assay for terminal deoxynucleotidyl transferase (terminal transferase, TdT) is described. The data indicate that the single-cell immunofluorescence assay is highly efficient and specific for the detection of cells containing TdT. Using this assay, we have examined 124 marrow or peripheral-blood samples from 104 patients with or without haematological malignancies. Results indicate that TdT(+) cells from 6% to 100% were found in the following patients: 34/40 samples from patients with ALL at the time of diagnosis or during relapse; 2/3 patients with acute undifferentiated leukaemia; 2/3 patients with acute myelomonocytic leukaemia; 1/24 patients with acute myeloblastic leukaemia; 1/5 patients with chronic myelocytic leukaemia (CML) in blastic crisis; and 2/2 patients with diffuse lymphoblastic lymphoma. In contrast less than 1% of TdT(+) cells were found in 20 marrow or peripheral-blood samples from ALL patients in complete remission; 8 patients with CML in chronic phase; 2 patients with myeloma; 1 sample from a patient with Hodgkin's disease, peripheral-blood samples from 7 normal donors and marrow samples from 6 patients without haematological malignancies. TdT(+) cells were also found in association with cells with lymphoblast morphology. The TdT(+) cells in marrow were shown to be directly correlated with the percentage of morphological lymphoblasts, with a Spearman rank coefficient of 0·81, significant at a 0·001 level. In 2 longitudinal studies of 2 ALL patients with TdT(+) cells at diagnosis, the percentage TdT(+) cells also changed in parallel with the proportion of lymphoblasts. However, studies of 2 other patients with morphologically diagnosed ALL with < 1% TdT(+) cells at diagnosis also showed < 1% TdT(+) cells throughout the period studied, indicating a stable phenotype of blast cells in these patients. The single-cell immunofluorescence assay for TdT, which requires < 0·1% of the cells used in

Mouse hepatitis virus immunofluorescence in formalin- or Bouin's-fixed tissues using trypsin digestion.

PubMed

Brownstein, D G; Barthold, S W

1982-02-01

Mouse hepatitis viral antigens were demonstrated by immunofluorescence in formalin- and Bouin's-fixed tissues processed routinely for histopathology followed by partial digestion with trypsin. Staining was superior in tissues fixed in formalin and was not diminished in tissue sections from paraffin blocks stored at room temperature as long as 2 years. The relative ease of this procedure and the commercial availability of reagents makes this a useful technique for the definitive diagnosis of mouse hepatitis virus infection.

Guidelines for improving the reproducibility of quantitative multiparameter immunofluorescence measurements by laser scanning cytometry on fixed cell suspensions from human solid tumors.

PubMed

Shackney, Stanley; Emlet, David R; Pollice, Agnese; Smith, Charles; Brown, Kathryn; Kociban, Deborah

2006-01-01

Laser scanning Cytometry (LSC) is a versatile technology that makes it possible to perform multiple measurements on individual cells and correlate them cell by cell with other cellular features. It would be highly desirable to be able to perform reproducible, quantitative, correlated cell-based immunofluorescence studies on individual cells from human solid tumors. However, such studies can be challenging because of the presence of large numbers of cell aggregates and other confounding factors. Techniques have been developed to deal with cell aggregates in data sets collected by LSC. Experience has also been gained in addressing other key technical and methodological issues that can affect the reproducibility of such cell-based immunofluorescence measurements. We describe practical aspects of cell sample collection, cell fixation and staining, protocols for performing multiparameter immunofluorescence measurements by LSC, use of controls and reference samples, and approaches to data analysis that we have found useful in improving the accuracy and reproducibility of LSC data obtained in human tumor samples. We provide examples of the potential advantages of LSC in examining quantitative aspects of cell-based analysis. Improvements in the quality of cell-based multiparameter immunofluorescence measurements make it possible to extract useful information from relatively small numbers of cells. This, in turn, permits the performance of multiple multicolor panels on each tumor sample. With links among the different panels that are provided by overlapping measurements, it is possible to develop increasingly more extensive profiles of intracellular expression of multiple proteins in clinical samples of human solid tumors. Examples of such linked panels of measurements are provided. Advances in methodology can improve cell-based multiparameter immunofluorescence measurements on cell suspensions from human solid tumors by LSC for use in prognostic and predictive clinical

19 CFR 10.879 - Indirect materials.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 19 Customs Duties 1 2011-04-01 2011-04-01 false Indirect materials. 10.879 Section 10.879 Customs... of Origin § 10.879 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.816 - Indirect materials.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 19 Customs Duties 1 2010-04-01 2010-04-01 false Indirect materials. 10.816 Section 10.816 Customs... Rules of Origin § 10.816 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.816 - Indirect materials.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 19 Customs Duties 1 2011-04-01 2011-04-01 false Indirect materials. 10.816 Section 10.816 Customs... Rules of Origin § 10.816 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.776 - Indirect materials.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 19 Customs Duties 1 2011-04-01 2011-04-01 false Indirect materials. 10.776 Section 10.776 Customs... Rules of Origin § 10.776 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.816 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.816 Section 10.816 Customs... Rules of Origin § 10.816 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.776 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.776 Section 10.776 Customs... Rules of Origin § 10.776 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

19 CFR 10.776 - Indirect materials.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 19 Customs Duties 1 2013-04-01 2013-04-01 false Indirect materials. 10.776 Section 10.776 Customs... Rules of Origin § 10.776 Indirect materials. Indirect materials are to be disregarded in determining..., except that the cost of such indirect materials may be included in meeting the value-content requirement...

Uncompacted Myelin Lamellae and Nodal Ion Channel Disruption in POEMS Syndrome.

PubMed

Hashimoto, Rina; Koike, Haruki; Takahashi, Mie; Ohyama, Ken; Kawagashira, Yuichi; Iijima, Masahiro; Sobue, Gen

2015-12-01

To elucidate the significance of uncompacted myelin lamellae (UML) and ion channel disruption at the nodes of Ranvier in the polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes (POEMS) syndrome, we evaluated sural nerve biopsy specimens from 33 patients with POEMS syndrome and from 7 control patients. Uncompacted myelin lamellae distribution was assessed by electron microscopy and immunofluorescence microscopy. In the POEMS patient biopsies, UML were seen more frequently in small versus large myelinated fibers. Paranodes and Schmidt-Lanterman incisures, where normal physiologic UM is located, were frequently associated with UM. Widening of the nodes of Ranvier (i.e. segmental demyelination) was not associated with UML. There was axonal hollowing with neurofilament condensation at Schmidt-Lanterman incisures with abnormal UML, suggesting axonal damage at those sites in the POEMS patient biopsies. Myelin sheath irregularity was conspicuous in large myelinated fibers and was associated with abnormally widened bizarrely shaped Schmidt-Lanterman incisures. Indirect immunofluorescent studies revealed abnormalities of sodium (pan sodium) and potassium (KCNQ2) channels, even at nonwidened nodes of Ranvier. Thus, UML was not apparently associated with segmental demyelination but seemed to be associated with axonal damage. These observations suggest that nodal ion channel disruption may be associated with functional deficits in POEMS syndrome patient nerves.

Autoantibodies to full body vascular cell junctions colocalize with MYZAP, ARVCF, desmoplakins I and II and p0071 in endemic pemphigus in Colombia, South America.

PubMed

Abreu Velez, Ana M; Yi, Hong; Warfvinge, Gunnar; Howard, Michael S

2018-03-01

We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre-EPF). Here we aimed to investigate disease autoreactivity to vessels in all body organs/systems. We compared 57 patients and 57 controls from the endemic area, matched by demographics, age, sex, and work activity. We performed immunofluorescence, immunohistochemistry, confocal microscopy, immunoblotting, indirect immune electron microscopy studies, and autometallographic studies. We performed ultrasonography on large patient arteries, investigating for vascular anomalies. In addition, we reviewed autopsies on seven patients who died affected by El Bagre-EPF. We immunoadsorbed any positive vessel immunofluorescence with desmoglein (Dsg1), investigating for new autoantigens. Overall, 57/57 patients affected by El Bagre-EPF displayed autoantibodies to vessels in all the organs/systems of the body via all methods (P < 0.01). The autoreactivity was polyclonal, and the patient's antibodies colocalized with commercial antibodies to desmoplakins I and II, p0071, ARVCF, and MYZAP (all from Progen Biotechnik, Germany; P < 0.01; all present at cell junctions). Immunoadsorption with Dsg1 on positive vessel immunofluorescence showed that the immune response against the vessels was directed against non-Dsg1 antigen(s). Autometallographic studies showed deposits of metals and metalloids in vessel cell junctions and in erythrocytes of 85% of patients (P < 0.01). Immune response to these vascular antigens is likely altering endothelial cells and vessel shapes, thus disturbing hemodynamic flow. The flow alterations likely lead to inflammation and may play a role in the atherogenesis often seen in these patients. © 2017 The International Society of Dermatology.

Immunofluorescence Technique Using HeLa Cells Expressing Recombinant Nucleoprotein for Detection of Immunoglobulin G Antibodies to Crimean-Congo Hemorrhagic Fever Virus

PubMed Central

Saijo, Masayuki; Qing, Tang; Niikura, Masahiro; Maeda, Akihiko; Ikegami, Tetsuro; Sakai, Koji; Prehaud, Christophe; Kurane, Ichiro; Morikawa, Shigeru

2002-01-01

A HeLa cell line continuously expressing recombinant nucleoprotein (rNP) of the Crimean-Congo hemorrhagic fever virus (CCHFV) was established by transfection with an expression vector containing the cDNA of CCHFV NP (pKS336-CCHFV-NP). These cells were used as antigens for indirect immunofluorescence (IF) to detect immunoglobulin G antibodies to CCHFV. The sensitivity and specificity of this IF technique were examined by using serum samples and were compared to those of the IF technique using CCHFV-infected Vero E6 cells (authentic antigen). Staining of the CCHFV rNP expressed in HeLa cells showed a unique granular pattern similar to that of CCHFV-infected Vero E6 cells. Positive staining could easily be distinguished from a negative result. All 13 serum samples determined to be positive by using the authentic antigen were also determined to be positive by using CCHFV rNP-expressing HeLa cells (recombinant antigen). The 108 serum samples determined to be negative by using the authentic antigen were also determined to be negative by using the recombinant antigen. Thus, both the sensitivity and the specificity of this IF technique were 100% compared to the IF with authentic antigen. The novel IF technique using CCHFV rNP-expressing HeLa cells can be used not only for diagnosis of CCHF but also for epidemiological studies on CCHFV infections. PMID:11825944

Imaging of single retinal ganglion cell with differential interference contrast microscopy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Oh, Juyeong; Kim, Yu Jeong; Kim, Chul-Ki; Lee, Taik Jin; Seo, Mina; Lee, Seok; Woo, Deok Ha; Jun, Seong Chan; Park, Ki-Ho; Kim, Seok Hwan; Kim, Jae Hun

2017-02-01

Glaucoma is a progressive optic neuropathy, characterized by the selective loss of retinal ganglion cells (RGCs). Therefore, monitoring the change of number or morphology of RGC is essential for the early detection as well as investigation of pathophysiology of glaucoma. Since RGC layer is transparent and hyporeflective, the direct optical visualization of RGCs has not been successful so far. Therefore, glaucoma evaluation mostly depends on indirect diagnostic methods such as the evaluation of optic disc morphology or retinal nerve fiber layer thickness measurement by optical coherence tomography. We have previously demonstrated single photoreceptor cell imaging with differential interference contrast (DIC) microscopy. Herein, we successfully visualized single RGC using DIC microscopy. Since RGC layer is much less reflective than photoreceptor layer, various techniques including the control of light wavelength and bandwidth using a tunable band pass filter were adopted to reduce the chromatic aberration in z-axis for higher and clearer resolution. To verify that the imaged cells were the RGCs, the flat-mounted retina of Sprague-Dawley rat, in which the RGCs were retrogradely labeled with fluorescence, was observed by both fluorescence and DIC microscopies for direct comparison. We have confirmed that the cell images obtained by fluorescence microscopy were perfectly matched with cell images by DIC microscopy. As conclusion, we have visualized single RGC with DIC microscopy, and confirmed with fluorescence microscopy.

Characterization of the host response to the myxosporean parasite, Ceratomyxa shasta (Noble), by histology, scanning electron microscopy, and immunological techniques

USGS Publications Warehouse

Bartholomew, J.L.; Smith, C.E.; Rohovec, J.S.; Fryer, J.L.

1989-01-01

The tissue response of Salmo gairdneri Richardson, against the myxosporean parasite. Ceratomyxa shasta (Noble), was investigated using histological techniques, scanning electron microscopy and immunological methods. The progress of infection in C. shasta-susceptible and resistant steelhead and rainbow trout was examined by standard histological techniques and by indirect fluorescent antibody methods using monoclonal antibodies directed against C. shasta antigens. Trophozoite stages were first observed in the posterior intestine and there was indication that resistance was due to the inability of the parasite to penetrate this tissue rather than to an inflammatory response. Examination of a severely infected intestine by scanning electron microscopy showed extensive destruction of the mucosal folds of the posterior intestine. Western blotting and indirect fluorescent antibody techniques were used to investigate the immunological component of the host response. No antibodies specific for C. shasta were detected by either method.

Indirect decentralized learning control

NASA Technical Reports Server (NTRS)

Longman, Richard W.; Lee, Soo C.; Phan, M.

1992-01-01

The new field of learning control develops controllers that learn to improve their performance at executing a given task, based on experience performing this specific task. In a previous work, the authors presented a theory of indirect learning control based on use of indirect adaptive control concepts employing simultaneous identification and control. This paper develops improved indirect learning control algorithms, and studies the use of such controllers in decentralized systems. The original motivation of the learning control field was learning in robots doing repetitive tasks such as on an assembly line. This paper starts with decentralized discrete time systems, and progresses to the robot application, modeling the robot as a time varying linear system in the neighborhood of the nominal trajectory, and using the usual robot controllers that are decentralized, treating each link as if it is independent of any coupling with other links. The basic result of the paper is to show that stability of the indirect learning controllers for all subsystems when the coupling between subsystems is turned off, assures convergence to zero tracking error of the decentralized indirect learning control of the coupled system, provided that the sample time in the digital learning controller is sufficiently short.

Indirect Self-Destructiveness and Emotional Intelligence.

PubMed

Tsirigotis, Konstantinos

2016-06-01

While emotional intelligence may have a favourable influence on the life and psychological and social functioning of the individual, indirect self-destructiveness exerts a rather negative influence. The aim of this study has been to explore possible relations between indirect self-destructiveness and emotional intelligence. A population of 260 individuals (130 females and 130 males) aged 20-30 (mean age of 24.5) was studied by using the Polish version of the chronic self-destructiveness scale and INTE, i.e., the Polish version of the assessing emotions scale. Indirect self-destructiveness has significant correlations with all variables of INTE (overall score, factor I, factor II), and these correlations are negative. The intensity of indirect self-destructiveness differentiates significantly the height of the emotional intelligence and vice versa: the height of the emotional intelligence differentiates significantly the intensity of indirect self-destructiveness. Indirect self-destructiveness has negative correlations with emotional intelligence as well as its components: the ability to recognize emotions and the ability to utilize emotions. The height of emotional intelligence differentiates the intensity of indirect self-destructiveness, and vice versa: the intensity of indirect self-destructiveness differentiates the height of emotional intelligence. It seems advisable to use emotional intelligence in the prophylactic and therapeutic work with persons with various types of disorders, especially with the syndrome of indirect self-destructiveness.

Impact of immunofluorescence on the histological pattern of pediatric kidney biopsies from northern Pakistan.

PubMed

Ali, Akhtar; Ali, Mohammad Usman; Ali, Mahrukh Ayesha

2011-01-01

Kidney biopsy is an investigation for diagnosis and prognosis of a variety of nephritides. It also influences therapeutic options. Immunofluorescence (IMF) greatly adds in identifying the pathologies which may not be obvious on light microscopy (L/M), such as IgM, IgA nephropathy, pauci-immune glomerulonephritis, and anti-glomerular basement membrane disease. We present here data of 170 pediatric kidney biopsies from July 2005 to December 2009 from Department of Nephrology and Hypertension, Lady Reading Hospital, Peshawar, Pakistan. The study was undertaken to see whether IMF would alter the histological pattern of pediatric kidney biopsies and to compare these data with an earlier data from our department of 415 pediatric kidney biopsies done over 7-year period from 1998 to 2005, which were analyzed with L/M alone. Out of 170 kidney biopsies using L/M and IMF, IgM turns out to be most common pattern (20%), followed by minimal change disease (MCD) (17.05%), focal and segmental glomerulosclerosis (FSGS) (15.88%), chronic sclerosing glomerulonephritis (Chr. sclerosing GN) (12.35%), mesangio proliferative glomerulonephritis (MPGN) (7.65%), mesangio capillary glomerulonephritis (MCGN) (6.47%), membranous glomerulonephritis (Mem. GN) (5.29%), IgA nephropathy (5.29%), cresentic glomerulonephritis (Cres. GN) (3.53%), lupus nephritis (2.96%), and others (3.53%). Comparing these results of 170 cases with 415 renal biopsies without IMF, IgM dominated the histological pattern in IMF group whereas MCD followed by FSGS and MPGN were prominent in group without IMF. Therefore, variation in the overall histological pattern with IMF technique proved statistically significant (p < 0.0001). Addition of IMF has altered the frequency of MCD, a change from 24% (100/415) to 17% (29/170), FSGS from 18.3% (76/415) to 15.88% (27/170), and MPGN from 17.35% (72/415) to 7.65% (13/170).

Impact of gluten-friendly™ technology on wheat kernel endosperm and gluten protein structure in seeds by light and electron microscopy.

PubMed

Landriscina, L; D'Agnello, P; Bevilacqua, A; Corbo, M R; Sinigaglia, M; Lamacchia, C

2017-04-15

The main aim of this paper was to assess the impact of Gluten-Friendly™ (GF) technology (Italian priority patent n° 102015000084813 filed on 17th December 2015) on wheat kernel endosperm morphology and gluten protein structure, using SEM, light and immunofluorescent microscopy. Microscopy was combined with immunodetection with specific antibodies for gliadins, γ-gliadins, LMW subunits and antigenic epitopes to gain a better understanding of the technology at a molecular level. The results showed significant changes to gluten proteins after GF treatment; cross-reactivity towards the antibodies recognizing almost the entire range of gluten proteins as well as the antigenic epitopes through the sequences QQSF, QQSY, PEQPFPQGC and QQPFP was significantly reduced. The present study confirms the results from our previous work and shows, for the first time, the mechanism by which a chemical-physical treatment abolishes the antigenic capacity of gluten. Copyright © 2016 Elsevier Ltd. All rights reserved.

A Comparative study for striatal-direct and -indirect pathway neurons to DA depletion-induced lesion in a PD rat model.

PubMed

Zheng, Xuefeng; Wu, Jiajia; Zhu, Yaofeng; Chen, Si; Chen, Zhi; Chen, Tao; Huang, Ziyun; Wei, Jiayou; Li, Yanmei; Lei, Wanlong

2018-04-16

Striatal-direct and -indirect Pathway Neurons showed different vulnerability in basal ganglia disorders. Therefore, present study aimed to examine and compare characteristic changes of densities, protein and mRNA levels of soma, dendrites, and spines between striatal-direct and -indirect pathway neurons after DA depletion by using immunohistochemistry, Western blotting, real-time PCR and immunoelectron microscopy techniques. Experimental results showed that: 1) 6OHDA-induced DA depletion decreased the soma density of striatal-direct pathway neurons (SP+), but no significant changes for striatal-indirect pathway neurons (ENK+). 2) DA depletion resulted in a decline of dendrite density for both striatal-direct (D1+) and -indirect (D2+) pathway neurons, and D2+ dendritic density declined more obviously. At the ultrastructure level, the densities of D1+ and D2+ dendritic spines reduced in the 6OHDA groups compared with their control groups, but the density of D2+ dendritic spines reduced more significant than that of D1. 3) Striatal DA depletion down-regulated protein and mRNA expression levels of SP and D1, on the contrary, ENK and D2 protein and mRNA levels of indirect pathway neurons were up-regulated significantly. Present results suggested that indirect pathway neurons be more sensitive to 6OHDA-induced DA depletion. Copyright © 2018 Elsevier Ltd. All rights reserved.

Full-field optical coherence microscopy is a novel technique for imaging enteric ganglia in the gastrointestinal tract

PubMed Central

CORON, E.; AUKSORIUS, E.; PIERETTI, A.; MAHÉ, M. M.; LIU, L.; STEIGER, C.; BROMBERG, Y.; BOUMA, B.; TEARNEY, G.; NEUNLIST, M.; GOLDSTEIN, A. M.

2013-01-01

Background Noninvasive methods are needed to improve the diagnosis of enteric neuropathies. Full-field optical coherence microscopy (FFOCM) is a novel optical microscopy modality that can acquire 1 μm resolution images of tissue. The objective of this research was to demonstrate FFOCM imaging for the characterization of the enteric nervous system (ENS). Methods Normal mice and EdnrB−/− mice, a model of Hirschsprung’s disease (HD), were imaged in three-dimensions ex vivo using FFOCM through the entire thickness and length of the gut. Quantitative analysis of myenteric ganglia was performed on FFOCM images obtained from whole-mount tissues and compared with immunohistochemistry imaged by confocal microscopy. Key Results Full-field optical coherence microscopy enabled visualization of the full thickness gut wall from serosa to mucosa. Images of the myenteric plexus were successfully acquired from the stomach, duodenum, colon, and rectum. Quantification of ganglionic neuronal counts on FFOCM images revealed strong interobserver agreement and identical values to those obtained by immunofluorescence microscopy. In EdnrB−/− mice, FFOCM analysis revealed a significant decrease in ganglia density along the colorectum and a significantly lower density of ganglia in all colorectal segments compared with normal mice. Conclusions & Inferences Full-field optical coherence microscopy enables optical microscopic imaging of the ENS within the bowel wall along the entire intestine. FFOCM is able to differentiate ganglionic from aganglionic colon in a mouse model of HD, and can provide quantitative assessment of ganglionic density. With further refinements that enable bowel wall imaging in vivo, this technology has the potential to revolutionize the characterization of the ENS and the diagnosis of enteric neuropathies. PMID:23106847

Ultrafast Optical Microscopy of Single Monolayer Molybdenum Disulfide Flakes

DOE PAGES

Seo, Minah; Yamaguchi, Hisato; Mohite, Aditya D.; ...

2016-02-15

We performed ultrafast optical microscopy on single flakes of atomically thin CVD-grown molybdenum disulfide, using non-degenerate femtosecond pump-probe spectroscopy to excite and probe carriers above and below the indirect and direct band gaps. These measurements reveal the influence of layer thickness on carrier dynamics when probing near the band gap. Furthermore, fluence-dependent measurements indicate that carrier relaxation is primarily influenced by surface-related defect and trap states after above-bandgap photoexcitation. Furthermore, the ability to probe femtosecond carrier dynamics in individual flakes can thus give much insight into light-matter interactions in these two-dimensional nanosystems.

Direct immunofluorescence of normal skin in rheumatoid arthritis.

PubMed

Fitzgerald, O M; Barnes, L; Woods, R; McHugh, L; Barry, C; O'Loughlin, S

1985-11-01

The clinical significance of previously described immunoglobulin and complement deposition in the superficial dermal vessel walls of patients with rheumatoid arthritis is unknown. In the present study, skin biopsies were obtained from the normal forearm and buttock of 48 unselected patients with rheumatoid arthritis and were examined by direct immunofluorescence (IF) for the presence of immunoglobulin (IgG,A,M) and complement (C3) in the vessel walls. Deposits of C3, IgM or IgG were detected in 10 patients. Five patients had deposits at the forearm sample alone, four patients had deposits at both biopsy sites, while one patient was positive at the buttock alone. Clinical features were similar in patients with and without vessel IF. However, patients with IF were significantly more seropositive with lower levels of complement and raised levels of serum IgA and IgM. There was also an increased level of circulating IgG immune complexes in these patients. Further analysis following exclusion of seronegative patients revealed similar results. This study suggests that the presence of vessel IF identifies a subgroup of patients who have evidence of more severe immunological disturbance.

Use of direct versus indirect preparation data for assessing risk associated with airborne exposures at asbestos-contaminated sites.

PubMed

Goldade, Mary Patricia; O'Brien, Wendy Pott

2014-01-01

At asbestos-contaminated sites, exposure assessment requires measurement of airborne asbestos concentrations; however, the choice of preparation steps employed in the analysis has been debated vigorously among members of the asbestos exposure and risk assessment communities for many years. This study finds that the choice of preparation technique used in estimating airborne amphibole asbestos exposures for risk assessment is generally not a significant source of uncertainty. Conventionally, the indirect preparation method has been less preferred by some because it is purported to result in false elevations in airborne asbestos concentrations, when compared to direct analysis of air filters. However, airborne asbestos sampling in non-occupational settings is challenging because non-asbestos particles can interfere with the asbestos measurements, sometimes necessitating analysis via indirect preparation. To evaluate whether exposure concentrations derived from direct versus indirect preparation techniques differed significantly, paired measurements of airborne Libby-type amphibole, prepared using both techniques, were compared. For the evaluation, 31 paired direct and indirect preparations originating from the same air filters were analyzed for Libby-type amphibole using transmission electron microscopy. On average, the total Libby-type amphibole airborne exposure concentration was 3.3 times higher for indirect preparation analysis than for its paired direct preparation analysis (standard deviation = 4.1), a difference which is not statistically significant (p = 0.12, two-tailed, Wilcoxon signed rank test). The results suggest that the magnitude of the difference may be larger for shorter particles. Overall, neither preparation technique (direct or indirect) preferentially generates more precise and unbiased data for airborne Libby-type amphibole concentration estimates. The indirect preparation method is reasonable for estimating Libby-type amphibole exposure and

The logic of indirect speech

PubMed Central

Pinker, Steven; Nowak, Martin A.; Lee, James J.

2008-01-01

When people speak, they often insinuate their intent indirectly rather than stating it as a bald proposition. Examples include sexual come-ons, veiled threats, polite requests, and concealed bribes. We propose a three-part theory of indirect speech, based on the idea that human communication involves a mixture of cooperation and conflict. First, indirect requests allow for plausible deniability, in which a cooperative listener can accept the request, but an uncooperative one cannot react adversarially to it. This intuition is supported by a game-theoretic model that predicts the costs and benefits to a speaker of direct and indirect requests. Second, language has two functions: to convey information and to negotiate the type of relationship holding between speaker and hearer (in particular, dominance, communality, or reciprocity). The emotional costs of a mismatch in the assumed relationship type can create a need for plausible deniability and, thereby, select for indirectness even when there are no tangible costs. Third, people perceive language as a digital medium, which allows a sentence to generate common knowledge, to propagate a message with high fidelity, and to serve as a reference point in coordination games. This feature makes an indirect request qualitatively different from a direct one even when the speaker and listener can infer each other's intentions with high confidence. PMID:18199841

Indirect Cost Reimbursement: An Industrial View.

ERIC Educational Resources Information Center

Bolton, Robert

1987-01-01

The meaning of indirect costs in an industrial environment is discussed. Other factors considered are corporate policies; nature of work being supported; the uniqueness of the work; who is doing the negotiating for industry; and indirect rates. Suggestions are offered for approaches to indirect cost reimbursement. (Author/MLW)

46 CFR 154.1720 - Indirect refrigeration.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 46 Shipping 5 2010-10-01 2010-10-01 false Indirect refrigeration. 154.1720 Section 154.1720... § 154.1720 Indirect refrigeration. A refrigeration system that is used to cool acetaldehyde, ethylene oxide, or methyl bromide, must be an indirect refrigeration system that does not use vapor compression. ...

46 CFR 154.1720 - Indirect refrigeration.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 46 Shipping 5 2011-10-01 2011-10-01 false Indirect refrigeration. 154.1720 Section 154.1720... § 154.1720 Indirect refrigeration. A refrigeration system that is used to cool acetaldehyde, ethylene oxide, or methyl bromide, must be an indirect refrigeration system that does not use vapor compression. ...

46 CFR 154.1720 - Indirect refrigeration.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 46 Shipping 5 2012-10-01 2012-10-01 false Indirect refrigeration. 154.1720 Section 154.1720... § 154.1720 Indirect refrigeration. A refrigeration system that is used to cool acetaldehyde, ethylene oxide, or methyl bromide, must be an indirect refrigeration system that does not use vapor compression. ...

46 CFR 154.1720 - Indirect refrigeration.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 46 Shipping 5 2014-10-01 2014-10-01 false Indirect refrigeration. 154.1720 Section 154.1720... § 154.1720 Indirect refrigeration. A refrigeration system that is used to cool acetaldehyde, ethylene oxide, or methyl bromide, must be an indirect refrigeration system that does not use vapor compression. ...

46 CFR 154.1720 - Indirect refrigeration.

Code of Federal Regulations, 2013 CFR

2013-10-01

... 46 Shipping 5 2013-10-01 2013-10-01 false Indirect refrigeration. 154.1720 Section 154.1720... § 154.1720 Indirect refrigeration. A refrigeration system that is used to cool acetaldehyde, ethylene oxide, or methyl bromide, must be an indirect refrigeration system that does not use vapor compression. ...

Tackling the Challenges of Dynamic Experiments Using Liquid-Cell Transmission Electron Microscopy.

PubMed

Parent, Lucas R; Bakalis, Evangelos; Proetto, Maria; Li, Yiwen; Park, Chiwoo; Zerbetto, Francesco; Gianneschi, Nathan C

2018-01-16

Revolutions in science and engineering frequently result from the development, and wide adoption, of a new, powerful characterization or imaging technique. Beginning with the first glass lenses and telescopes in astronomy, to the development of visual-light microscopy, staining techniques, confocal microscopy, and fluorescence super-resolution microscopy in biology, and most recently aberration-corrected, cryogenic, and ultrafast (4D) electron microscopy, X-ray microscopy, and scanning probe microscopy in nanoscience. Through these developments, our perception and understanding of the physical nature of matter at length-scales beyond ordinary perception have been fundamentally transformed. Despite this progression in microscopy, techniques for observing nanoscale chemical processes and solvated/hydrated systems are limited, as the necessary spatial and temporal resolution presents significant technical challenges. However, the standard reliance on indirect or bulk phase characterization of nanoscale samples in liquids is undergoing a shift in recent times with the realization ( Williamson et al. Nat. Mater . 2003 , 2 , 532 - 536 ) of liquid-cell (scanning) transmission electron microscopy, LC(S)TEM, where picoliters of solution are hermetically sealed between electron-transparent "windows," which can be directly imaged or videoed at the nanoscale using conventional transmission electron microscopes. This Account seeks to open a discussion on the topic of standardizing strategies for conducting imaging experiments with a view to characterizing dynamics and motion of nanoscale materials. This is a challenge that could be described by critics and proponents alike, as analogous to doing chemistry in a lightning storm; where the nature of the solution, the nanomaterial, and the dynamic behaviors are all potentially subject to artifactual influence by the very act of our observation.

Two stage indirect evaporative cooling system

DOEpatents

Bourne, Richard C.; Lee, Brian E.; Callaway, Duncan

2005-08-23

A two stage indirect evaporative cooler that moves air from a blower mounted above the unit, vertically downward into dry air passages in an indirect stage and turns the air flow horizontally before leaving the indirect stage. After leaving the dry passages, a major air portion travels into the direct stage and the remainder of the air is induced by a pressure drop in the direct stage to turn 180.degree. and returns horizontally through wet passages in the indirect stage and out of the unit as exhaust air.

19 CFR 10.2024 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.2024 Section 10.2024... Agreement Rules of Origin § 10.2024 Indirect materials. An indirect material, as defined in § 10.2013(i), will be considered to be an originating material without regard to where it is produced. Example...

19 CFR 10.3024 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.3024 Section 10.3024... Promotion Agreement Rules of Origin § 10.3024 Indirect materials. An indirect material, as defined in § 10.3013(h), will be considered to be an originating material without regard to where it is produced...

19 CFR 10.460 - Indirect materials.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 19 Customs Duties 1 2011-04-01 2011-04-01 false Indirect materials. 10.460 Section 10.460 Customs... of Origin § 10.460 Indirect materials. An indirect material, as defined in § 10.402(o), will be considered to be an originating material without regard to where it is produced. Example. Chilean Producer C...

19 CFR 10.460 - Indirect materials.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 19 Customs Duties 1 2013-04-01 2013-04-01 false Indirect materials. 10.460 Section 10.460 Customs... of Origin § 10.460 Indirect materials. An indirect material, as defined in § 10.402(o), will be considered to be an originating material without regard to where it is produced. Example. Chilean Producer C...

19 CFR 10.1024 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.1024 Section 10.1024... Agreement Rules of Origin § 10.1024 Indirect materials. An indirect material, as defined in § 10.1002(n) of.... Korean Producer A produces good C using non-originating material B. Producer A imports non-originating...

19 CFR 10.460 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.460 Section 10.460 Customs... of Origin § 10.460 Indirect materials. An indirect material, as defined in § 10.402(o), will be considered to be an originating material without regard to where it is produced. Example. Chilean Producer C...

Indirect decentralized repetitive control

NASA Technical Reports Server (NTRS)

Lee, Soo Cheol; Longman, Richard W.

1993-01-01

Learning control refers to controllers that learn to improve their performance at executing a given task, based on experience performing this specific task. In a previous work, the authors presented a theory of indirect decentralized learning control based on use of indirect adaptive control concepts employing simultaneous identification and control. This paper extends these results to apply to the indirect repetitive control problem in which a periodic (i.e., repetitive) command is given to a control system. Decentralized indirect repetitive control algorithms are presented that have guaranteed convergence to zero tracking error under very general conditions. The original motivation of the repetitive control and learning control fields was learning in robots doing repetitive tasks such as on an assembly line. This paper starts with decentralized discrete time systems, and progresses to the robot application, modeling the robot as a time varying linear system in the neighborhood of the desired trajectory. Decentralized repetitive control is natural for this application because the feedback control for link rotations is normally implemented in a decentralized manner, treating each link as if it is independent of the other links.

19 CFR 10.541 - Indirect materials.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 19 Customs Duties 1 2011-04-01 2011-04-01 false Indirect materials. 10.541 Section 10.541 Customs... Rules of Origin § 10.541 Indirect materials. An indirect material, as defined in § 10.502(j) of this subpart, will be considered to be an originating material without regard to where it is produced, and its...

19 CFR 10.541 - Indirect materials.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 19 Customs Duties 1 2010-04-01 2010-04-01 false Indirect materials. 10.541 Section 10.541 Customs... Rules of Origin § 10.541 Indirect materials. An indirect material, as defined in § 10.502(j) of this subpart, will be considered to be an originating material without regard to where it is produced, and its...

19 CFR 10.924 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.924 Section 10.924 Customs... Rules of Origin § 10.924 Indirect materials. An indirect material, as defined in § 10.902(m) of this subpart, will be considered to be an originating material without regard to where it is produced. Example...

19 CFR 10.541 - Indirect materials.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 19 Customs Duties 1 2013-04-01 2013-04-01 false Indirect materials. 10.541 Section 10.541 Customs... Rules of Origin § 10.541 Indirect materials. An indirect material, as defined in § 10.502(j) of this subpart, will be considered to be an originating material without regard to where it is produced, and its...

19 CFR 10.541 - Indirect materials.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 19 Customs Duties 1 2014-04-01 2014-04-01 false Indirect materials. 10.541 Section 10.541 Customs... Rules of Origin § 10.541 Indirect materials. An indirect material, as defined in § 10.502(j) of this subpart, will be considered to be an originating material without regard to where it is produced, and its...

14 CFR 296.3 - Indirect cargo air carrier.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 14 Aeronautics and Space 4 2012-01-01 2012-01-01 false Indirect cargo air carrier. 296.3 Section... PROCEEDINGS) ECONOMIC REGULATIONS INDIRECT AIR TRANSPORTATION OF PROPERTY General § 296.3 Indirect cargo air carrier. An indirect cargo air carrier is any U.S. citizen who undertakes to engage indirectly in air...

14 CFR 296.3 - Indirect cargo air carrier.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 14 Aeronautics and Space 4 2013-01-01 2013-01-01 false Indirect cargo air carrier. 296.3 Section... PROCEEDINGS) ECONOMIC REGULATIONS INDIRECT AIR TRANSPORTATION OF PROPERTY General § 296.3 Indirect cargo air carrier. An indirect cargo air carrier is any U.S. citizen who undertakes to engage indirectly in air...

14 CFR 296.3 - Indirect cargo air carrier.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 14 Aeronautics and Space 4 2014-01-01 2014-01-01 false Indirect cargo air carrier. 296.3 Section... PROCEEDINGS) ECONOMIC REGULATIONS INDIRECT AIR TRANSPORTATION OF PROPERTY General § 296.3 Indirect cargo air carrier. An indirect cargo air carrier is any U.S. citizen who undertakes to engage indirectly in air...

14 CFR 296.3 - Indirect cargo air carrier.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 14 Aeronautics and Space 4 2011-01-01 2011-01-01 false Indirect cargo air carrier. 296.3 Section... PROCEEDINGS) ECONOMIC REGULATIONS INDIRECT AIR TRANSPORTATION OF PROPERTY General § 296.3 Indirect cargo air carrier. An indirect cargo air carrier is any U.S. citizen who undertakes to engage indirectly in air...

In situ detection of GM1 and GM2 gangliosides using immunohistochemical and immunofluorescent techniques for auxiliary diagnosis of canine and feline gangliosidoses.

PubMed

Kohyama, Moeko; Yabuki, Akira; Ochiai, Kenji; Nakamoto, Yuya; Uchida, Kazuyuki; Hasegawa, Daisuke; Takahashi, Kimimasa; Kawaguchi, Hiroaki; Tsuboi, Masaya; Yamato, Osamu

2016-03-31

GM1 and GM2 gangliosidoses are progressive neurodegenerative lysosomal storage diseases resulting from the excessive accumulation of GM1 and GM2 gangliosides in the lysosomes, respectively. The diagnosis of gangliosidosis is carried out based on comprehensive findings using various types of specimens for histological, ultrastructural, biochemical and genetic analyses. Therefore, the partial absence or lack of specimens might have resulted in many undiagnosed cases. The aim of the present study was to establish immunohistochemical and immunofluorescent techniques for the auxiliary diagnosis of canine and feline gangliosidoses, using paraffin-embedded brain specimens stored for a long period. Using hematoxylin and eosin staining, cytoplasmic accumulation of pale to eosinophilic granular materials in swollen neurons was observed in animals previously diagnosed with GM1 or GM2 gangliosidosis. The immunohistochemical and immunofluorescent techniques developed in this study clearly demonstrated the accumulated material to be either GM1 or GM2 ganglioside. Immunohistochemical and immunofluorescent techniques using stored paraffin-embedded brain specimens are useful for the retrospective diagnosis of GM1 and GM2 gangliosidoses in dogs and cats.

A novel fiber laser development for photoacoustic microscopy

NASA Astrophysics Data System (ADS)

Yavas, Seydi; Aytac-Kipergil, Esra; Arabul, Mustafa U.; Erkol, Hakan; Akcaalan, Onder; Eldeniz, Y. Burak; Ilday, F. Omer; Unlu, Mehmet B.

2013-03-01

Photoacoustic microscopy, as an imaging modality, has shown promising results in imaging angiogenesis and cutaneous malignancies like melanoma, revealing systemic diseases including diabetes, hypertension, tracing drug efficiency and assessment of therapy, monitoring healing processes such as wound cicatrization, brain imaging and mapping. Clinically, photoacoustic microscopy is emerging as a capable diagnostic tool. Parameters of lasers used in photoacoustic microscopy, particularly, pulse duration, energy, pulse repetition frequency, and pulse-to-pulse stability affect signal amplitude and quality, data acquisition speed and indirectly, spatial resolution. Lasers used in photoacoustic microscopy are typically Q-switched lasers, low-power laser diodes, and recently, fiber lasers. Significantly, the key parameters cannot be adjusted independently of each other, whereas microvasculature and cellular imaging, e.g., have different requirements. Here, we report an integrated fiber laser system producing nanosecond pulses, covering the spectrum from 600 nm to 1100 nm, developed specifically for photoacoustic excitation. The system comprises of Yb-doped fiber oscillator and amplifier, an acousto-optic modulator and a photonic-crystal fiber to generate supercontinuum. Complete control over the pulse train, including generation of non-uniform pulse trains, is achieved via the AOM through custom-developed field-programmable gate-array electronics. The system is unique in that all the important parameters are adjustable: pulse duration in the range of 1-3 ns, pulse energy up to 10 μJ, repetition rate from 50 kHz to 3 MHz. Different photocoustic imaging probes can be excited with the ultrabroad spectrum. The entire system is fiber-integrated; guided-beam-propagation rendersit misalignment free and largely immune to mechanical perturbations. The laser is robust, low-cost and built using readily available components.

Radiative capture reactions via indirect methods

NASA Astrophysics Data System (ADS)

Mukhamedzhanov, A. M.; Rogachev, G. V.

2017-10-01

Many radiative capture reactions of astrophysical interest occur at such low energies that their direct measurement is hardly possible. Until now the only indirect method, which was used to determine the astrophysical factor of the astrophysical radiative capture process, was the Coulomb dissociation. In this paper we address another indirect method, which can provide information about resonant radiative capture reactions at astrophysically relevant energies. This method can be considered an extension of the Trojan horse method for resonant radiative capture reactions. The idea of the suggested indirect method is to use the indirect reaction A (a ,s γ )F to obtain information about the radiative capture reaction A (x ,γ )F , where a =(s x ) and F =(x A ) . The main advantage of using the indirect reactions is the absence of the penetrability factor in the channel x +A , which suppresses the low-energy cross sections of the A (x ,γ )F reactions and does not allow one to measure these reactions at astrophysical energies. A general formalism to treat indirect resonant radiative capture reactions is developed when only a few intermediate states contribute and a statistical approach cannot be applied. The indirect method requires coincidence measurements of the triple differential cross section, which is a function of the photon scattering angle, energy, and the scattering angle of the outgoing spectator particle s . Angular dependence of the triple differential cross section at fixed scattering angle of the spectator s is the angular γ -s correlation function. Using indirect resonant radiative capture reactions, one can obtain information about important astrophysical resonant radiative capture reactions such as (p ,γ ) , (α ,γ ) , and (n ,γ ) on stable and unstable isotopes. The indirect technique makes accessible low-lying resonances, which are close to the threshold, and even subthreshold bound states located at negative energies. In this paper, after

Ultrasonic force microscopy: detection and imaging of ultra-thin molecular domains.

PubMed

Dinelli, Franco; Albonetti, Cristiano; Kolosov, Oleg V

2011-03-01

The analysis of the formation of ultra-thin organic films is a very important issue. In fact, it is known that the properties of organic light emitting diodes and field effect transistors are strongly affected by the early growth stages. For instance, in the case of sexithiophene, the presence of domains made of molecules with the backbone parallel to the substrate surface has been indirectly evidenced by photoluminescence spectroscopy and confocal microscopy. On the contrary, conventional scanning force microscopy both in contact and intermittent contact modes have failed to detect such domains. In this paper, we show that Ultrasonic Force Microscopy (UFM), sensitive to nanomechanical properties, allows one to directly identify the structure of sub-monolayer thick films. Sexithiophene flat domains have been imaged for the first time with nanometer scale spatial resolution. A comparison with lateral force and intermittent contact modes has been carried out in order to explain the origins of the UFM contrast and its advantages. In particular, it indicates that UFM is highly suitable for investigations where high sensitivity to material properties, low specimen damage and high spatial resolution are required. Copyright © 2010 Elsevier B.V. All rights reserved.

Automated analysis of siRNA screens of cells infected by hepatitis C and dengue viruses based on immunofluorescence microscopy images

NASA Astrophysics Data System (ADS)

Matula, Petr; Kumar, Anil; Wörz, Ilka; Harder, Nathalie; Erfle, Holger; Bartenschlager, Ralf; Eils, Roland; Rohr, Karl

2008-03-01

We present an image analysis approach as part of a high-throughput microscopy siRNA-based screening system using cell arrays for the identification of cellular genes involved in hepatitis C and dengue virus replication. Our approach comprises: cell nucleus segmentation, quantification of virus replication level in the neighborhood of segmented cell nuclei, localization of regions with transfected cells, cell classification by infection status, and quality assessment of an experiment and single images. In particular, we propose a novel approach for the localization of regions of transfected cells within cell array images, which combines model-based circle fitting and grid fitting. By this scheme we integrate information from single cell array images and knowledge from the complete cell arrays. The approach is fully automatic and has been successfully applied to a large number of cell array images from screening experiments. The experimental results show a good agreement with the expected behaviour of positive as well as negative controls and encourage the application to screens from further high-throughput experiments.

A study of the aetiology of gastritis following gastric surgery. I. Immunofluorescent studies of the gastric mucosa.

PubMed Central

Chapel, H M; Hoare, A M

1979-01-01

The gastritis which follows surgical trauma is probably not of autoimmune origin. Although identical with the gastritis of pernicious anaemia (PA) upon routine histological examination, immunofluorescent examination of post-operative gastritis differs in that IgA-containing plasma cells alone are found, in contrast to the predominantly IgG-containing plasma cells in PA. PMID:389496

Indirect Reciprocity; A Field Experiment.

PubMed

van Apeldoorn, Jacobien; Schram, Arthur

2016-01-01

Indirect reciprocity involves cooperative acts towards strangers, either in response to their kindness to third parties (downstream) or after receiving kindness from others oneself (upstream). It is considered to be important for the evolution of cooperative behavior amongst humans. Though it has been widely studied theoretically, the empirical evidence of indirect reciprocity has thus far been limited and based solely on behavior in laboratory experiments. We provide evidence from an online environment where members can repeatedly ask and offer services to each other, free of charge. For the purpose of this study we created several new member profiles, which differ only in terms of their serving history. We then sent out a large number of service requests to different members from all over the world. We observe that a service request is more likely to be rewarded for those with a profile history of offering the service (to third parties) in the past. This provides clear evidence of (downstream) indirect reciprocity. We find no support for upstream indirect reciprocity (in this case, rewarding the service request after having previously received the service from third parties), however. Our evidence of downstream indirect reciprocity cannot be attributed to reputational effects concerning one's trustworthiness as a service user.

Indirect Reciprocity; A Field Experiment

PubMed Central

van Apeldoorn, Jacobien; Schram, Arthur

2016-01-01

Indirect reciprocity involves cooperative acts towards strangers, either in response to their kindness to third parties (downstream) or after receiving kindness from others oneself (upstream). It is considered to be important for the evolution of cooperative behavior amongst humans. Though it has been widely studied theoretically, the empirical evidence of indirect reciprocity has thus far been limited and based solely on behavior in laboratory experiments. We provide evidence from an online environment where members can repeatedly ask and offer services to each other, free of charge. For the purpose of this study we created several new member profiles, which differ only in terms of their serving history. We then sent out a large number of service requests to different members from all over the world. We observe that a service request is more likely to be rewarded for those with a profile history of offering the service (to third parties) in the past. This provides clear evidence of (downstream) indirect reciprocity. We find no support for upstream indirect reciprocity (in this case, rewarding the service request after having previously received the service from third parties), however. Our evidence of downstream indirect reciprocity cannot be attributed to reputational effects concerning one’s trustworthiness as a service user. PMID:27043712

Comparing the shear bond strength of direct and indirect composite inlays in relation to different surface conditioning and curing techniques

PubMed Central

Zorba, Yahya Orcun; Ilday, Nurcan Ozakar; Bayındır, Yusuf Ziya; Demirbuga, Sezer

2013-01-01

Objective: The aim of this study was to test the null hypothesis that different surface conditioning (etch and rinse and self-etch) and curing techniques (light cure/dual cure) had no effect on the shear bond strength of direct and indirect composite inlays. Materials and Methods: A total of 112 extracted human molar teeth were horizontally sectioned and randomly divided into two groups according to restoration technique (direct and indirect restorations). Each group was further subdivided into seven subgroups (n = 8) according to bonding agent (etch and rinse adhesives Scotchbond multi-purpose plus, All-Bond 3, Adper Single Bond and Prime Bond NT; and self-etch adhesives Clearfil Liner Bond, Futurabond DC and G bond). Indirect composites were cemented to dentin surfaces using dual-curing luting cement. Shear bond strength of specimens was tested using a Universal Testing Machine. Two samples from each subgroup were evaluated under Scanning electron microscopy to see the failing modes. Data was analyzed using independent sample t-tests and Tukey's tests. Results: Surface conditioning and curing of bonding agents were all found to have significant effects on shear bond strength (P < 0.05) of both direct and indirect composite inlays. With direct restoration, etch and rinse systems and dual-cured bonding agents yielded higher bond strengths than indirect restoration, self-etch systems and light-cured bonding agents. Conclusions: The results of the present study indicated that direct restoration to be a more reliable method than indirect restoration. Although etch and rinse bonding systems showed higher shear bond strength to dentin than self-etch systems, both systems can be safely used for the adhesion of direct as well as indirect restorations. PMID:24932118

Comparing the shear bond strength of direct and indirect composite inlays in relation to different surface conditioning and curing techniques.

PubMed

Zorba, Yahya Orcun; Ilday, Nurcan Ozakar; Bayındır, Yusuf Ziya; Demirbuga, Sezer

2013-10-01

The aim of this study was to test the null hypothesis that different surface conditioning (etch and rinse and self-etch) and curing techniques (light cure/dual cure) had no effect on the shear bond strength of direct and indirect composite inlays. A total of 112 extracted human molar teeth were horizontally sectioned and randomly divided into two groups according to restoration technique (direct and indirect restorations). Each group was further subdivided into seven subgroups (n = 8) according to bonding agent (etch and rinse adhesives Scotchbond multi-purpose plus, All-Bond 3, Adper Single Bond and Prime Bond NT; and self-etch adhesives Clearfil Liner Bond, Futurabond DC and G bond). Indirect composites were cemented to dentin surfaces using dual-curing luting cement. Shear bond strength of specimens was tested using a Universal Testing Machine. Two samples from each subgroup were evaluated under Scanning electron microscopy to see the failing modes. Data was analyzed using independent sample t-tests and Tukey's tests. Surface conditioning and curing of bonding agents were all found to have significant effects on shear bond strength (P < 0.05) of both direct and indirect composite inlays. With direct restoration, etch and rinse systems and dual-cured bonding agents yielded higher bond strengths than indirect restoration, self-etch systems and light-cured bonding agents. The results of the present study indicated that direct restoration to be a more reliable method than indirect restoration. Although etch and rinse bonding systems showed higher shear bond strength to dentin than self-etch systems, both systems can be safely used for the adhesion of direct as well as indirect restorations.

The ultrastructure of mono- and holocentric plant centromeres: an immunological investigation by structured illumination microscopy and scanning electron microscopy.

PubMed

Wanner, Gerhard; Schroeder-Reiter, Elizabeth; Ma, Wei; Houben, Andreas; Schubert, Veit

2015-12-01

The spatial distribution of the three centromere-associated proteins α-tubulin, CENH3, and phosphorylated histone H2A (at threonine 120, H2AThr120ph) was analysed by indirect immunodetection at monocentric cereal chromosomes and at the holocentric chromosomes of Luzula elegans by super-resolution light microscopy and scanning electron microscopy (SEM). Using structured illumination microscopy (SIM) as the super-resolution technique on squashed specimens and SEM on uncoated isolated specimens, the three-dimensional (3D) distribution of the proteins was visualized at the centromeres. Technical aspects of 3D SEM are explained in detail. We show that CENH3 forms curved "pads" mainly around the lateral centromeric region in the primary constriction of metacentric chromosomes. H2AThr120ph is present in both the primary constriction and in the pericentromere. α-tubulin-labeled microtubule bundles attach to CENH3-containing chromatin structures, either in single bundles with a V-shaped attachment to the centromere or in split bundles to bordering pericentromeric flanks. In holocentric L. elegans chromosomes, H2AThr120ph is located predominantly in the centromeric groove of each chromatid as proven by subsequent FIB/FESEM ablation and 3D reconstruction. α-tubulin localizes to the edges of the groove. In both holocentric and monocentric chromosomes, no additional intermediate structures between microtubules and the centromere were observed. We established models of the distribution of CENH3, H2AThr120ph and the attachment sites of microtubules for metacentric and holocentric plant chromosomes.

48 CFR 342.705 - Final indirect cost rates.

Code of Federal Regulations, 2012 CFR

2012-10-01

...) The Division of Financial Advisory Services, NIH, shall establish indirect cost rates, fringe benefit... 48 Federal Acquisition Regulations System 4 2012-10-01 2012-10-01 false Final indirect cost rates... MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 342.705 Final indirect cost rates. (a) The Division of...

48 CFR 342.705 - Final indirect cost rates.

Code of Federal Regulations, 2014 CFR

2014-10-01

...) The Division of Financial Advisory Services, NIH, shall establish indirect cost rates, fringe benefit... 48 Federal Acquisition Regulations System 4 2014-10-01 2014-10-01 false Final indirect cost rates... MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 342.705 Final indirect cost rates. (a) The Division of...

48 CFR 342.705 - Final indirect cost rates.

Code of Federal Regulations, 2011 CFR

2011-10-01

...) The Division of Financial Advisory Services, NIH, shall establish indirect cost rates, fringe benefit... 48 Federal Acquisition Regulations System 4 2011-10-01 2011-10-01 false Final indirect cost rates... MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 342.705 Final indirect cost rates. (a) The Division of...

48 CFR 342.705 - Final indirect cost rates.

Code of Federal Regulations, 2013 CFR

2013-10-01

...) The Division of Financial Advisory Services, NIH, shall establish indirect cost rates, fringe benefit... 48 Federal Acquisition Regulations System 4 2013-10-01 2013-10-01 false Final indirect cost rates... MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 342.705 Final indirect cost rates. (a) The Division of...

48 CFR 342.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

...) The Division of Financial Advisory Services, NIH, shall establish indirect cost rates, fringe benefit... 48 Federal Acquisition Regulations System 4 2010-10-01 2010-10-01 false Final indirect cost rates... MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 342.705 Final indirect cost rates. (a) The Division of...

Increasing the accuracy and scalability of the Immunofluorescence Assay for Epstein Barr Virus by inferring continuous titers from a single sample dilution.

PubMed

Goh, Sherry Meow Peng; Swaminathan, Muthukaruppan; Lai, Julian U-Ming; Anwar, Azlinda; Chan, Soh Ha; Cheong, Ian

2017-01-01

High Epstein Barr Virus (EBV) titers detected by the indirect Immunofluorescence Assay (IFA) are a reliable predictor of Nasopharyngeal Carcinoma (NPC). Despite being the gold standard for serological detection of NPC, the IFA is limited by scaling bottlenecks. Specifically, 5 serial dilutions of each patient sample must be prepared and visually matched by an evaluator to one of 5 discrete titers. Here, we describe a simple method for inferring continuous EBV titers from IFA images acquired from NPC-positive patient sera using only a single sample dilution. In the first part of our study, 2 blinded evaluators used a set of reference titer standards to perform independent re-evaluations of historical samples with known titers. Besides exhibiting high inter-evaluator agreement, both evaluators were also in high concordance with historical titers, thus validating the accuracy of the reference titer standards. In the second part of the study, the reference titer standards were IFA-processed and assigned an 'EBV Score' using image analysis. A log-linear relationship between titers and EBV Score was observed. This relationship was preserved even when images were acquired and analyzed 3days post-IFA. We conclude that image analysis of IFA-processed samples can be used to infer a continuous EBV titer with just a single dilution of NPC-positive patient sera. This work opens new possibilities for improving the accuracy and scalability of IFA in the context of clinical screening. Copyright © 2016. Published by Elsevier B.V.

Bacterial adhesion on direct and indirect dental restorative composite resins: An in vitro study on a natural biofilm.

PubMed

Derchi, Giacomo; Vano, Michele; Barone, Antonio; Covani, Ugo; Diaspro, Alberto; Salerno, Marco

2017-05-01

Both direct and indirect techniques are used for dental restorations. Which technique should be preferred or whether they are equivalent with respect to bacterial adhesion is unclear. The purpose of this in vitro study was to determine the affinity of bacterial biofilm to dental restorative composite resins placed directly and indirectly. Five direct composite resins for restorations (Venus Diamond, Adonis, Optifil, Enamel Plus HRi, Clearfil Majesty Esthetic) and 3 indirect composite resins (Gradia, Estenia, Signum) were selected. The materials were incubated in unstimulated whole saliva for 1 day. The biofilms grown were collected and their bacterial cells counted. In parallel, the composite resin surface morphology was analyzed with atomic force microscopy. Both bacterial cell count and surface topography parameters were subjected to statistical analysis (α=.05). Indirect composite resins showed significantly lower levels than direct composite resins for bacterial cell adhesion, (P<.001). No significant differences were observed within the direct composite resins (P>.05). However, within the indirect composite resins a significantly lower level was found for Gradia than Estenia or Signum (P<.01). A partial correlation was observed between composite resin roughness and bacterial adhesion when the second and particularly the third-order statistical moments of the composite resin height distributions were considered. Indirect dental restorative composite resins were found to be less prone to biofilm adhesion than direct composite resins. A correlation of bacterial adhesion to surface morphology exists that is described by kurtosis; thus, advanced data analysis is required to discover possible insights into the biologic effects of morphology. Copyright © 2016 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.

7 CFR 2903.4 - Indirect costs.

Code of Federal Regulations, 2013 CFR

2013-01-01

... AGRICULTURE BIODIESEL FUEL EDUCATION PROGRAM General Information § 2903.4 Indirect costs. (a) For the Biodiesel Fuel Education Program, applicants should use the current indirect cost rate negotiated with the...

7 CFR 2903.4 - Indirect costs.

Code of Federal Regulations, 2011 CFR

2011-01-01

... AGRICULTURE BIODIESEL FUEL EDUCATION PROGRAM General Information § 2903.4 Indirect costs. (a) For the Biodiesel Fuel Education Program, applicants should use the current indirect cost rate negotiated with the...

7 CFR 2903.4 - Indirect costs.

Code of Federal Regulations, 2014 CFR

2014-01-01

... AGRICULTURE BIODIESEL FUEL EDUCATION PROGRAM General Information § 2903.4 Indirect costs. (a) For the Biodiesel Fuel Education Program, applicants should use the current indirect cost rate negotiated with the...

7 CFR 2903.4 - Indirect costs.

Code of Federal Regulations, 2012 CFR

2012-01-01

... AGRICULTURE BIODIESEL FUEL EDUCATION PROGRAM General Information § 2903.4 Indirect costs. (a) For the Biodiesel Fuel Education Program, applicants should use the current indirect cost rate negotiated with the...

7 CFR 2903.4 - Indirect costs.

Code of Federal Regulations, 2010 CFR

2010-01-01

... AGRICULTURE BIODIESEL FUEL EDUCATION PROGRAM General Information § 2903.4 Indirect costs. (a) For the Biodiesel Fuel Education Program, applicants should use the current indirect cost rate negotiated with the...

Correlative Stochastic Optical Reconstruction Microscopy and Electron Microscopy

PubMed Central

Kim, Doory; Deerinck, Thomas J.; Sigal, Yaron M.; Babcock, Hazen P.; Ellisman, Mark H.; Zhuang, Xiaowei

2015-01-01

Correlative fluorescence light microscopy and electron microscopy allows the imaging of spatial distributions of specific biomolecules in the context of cellular ultrastructure. Recent development of super-resolution fluorescence microscopy allows the location of molecules to be determined with nanometer-scale spatial resolution. However, correlative super-resolution fluorescence microscopy and electron microscopy (EM) still remains challenging because the optimal specimen preparation and imaging conditions for super-resolution fluorescence microscopy and EM are often not compatible. Here, we have developed several experiment protocols for correlative stochastic optical reconstruction microscopy (STORM) and EM methods, both for un-embedded samples by applying EM-specific sample preparations after STORM imaging and for embedded and sectioned samples by optimizing the fluorescence under EM fixation, staining and embedding conditions. We demonstrated these methods using a variety of cellular targets. PMID:25874453

IFDOTMETER: A New Software Application for Automated Immunofluorescence Analysis.

PubMed

Rodríguez-Arribas, Mario; Pizarro-Estrella, Elisa; Gómez-Sánchez, Rubén; Yakhine-Diop, S M S; Gragera-Hidalgo, Antonio; Cristo, Alejandro; Bravo-San Pedro, Jose M; González-Polo, Rosa A; Fuentes, José M

2016-04-01

Most laboratories interested in autophagy use different imaging software for managing and analyzing heterogeneous parameters in immunofluorescence experiments (e.g., LC3-puncta quantification and determination of the number and size of lysosomes). One solution would be software that works on a user's laptop or workstation that can access all image settings and provide quick and easy-to-use analysis of data. Thus, we have designed and implemented an application called IFDOTMETER, which can run on all major operating systems because it has been programmed using JAVA (Sun Microsystems). Briefly, IFDOTMETER software has been created to quantify a variety of biological hallmarks, including mitochondrial morphology and nuclear condensation. The program interface is intuitive and user-friendly, making it useful for users not familiar with computer handling. By setting previously defined parameters, the software can automatically analyze a large number of images without the supervision of the researcher. Once analysis is complete, the results are stored in a spreadsheet. Using software for high-throughput cell image analysis offers researchers the possibility of performing comprehensive and precise analysis of a high number of images in an automated manner, making this routine task easier. © 2015 Society for Laboratory Automation and Screening.

PEDOT:PSS/GO nanocomposites: Determination of the aspect ratio by indirect measurements

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

Giuri, Antonella; Colella, Silvia; Listorti, Andrea

2016-05-18

Polymer nanocomposites properties significantly depend on the average size of the fillers dispersed into the matrix and on the grade of the dispersion, the latter influenced by the process techniques. In this work, we determined the aspect ratio of graphene oxide (GO) dispersed into Poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS), starting from the indirect measurement of the rheological behavior of polymer/filler mixtures, as a function of the shear rate and the volumetric composition. PEDOT:PSS+GO nanocomposite films were also realized by spin coating on different substrates and characterized by Scanning electron microscopy (SEM) and X-ray diffraction (XRD), in order to analyze the quality of themore » dispersion, even by direct measurements.« less

Investigation of skin structures based on infrared wave parameter indirect microscopic imaging

NASA Astrophysics Data System (ADS)

Zhao, Jun; Liu, Xuefeng; Xiong, Jichuan; Zhou, Lijuan

2017-02-01

Detailed imaging and analysis of skin structures are becoming increasingly important in modern healthcare and clinic diagnosis. Nanometer resolution imaging techniques such as SEM and AFM can cause harmful damage to the sample and cannot measure the whole skin structure from the very surface through epidermis, dermis to subcutaneous. Conventional optical microscopy has the highest imaging efficiency, flexibility in onsite applications and lowest cost in manufacturing and usage, but its image resolution is too low to be accepted for biomedical analysis. Infrared parameter indirect microscopic imaging (PIMI) uses an infrared laser as the light source due to its high transmission in skins. The polarization of optical wave through the skin sample was modulated while the variation of the optical field was observed at the imaging plane. The intensity variation curve of each pixel was fitted to extract the near field polarization parameters to form indirect images. During the through-skin light modulation and image retrieving process, the curve fitting removes the blurring scattering from neighboring pixels and keeps only the field variations related to local skin structures. By using the infrared PIMI, we can break the diffraction limit, bring the wide field optical image resolution to sub-200nm, in the meantime of taking advantage of high transmission of infrared waves in skin structures.

Rapid Diagnosis of Arbovirus and Arenavirus Infections by Immunofluorescence.

DTIC Science & Technology

1981-01-01

in the indirect imunofluorescence test, have been prepared. The viruses in the slide were: lanai, Japanese encephalitis, Langat , Rocio and yellow fever... Langat , Rocio and yellow fever. The slides gave positive reactions when tested with hyperimmune mouse sera including 6 of 7 sera for viruses not...Nakayama; Langat , TP21; Pongola, MP 781; Rocio, San Paulo ; Sicilian phlebotomus fever, Sabin; West Nile, EgIO; yellow fever, Asibi. the viruses are

An expert protocol for immunofluorescent detection of calcium channels in tsA-201 cells.

PubMed

Koch, Peter; Herzig, Stefan; Matthes, Jan

Pore-forming subunits of voltage gated calcium channels (VGCC) are large membrane proteins (260kDa) containing 24 transmembrane domains. Despite transfection with viral promoter driven vectors, biochemical analysis of VGCC is often hampered by rather low expression levels in heterologous systems rendering VGCC challenging targets. Especially in immunofluorescent detection, calcium channels are demanding proteins. We provide an expert step-by-step protocol with adapted conditions for handling procedures (tsA-201 cell culture, transient transfection, incubation time and temperature at 28°C or 37°C and immunostaining) to address the L-type calcium-channel pore Ca v 1.2 in an immunofluorescent approach. We performed immunocytochemical analysis of Ca v 1.2 expression at single-cell level in combination with detection of different markers for cellular organelles. We show confluency levels and shapes of tsA-201 cells at different time points during an experiment. Our experiments reveal sufficient levels of Ca v 1.2 protein and a correct Ca v 1.2 expression pattern in polygonal shaped cells already 12h after transfection. A sequence of elaborated protocol modifications allows subcellular localization analysis of Ca v 1.2 in an immunocytochemical approach. We provide a protocol that may be used to achieve insights into physiological and pathophysiological processes involving voltage gated calcium channels. Our protocol may be used for expression analysis of other challenging proteins and efficient overexpression may be exploited in related biochemical techniques requiring immunolabels. Copyright © 2016 Elsevier Inc. All rights reserved.

19 CFR 10.460 - Indirect materials.

Code of Federal Regulations, 2010 CFR

2010-04-01

... ARTICLES CONDITIONALLY FREE, SUBJECT TO A REDUCED RATE, ETC. United States-Chile Free Trade Agreement Rules of Origin § 10.460 Indirect materials. An indirect material, as defined in § 10.402(o), will be...

You can't measure what you can't see - detectors for microscopies

NASA Astrophysics Data System (ADS)

Denes, Peter

For centuries, the human eye has been the imaging detector of choice thanks to its high sensitivity, wide dynamic range, and direct connection to a built-in data recording and analysis system. The eye, however, is limited to visible light, which excludes microscopies with electrons and X-rays, and the built-in recording system stores archival information at very low rates. The former limitation has been overcome by ``indirect'' detectors, which convert probe particles to visible light, and the latter by a variety of recording techniques, from photographic film to semiconductor-based imagers. Semiconductor imagers have been used for decades as ``direct'' detectors in particle physics, and almost as long for hard X-rays. For soft X-ray microscopy, the challenge has been the small signal levels - plus getting the X-rays into the detector itself, given how quickly they are absorbed in inert layers. For electron microscopy, the challenge has been reconciling detector spatial resolution and pixel count with the large multiple scattering of electrons with energies used for microscopy. Further, a high recording rate (``movies'' rather than ``snapshots'') enables time-resolved studies, time-dependent corrections, shot-by-shot experiments and scanning techniques - at the expense of creating large data volumes. This talk will discuss solutions to these challenges, as well as an outlook towards future developments.

Simian virus 40 T-antigen-related cell surface antigen: serological demonstration on simian virus 40-transformed monolayer cells in situ.

PubMed Central

Deppert, W; Hanke, K; Henning, R

1980-01-01

Simian virus 40 (SV40)-transformed monolayer cells were analyzed in situ by indirect immunofluorescence microscopy for the postulated cell surface location of SV40 T-antigen-related molecules. With antisera prepared against purified, sodium dodecyl sulfate-denatured SV40 T-antigen, positive surface staining was obtained when the cells had been treated with formaldehyde before immunofluorescence analysis. In contrast, living SV40-transformed cells analyzed in monolayer were surface fluorescence negative. The fixation procedure developed in this study combined with a double staining immunofluorescence technique allowed the simultaneous analysis of the same cells for the expression of both SV40 T-antigen-related surface antigen and nuclear T-antigen. The localization of SV40 T-antigen-related surface antigen on the outer surface of the plasma membrane of formaldehyde-fixed SV40-transformed cells was demonstrated directly by the protein A-mediated binding of Staphylococcus aureus bacteria on formaldehyde-fixed SV40-transformed cells precoated with antiserum against sodium dodecyl sulfate-denatured T-antigen. Both cell surface staining and S. aureus binding were found to be highly specific for SV40 T-antigen-related binding sites. These results indicate that T-antigen-related molecules in a cryptic form are located on the surface of SV40-transformed monolayer cells and can be detected in situ after modification of the cell surface architecture. Images PMID:6255189

Increased centrosome number in BRCA-related breast cancer specimens determined by immunofluorescence analysis.

PubMed

Watanabe, Gou; Chiba, Natsuko; Nomizu, Tadashi; Furuta, Akihiko; Sato, Kaolu; Miyashita, Minoru; Tada, Hiroshi; Suzuki, Akihiko; Ohuchi, Noriaki; Ishida, Takanori

2018-06-01

BRCA-related breast carcinoma can be prevented through prophylactic surgery and an intensive follow-up regimen. However, BRCA genetic tests cannot be routinely performed, and some BRCA mutations could not be defined as deleterious mutations or normal variants. Therefore, an easy functional assay of BRCA will be useful to evaluate BRCA status. As it has been reported that BRCA functions in the regulation of centrosome number, we focused on centrosome number in cancer tissues. Here, 70 breast cancer specimens with known BRCA status were analyzed using immunofluorescence of γ-tubulin (a marker of centrosome) foci. The number of foci per cell was higher in cases with BRCA mutation compared to wild-type cases, that is, 1.9 (95% confidence interval [CI], 1.5-2.3) vs 0.5 (95% CI, 0.2-0.8) (P < .001). Specifically, foci numbers per cell in BRCA1 and BRCA2 mutation cases were 1.2 (95% CI, 0.6-1.8) and 2.2 (95% CI, 1.7-2.6), respectively, both higher than those in wild-type cases (P = .042 and P < .0001, respectively). The predictive value of γ-tubulin foci as determined by area under the curve (AUC = 0.86) for BRCA status was superior to BRCAPRO (AUC = 0.69), Myriad Table (AUC = 0.61), and KOHBRA BRCA risk calculator (AUC = 0.65) pretest values. The use of γ-tubulin foci to predict BRCA status had sensitivity = 83% (19/23), specificity = 89% (42/47), and positive predictive value = 77% (20/26). Thus, γ-tubulin immunofluorescence, a functional assessment of BRCA, can be used as a new prospective test of BRCA status. © 2018 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Multiplexed Immunofluorescence Reveals Potential PD-1/PD-L1 Pathway Vulnerabilities in Craniopharyngioma.

PubMed

Coy, Shannon; Rashid, Rumana; Lin, Jia-Ren; Du, Ziming; Donson, Andrew M; Hankinson, Todd C; Foreman, Nicholas K; Manley, Peter E; Kieran, Mark W; Reardon, David A; Sorger, Peter K; Santagata, Sandro

2018-03-02

Craniopharyngiomas are neoplasms of the sellar/parasellar region that are classified into adamantinomatous (ACP) and papillary (PCP) subtypes. Surgical resection of craniopharyngiomas is challenging, and recurrence is common, frequently leading to profound morbidity. BRAF V600E mutations render PCP susceptible to BRAF/MEK inhibitors, but effective targeted therapies are needed for ACP. We explored the feasibility of targeting the PD-1/PD-L1 immune checkpoint pathway in ACP and PCP. We mapped and quantified PD-L1 and PD-1 expression in ACP and PCP resections using immunohistochemistry, immunofluorescence, and RNA in situ hybridization. We used tissue-based cyclic immunofluorescence (t-CyCIF) to map the spatial distribution of immune cells and characterize cell cycle and signaling pathways in ACP tumor cells which intrinsically express PD-1. All ACP (15±14% of cells, n=23, average±S.D.) and PCP (35±22% of cells, n=18) resections expressed PD-L1. In ACP, PD-L1 was predominantly expressed by tumor cells comprising the cyst-lining. In PCP, PD-L1 was highly-expressed by tumor cells surrounding the stromal fibrovascular cores. ACP also exhibited tumor cell-intrinsic PD-1 expression in whorled epithelial cells with nuclear-localized beta-catenin. These cells exhibited evidence of elevated mTOR and MAPK signaling. Profiling of immune populations in ACP and PCP showed a modest density of CD8+ T-cells. ACP exhibit PD-L1 expression in the tumor cyst-lining and intrinsic PD-1 expression in cells proposed to comprise an oncogenic stem-like population. In PCP, proliferative tumor cells express PD-L1 in a continuous band at the stromal-epithelial interface. Targeting PD-L1 and/or PD-1 in both subtypes of craniopharyngioma might therefore be an effective therapeutic strategy.

Fluorescence microscopy.

PubMed

Sanderson, Michael J; Smith, Ian; Parker, Ian; Bootman, Martin D

2014-10-01

Fluorescence microscopy is a major tool with which to monitor cell physiology. Although the concepts of fluorescence and its optical separation using filters remain similar, microscope design varies with the aim of increasing image contrast and spatial resolution. The basics of wide-field microscopy are outlined to emphasize the selection, advantages, and correct use of laser scanning confocal microscopy, two-photon microscopy, scanning disk confocal microscopy, total internal reflection, and super-resolution microscopy. In addition, the principles of how these microscopes form images are reviewed to appreciate their capabilities, limitations, and constraints for operation. © 2014 Cold Spring Harbor Laboratory Press.

Fluorescence Microscopy

PubMed Central

Sanderson, Michael J.; Smith, Ian; Parker, Ian; Bootman, Martin D.

2016-01-01

Fluorescence microscopy is a major tool with which to monitor cell physiology. Although the concepts of fluorescence and its optical separation using filters remain similar, microscope design varies with the aim of increasing image contrast and spatial resolution. The basics of wide-field microscopy are outlined to emphasize the selection, advantages, and correct use of laser scanning confocal microscopy, two-photon microscopy, scanning disk confocal microscopy, total internal reflection, and super-resolution microscopy. In addition, the principles of how these microscopes form images are reviewed to appreciate their capabilities, limitations, and constraints for operation. PMID:25275114

Multiplexed immunofluorescence delineates proteomic cancer cell states associated with metabolism

PubMed Central

Sood, Anup; Miller, Alexandra M.; Brogi, Edi; Sui, Yunxia; Armenia, Joshua; McDonough, Elizabeth; Santamaria-Pang, Alberto; Stamper, Aleksandra; Campos, Carl; Pang, Zhengyu; Li, Qing; Port, Elisa; Graeber, Thomas G.; Schultz, Nikolaus; Ginty, Fiona; Larson, Steven M.

2016-01-01

The phenotypic diversity of cancer results from genetic and nongenetic factors. Most studies of cancer heterogeneity have focused on DNA alterations, as technologies for proteomic measurements in clinical specimen are currently less advanced. Here, we used a multiplexed immunofluorescence staining platform to measure the expression of 27 proteins at the single-cell level in formalin-fixed and paraffin-embedded samples from treatment-naive stage II/III human breast cancer. Unsupervised clustering of protein expression data from 638,577 tumor cells in 26 breast cancers identified 8 clusters of protein coexpression. In about one-third of breast cancers, over 95% of all neoplastic cells expressed a single protein coexpression cluster. The remaining tumors harbored tumor cells representing multiple protein coexpression clusters, either in a regional distribution or intermingled throughout the tumor. Tumor uptake of the radiotracer 18F-fluorodeoxyglucose was associated with protein expression clusters characterized by hormone receptor loss, PTEN alteration, and HER2 gene amplification. Our study demonstrates an approach to generate cellular heterogeneity metrics in routinely collected solid tumor specimens and integrate them with in vivo cancer phenotypes. PMID:27182557

Immunofluorescence assay method to detect dengue virus in Paniai-Papua

NASA Astrophysics Data System (ADS)

Sucipto, Teguh Hari; Ahwanah, Nur Laila Fitriati; Churrotin, Siti; Matake, Norifumi; Kotaki, Tomohiro; Soegijanto, Soegeng

2016-03-01

The dengue viruses (DENV), which include in the family Flaviviridae and the genus Flavivirus, was endemic in tropical areas and had been transmitted to humans by Aedes aegypti. An increasing number of immigrants from endemic areas to the non-endemic areas have emphasized the need for a simple and reliable test for the diagnosis of dengue virus infection. The purpose of this study was to detect the dengue virus by immunofluorescence assay (IFA) in the general population at Paniai-Papua. The results obtained from this study had showed a significantly better discrimination for DENV specific IgG antibodies. A total of 158 samples, 116 samples were IgG antibodies positive and 42 samples were negative. The conclusion of this study, Papua is not only a malaria endemic area, but also dengue virus infections were detected by IFA method. Therefore, the IFA can be used as an important diagnostic tool, which is a quick and an easy way to test samples from immigrants who come to the non-endemic areas.

Synergistic Interactions Between Direct and Indirect Bypasses in Combined Procedures: The Significance of Indirect Bypasses in Moyamoya Disease.

PubMed

Uchino, Haruto; Kim, Jae-Hoon; Fujima, Noriyuki; Kazumata, Ken; Ito, Masaki; Nakayama, Naoki; Kuroda, Satoshi; Houkin, Kiyohiro

2017-02-01

Whether additional indirect bypasses effectively contribute to revascularization in combined procedures remains unclear in patients with moyamoya disease. To evaluate the longitudinal changes associated with combined procedures while following up pediatric and adult patients long term and to assess whether any other clinical factors or hemodynamic parameters affected these changes to determine an optimal surgical strategy. We studied 58 hemispheres in 43 adults and 39 hemispheres in 26 children who underwent combined revascularization for moyamoya disease. To evaluate bypass development, we assessed the sizes of the superficial temporal artery and middle meningeal artery using magnetic resonance angiography. Multivariate analysis determined the effects of multiple variables on bypass development. Indirect bypass (middle meningeal artery) development occurred in 95% and 78% of the pediatric and adult hemispheres, respectively. Of these, dual development of direct and indirect bypasses occurred in 54% of the pediatric hemispheres and in 47% of the adult hemispheres. Reciprocal superficial temporal artery regression occurred in 28% of the hemispheres during the transition from the postoperative acute phase to the chronic phase during indirect bypass development. Good indirect bypass development was associated with adult hemispheres at Suzuki stage 4 or greater (odds ratio, 7.4; 95% confidence interval, 1.4-39.4; P = .02). Disease onset type and preoperative hemodynamic parameters were not considered predictors for the development of surgical revascularization. Simultaneous direct and indirect bypass development was most frequently observed, regardless of patient age and hemodynamic status. Applying indirect bypass as an adjunct to direct bypass could maximize revascularization in adults and children.

Indirect Effects of Environmental Change in Resource Competition Models.

PubMed

Kleinhesselink, Andrew R; Adler, Peter B

2015-12-01

Anthropogenic environmental change can affect species directly by altering physiological rates or indirectly by changing competitive outcomes. The unknown strength of competition-mediated indirect effects makes it difficult to predict species abundances in the face of ongoing environmental change. Theory developed with phenomenological competition models shows that indirect effects are weak when coexistence is strongly stabilized, but these models lack a mechanistic link between environmental change and species performance. To extend existing theory, we examined the relationship between coexistence and indirect effects in mechanistic resource competition models. We defined environmental change as a change in resource supply points and quantified the resulting competition-mediated indirect effects on species abundances. We found that the magnitude of indirect effects increases in proportion to niche overlap. However, indirect effects also depend on differences in how competitors respond to the change in resource supply, an insight hidden in nonmechanistic models. Our analysis demonstrates the value of using niche overlap to predict the strength of indirect effects and clarifies the types of indirect effects that global change can have on competing species.

Ballistic fractures: indirect fracture to bone.

PubMed

Dougherty, Paul J; Sherman, Don; Dau, Nathan; Bir, Cynthia

2011-11-01

Two mechanisms of injury, the temporary cavity and the sonic wave, have been proposed to produce indirect fractures as a projectile passes nearby in tissue. The purpose of this study is to evaluate the temporal relationship of pressure waves using strain gauge technology and high-speed video to elucidate whether the sonic wave, the temporary cavity, or both are responsible for the formation of indirect fractures. Twenty-eight fresh frozen cadaveric diaphyseal tibia (2) and femurs (26) were implanted into ordnance gelatin blocks. Shots were fired using 9- and 5.56-mm bullets traversing through the gelatin only, passing close to the edge of the bone, but not touching, to produce an indirect fracture. High-speed video of the impact event was collected at 20,000 frames/s. Acquisition of the strain data were synchronized with the video at 20,000 Hz. The exact time of fracture was determined by analyzing and comparing the strain gauge output and video. Twenty-eight shots were fired, 2 with 9-mm bullets and 26 with 5.56-mm bullets. Eight indirect fractures that occurred were of a simple (oblique or wedge) pattern. Comparison of the average distance of the projectile from the bone was 9.68 mm (range, 3-20 mm) for fractured specimens and 15.15 mm (range, 7-28 mm) for nonfractured specimens (Student's t test, p = 0.036). In this study, indirect fractures were produced after passage of the projectile. Thus, the temporary cavity, not the sonic wave, was responsible for the indirect fractures.

Tip/tilt-compensated through-focus scanning optical microscopy

NASA Astrophysics Data System (ADS)

Lee, Jun Ho; Park, Jun Hyung; Jeong, Dohwan; Shin, Eun Ji; Park, Chris

2016-11-01

Through-Focus Optical Microscopy (TSOM), with nanometer scale lateral and vertical sensitivity matching those of scanning electron microscopy, has been demonstrated to be utilized for 3D inspection and metrology. There have been sensitivity and instability issues in acquiring through-focus images because TSOM 3D information is indirectly extracted by differentiating a target TSOM image from reference TSOM images. This paper first reports on the optical axis instability that occurs during the scanning process of TSOM when implemented in an existing patterned wafer inspection tool by moving the wafer plane; this is followed by quantitative confirmation of the optical/mechanical instability using a new TSOM tool on an optical bench with a Shack-Hartmann wavefront sensor and a tip/tilt sensor. Then, this paper proposes two tip/tilt compensated TSOM optical acquisition methods that can be applied with adaptive optics. The first method simply adopts a tip/tilt mirror with a quad cell in a simple closed loop, while the second method adopts a highorder deformable mirror with a Shack-Hartmann sensor. The second method is able to correct high-order residual aberrations as well as to perform through-focus scanning without z-axis movement, while the first method is easier to implement in pre-existing wafer inspection systems with only minor modification.

19 CFR 10.603 - Indirect materials.

Code of Federal Regulations, 2010 CFR

2010-04-01

... Duties U.S. CUSTOMS AND BORDER PROTECTION, DEPARTMENT OF HOMELAND SECURITY; DEPARTMENT OF THE TREASURY ARTICLES CONDITIONALLY FREE, SUBJECT TO A REDUCED RATE, ETC. Dominican Republic-Central America-United States Free Trade Agreement Rules of Origin § 10.603 Indirect materials. An indirect material, as defined...

DESENSITIZING BIOACTIVE AGENTS IMPROVES BOND STRENGTH OF INDIRECT RESIN-CEMENTED RESTORATIONS: PRELIMINARY RESULTS

PubMed Central

Pires-De-Souza, Fernanda de Carvalho Panzeri; de Marco, Fabíola Fiorezi; Casemiro, Luciana Assirati; Panzeri, Heitor

2007-01-01

Objective: The aim of this study was to assess the bond strength of indirect composite restorations cemented with a resin-based cement associated with etch-and-rinse and self-etching primer adhesive systems to dentin treated or not with a bioactive material. Materials and Method: Twenty bovine incisor crowns had the buccal enamel removed and the dentin ground flat. The teeth were assigned to 4 groups (n=5): Group I: acid etching + Prime & Bond NT (Dentsply); Group II: application of a bioactive glass (Biosilicato®)+ acid etching + Prime & Bond NT; Group III: One-up Bond F (J Morita); Group IV: Biosilicato® + One-up Bond F. Indirect composite resin (Artglass, Kulzer) cylinders (6x10mm) were fabricated and cemented to the teeth with a dualcure resin-based cement (Enforce, Dentsply). After cementation, the specimens were stored in artificial saliva at 37oC for 30 days and thereafter tested in tensile strength in a universal testing machine (EMIC) with 50 kgf load cell at a crosshead speed of 1 mm/min. Failure modes were assessed under scanning electron microscopy. Data were analyzed statistically by ANOVA and Tukey's test (95% level of confidence). Results: Groups I, II and III had statistically similar results (p>0.05). Group IV had statistically significant higher bond strength means (p<0.05) than the other groups. The analysis of the debonded surfaces showed a predominance of adhesive failure mode for Group III and mixed failure mode for the other groups. Conclusion: The use of desensitizing agent did not affect negatively the bonding of the indirect composite restorations to dentin, independently of the tested adhesive systems. PMID:19089114

48 CFR 942.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 5 2010-10-01 2010-10-01 false Final indirect cost rates. 942.705 Section 942.705 Federal Acquisition Regulations System DEPARTMENT OF ENERGY CONTRACT MANAGEMENT CONTRACT ADMINISTRATION AND AUDIT SERVICES Indirect Cost Rates 942.705 Final indirect cost rates. ...

48 CFR 3442.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 7 2010-10-01 2010-10-01 false Final indirect cost rates. 3442.705 Section 3442.705 Federal Acquisition Regulations System DEPARTMENT OF EDUCATION ACQUISITION REGULATION CONTRACT MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 3442.705 Final indirect cost rates...

27 CFR 6.26 - Indirect interest.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2013-04-01 2013-04-01 false Indirect interest. 6.26... OF THE TREASURY ALCOHOL âTIED-HOUSEâ Unlawful Inducements Interest in Retail License § 6.26 Indirect interest. Industry member interest in retail licenses includes any interest acquired by corporate officials...

27 CFR 6.26 - Indirect interest.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2012-04-01 2012-04-01 false Indirect interest. 6.26... OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail License § 6.26 Indirect interest. Industry member interest in retail licenses includes any interest acquired by corporate officials...

27 CFR 6.32 - Indirect interest.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2012-04-01 2012-04-01 false Indirect interest. 6.32... OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail Property § 6.32 Indirect interest. Industry member interest in retail property includes any interest acquired by corporate officials...

27 CFR 6.32 - Indirect interest.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2013-04-01 2013-04-01 false Indirect interest. 6.32... OF THE TREASURY ALCOHOL âTIED-HOUSEâ Unlawful Inducements Interest in Retail Property § 6.32 Indirect interest. Industry member interest in retail property includes any interest acquired by corporate officials...

48 CFR 842.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 5 2010-10-01 2010-10-01 false Final indirect cost rates. 842.705 Section 842.705 Federal Acquisition Regulations System DEPARTMENT OF VETERANS AFFAIRS CONTRACT MANAGEMENT CONTRACT ADMINISTRATION AND AUDIT SERVICES Indirect Cost Rates 842.705 Final indirect cost rates...

Development of an indirect enzyme-linked immunosorbent assay test for detecting antibodies to chicken astrovirus in chicken sera.

PubMed

Skibinska, A; Lee, A; Wylie, M; Smyth, V J; Welsh, M D; Todd, D

2015-01-01

The development of an indirect enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of Group B chicken astrovirus (CAstV) infections is described. The test was based on the use of an affinity-purified capsid antigen, specific to CAstV isolate 11672, produced as a glutathione-S-transferase N-terminal fusion protein by a recombinant baculovirus. Strongly positive ELISA signals were elicited against experimentally produced antisera raised to CAstVs from Group B (subgroups i and ii) but were negative for antisera raised to a Group A CAstV. Using a panel of 240 selected serum samples, 99% agreement was observed when the results obtained by ELISA were compared to those from an indirect immunofluorescence test for CAstV 11672. The ELISA test was applied to 68 serum sets comprising 1864 samples, which were obtained from parent and grandparent flocks originating mainly in the UK. Of the 52 sets containing ELISA-positive samples, 24 sets had >75% samples positive and nine sets had <25% samples positive and were regarded as having high and low seropositivities, respectively. Of the 1864 serum samples tested 1090 (58.5%) were ELISA positive and of these, 234 sera (21.5%) produced strongly positive signals, whereas moderately positive and weakly positive signals were produced by 562 (51.5%) and 294 (27%) sera. When used for flock screening purposes, this ELISA test can be used to (i) investigate the occurrence of first-time CAstV infections of parent flocks during lay and the possible adverse effects caused by vertically transmitted CAstV infections on broiler hatchability and performance and (ii) diagnose Group B CAstV infections within specific pathogen free flocks.

Forecasting of indirect consumables for a Job Shop

NASA Astrophysics Data System (ADS)

Shakeel, M.; Khan, S.; Khan, W. A.

2016-08-01

A job shop has an arrangement where similar machines (Direct consumables) are grouped together and use indirect consumables to produce a product. The indirect consumables include hack saw blades, emery paper, painting brush etc. The job shop is serving various orders at a particular time for the optimal operation of job shop. Forecasting is required to predict the demand of direct and indirect consumables in a job shop. Forecasting is also needed to manage lead time, optimize inventory cost and stock outs. The objective of this research is to obtain the forecast for indirect consumables. The paper shows how job shop can manage their indirect consumables more accurately by establishing a new technique of forecasting. This results in profitable use of job shop by multiple users.

Indirect Comparisons: A Review of Reporting and Methodological Quality

PubMed Central

Donegan, Sarah; Williamson, Paula; Gamble, Carrol; Tudur-Smith, Catrin

2010-01-01

Background The indirect comparison of two interventions can be valuable in many situations. However, the quality of an indirect comparison will depend on several factors including the chosen methodology and validity of underlying assumptions. Published indirect comparisons are increasingly more common in the medical literature, but as yet, there are no published recommendations of how they should be reported. Our aim is to systematically review the quality of published indirect comparisons to add to existing empirical data suggesting that improvements can be made when reporting and applying indirect comparisons. Methodology/Findings Reviews applying statistical methods to indirectly compare the clinical effectiveness of two interventions using randomised controlled trials were eligible. We searched (1966–2008) Database of Abstracts and Reviews of Effects, The Cochrane library, and Medline. Full review publications were assessed for eligibility. Specific criteria to assess quality were developed and applied. Forty-three reviews were included. Adequate methodology was used to calculate the indirect comparison in 41 reviews. Nineteen reviews assessed the similarity assumption using sensitivity analysis, subgroup analysis, or meta-regression. Eleven reviews compared trial-level characteristics. Twenty-four reviews assessed statistical homogeneity. Twelve reviews investigated causes of heterogeneity. Seventeen reviews included direct and indirect evidence for the same comparison; six reviews assessed consistency. One review combined both evidence types. Twenty-five reviews urged caution in interpretation of results, and 24 reviews indicated when results were from indirect evidence by stating this term with the result. Conclusions This review shows that the underlying assumptions are not routinely explored or reported when undertaking indirect comparisons. We recommend, therefore, that the quality of indirect comparisons should be improved, in particular, by assessing

Super-resolution microscopy as a potential approach to diagnosis of platelet granule disorders.

PubMed

Westmoreland, D; Shaw, M; Grimes, W; Metcalf, D J; Burden, J J; Gomez, K; Knight, A E; Cutler, D F

2016-04-01

Many platelet functions are dependent on bioactive molecules released from their granules. Deficiencies of these granules in number, shape or content are associated with bleeding. The small size of these granules is such that imaging them for diagnosis has traditionally required electron microscopy. However, recently developed super-resolution microscopes provide sufficient spatial resolution to effectively image platelet granules. When combined with automated image analysis, these methods provide a quantitative, unbiased, rapidly acquired dataset that can readily and reliably reveal differences in platelet granules between individuals. To demonstrate the ability of structured illumination microscopy (SIM) to efficiently differentiate between healthy volunteers and three patients with Hermansky-Pudlak syndrome. Blood samples were taken from three patients with Hermansky-Pudlak syndrome and seven controls. Patients 1-3 have gene defects in HPS1, HPS6 and HPS5, respectively; all controls were healthy volunteers. Platelet-rich plasma was isolated from blood and the platelets fixed, stained for CD63 and processed for analysis by immunofluorescence microscopy, using a custom-built SIM microscope. SIM can successfully resolve CD63-positive structures in fixed platelets. A determination of the number of CD63-positive structures per platelet allowed us to conclude that each patient was significantly different from all of the controls with 99% confidence. A super-resolution imaging approach is effective and rapid in objectively differentiating between patients with a platelet bleeding disorder and healthy volunteers. CD63 is a useful marker for predicting Hermansky-Pudlak syndrome and could be used in the diagnosis of patients suspected of other platelet granule disorders. © 2016 The Authors. Journal of Thrombosis and Haemostasis published by Wiley Periodicals, Inc. on behalf of International Society on Thrombosis and Haemostasis.

14 CFR 296.3 - Indirect cargo air carrier.

Code of Federal Regulations, 2010 CFR

2010-01-01

... PROCEEDINGS) ECONOMIC REGULATIONS INDIRECT AIR TRANSPORTATION OF PROPERTY General § 296.3 Indirect cargo air... transportation of property, and uses for the whole or any part of such transportation the services of an air... 14 Aeronautics and Space 4 2010-01-01 2010-01-01 false Indirect cargo air carrier. 296.3 Section...

A novel histochemical method of simultaneous detection by a single- or double-immunofluorescence and Bielschowsky's silver staining in teased rat sciatic nerves.

PubMed

Segura-Anaya, Edith; Flores-Miranda, Rommel; Martínez-Gómez, Alejandro; Dent, Myrna A R

2018-07-01

The Golgi silver method has been widely used in neuroscience for the study of normal and pathological morphology of neurons. The method has been steadily improved and Bielschowsky's silver staining method (BSSM) is widely used in various pathological conditions, like Alzheimer's disease. In this work, teased sciatic nerves were silver impregnated using BSSM. We also developed simultaneous staining by silver impregnation and single- or double-immunofluorescence of the same section in teased nerve preparations. We immunostained against non-myelinating Schwann cells and different myelinating Schwann cell domains. BSSM teased nerves show a strong staining of axons (black) and a gold-brown staining of myelinating and non-myelinating Schwann cells. We were also able to stain by immunofluorescence these BSSM teased nerves with specific molecular markers against non-myelinating Schwann cells, also against non-compact myelin such as the Schmidt-Lanterman incisures or paranodal regions and compact myelin, but not axons. In peripheral nerves, several silver impregnation methods have been used to stain nerves in paraffin sections, but not in teased nerves to enable the assessment of isolated nerve fibers. In conclusion, BSSM gives accurate information of nerve morphology and combining the procedure with immunofluorescence it would be very useful to study the molecular nerve domain organization of the nerve fibers, and to study the molecular pathology of axon degeneration, or myelin disorders, or of any peripheral neuropathy, also to study demyelination diseases in the central nervous system. Copyright © 2018. Published by Elsevier B.V.

Dictionary of Microscopy

NASA Astrophysics Data System (ADS)

Heath, Julian

2005-10-01

The past decade has seen huge advances in the application of microscopy in all areas of science. This welcome development in microscopy has been paralleled by an expansion of the vocabulary of technical terms used in microscopy: terms have been coined for new instruments and techniques and, as microscopes reach even higher resolution, the use of terms that relate to the optical and physical principles underpinning microscopy is now commonplace. The Dictionary of Microscopy was compiled to meet this challenge and provides concise definitions of over 2,500 terms used in the fields of light microscopy, electron microscopy, scanning probe microscopy, x-ray microscopy and related techniques. Written by Dr Julian P. Heath, Editor of Microscopy and Analysis, the dictionary is intended to provide easy navigation through the microscopy terminology and to be a first point of reference for definitions of new and established terms. The Dictionary of Microscopy is an essential, accessible resource for: students who are new to the field and are learning about microscopes equipment purchasers who want an explanation of the terms used in manufacturers' literature scientists who are considering using a new microscopical technique experienced microscopists as an aide mémoire or quick source of reference librarians, the press and marketing personnel who require definitions for technical reports.

Direct and indirect punishment among strangers in the field

PubMed Central

Balafoutas, Loukas; Nikiforakis, Nikos; Rockenbach, Bettina

2014-01-01

Many interactions in modern human societies are among strangers. Explaining cooperation in such interactions is challenging. The two most prominent explanations critically depend on individuals’ willingness to punish defectors: In models of direct punishment, individuals punish antisocial behavior at a personal cost, whereas in models of indirect reciprocity, they punish indirectly by withholding rewards. We investigate these competing explanations in a field experiment with real-life interactions among strangers. We find clear evidence of both direct and indirect punishment. Direct punishment is not rewarded by strangers and, in line with models of indirect reciprocity, is crowded out by indirect punishment opportunities. The existence of direct and indirect punishment in daily life indicates the importance of both means for understanding the evolution of cooperation. PMID:25349390

Direct and indirect punishment among strangers in the field.

PubMed

Balafoutas, Loukas; Nikiforakis, Nikos; Rockenbach, Bettina

2014-11-11

Many interactions in modern human societies are among strangers. Explaining cooperation in such interactions is challenging. The two most prominent explanations critically depend on individuals' willingness to punish defectors: In models of direct punishment, individuals punish antisocial behavior at a personal cost, whereas in models of indirect reciprocity, they punish indirectly by withholding rewards. We investigate these competing explanations in a field experiment with real-life interactions among strangers. We find clear evidence of both direct and indirect punishment. Direct punishment is not rewarded by strangers and, in line with models of indirect reciprocity, is crowded out by indirect punishment opportunities. The existence of direct and indirect punishment in daily life indicates the importance of both means for understanding the evolution of cooperation.

Histopathologic and direct immunofluorescence findings of the papulopustular lesions in Behçet's disease.

PubMed

Ilknur, Turna; Pabuççuoglu, Uğur; Akin, Ciler; Lebe, Banu; Gunes, Ali Tahsin

2006-01-01

Although papulopustular lesions are common in patients with Behçet's disease (BD), clinically they may not be differentiated from other diseases with papulopustular presentation such as acne vulgaris or folliculitis. Therefore, there is disagreement as to whether they should be used as a diagnostic criterion in BD. The aim of this study was to determine whether the histopathologic evaluation of the papulopustular lesions may assist in the diagnosis of BD. Eighteen patients with BD and 16 control patients consisting of eleven patients with bacterial folliculitis and five patients with acne vulgaris were included in the study. After the detailed histopathologic evaluation by two pathologists who were blinded to the clinical diagnoses, the histopathologic findings were classified into three patterns as follows; pattern I: vasculitis (lymphocytic or leucocytoclastic); pattern II: folliculitis and/or perifolliculitis; pattern III: superficial and/or deep perivascular, and/or interstitial dermatitis. In addition, direct immunofluorescence studies were performed in order to evaluate the deposition of IgM, IgG, IgA, C3, or fibrinogen in dermal blood vessels. 27.8% of the patients with BD (5 patients) revealed lymphocytic vasculitis, while none of the control group did; and the difference was found statistically significant (P=0.046). The rate of pattern II which included folliculitis and/or perifolliculitis was 50.0% in control patients and 16.7% in the patients with BD; and the difference was found statistically significant (P=0.038). No difference was found between the two groups with regard to pattern III or direct immunofluorescence findings (P>0.05). Our results indicate that only vasculitic changes can be useful when histopathological features of papulopustular lesions are to be employed as a diagnostic criterion in patients with suspected BD.

27 CFR 6.26 - Indirect interest.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Indirect interest. 6.26 Section 6.26 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail License § 6.26 Indirect interest. Industry member interest in retail...

27 CFR 6.32 - Indirect interest.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Indirect interest. 6.32 Section 6.32 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail Property § 6.32 Indirect interest. Industry member interest in retail...

27 CFR 6.32 - Indirect interest.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Indirect interest. 6.32 Section 6.32 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail Property § 6.32 Indirect interest. Industry member interest in retail...

27 CFR 6.26 - Indirect interest.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Indirect interest. 6.26 Section 6.26 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Interest in Retail License § 6.26 Indirect interest. Industry member interest in retail...

Direct and indirect measurement of the magnetocaloric effect in bulk and nanostructured Ni-Mn-In Heusler alloy

NASA Astrophysics Data System (ADS)

Ghahremani, Mohammadreza; Aslani, Amir; Hosseinnia, Marjan; Bennett, Lawrence H.; Della Torre, Edward

2018-05-01

A systematic study of the magnetocaloric effect of a Ni51Mn33.4In15.6 Heusler alloy converted to nanoparticles via high energy ball-milling technique in the temperature range of 270 to 310 K has been performed. The properties of the particles were characterized by x-ray diffraction, electron microscopy, and magnetometer techniques. Isothermal magnetic field variation of magnetization exhibits field hysteresis in bulk Ni51Mn33.4In15.6 alloy across the martensitic transition which significantly lessened in the nanoparticles. The magnetocaloric effects of the bulk and nanoparticle samples were measured both with direct method, through our state of the art direct test bed apparatus with controllability over the applied fields and temperatures, as well as an indirect method through Maxwell and thermodynamic equations. In direct measurements, nanoparticle sample's critical temperature decreased by 6 K, but its magnetocaloric effect enhanced by 17% over the bulk counterpart. Additionally, when comparing the direct and indirect magnetocaloric curves, the direct method showed 14% less adiabatic temperature change in the bulk and 5% less adiabatic temperature change in the nanostructured sample.

Cross-reactive antibodies in convalescent SARS patients' sera against the emerging novel human coronavirus EMC (2012) by both immunofluorescent and neutralizing antibody tests.

PubMed

Chan, Kwok-Hung; Chan, Jasper Fuk-Woo; Tse, Herman; Chen, Honglin; Lau, Candy Choi-Yi; Cai, Jian-Piao; Tsang, Alan Ka-Lun; Xiao, Xincai; To, Kelvin Kai-Wang; Lau, Susanna Kar-Pui; Woo, Patrick Chiu-Yat; Zheng, Bo-Jiang; Wang, Ming; Yuen, Kwok-Yung

2013-08-01

A severe acute respiratory syndrome (SARS)-like disease due to a novel betacoronavirus, human coronavirus EMC (HCoV-EMC), has emerged recently. HCoV-EMC is phylogenetically closely related to Tylonycteris-bat-coronavirus-HKU4 and Pipistrellus-bat-coronavirus-HKU5 in Hong Kong. We conducted a seroprevalence study on archived sera from 94 game-food animal handlers at a wild life market, 28 SARS patients, and 152 healthy blood donors in Southern China to assess the zoonotic potential and evidence for intrusion of HCoV-EMC and related viruses into humans. Anti-HCoV-EMC and anti-SARS-CoV antibodies were detected using screening indirect immunofluorescence (IF) and confirmatory neutralizing antibody tests. Two (2.1%) animal handlers had IF antibody titer of ≥ 1:20 against both HCoV-EMC and SARS-CoV with neutralizing antibody titer of <1:10. No blood donor had antibody against either virus. Surprisingly, 17/28 (60.7%) of SARS patients had significant IF antibody titers with 7/28 (25%) having anti-HCoV-EMC neutralizing antibodies at low titers which significantly correlated with that of HCoV-OC43. Bioinformatics analysis demonstrated a significant B-cell epitope overlapping the heptad repeat-2 region of Spike protein. Virulence of SARS-CoV over other betacoronaviruses may boost cross-reactive neutralizing antibodies against other betacoronaviruses. Convalescent SARS sera may contain cross-reactive antibodies against other betacoronaviruses and confound seroprevalence study for HCoV-EMC. Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Indirect combustion noise of auxiliary power units

NASA Astrophysics Data System (ADS)

Tam, Christopher K. W.; Parrish, Sarah A.; Xu, Jun; Schuster, Bill

2013-08-01

Recent advances in noise suppression technology have significantly reduced jet and fan noise from commercial jet engines. This leads many investigators in the aeroacoustics community to suggest that core noise could well be the next aircraft noise barrier. Core noise consists of turbine noise and combustion noise. There is direct combustion noise generated by the combustion processes, and there is indirect combustion noise generated by the passage of combustion hot spots, or entropy waves, through constrictions in an engine. The present work focuses on indirect combustion noise. Indirect combustion noise has now been found in laboratory experiments. The primary objective of this work is to investigate whether indirect combustion noise is also generated in jet and other engines. In a jet engine, there are numerous noise sources. This makes the identification of indirect combustion noise a formidable task. Here, our effort concentrates exclusively on auxiliary power units (APUs). This choice is motivated by the fact that APUs are relatively simple engines with only a few noise sources. It is, therefore, expected that the chance of success is higher. Accordingly, a theoretical model study of the generation of indirect combustion noise in an Auxiliary Power Unit (APU) is carried out. The cross-sectional areas of an APU from the combustor to the turbine exit are scaled off to form an equivalent nozzle. A principal function of a turbine in an APU is to extract mechanical energy from the flow stream through the exertion of a resistive force. Therefore, the turbine is modeled by adding a negative body force to the momentum equation. This model is used to predict the ranges of frequencies over which there is a high probability for indirect combustion noise generation. Experimental spectra of internal pressure fluctuations and far-field noise of an RE220 APU are examined to identify anomalous peaks. These peaks are possible indirection combustion noise. In the case of the

Cell differentiation in cardiac myxomas: confocal microscopy and gene expression analysis after laser capture microdissection.

PubMed

Pucci, Angela; Mattioli, Claudia; Matteucci, Marco; Lorenzini, Daniele; Panvini, Francesca; Pacini, Simone; Ippolito, Chiara; Celiento, Michele; De Martino, Andrea; Dolfi, Amelio; Belgio, Beatrice; Bortolotti, Uberto; Basolo, Fulvio; Bartoloni, Giovanni

2018-05-22

Cardiac myxomas are rare tumors with a heterogeneous cell population including properly neoplastic (lepidic), endothelial and smooth muscle cells. The assessment of neoplastic (lepidic) cell differentiation pattern is rather difficult using conventional light microscopy immunohistochemistry and/or whole tissue extracts for mRNA analyses. In a preliminary study, we investigated 20 formalin-fixed and paraffin-embedded cardiac myxomas by means of conventional immunohistochemistry; in 10/20 cases, cell differentiation was also analyzed by real-time RT-PCR after laser capture microdissection of the neoplastic cells, whereas calretinin and endothelial antigen CD31 immunoreactivity was localized in 4/10 cases by double immunofluorescence confocal microscopy. Gene expression analyses of α-smooth muscle actin, endothelial CD31 antigen, alpha-cardiac actin, matrix metalloprotease-2 (MMP2) and tissue inhibitor of matrix metalloprotease-1 (TIMP1) was performed on cDNA obtained from either microdissected neoplastic cells or whole tumor sections. We found very little or absent CD31 and α-Smooth Muscle Actin expression in the microdissected cells as compared to the whole tumors, whereas TIMP1 and MMP2 genes were highly expressed in both ones, greater levels being found in patients with embolic phenomena. α-Cardiac Actin was not detected. Confocal microscopy disclosed two different signals corresponding to calretinin-positive myxoma cells and to endothelial CD31-positive cells, respectively. In conclusion, the neoplastic (lepidic) cells showed a distinct gene expression pattern and no consistent overlapping with endothelial and smooth muscle cells or cardiac myocytes; the expression of TIMP1 and MMP2 might be related to clinical presentation; larger series studies using also systematic transcriptome analysis might be useful to confirm the present results.

An erythrodermic variant of pemphigus foliaceus with puzzling histologic and immunopathologic features.

PubMed

Peterson, Jennifer D; Worobec, Sophie M; Chan, Lawrence S

2007-01-01

Pemphigus foliaceus is an autoimmune blistering disorder that affects the skin owing to autoantibodies against desmoglein 1. We employed clinical, histologic, immunopathologic, and serum laboratory studies to investigate a case of an erythrodermic variant of pemphigus foliaceus in an elderly man following treatment with bisoprolol-hydrochlorothiazide. Early histopathology revealed psoriasiform dermatitis, but later biopsies showed subcorneal and granular layer separation with neutrophilic infiltrate. Direct immunofluorescence showed intercellular deposits of immunoglobulin G throughout the epidermis, granular staining of C3 along the basement membrane zone, and fibrin and C3 deposition around the blood vessels. Indirect immunofluorescence on monkey esophagus showed a titer of greater than 1:1,280. Indirect immunofluorescence on rat bladder, antinuclear antibody, lupus panel, and kidney function panel were all negative. There are no reports in the literature of pemphigus foliaceus being induced by bisoprolol, but reports exist of propanolol resulting in drug-induced pemphigus foliaceus.

Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets

PubMed Central

Hiromitsu, Shirasawa; Jin, Kumagai; Emiko, Sato; Katsuya, Kabashima; Yukiyo, Kumazawa; Wataru, Sato; Hiroshi, Miura; Ryuta, Nakamura; Hiroshi, Nanjo; Yoshihiro, Minamiya; Yoichi, Akagami; Yukihiro, Terada

2015-01-01

Recently, a new technique was developed for non-catalytically mixing microdroplets. In this method, an alternating-current (AC) electric field is used to promote the antigen–antibody reaction within the microdroplet. Previously, this technique has only been applied to histological examinations of flat structures, such as surgical specimens. In this study, we applied this technique for the first time to immunofluorescence staining of three-dimensional structures, specifically, mammalian eggs. We diluted an antibody against microtubules from 1:1,000 to 1:16,000, and compared the chromatic degree and extent of fading across dilutions. In addition, we varied the frequency of AC electric-field mixing from 5 Hz to 46 Hz and evaluated the effect on microtubule staining. Microtubules were more strongly stained after AC electric-field mixing for only 5 minutes, even when the concentration of primary antibody was 10 times lower than in conventional methods. AC electric-field mixing also alleviated microtubule fading. At all frequencies tested, AC electric-field mixing resulted in stronger microtubule staining than in controls. There was no clear difference in a microtubule staining between frequencies. These results suggest that the novel method could reduce antibody consumption and shorten immunofluorescence staining time. PMID:26477850

Comparison of haemoglobin estimates using direct & indirect cyanmethaemoglobin methods.

PubMed

Bansal, Priyanka Gupta; Toteja, Gurudayal Singh; Bhatia, Neena; Gupta, Sanjeev; Kaur, Manpreet; Adhikari, Tulsi; Garg, Ashok Kumar

2016-10-01

Estimation of haemoglobin is the most widely used method to assess anaemia. Although direct cyanmethaemoglobin method is the recommended method for estimation of haemoglobin, but it may not be feasible under field conditions. Hence, the present study was undertaken to compare indirect cyanmethaemoglobin method against the conventional direct method for haemoglobin estimation. Haemoglobin levels were estimated for 888 adolescent girls aged 11-18 yr residing in an urban slum in Delhi by both direct and indirect cyanmethaemoglobin methods, and the results were compared. The mean haemoglobin levels for 888 whole blood samples estimated by direct and indirect cyanmethaemoglobin method were 116.1 ± 12.7 and 110.5 ± 12.5 g/l, respectively, with a mean difference of 5.67 g/l (95% confidence interval: 5.45 to 5.90, P<0.001); which is equivalent to 0.567 g%. The prevalence of anaemia was reported as 59.6 and 78.2 per cent by direct and indirect methods, respectively. Sensitivity and specificity of indirect cyanmethaemoglobin method were 99.2 and 56.4 per cent, respectively. Using regression analysis, prediction equation was developed for indirect haemoglobin values. The present findings revealed that indirect cyanmethaemoglobin method overestimated the prevalence of anaemia as compared to the direct method. However, if a correction factor is applied, indirect method could be successfully used for estimating true haemoglobin level. More studies should be undertaken to establish agreement and correction factor between direct and indirect cyanmethaemoglobin methods.

Segmentation of fluorescence microscopy images for quantitative analysis of cell nuclear architecture.

PubMed

Russell, Richard A; Adams, Niall M; Stephens, David A; Batty, Elizabeth; Jensen, Kirsten; Freemont, Paul S

2009-04-22

Considerable advances in microscopy, biophysics, and cell biology have provided a wealth of imaging data describing the functional organization of the cell nucleus. Until recently, cell nuclear architecture has largely been assessed by subjective visual inspection of fluorescently labeled components imaged by the optical microscope. This approach is inadequate to fully quantify spatial associations, especially when the patterns are indistinct, irregular, or highly punctate. Accurate image processing techniques as well as statistical and computational tools are thus necessary to interpret this data if meaningful spatial-function relationships are to be established. Here, we have developed a thresholding algorithm, stable count thresholding (SCT), to segment nuclear compartments in confocal laser scanning microscopy image stacks to facilitate objective and quantitative analysis of the three-dimensional organization of these objects using formal statistical methods. We validate the efficacy and performance of the SCT algorithm using real images of immunofluorescently stained nuclear compartments and fluorescent beads as well as simulated images. In all three cases, the SCT algorithm delivers a segmentation that is far better than standard thresholding methods, and more importantly, is comparable to manual thresholding results. By applying the SCT algorithm and statistical analysis, we quantify the spatial configuration of promyelocytic leukemia nuclear bodies with respect to irregular-shaped SC35 domains. We show that the compartments are closer than expected under a null model for their spatial point distribution, and furthermore that their spatial association varies according to cell state. The methods reported are general and can readily be applied to quantify the spatial interactions of other nuclear compartments.

Segmentation of Fluorescence Microscopy Images for Quantitative Analysis of Cell Nuclear Architecture

PubMed Central

Russell, Richard A.; Adams, Niall M.; Stephens, David A.; Batty, Elizabeth; Jensen, Kirsten; Freemont, Paul S.

2009-01-01

Abstract Considerable advances in microscopy, biophysics, and cell biology have provided a wealth of imaging data describing the functional organization of the cell nucleus. Until recently, cell nuclear architecture has largely been assessed by subjective visual inspection of fluorescently labeled components imaged by the optical microscope. This approach is inadequate to fully quantify spatial associations, especially when the patterns are indistinct, irregular, or highly punctate. Accurate image processing techniques as well as statistical and computational tools are thus necessary to interpret this data if meaningful spatial-function relationships are to be established. Here, we have developed a thresholding algorithm, stable count thresholding (SCT), to segment nuclear compartments in confocal laser scanning microscopy image stacks to facilitate objective and quantitative analysis of the three-dimensional organization of these objects using formal statistical methods. We validate the efficacy and performance of the SCT algorithm using real images of immunofluorescently stained nuclear compartments and fluorescent beads as well as simulated images. In all three cases, the SCT algorithm delivers a segmentation that is far better than standard thresholding methods, and more importantly, is comparable to manual thresholding results. By applying the SCT algorithm and statistical analysis, we quantify the spatial configuration of promyelocytic leukemia nuclear bodies with respect to irregular-shaped SC35 domains. We show that the compartments are closer than expected under a null model for their spatial point distribution, and furthermore that their spatial association varies according to cell state. The methods reported are general and can readily be applied to quantify the spatial interactions of other nuclear compartments. PMID:19383481

Indirectness in Discourse: Ethnicity as Conversational Style.

ERIC Educational Resources Information Center

Tannen, Deborah

1981-01-01

Examines social differences in expectations of indirectness in conversations between married couples, both Greek and Greek-American. Concludes that Greeks are more likely to expect indirectness in the context presented and that Greek-Americans have retained the Greek communicative patterns. (FL)

10 CFR 605.16 - Indirect cost limitations.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 4 2010-01-01 2010-01-01 false Indirect cost limitations. 605.16 Section 605.16 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS THE OFFICE OF ENERGY RESEARCH FINANCIAL ASSISTANCE.../technical meetings will not include payment for indirect costs. ...

Smoking Cessation Is Associated With Lower Indirect Costs.

PubMed

Baker, Christine L; Bruno, Marianna; Emir, Birol; Li, Vicky W; Goren, Amir

2018-06-01

This study quantified differences in indirect costs due to decreased work productivity between current and former smokers. Former smokers were further categorized by number of years since quitting to assess corresponding differences. Data on employed individuals were obtained from the 2013 US National Health and Wellness Survey (NHWS; N = 75,000). Indirect costs were calculated for current smokers and former smokers from weekly wages based on age and sex. The annual total indirect costs for current smokers were $1327.53, $1560.18, and $1839.87 higher than for those who quit 0 to 4 years, 5 to 10 years, and more than or equal to 11 years prior, respectively. There were no significant differences in mean total indirect costs between the former smoker groups. Current smokers showed significantly higher total annual indirect costs compared with former smokers, independently of the number of years since quitting smoking.

Robust tumor morphometry in multispectral fluorescence microscopy

NASA Astrophysics Data System (ADS)

Tabesh, Ali; Vengrenyuk, Yevgen; Teverovskiy, Mikhail; Khan, Faisal M.; Sapir, Marina; Powell, Douglas; Mesa-Tejada, Ricardo; Donovan, Michael J.; Fernandez, Gerardo

2009-02-01

Morphological and architectural characteristics of primary tissue compartments, such as epithelial nuclei (EN) and cytoplasm, provide important cues for cancer diagnosis, prognosis, and therapeutic response prediction. We propose two feature sets for the robust quantification of these characteristics in multiplex immunofluorescence (IF) microscopy images of prostate biopsy specimens. To enable feature extraction, EN and cytoplasm regions were first segmented from the IF images. Then, feature sets consisting of the characteristics of the minimum spanning tree (MST) connecting the EN and the fractal dimension (FD) of gland boundaries were obtained from the segmented compartments. We demonstrated the utility of the proposed features in prostate cancer recurrence prediction on a multi-institution cohort of 1027 patients. Univariate analysis revealed that both FD and one of the MST features were highly effective for predicting cancer recurrence (p <= 0.0001). In multivariate analysis, an MST feature was selected for a model incorporating clinical and image features. The model achieved a concordance index (CI) of 0.73 on the validation set, which was significantly higher than the CI of 0.69 for the standard multivariate model based solely on clinical features currently used in clinical practice (p < 0.0001). The contributions of this work are twofold. First, it is the first demonstration of the utility of the proposed features in morphometric analysis of IF images. Second, this is the largest scale study of the efficacy and robustness of the proposed features in prostate cancer prognosis.

Characterization of Polymer Blends: Optical Microscopy (*Polarized, Interference and Phase Contrast Microscopy*) and Confocal Microscopy

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

Ramanathan, Nathan Muruganathan; Darling, Seth B.

2015-01-01

Chapter 15 surveys the characterization of macro, micro and meso morphologies of polymer blends by optical microscopy. Confocal Microscopy offers the ability to view the three dimensional morphology of polymer blends, popular in characterization of biological systems. Confocal microscopy uses point illumination and a spatial pinhole to eliminate out-of focus light in samples that are thicker than the focal plane.

21 CFR 870.3640 - Indirect pacemaker generator function analyzer.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Indirect pacemaker generator function analyzer... Indirect pacemaker generator function analyzer. (a) Identification. An indirect pacemaker generator function analyzer is an electrically powered device that is used to determine pacemaker function or...

21 CFR 870.3640 - Indirect pacemaker generator function analyzer.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Indirect pacemaker generator function analyzer... Indirect pacemaker generator function analyzer. (a) Identification. An indirect pacemaker generator function analyzer is an electrically powered device that is used to determine pacemaker function or...

21 CFR 870.3640 - Indirect pacemaker generator function analyzer.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Indirect pacemaker generator function analyzer... Indirect pacemaker generator function analyzer. (a) Identification. An indirect pacemaker generator function analyzer is an electrically powered device that is used to determine pacemaker function or...

21 CFR 870.3640 - Indirect pacemaker generator function analyzer.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Indirect pacemaker generator function analyzer... Indirect pacemaker generator function analyzer. (a) Identification. An indirect pacemaker generator function analyzer is an electrically powered device that is used to determine pacemaker function or...

Are Indirect Benefits Relevant to Health Care Allocation Decisions?

PubMed Central

Du Toit, Jessica; Millum, Joseph

2016-01-01

Abstract When allocating scarce healthcare resources, the expected benefits of alternative allocations matter. But, there are different kinds of benefits. Some are direct benefits to the recipient of the resource such as the health improvements of receiving treatment. Others are indirect benefits to third parties such as the economic gains from having a healthier workforce. This article considers whether only the direct benefits of alternative healthcare resource allocations are relevant to allocation decisions, or whether indirect benefits are relevant too. First, we distinguish different conceptions of direct and indirect benefits and argue that only a recipient conception could be morally relevant. We analyze four arguments for thinking that indirect benefits should not count and argue that none is successful in showing that the indirectness of a benefit is a good reason not to count it. We conclude that direct and indirect benefits should be evaluated in the same way. PMID:27465773

48 CFR 1542.703-2 - Certificate of indirect costs.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 6 2010-10-01 2010-10-01 true Certificate of indirect costs. 1542.703-2 Section 1542.703-2 Federal Acquisition Regulations System ENVIRONMENTAL PROTECTION AGENCY CONTRACT MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 1542.703-2 Certificate of indirect...

Innovative Strategies for Clinical Microscopy Instruction: Virtual Versus Light Microscopy.

PubMed

McDaniel, M Jane; Russell, Gregory B; Crandall, Sonia J

2018-06-01

The purpose of the study was to compare virtual microscopy with light microscopy to determine differences in learning outcomes and learner attitudes in teaching clinical microscopy to physician assistant (PA) students. A prospective, randomized, crossover design study was conducted with a convenience sample of 67 first-year PA students randomized to 2 groups. One group used light microscopes to find microscopic structures, whereas the other group used instructor-directed video streaming of microscopic elements. At the midpoint of the study, the groups switched instructional strategies. Learning outcomes were assessed via posttest after each section of the study, with comparison of final practical examination results to previous cohorts. Attitudes about the 2 educational strategies were assessed through a postcourse questionnaire with a Likert scale. Analysis of the first posttest demonstrated that students in the video-streamed group had significantly better learning outcomes than those in the light microscopy group (P = .004; Cohen's d = 0.74). Analysis of the posttest after crossover showed no differences between the 2 groups (P = .48). Between the 2 posttests, students first assigned to the light microscopy group scored a 6.6 mean point increase (±10.4 SD; p = .0011), whereas students first assigned to the virtual microscopy group scored a 1.3 mean point increase (±7.1 SD; p = .29). The light microscopy group improved more than the virtual microscopy group (P = .019). Analysis of practical examination data revealed higher scores for the study group compared with 5 previous cohorts of first-year students (P < .0001; Cohen's d = 0.66). Students preferred virtual microscopy to traditional light microscopy. Virtual microscopy is an effective educational strategy, and students prefer this method when learning to interpret images of clinical specimens.

The Minicommunity Design to Assess Indirect Effects of Vaccination.

PubMed

Halloran, M Elizabeth

2012-08-01

We propose the minicommunity design to estimate indirect effects of vaccination. Establishing indirect effects of vaccination in unvaccinated subpopulations could have important implications for global vaccine policies. In the minicommunity design, the household or other small transmission unit serves as the cluster in which to estimate indirect effects of vaccination, similar to studies in larger communities to estimate indirect, total, and overall effects. Examples from the literature include studies in small transmission units to estimate indirect effects of pertussis, pneumococcal, influenza, and cholera vaccines. We characterize the minicommunity design by several methodologic considerations, including the assignment mechanism, ascertainment, the role of transmission outside the transmission unit, and the relation of the size of the transmission unit to number of people vaccinated. The minicommunity study for indirect effects is contrasted with studies to estimate vaccine effects on infectiousness and protective effects under conditions of household exposure within small transmission units. The minicommunity design can be easily implemented in individually randomized studies by enrolling and following-up members of households of the randomized individuals. The methodology for the minicommunity design for estimating indirect effects of vaccination deserves much future research.

A flow-cytometry-based method to simplify the analysis and quantification of protein association to chromatin in mammalian cells

PubMed Central

Forment, Josep V.; Jackson, Stephen P.

2016-01-01

Protein accumulation on chromatin has traditionally been studied using immunofluorescence microscopy or biochemical cellular fractionation followed by western immunoblot analysis. As a way to improve the reproducibility of this kind of analysis, make it easier to quantify and allow a stream-lined application in high-throughput screens, we recently combined a classical immunofluorescence microscopy detection technique with flow cytometry1. In addition to the features described above, and by combining it with detection of both DNA content and DNA replication, this method allows unequivocal and direct assignment of cell-cycle distribution of protein association to chromatin without the need for cell culture synchronization. Furthermore, it is relatively quick (no more than a working day from sample collection to quantification), requires less starting material compared to standard biochemical fractionation methods and overcomes the need for flat, adherent cell types that are required for immunofluorescence microscopy. PMID:26226461

48 CFR 642.703-2 - Certificate of indirect costs.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 4 2010-10-01 2010-10-01 false Certificate of indirect costs. 642.703-2 Section 642.703-2 Federal Acquisition Regulations System DEPARTMENT OF STATE CONTRACT MANAGEMENT CONTRACT ADMINISTRATION AND AUDIT SERVICES Indirect Cost Rates 642.703-2 Certificate of indirect...

Transport of Indirect Excitons in High Magnetic Fields

NASA Astrophysics Data System (ADS)

Dorow, C. J.; Kuznetsova, Y. Y.; Calman, E. V.; Butov, L. V.; Wilkes, J.; Campman, K. L.; Gossard, A. C.

Spatially- and spectrally-resolved photoluminescence measurements of indirect excitons in high magnetic fields are presented. The high magnetic field regime for excitons is realized when the cyclotron splitting compares to the exciton binding energy. Due to small mass and binding energy, the high magnetic field regime for excitons is achievable in lab, requiring a few Tesla. Long indirect exciton lifetimes allow large exciton transport distances before recombination, giving an opportunity to study transport and relaxation kinetics of indirect magnetoexcitons via optical imaging. Indirect excitons in several Landau level states are realized. 0e -0h indirect magnetoexcitons (formed from electrons and holes at zeroth Landau levels) travel over large distances and form an emission ring around the excitation spot. In contrast, the 1e -1h and 2e -2h states do not exhibit long transport distances, and the spatial profiles of the emission closely follow the laser excitation. The 0e -0h indirect magnetoexciton transport distance reduces with increasing magnetic field. Accompanying theoretical work explains these effects in terms of magnetoexciton energy relaxation and effective mass enhancement. Supported by NSF Grant No. 1407277. J.W. was supported by the EPSRC (Grant EP/L022990/1). C.J.D. was supported by the NSF Graduate Research Fellowship Program under Grant No. DGE-1144086.

10 CFR 602.15 - Indirect cost limitations.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 10 Energy 4 2013-01-01 2013-01-01 false Indirect cost limitations. 602.15 Section 602.15 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS EPIDEMIOLOGY AND OTHER HEALTH STUDIES FINANCIAL ASSISTANCE PROGRAM § 602.15 Indirect cost limitations. Awards issued under this part for conferences and...

10 CFR 602.15 - Indirect cost limitations.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 10 Energy 4 2012-01-01 2012-01-01 false Indirect cost limitations. 602.15 Section 602.15 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS EPIDEMIOLOGY AND OTHER HEALTH STUDIES FINANCIAL ASSISTANCE PROGRAM § 602.15 Indirect cost limitations. Awards issued under this part for conferences and...

10 CFR 602.15 - Indirect cost limitations.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 10 Energy 4 2014-01-01 2014-01-01 false Indirect cost limitations. 602.15 Section 602.15 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS EPIDEMIOLOGY AND OTHER HEALTH STUDIES FINANCIAL ASSISTANCE PROGRAM § 602.15 Indirect cost limitations. Awards issued under this part for conferences and...

10 CFR 602.15 - Indirect cost limitations.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 10 Energy 4 2011-01-01 2011-01-01 false Indirect cost limitations. 602.15 Section 602.15 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS EPIDEMIOLOGY AND OTHER HEALTH STUDIES FINANCIAL ASSISTANCE PROGRAM § 602.15 Indirect cost limitations. Awards issued under this part for conferences and...

10 CFR 602.15 - Indirect cost limitations.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 10 Energy 4 2010-01-01 2010-01-01 false Indirect cost limitations. 602.15 Section 602.15 Energy DEPARTMENT OF ENERGY (CONTINUED) ASSISTANCE REGULATIONS EPIDEMIOLOGY AND OTHER HEALTH STUDIES FINANCIAL ASSISTANCE PROGRAM § 602.15 Indirect cost limitations. Awards issued under this part for conferences and...

Are Indirect Benefits Relevant to Health Care Allocation Decisions?

PubMed

Du Toit, Jessica; Millum, Joseph

2016-10-01

When allocating scarce healthcare resources, the expected benefits of alternative allocations matter. But, there are different kinds of benefits. Some are direct benefits to the recipient of the resource such as the health improvements of receiving treatment. Others are indirect benefits to third parties such as the economic gains from having a healthier workforce. This article considers whether only the direct benefits of alternative healthcare resource allocations are relevant to allocation decisions, or whether indirect benefits are relevant too. First, we distinguish different conceptions of direct and indirect benefits and argue that only a recipient conception could be morally relevant. We analyze four arguments for thinking that indirect benefits should not count and argue that none is successful in showing that the indirectness of a benefit is a good reason not to count it. We conclude that direct and indirect benefits should be evaluated in the same way. Published by Oxford University Press on behalf of the Journal of Medicine and Philosophy, Inc. 2016.

Correlative Single-Molecule Localization Microscopy and Confocal Microscopy.

PubMed

Soeller, Christian; Hou, Yufeng; Jayasinghe, Isuru D; Baddeley, David; Crossman, David

2017-01-01

Single-molecule localization microscopy allows the ability to image fluorescence labeled molecular targets at nanoscale resolution. However, for many biological questions the ability to provide tissue and cellular context in addition to these high resolution data is eminently informative. Here, we describe a procedure to achieve this aim by correlatively imaging human cardiac tissue first at the nanoscale with direct stochastic optical reconstruction microscopy (dSTORM) and then at the diffraction limit with conventional confocal microscopy.

Reflection imaging of China ink-perfused brain vasculature using confocal laser-scanning microscopy after clarification of brain tissue by the Spalteholz method.

PubMed

Gutierre, R C; Vannucci Campos, D; Mortara, R A; Coppi, A A; Arida, R M

2017-04-01

Confocal laser-scanning microscopy is a useful tool for visualizing neurons and glia in transparent preparations of brain tissue from laboratory animals. Currently, imaging capillaries and venules in transparent brain tissues requires the use of fluorescent proteins. Here, we show that vessels can be imaged by confocal laser-scanning microscopy in transparent cortical, hippocampal and cerebellar preparations after clarification of China ink-injected specimens by the Spalteholz method. This method may be suitable for global, three-dimensional, quantitative analyses of vessels, including stereological estimations of total volume and length and of surface area of vessels, which constitute indirect approaches to investigate angiogenesis. © 2017 Anatomical Society.

The chemotaxis-like Che1 pathway has an indirect role in adhesive cell properties of Azospirillum brasilense.

PubMed

Siuti, Piro; Green, Calvin; Edwards, Amanda Nicole; Doktycz, Mitchel J; Alexandre, Gladys

2011-10-01

The Azospirillum brasilense chemotaxis-like Che1 signal transduction pathway was recently shown to modulate changes in adhesive cell surface properties that, in turn, affect cell-to-cell aggregation and flocculation behaviors rather than flagellar-mediated chemotaxis. Attachment to surfaces and root colonization may be functions related to flocculation. Here, the conditions under which A. brasilense wild-type Sp7 and che1 mutant strains attach to abiotic and biotic surfaces were examined using in vitro attachment and biofilm assays combined with atomic force microscopy and confocal microscopy. The nitrogen source available for growth is found to be a major modulator of surface attachment by A. brasilense and could be promoted in vitro by lectins, suggesting that it depends on interaction with surface-exposed residues within the extracellular matrix of cells. However, Che1-dependent signaling is shown to contribute indirectly to surface attachment, indicating that distinct mechanisms are likely underlying flocculation and attachment to surfaces in A. brasilense. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Immunocytochemical methods to study the distribution of Orientia tsutsugamushi in Leptotrombidium (Acari: Trombiculidae) chiggers.

PubMed

Myint, K S; Linthicum, K J; Tanskul, P; Lerdthusnee, K; Vaughn, D W; Manomuth, C; Mongkolsirichaikul, D; Hansukjariya, P; Hastriter, M W

1998-07-01

Immunocytochemical methods were developed and tested for their ability to detect the distribution of Orientia tsutsugamushi in paraffin sections of adult chiggers (Leptotrombidium imphalum Vercammen-Grandjean & Langston). Rickettsial antigen was detected by application of a simple direct or amplified immunocytochemistry procedure and an indirect immunofluorescent procedure. In the direct procedure alkaline phosphatase conjugation to the mouse polyclonal antibody to the Karp strain was followed by the HistoMark Red test system to detect rickettsial antigen. The amplification procedure used a similar method but used an unlabeled primary antibody followed by secondary biotinylated antimouse IgG, streptavidin-alkaline phosphatase, and the HistoMark Red test system. The immunofluorescent procedure included a biotinylated secondary antibody followed by addition of a streptavidin-FITC conjugate. Specific tissue tropisms in infected chiggers were observed in the salivary glands, nervous tissue, and ovaries of adult female mites in all procedures; however, nonspecific fluorescence of the chigger limited definitive identification of tissue tropisms with the indirect immunofluorescent procedure.

Characterization and possible function of glyceraldehyde-3-phosphate dehydrogenase-spermatogenic protein GAPDHS in mammalian sperm.

PubMed

Margaryan, Hasmik; Dorosh, Andriy; Capkova, Jana; Manaskova-Postlerova, Pavla; Philimonenko, Anatoly; Hozak, Pavel; Peknicova, Jana

2015-03-08

Sperm proteins are important for the sperm cell function in fertilization. Some of them are involved in the binding of sperm to the egg. We characterized the acrosomal sperm protein detected by a monoclonal antibody (MoAb) (Hs-8) that was prepared in our laboratory by immunization of BALB/c mice with human ejaculated sperms and we tested the possible role of this protein in the binding assay. Indirect immunofluorescence and immunogold labelling, gel electrophoresis, Western blotting and protein sequencing were used for Hs-8 antigen characterization. Functional analysis of GAPDHS from the sperm acrosome was performed in the boar model using sperm/zona pellucida binding assay. Monoclonal antibody Hs-8 is an anti-human sperm antibody that cross-reacts with the Hs-8-related protein in spermatozoa of other mammalian species (boar, mouse). In the immunofluorescence test, Hs-8 antibody recognized the protein localized in the acrosomal part of the sperm head and in the principal piece of the sperm flagellum. In immunoblotting test, MoAb Hs-8 labelled a protein of 45 kDa in the extract of human sperm. Sequence analysis identified protein Hs-8 as GAPDHS (glyceraldehyde 3-phosphate dehydrohenase-spermatogenic). For this reason, commercial mouse anti-GAPDHS MoAb was applied in control tests. Both antibodies showed similar staining patterns in immunofluorescence tests, in electron microscopy and in immunoblot analysis. Moreover, both Hs-8 and anti-GAPDHS antibodies blocked sperm/zona pellucida binding. GAPDHS is a sperm-specific glycolytic enzyme involved in energy production during spermatogenesis and sperm motility; its role in the sperm head is unknown. In this study, we identified the antigen with Hs8 antibody and confirmed its localization in the apical part of the sperm head in addition to the principal piece of the flagellum. In an indirect binding assay, we confirmed the potential role of GAPDHS as a binding protein that is involved in the secondary sperm

Predictors of negotiated NIH indirect rates at US institutions.

PubMed

Johnston, S Claiborne; Desmond-Hellmann, Susan; Hauser, Stewart; Vermillion, Eric; Mia, Nilo

2015-01-01

The United States (US) Department of Health and Human Services and the Office of Naval Research negotiate institutional rates for payments of overhead costs associated with administration and space usage, commonly known as indirect rates. Such payments account for a large proportion of spending by the National Institutes of Health (NIH). Little has been published about differences in rates and their predictors. Negotiated indirect rates for on-campus research grants were requested from the Council on Governmental Relations for the 100 institutions with greatest NIH funding in 2010. NIH funding, cost of living (ACCRA Index for 2008), public vs. private status, negotiating governmental organization (Department of Health and Human Services or Office of Naval Research), US Census Region, and year were assessed as predictors of institutional indirect rates using generalized estimating equations with all variables included in the model. Overall, 72 institutions participated, with 207 reported indirect rates for the years 2006, 2008, and 2010. Indirect rates ranged from 36.3% to 78%, with an average of 54.5%. Mean rates increased from 53.6% in 2006 to 55.4% in 2010 (p<0.001). In multivariable models, private institutions had 6.2% (95% CI 3.7%-8.7%; p<0.001) higher indirect rates than public institutions. Rates in the Northeast were highest (Midwest 4.0% lower; West 4.9% lower; South 5.2% lower). Greater NIH funding (p = 0.025) and cost of living (p = 0.034) also predicted indirect rates while negotiating governmental organization did not (p = 0.414). Negotiated indirect rates for governmental research grants to academic centers vary widely. Although the association between indirect rates and cost of living may be justified, the cause of variation in rates by region, public-private status, and NIH funding levels is unclear.

Rapid multiple immunofluorescent staining for the simultaneous detection of cytokeratin and vimentin in the cytology of canine tumors.

PubMed

Sawa, Mariko; Yabuki, Akira; Kohyama, Moeko; Miyoshi, Noriaki; Yamato, Osamu

2018-06-01

Immunocytochemistry (ICC) is utilized as an advanced technique in veterinary cytology. In tumor diagnosis, cytokeratin and vimentin are markers used to distinguish the origin of tumor cells. Standard enzyme-based ICC has limitations in clinical use; and therefore, more convenient and reliable methods are needed. The purpose of this study was to develop a rapid multiple immunofluorescent (RMIF) detection method for dual cytokeratin and vimentin staining on cytology slides in dogs. Air-dried smear samples from solid tumors and sediments of pleural effusions were prepared from dogs (n = 14) that were admitted to the Veterinary Teaching Hospital, Kagoshima University, Japan. Mouse monoclonal anti-human cytokeratin (AE1/AE3) and rabbit monoclonal anti-human vimentin (SP20) antibodies were used as primary antibodies, followed by staining with Alexa Fluor-conjugated secondary antibodies. Staining using the RMIF method was compared with enzyme-based ICC staining. Rapid multiple immunofluorescent immunostaining was clear and specific in the evaluated smears, whereas the enzyme-based ICC showed nonspecific signals. By using the RMIF staining method, epithelial cells, mesenchymal cells, and mesothelial cells could be classified on a single smear of a pleural effusion. In smears of lymph nodes with epithelial tumor metastases, the RMIF method successfully detected metastatic epithelial tumor cells. The RMIF method might be a useful tool for diagnostic cytology in veterinary medicine. © 2018 American Society for Veterinary Clinical Pathology.

Cross-reactions between alpha-streptococci and Omniserum, a polyvalent pneumococcal serum, demonstrated by direct immunofluorescence, immunoelectroosmophoresis, and latex agglutination.

PubMed Central

Holmberg, H; Danielsson, D; Hardie, J; Krook, A; Whiley, R

1985-01-01

In recent years several groups have used serological methods to demonstrate pneumococcal capsular antigens in sputum. In the present study 123 strains of alpha-hemolytic streptococci (including 97 strains from sputum or pharyngeal specimens) were tested for cross-reactions with a polyvalent antipneumococcal serum (Omniserum). Representatives of the following species were included: Streptococcus bovis, S. equinus, S. intermedius, S. lactis, S. milleri, S. mitis, S. mutans, S. sobrinus, S. salivarius, S. sanguis, S. suis, and Aerococcus viridans. Serological reactions were detected by direct immunofluorescence, immunoelectroosmophoresis, and latex agglutination. Fifteen (12%) of the strains gave positive reactions by all three methods. Positive reactions were also observed with another 32 strains (26%) with two of the methods, whereas 37 strains (30%) gave positive reactions by just one technique. Altogether 84 (68%) strains gave positive reactions with one or more of the methods. Latex agglutination gave positive reactions with 26 (21%) strains compared with 57 (46%) in immunofluorescence and 63 (51%) in immunoelectroosmophoresis. Absorption of the antiserum with one alpha-hemolytic strain reduced but did not entirely eliminate the cross-reactions with five tested strains. These findings indicate a potential risk of cross-reactions with polyvalent antipneumococcal serum in tests carried out on sputa or other specimens which may be contaminated with alpha-hemolytic streptococci. PMID:3889046

Indirect Dietary Residential Exposure Assessment Model (IDREAM) Implementation

EPA Pesticide Factsheets

The Indirect Dietary Residential Exposure Assessment Model (IDREAM) estimates indirect ingestion exposure to disinfectants used in residential settings on hard surfaces where there may be inadvertent transfer to edible items prepared on those surfaces.

Progress in the Correlative Atomic Force Microscopy and Optical Microscopy

PubMed Central

Zhou, Lulu; Cai, Mingjun; Tong, Ti; Wang, Hongda

2017-01-01

Atomic force microscopy (AFM) has evolved from the originally morphological imaging technique to a powerful and multifunctional technique for manipulating and detecting the interactions between molecules at nanometer resolution. However, AFM cannot provide the precise information of synchronized molecular groups and has many shortcomings in the aspects of determining the mechanism of the interactions and the elaborate structure due to the limitations of the technology, itself, such as non-specificity and low imaging speed. To overcome the technical limitations, it is necessary to combine AFM with other complementary techniques, such as fluorescence microscopy. The combination of several complementary techniques in one instrument has increasingly become a vital approach to investigate the details of the interactions among molecules and molecular dynamics. In this review, we reported the principles of AFM and optical microscopy, such as confocal microscopy and single-molecule localization microscopy, and focused on the development and use of correlative AFM and optical microscopy. PMID:28441775

Resonant indirect optical absorption in germanium

NASA Astrophysics Data System (ADS)

Menéndez, José; Noël, Mario; Zwinkels, Joanne C.; Lockwood, David J.

2017-09-01

The optical absorption coefficient of pure Ge has been determined from high-accuracy, high-precision optical measurements at photon energies covering the spectral range between the indirect and direct gaps. The results are compared with a theoretical model that fully accounts for the resonant nature of the energy denominators that appear in perturbation-theory expansions of the absorption coefficient. The model generalizes the classic Elliott approach to indirect excitons, and leads to a predicted optical absorption that is in excellent agreement with the experimental values using just a single adjustable parameter: the average deformation potential DΓ L coupling electrons at the bottom of the direct and indirect valleys in the conduction band. Remarkably, the fitted value, DΓ L=4.3 ×108eV /cm , is in nearly perfect agreement with independent measurements and ab initio predictions of this parameter, confirming the validity of the proposed theory, which has general applicability.

L'indirection: Procede d'expression et de persuasion en communication publique (Indirection: Process of Expression and Persuasion in Public Communication).

ERIC Educational Resources Information Center

Gauthier, Gilles

2001-01-01

Focuses on the indirection process presented in Searle's and Vanderveken's theory of speech acts: the performance of a primary speech act by means of the accomplishment of a secondary speech act. Discusses indirection mechanisms used in advertising and in political communication. (Author/VWL)

Immunofluorescence-based methods to monitor DNA end resection

PubMed Central

Mukherjee, Bipasha; Tomimatsu, Nozomi; Burma, Sandeep

2017-01-01

Summary Double-strand breaks (DSBs) are the most deleterious amongst all types of DNA damage that can occur in the cell. These breaks arise from both endogenous (for example, DNA replication stress) as well as exogenous insults (for example, ionizing radiation). DSBs are principally repaired by one of two major pathways: non-homologous end joining (NHEJ) or homologous recombination (HR). NHEJ is an error-prone process that can occur in all phases of the cell cycle, while HR is limited to the S and G2 phases of the cell cycle when a sister chromatid is available as a template for error-free repair. The first step in HR is “DNA end resection”, a process during which the broken DNA end is converted into a long stretch of 3′-ended single-stranded DNA (ssDNA). In recent years, DNA end resection has been identified as a pivotal step that controls “repair pathway choice” i.e., the appropriate choice between NHEJ and HR for DSB repair. Therefore, methods to quantitatively or semi-quantitatively assess DNA end resection have gained importance in laboratories working on DNA repair. In this chapter, we describe two simple immunofluorescence-based techniques to monitor DNA end resection in mammalian cells. The first technique involves immuno-detection of Replication Protein A (RPA), a ssDNA-binding protein that binds to resected DNA. The second technique involves labeling of genomic DNA with 5-bromo-2′-deoxyuridine (BrdU) that can be detected by anti-BrdU antibody only after the DNA becomes single stranded due to resection. These methods are not complicated, do not involve sophisticated instrumentation or reporter constructs, and can be applied to most mammalian cell lines, and therefore, should be of broad utility as simple ways of monitoring DNA end resection in vivo. PMID:25804748

Incomplete caries removal and indirect pulp capping in primary molars: a randomized controlled trial.

PubMed

Bressani, Ana Eliza Lemes; Mariath, Adriela Azevedo Souza; Haas, Alex Nogueira; Garcia-Godoy, Franklin; de Araujo, Fernando Borba

2013-08-01

To compare the effect of incomplete caries removal (ICR) and indirect pulp capping (IPC) with calcium hydroxide (CH) or an inert material (wax) on color, consistency and contamination of the remaining dentin of primary molars. This double-blind, parallel-design, randomized controlled trial included 30 children presenting one primary molar with deep caries lesion. Children were randomly assigned after ICR to receive IPC with CH or wax. All teeth were then restored with resin composite. Baseline dentin color and consistency were evaluated after ICR, and dentin samples were collected for contamination analyses using scanning electron microscopy. After 3 months, restorations were removed and the three parameters were re-evaluated. In both groups, dentin became significantly darker after 3 months. No cases of yellow dentin were observed after 3 months with CH compared to 33.3% of the wax cases (P < 0.05). A statistically significant difference over time was observed only for CH regarding consistency. CH stimulated a dentin hardening process in a statistically higher number of cases than wax (86.7% vs. 33.3%; P = 0.008). Contamination changed significantly over time in CH and wax without significant difference between groups. It was concluded that CH and wax arrested the carious process of the remaining carious dentin after indirect pulp capping, but CH showed superior dentin color and consistency after 3 months.

Quantification of Bacterial Twitching Motility in Dense Colonies Using Transmitted Light Microscopy and Computational Image Analysis.

PubMed

Smith, Benjamin; Li, Jianfang; Metruccio, Matteo; Wan, Stephanie; Evans, David; Fleiszig, Suzanne

2018-04-20

A method was developed to allow the quantification and mapping of relative bacterial twitching motility in dense samples, where tracking of individual bacteria was not feasible. In this approach, movies of bacterial films were acquired using differential interference contrast microscopy (DIC), and bacterial motility was then indirectly quantified by the degree to which the bacteria modulated the intensity of light in the field-of-view over time. This allowed the mapping of areas of relatively high and low motility within a single field-of-view, and comparison of the total distribution of motility between samples.

Multi-color localization microscopy of fixed cells as a promising tool to study organization of bacterial cytoskeleton

NASA Astrophysics Data System (ADS)

Vedyaykin, A. D.; Gorbunov, V. V.; Sabantsev, A. V.; Polinovskaya, V. S.; Vishnyakov, I. E.; Melnikov, A. S.; Serdobintsev, P. Yu; Khodorkovskii, M. A.

2015-11-01

Localization microscopy allows visualization of biological structures with resolution well below the diffraction limit. Localization microscopy was used to study FtsZ organization in Escherichia coli previously in combination with fluorescent protein labeling, but the fact that fluorescent chimeric protein was unable to rescue temperature-sensitive ftsZ mutants suggests that obtained images may not represent native FtsZ structures faithfully. Indirect immunolabeling of FtsZ not only overcomes this problem, but also allows the use of the powerful visualization methods arsenal available for different structures in fixed cells. In this work we simultaneously obtained super-resolution images of FtsZ structures and diffraction-limited or super-resolution images of DNA and cell surface in E. coli, which allows for the study of the spatial arrangement of FtsZ structures with respect to the nucleoid positions and septum formation.

Automobile Industry Retail Price Equivalent and Indirect Cost ...

EPA Pesticide Factsheets

This report develops a modified multiplier, referred to as an indirect cost (IC) multiplier, which specifically evaluates the components of indirect costs that are likely to be affected by vehicle modifications associated with environmental regulation. A range of IC multipliers are developed that 1) account for differences in the technical complexity of required vehicle modifications and 2) adjust over time as new technologies become assimilated into the automotive production process. To develop an improved methodology for estimating indirect costs of new environmental regulations on automobile manufacturers.

Confocal imaging of butterfly tissue.

PubMed

Brunetti, Craig R

2014-01-01

To understand the molecular events responsible for morphological change requires the ability to examine gene expression in a wide range of organisms in addition to model systems to determine how the differences in gene expression correlate with phenotypic differences. There are approximately 12,000 species of butterflies, most, with distinct patterns on their wings. The most important tool for studying gene expression in butterflies is confocal imaging of butterfly tissue by indirect immunofluorescence using either cross-reactive antibodies from closely related species such as Drosophila or developing butterfly-specific antibodies. In this report, we describe how indirect immunofluorescence protocols can be used to visualize protein expression patterns on the butterfly wing imaginal disc and butterfly embryo.

Cell junction protein armadillo repeat gene deleted in velo-cardio-facial syndrome is expressed in the skin and colocalizes with autoantibodies of patients affected by a new variant of endemic pemphigus foliaceus in Colombia.

PubMed

Abreu-Velez, Ana Maria; Yi, Hong; Howard, Michael S

2017-10-01

We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America (El Bagre-EPF, or pemphigus Abreu-Manu). El Bagre-EPF differs from other types of EPF clinically, epidemiologically, immunologically and in its target antigens. We reported the presence of patient autoantibodies colocalizing with armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), a catenin cell junction protein colocalizing with El Bagre-EPF autoantibodies in the heart and within pilosebaceous units along their neurovascular supply routes. Here we investigate the presence of ARVCF in skin and its possibility as a cutaneous El Bagre-EPF antigen. We used a case-control study, testing sera of 45 patients and 45 controls via direct and indirect immunofluorescence (DIF/IIF), confocal microscopy, immunoelectron microscopy and immunoblotting for the presence of ARVCF and its relationship with El Bagre-EPF autoantibodies in the skin. We also immunoadsorbed samples with desmoglein 1 (Dsg1) ectodomain (El Bagre-EPF antigen) by incubating with the positive ARVCF samples from DIF and IIF. ARVCF was expressed in all the samples from the cases and controls. Immunoadsorption with Dsg1 on positive ARVCF immunofluorescence DIF/IIF cases showed that the immune response was present against non-desmoglein 1 antigen(s). Overall, 40/45 patients showed colocalization of their autoantibodies with ARVCF in the epidermis; no controls from the endemic area displayed colocalization. We demonstrate that ARVCF is expressed in many areas of human skin, and colocalizes with the majority of El Bagre-EPF autoantibodies as a putative antigen.

Cell junction protein armadillo repeat gene deleted in velo-cardio-facial syndrome is expressed in the skin and colocalizes with autoantibodies of patients affected by a new variant of endemic pemphigus foliaceus in Colombia

PubMed Central

Abreu-Velez, Ana Maria; Yi, Hong; Howard, Michael S.

2017-01-01

Background We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America (El Bagre-EPF, or pemphigus Abreu-Manu). El Bagre-EPF differs from other types of EPF clinically, epidemiologically, immunologically and in its target antigens. We reported the presence of patient autoantibodies colocalizing with armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), a catenin cell junction protein colocalizing with El Bagre-EPF autoantibodies in the heart and within pilosebaceous units along their neurovascular supply routes. Here we investigate the presence of ARVCF in skin and its possibility as a cutaneous El Bagre-EPF antigen. Methods We used a case-control study, testing sera of 45 patients and 45 controls via direct and indirect immunofluorescence (DIF/IIF), confocal microscopy, immunoelectron microscopy and immunoblotting for the presence of ARVCF and its relationship with El Bagre-EPF autoantibodies in the skin. We also immunoadsorbed samples with desmoglein 1 (Dsg1) ectodomain (El Bagre-EPF antigen) by incubating with the positive ARVCF samples from DIF and IIF. Results ARVCF was expressed in all the samples from the cases and controls. Immunoadsorption with Dsg1 on positive ARVCF immunofluorescence DIF/IIF cases showed that the immune response was present against non-desmoglein 1 antigen(s). Overall, 40/45 patients showed colocalization of their autoantibodies with ARVCF in the epidermis; no controls from the endemic area displayed colocalization. Conclusions We demonstrate that ARVCF is expressed in many areas of human skin, and colocalizes with the majority of El Bagre-EPF autoantibodies as a putative antigen. PMID:29214101

Double-Stranded RNA Is Detected by Immunofluorescence Analysis in RNA and DNA Virus Infections, Including Those by Negative-Stranded RNA Viruses.

PubMed

Son, Kyung-No; Liang, Zhiguo; Lipton, Howard L

2015-09-01

Early biochemical studies of viral replication suggested that most viruses produce double-stranded RNA (dsRNA), which is essential for the induction of the host immune response. However, it was reported in 2006 that dsRNA could be detected by immunofluorescence antibody staining in double-stranded DNA and positive-strand RNA virus infections but not in negative-strand RNA virus infections. Other reports in the literature seemed to support these observations. This suggested that negative-strand RNA viruses produce little, if any, dsRNA or that more efficient viral countermeasures to mask dsRNA are mounted. Because of our interest in the use of dsRNA antibodies for virus discovery, particularly in pathological specimens, we wanted to determine how universal immunostaining for dsRNA might be in animal virus infections. We have detected the in situ formation of dsRNA in cells infected with vesicular stomatitis virus, measles virus, influenza A virus, and Nyamanini virus, which represent viruses from different negative-strand RNA virus families. dsRNA was also detected in cells infected with lymphocytic choriomeningitis virus, an ambisense RNA virus, and minute virus of mice (MVM), a single-stranded DNA (ssDNA) parvovirus, but not hepatitis B virus. Although dsRNA staining was primarily observed in the cytoplasm, it was also seen in the nucleus of cells infected with influenza A virus, Nyamanini virus, and MVM. Thus, it is likely that most animal virus infections produce dsRNA species that can be detected by immunofluorescence staining. The apoptosis induced in several uninfected cell lines failed to upregulate dsRNA formation. An effective antiviral host immune response depends on recognition of viral invasion and an intact innate immune system as a first line of defense. Double-stranded RNA (dsRNA) is a viral product essential for the induction of innate immunity, leading to the production of type I interferons (IFNs) and the activation of hundreds of IFN

Two distinct neural mechanisms underlying indirect reciprocity.

PubMed

Watanabe, Takamitsu; Takezawa, Masanori; Nakawake, Yo; Kunimatsu, Akira; Yamasue, Hidenori; Nakamura, Mitsuhiro; Miyashita, Yasushi; Masuda, Naoki

2014-03-18

Cooperation is a hallmark of human society. Humans often cooperate with strangers even if they will not meet each other again. This so-called indirect reciprocity enables large-scale cooperation among nonkin and can occur based on a reputation mechanism or as a succession of pay-it-forward behavior. Here, we provide the functional and anatomical neural evidence for two distinct mechanisms governing the two types of indirect reciprocity. Cooperation occurring as reputation-based reciprocity specifically recruited the precuneus, a region associated with self-centered cognition. During such cooperative behavior, the precuneus was functionally connected with the caudate, a region linking rewards to behavior. Furthermore, the precuneus of a cooperative subject had a strong resting-state functional connectivity (rsFC) with the caudate and a large gray matter volume. In contrast, pay-it-forward reciprocity recruited the anterior insula (AI), a brain region associated with affective empathy. The AI was functionally connected with the caudate during cooperation occurring as pay-it-forward reciprocity, and its gray matter volume and rsFC with the caudate predicted the tendency of such cooperation. The revealed difference is consistent with the existing results of evolutionary game theory: although reputation-based indirect reciprocity robustly evolves as a self-interested behavior in theory, pay-it-forward indirect reciprocity does not on its own. The present study provides neural mechanisms underlying indirect reciprocity and suggests that pay-it-forward reciprocity may not occur as myopic profit maximization but elicit emotional rewards.

Two distinct neural mechanisms underlying indirect reciprocity

PubMed Central

Watanabe, Takamitsu; Takezawa, Masanori; Nakawake, Yo; Kunimatsu, Akira; Yamasue, Hidenori; Nakamura, Mitsuhiro; Miyashita, Yasushi; Masuda, Naoki

2014-01-01

Cooperation is a hallmark of human society. Humans often cooperate with strangers even if they will not meet each other again. This so-called indirect reciprocity enables large-scale cooperation among nonkin and can occur based on a reputation mechanism or as a succession of pay-it-forward behavior. Here, we provide the functional and anatomical neural evidence for two distinct mechanisms governing the two types of indirect reciprocity. Cooperation occurring as reputation-based reciprocity specifically recruited the precuneus, a region associated with self-centered cognition. During such cooperative behavior, the precuneus was functionally connected with the caudate, a region linking rewards to behavior. Furthermore, the precuneus of a cooperative subject had a strong resting-state functional connectivity (rsFC) with the caudate and a large gray matter volume. In contrast, pay-it-forward reciprocity recruited the anterior insula (AI), a brain region associated with affective empathy. The AI was functionally connected with the caudate during cooperation occurring as pay-it-forward reciprocity, and its gray matter volume and rsFC with the caudate predicted the tendency of such cooperation. The revealed difference is consistent with the existing results of evolutionary game theory: although reputation-based indirect reciprocity robustly evolves as a self-interested behavior in theory, pay-it-forward indirect reciprocity does not on its own. The present study provides neural mechanisms underlying indirect reciprocity and suggests that pay-it-forward reciprocity may not occur as myopic profit maximization but elicit emotional rewards. PMID:24591599

Indirect effects in community ecology: Their definition, study and importance.

PubMed

Strauss, S Y

1991-07-01

The diversity of indirect interactions that can occur within communities is large. Recent research on indirect interactions is scattered in the literature under numerous labels. The definition of indirect effects is an important aspect of their study, and clarifies some of the subtle differences among indirect effects found in natural communities. Choosing which species to study, how to manipulate species and for what duration, which attributes to measure and, finally, which analytical techniques to use are all problems facing the community ecologist. Ultimately, we are striving for the best means of determining the relative importance of direct and indirect effects in structuring communities. Copyright © 1991. Published by Elsevier Ltd.

Preschool children's behavioral tendency toward social indirect reciprocity.

PubMed

Kato-Shimizu, Mayuko; Onishi, Kenji; Kanazawa, Tadahiro; Hinobayashi, Toshihiko

2013-01-01

Social indirect reciprocity seems to be crucial in enabling large-scale cooperative networks among genetically unrelated individuals in humans. However, there are relatively few studies on social indirect reciprocity in children compared to adults. Investigating whether young children have a behavioral tendency toward social indirect reciprocity will help us understand how and when the fundamental ability to form cooperative relationships among adults is acquired. Using naturalistic observation at a nursery school, this study examined whether 5- to 6-year-olds show a behavioral tendency to engage in social indirect reciprocity in response to their peers' prosocial behavior toward a third party. The results revealed that bystander children tended to display prosocial behavior toward their peers more frequently after observing these peers' prosocial behavior toward third-party peers, compared with control situations; this suggests that 5- to 6-year-olds may have an essential behavioral tendency to establish social indirect reciprocity when interacting with peers in their daily lives. In addition, bystanders tended to display affiliative behavior after observing focal children's prosocial behavior. In other words, observing peers' prosocial behavior toward third-party peers evoked bystanders' positive emotions toward the helpers. Considering both the present results and previous findings, we speculate that in preschoolers, such positive emotions might mediate the increase in the bystander's prosocial behavior toward the helper. In addition, an intuitional emotional process plays an important role in the preschooler's behavioral tendency toward social indirect reciprocity in natural interactions with peers.

Estimating direct and indirect costs of premenstrual syndrome.

PubMed

Borenstein, Jeff; Chiou, Chiun-Fang; Dean, Bonnie; Wong, John; Wade, Sally

2005-01-01

To quantify the economic impact of premenstrual syndrome (PMS) on the employer. Data were collected from 374 women aged 18-45 with regular menses. Direct costs were quantified using administrative claims of these patients and the Medicare Fee Schedule. Indirect costs were quantified by both self-reported days of work missed and lost productivity at work. Regression analyses were used to develop a model to project PMS-related direct and indirect costs. A total of 29.6% (n = 111) of the participants were diagnosed with PMS. A PMS diagnosis was associated with an average annual increase of $59 in direct costs (P < 0.026) and $4333 in indirect costs per patient (P < 0.0001) compared with patients without PMS. A PMS diagnosis correlated with a modest increase in direct medical costs and a large increase in indirect costs.

Separating astrophysical sources from indirect dark matter signals

PubMed Central

Siegal-Gaskins, Jennifer M.

2015-01-01

Indirect searches for products of dark matter annihilation and decay face the challenge of identifying an uncertain and subdominant signal in the presence of uncertain backgrounds. Two valuable approaches to this problem are (i) using analysis methods which take advantage of different features in the energy spectrum and angular distribution of the signal and backgrounds and (ii) more accurately characterizing backgrounds, which allows for more robust identification of possible signals. These two approaches are complementary and can be significantly strengthened when used together. I review the status of indirect searches with gamma rays using two promising targets, the Inner Galaxy and the isotropic gamma-ray background. For both targets, uncertainties in the properties of backgrounds are a major limitation to the sensitivity of indirect searches. I then highlight approaches which can enhance the sensitivity of indirect searches using these targets. PMID:25304638

[Immunofluorescence assay with Crithidia luciliae for the detection of anti-DNA antibodies. Atypical images and their relationship with Chagas' disease and leishmaniasis].

PubMed

Griemberg, Gloria; Ferrarotti, Nidia F; Svibel, Graciela; Ravelli, Maria R; Taranto, Nestor J; Malchiodi, Emilio L; Pizzimenti, Maria C

2006-01-01

Anti-native DNA antibodies can be detected by indirect immunofluorescence assay with Crithidia luciliae, displaying an annular image due to a kinetoplast containing double stranded DNA. Other structures such as membrane, flagellum and basal corpuscle can be stained as well, showing what is called atypical fluorescent images. As C. luciliae belongs to the Trypanosomatidae family, which include the human pathogens Trypanosoma cruzi and Leishmania spp., it was considered that these atypical images could be caused by cross-reactions. Serological studies for Chagas' disease were performed in 105 serum samples displaying atypical images. Sixty four percent of the samples from non endemic and 78.3% from endemic areas for Chagas' disease showed fluorescence in both, membrane and flagellum (joint image). Fifty samples from normal blood donors and 57 samples from patients with conective tissue diseases were tested with C. luciliae. None of them presented the joint image except for two patients with lupus who were also chagasic. In addition, 54 samples from chagasic patients were studied and all of them presented the joint image. We also studied 46 samples from patients with leishmaniasis from whom 28 were coinfected with T. cruzi. The joint image was observed in 88.0% of the samples with leishmaniasis and in 89.3% of the co-infected samples. The results suggest that C. luciliae could be used as an economical, and of low risk, alternative substrate for the serological diagnosis of Chagas' disease, even though it does not discriminate for Leishmania spp. infection. This study also suggests that whenever atypical images are observed in C. luciliae during the search for anti-DNA antibodies, it would be convenient to submit the patient to clinical and serological tests for the diagnosis of leishmaniosis and Chagas' disease.

Impact of virtual microscopy with conventional microscopy on student learning in dental histology.

PubMed

Hande, Alka Harish; Lohe, Vidya K; Chaudhary, Minal S; Gawande, Madhuri N; Patil, Swati K; Zade, Prajakta R

2017-01-01

In dental histology, the assimilation of histological features of different dental hard and soft tissues is done by conventional microscopy. This traditional method of learning prevents the students from screening the entire slide and change of magnification. To address these drawbacks, modification in conventional microscopy has evolved and become motivation for changing the learning tool. Virtual microscopy is the technique in which there is complete digitization of the microscopic glass slide, which can be analyzed on a computer. This research is designed to evaluate the effectiveness of virtual microscopy with conventional microscopy on student learning in dental histology. A cohort of 105 students were included and randomized into three groups: A, B, and C. Group A students studied the microscopic features of oral histologic lesions by conventional microscopy, Group B by virtual microscopy, and Group C by both conventional and virtual microscopy. The students' understanding of the subject was evaluated by a prepared questionnaire. The effectiveness of the study designs on knowledge gains and satisfaction levels was assessed by statistical assessment of differences in mean test scores. The difference in score between Groups A, B, and C at pre- and post-test was highly significant. This enhanced understanding of the subject may be due to benefits of using virtual microscopy in teaching histology. The augmentation of conventional microscopy with virtual microscopy shows enhancement of the understanding of the subject as compared to the use of conventional microscopy and virtual microscopy alone.

Electron Microscopy.

ERIC Educational Resources Information Center

Beer, Michael

1980-01-01

Reviews technical aspects of structure determination in biological electron microscopy (EM). Discusses low dose EM, low temperature microscopy, electron energy loss spectra, determination of mass or molecular weight, and EM of labeled systems. Cites 34 references. (CS)

Epidemic spreading through direct and indirect interactions.

PubMed

Ganguly, Niloy; Krueger, Tyll; Mukherjee, Animesh; Saha, Sudipta

2014-09-01

In this paper we study the susceptible-infected-susceptible epidemic dynamics, considering a specialized setting where popular places (termed passive entities) are visited by agents (termed active entities). We consider two types of spreading dynamics: direct spreading, where the active entities infect each other while visiting the passive entities, and indirect spreading, where the passive entities act as carriers and the infection is spread via them. We investigate in particular the effect of selection strategy, i.e., the way passive entities are chosen, in the spread of epidemics. We introduce a mathematical framework to study the effect of an arbitrary selection strategy and derive formulas for prevalence, extinction probabilities, and epidemic thresholds for both indirect and direct spreading. We also obtain a very simple relationship between the extinction probability and the prevalence. We pay special attention to preferential selection and derive exact formulas. The analysis reveals that an increase in the diversity in the selection process lowers the epidemic thresholds. Comparing the direct and indirect spreading, we identify regions in the parameter space where the prevalence of the indirect spreading is higher than the direct one.

Epidemic spreading through direct and indirect interactions

NASA Astrophysics Data System (ADS)

Ganguly, Niloy; Krueger, Tyll; Mukherjee, Animesh; Saha, Sudipta

2014-09-01

In this paper we study the susceptible-infected-susceptible epidemic dynamics, considering a specialized setting where popular places (termed passive entities) are visited by agents (termed active entities). We consider two types of spreading dynamics: direct spreading, where the active entities infect each other while visiting the passive entities, and indirect spreading, where the passive entities act as carriers and the infection is spread via them. We investigate in particular the effect of selection strategy, i.e., the way passive entities are chosen, in the spread of epidemics. We introduce a mathematical framework to study the effect of an arbitrary selection strategy and derive formulas for prevalence, extinction probabilities, and epidemic thresholds for both indirect and direct spreading. We also obtain a very simple relationship between the extinction probability and the prevalence. We pay special attention to preferential selection and derive exact formulas. The analysis reveals that an increase in the diversity in the selection process lowers the epidemic thresholds. Comparing the direct and indirect spreading, we identify regions in the parameter space where the prevalence of the indirect spreading is higher than the direct one.

An indecent proposal: the dual functions of indirect speech.

PubMed

Chakroff, Aleksandr; Thomas, Kyle A; Haque, Omar S; Young, Liane

2015-01-01

People often use indirect speech, for example, when trying to bribe a police officer by asking whether there might be "a way to take care of things without all the paperwork." Recent game theoretic accounts suggest that a speaker uses indirect speech to reduce public accountability for socially risky behaviors. The present studies examine a secondary function of indirect speech use: increasing the perceived moral permissibility of an action. Participants report that indirect speech is associated with reduced accountability for unethical behavior, as well as increased moral permissibility and increased likelihood of unethical behavior. Importantly, moral permissibility was a stronger mediator of the effect of indirect speech on likelihood of action, for judgments of one's own versus others' unethical action. In sum, the motorist who bribes the police officer with winks and nudges may not only avoid public punishment but also maintain the sense that his actions are morally permissible. Copyright © 2014 Cognitive Science Society, Inc.

RELATION BETWEEN EPSTEIN-BARR VIRAL AND CELL MEMBRANE IMMUNOFLUORESCENCE OF BURKITT TUMOR CELLS

PubMed Central

Klein, G.; Pearson, G.; Nadkarni, J. S.; Nadkarni, J. J.; Klein, E.; Henle, G.; Henle, W.; Clifford, P.

1968-01-01

A comparison was made of the immunofluorescence tests for detection of cell membrane and Epstein-Barr virus antigens in cells from Burkitt tumor biopsies or continuous cultures derived therefrom. On the whole, cell membrane fluorescence in established lines appeared to depend not only upon the presence of EBV but to a considerable degree also upon the extent of the persistent viral infection. There was no constant relationship, however, between the results of the two tests and exceptions to the rule were noted. These observations indicate that different antigens are involved in the two tests. Biopsy cells in general and young cultures may reveal strong MIF activity but few, if any, EBV-positive cells. The reverse, the presence of relatively large numbers of EBV antigen-containing cells in the absence of significant MIF reactions, was also noted on occasion in a few established cultures. The possible interpretations of these findings have been discussed. PMID:4878906

Indirect techniques in nuclear astrophysics: a review.

PubMed

Tribble, R E; Bertulani, C A; Cognata, M La; Mukhamedzhanov, A M; Spitaleri, C

2014-10-01

In this review, we discuss the present status of three indirect techniques that are used to determine reaction rates for stellar burning processes, asymptotic normalization coefficients, the Trojan Horse method and Coulomb dissociation. A comprehensive review of the theory behind each of these techniques is presented. This is followed by an overview of the experiments that have been carried out using these indirect approaches.

Indirect Plant Defense against Insect Herbivores: A Review

USDA-ARS?s Scientific Manuscript database

Plants respond to herbivore attack by launching two types of defenses: direct defense and indirect defense. Direct defense includes all plant traits that increase the resistance of host plants to attacking insect herbivores by affecting the physiology and/or behavior of the attackers. Indirect defe...

48 CFR 242.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 3 2010-10-01 2010-10-01 false Final indirect cost rates. 242.705 Section 242.705 Federal Acquisition Regulations System DEFENSE ACQUISITION REGULATIONS SYSTEM, DEPARTMENT OF DEFENSE CONTRACT MANAGEMENT CONTRACT ADMINISTRATION AND AUDIT SERVICES Indirect Cost Rates 242...

48 CFR 1542.705 - Final indirect cost rates.

Code of Federal Regulations, 2010 CFR

2010-10-01

.... 1542.705 Section 1542.705 Federal Acquisition Regulations System ENVIRONMENTAL PROTECTION AGENCY CONTRACT MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 1542.705 Final indirect cost rates. (a) The... contracts when the Environmental Protection Agency (EPA) is the cognizant federal agency and a FACO will be...

On the comprehensibility and perceived privacy protection of indirect questioning techniques.

PubMed

Hoffmann, Adrian; Waubert de Puiseau, Berenike; Schmidt, Alexander F; Musch, Jochen

2017-08-01

On surveys that assess sensitive personal attributes, indirect questioning aims at increasing respondents' willingness to answer truthfully by protecting confidentiality. However, the assumption that subjects understand questioning procedures fully and trust them to protect their privacy is rarely tested. In a scenario-based design, we compared four indirect questioning procedures in terms of their comprehensibility and perceived privacy protection. All indirect questioning techniques were found to be less comprehensible by respondents than a conventional direct question used for comparison. Less-educated respondents experienced more difficulties when confronted with any indirect questioning technique. Regardless of education, the crosswise model was found to be the most comprehensible among the four indirect methods. Indirect questioning in general was perceived to increase privacy protection in comparison to a direct question. Unexpectedly, comprehension and perceived privacy protection did not correlate. We recommend assessing these factors separately in future evaluations of indirect questioning.

Gender differentiation in indirect self-destructiveness and suicide attempt methods (gender, indirect self-destructiveness, and suicide attempts).

PubMed

Tsirigotis, Konstantinos; Gruszczyński, Wojciech; Tsirigotis-Maniecka, Marta

2014-06-01

The objective of this study is to examine the gender (sex) differentiation of indirect self-destructiveness and its manifestations as well as its relationships with suicide attempt methods in females and males. The study was conducted among 147 persons (114 females, 33 males) who attempted suicide. The research instrument was the polish version of the Chronic Self-Destructiveness Scale including Transgression and Risk, Poor Health Maintenance, Personal and Social Neglects, Lack of Planfulness, and Helplessness and Passiveness in the face of problems. Differences testing and correlation analyses were applied. Females scored higher on poor health maintenance and males scored significantly higher on personal and social neglects, lack of planfulness, and helplessness. Noteworthy is that the intensity of indirect self-destructiveness in females reached the same magnitude as in males. A number of statistically significant correlations were found between indirect self-destructiveness, or its manifestations, and the methods of suicide attempt in the two groups. Among these categories, the highest contribution was of helplessness and passiveness (both of groups), poor health maintenance (males), and personal and social neglects (females). Results of this study can be useful in the therapeutic efforts and prevention of not only indirectly self-destructive behaviours but also possible suicide attempts. Both preventive and therapeutic activities can take into account the specificity of those phenomena resulting from one's sex/gender. It is important to adapt preventive and therapeutic measures to psychological (personal) features that arise from an individual's sex/gender.

Correlation mapping microscopy

NASA Astrophysics Data System (ADS)

McGrath, James; Alexandrov, Sergey; Owens, Peter; Subhash, Hrebesh M.; Leahy, Martin J.

2015-03-01

Changes in the microcirculation are associated with conditions such as Raynauds disease. Current modalities used to assess the microcirculation such as nailfold capillaroscopy are limited due to their depth ambiguity. A correlation mapping technique was recently developed to extend the capabilities of Optical Coherence Tomography to generate depth resolved images of the microcirculation. Here we present the extension of this technique to microscopy modalities, including confocal microscopy. It is shown that this correlation mapping microscopy technique can extend the capabilities of conventional microscopy to enable mapping of vascular networks in vivo with high spatial resolution.

34 CFR 75.563 - Restricted indirect cost rate-programs covered.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 34 Education 1 2014-07-01 2014-07-01 false Restricted indirect cost rate-programs covered. 75.563 Section 75.563 Education Office of the Secretary, Department of Education DIRECT GRANT PROGRAMS What Conditions Must Be Met by a Grantee? Indirect Cost Rates § 75.563 Restricted indirect cost rate—programs...

The association between reality-based beliefs and indirectly experienced traumatization.

PubMed

Shiri, Shimon; Wexler, Isaiah D; Schwartz, Isabella; Kadari, Michal; Kreitler, Shulamith

2010-12-01

The purpose of the study was to examine the association between belief types and the magnitude of indirect traumatization. Specific types of beliefs were defined in terms of the cognitive orientation theory, which is a cognitive-motivational approach to the understanding, predicting, and changing of behaviors. Belief types that were analyzed included beliefs about self, general beliefs, beliefs about norms, and goal beliefs as they relate to personal growth. Study participants included 38 rescuers (body handlers), 37 nurses, and 31 rehabilitation workers who treated injured civilians that had been exposed to politically motivated violence. The Cognitive Orientation for Posttraumatic Growth Scale was used to assess beliefs about personal growth. The Revised Posttraumatic Stress Disorder Inventory was administered to evaluate indirect traumatization. The results indicate that three of the four belief types related to personal growth were associated with the level of indirect traumatization. Optimistic and positive beliefs about self and general beliefs were associated with a lower level of indirect traumatization symptomatology, suggesting that these types of beliefs may counteract indirect traumatization. On the other hand, stronger goal beliefs were associated with greater indirect traumatization. The negative association between positive goal beliefs and indirect trauma may be related to the gap the individual perceives between the hoped-for ideals and the trauma-stricken reality. These results indicate the importance of cognitive beliefs and their possible role in determining the response to indirect traumatization.

Association of p60c-src with endosomal membranes in mammalian fibroblasts

PubMed Central

1992-01-01

We have examined the subcellular localization of p60c-src in mammalian fibroblasts. Analysis of indirect immunofluorescence by three- dimensional optical sectioning microscopy revealed a granular cytoplasmic staining that co-localized with the microtubule organizing center. Immunofluorescence experiments with antibodies against a number of membrane markers demonstrated a striking co-localization between p60c-src and the cation-dependent mannose-6-phosphate receptor (CI- MPR), a marker that identifies endosomes. Both p60c-src and the CI-MPR were found to cluster at the spindle poles throughout mitosis. In addition, treatment of interphase and mitotic cells with brefeldin A resulted in a clustering of p60c-src and CI-MPR at a peri-centriolar position. Biochemical fractionation of cellular membranes showed that a major proportion of p60c-src co-enriched with endocytic membranes. Treatment of membranes containing HRP to alter their apparent density also altered the density of p60c-src-containing membranes. Similar density shift experiments with total cellular membranes revealed that the majority of membrane-associated p60c-src in the cell is associated with endosomes, while very little is associated with plasma membranes. These results support a role for p60c-src in the regulation of endosomal membranes and protein trafficking. PMID:1378446

Binding of anti-basement membrane antibody to alveolar basement membrane after intratracheal gasoline instillation in rabbits.

PubMed Central

Yamamoto, T.; Wilson, C. B.

1987-01-01

A possible causal relationship has been suggested between hydrocarbon (gasoline, solvents, etc.) exposure and development of anti-basement membrane antibody-associated Goodpasture's syndrome in man. The authors evaluated the effect of hydrocarbons on pulmonary capillary permeability and binding of heterologous anti-basement membrane antibodies in the lungs after intratracheal instillation of minute amounts of unleaded gasoline into rabbits. The anti-glomerular basement membrane (GBM) antibodies used reacted with the alveolar basement membrane (ABM) in vitro by indirect immunofluorescence. The gasoline treatment altered pulmonary capillary permeability, judging from the increased accumulation of systemically administered radioiodinated bovine serum albumin in the alveolar and extravascular spaces of lungs; it also induced focal macroscopic and microscopic pulmonary histologic lesions. The gasoline caused focal in vivo binding of the anti-GBM antibodies to the ABM detectable by immunofluorescence microscopy. No binding was observed in lungs from control rabbits given saline instillations when assayed by immunofluorescence. The paired label radioisotope technique confirmed the increased antibody binding to lungs injured with gasoline (1.08 +/- 0.03 micrograms) versus 0.37 +/- 0.07 microgram after saline (P less than 0.001). These results indicate that gasoline exposure damages a pulmonary barrier that normally prevents binding of anti-GBM/ABM antibody to ABM and suggest that hydrocarbon exposure may be one of perhaps several pneumotoxic events that contribute to the episodic pulmonary hemorrhage in Goodpasture's syndrome by temporarily allowing ABM binding of anti-basement membrane antibodies. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:3548409

Indirect exposure to a family planning mass media campaign in Nepal.

PubMed

Boulay, Marc; Storey, J Douglas; Sood, Suruchi

2002-01-01

It is often noted that some individuals become aware of a mass media program's messages through discussions with other individuals. However, the extent to which indirect exposure occurs, and its influence on behavior, are somewhat unclear. This study examines the role of indirect exposure in extending the reach of a family planning mass media campaign in Nepal. Sociometric data, gathered from nearly all women between the ages of 15 and 49 years living in six villages in Dang District, Nepal (N = 667), assessed indirect exposure to the radio program. Indirect exposure was extensive; half of all respondents were indirectly exposed to the program's messages and the overall reach of the program increased from 50% to 75% when indirect exposure was considered. Members of community groups had higher levels of direct exposure to the radio program and more extensive and diverse social networks, allowing them to serve as a conduit for these messages into the wider community. While direct exposure to the radio program appeared to influence family planning knowledge, indirect exposure was more strongly associated with contraceptive use. These findings suggest that program evaluations that ignore indirect exposure underestimate the impact of a mass media program on behavior.

Mini-titins in striated and smooth molluscan muscles: structure, location and immunological crossreactivity.

PubMed

Vibert, P; Edelstein, S M; Castellani, L; Elliott, B W

1993-12-01

Invertebrate mini-titins are members of a class of myosin-binding proteins belonging to the immunoglobulin superfamily that may have structural and/or regulatory properties. We have isolated mini-titins from three molluscan sources: the striated and smooth adductor muscles of the scallop, and the smooth catch muscles of the mussel. Electron microscopy reveals flexible rod-like molecules about 0.2 micron long and 30 A wide with a distinctive polarity. Antibodies to scallop mini-titin label the A-band and especially the A/I junction of scallop striated muscle myofibrils by indirect immunofluorescence and immuno-electron microscopy. This antibody crossreacts with mini-titins in scallop smooth and Mytilus catch muscles, as well as with proteins in striated muscles from Limulus, Lethocerus (asynchronous flight muscle), and crayfish. It labels the A/I junction (I-region in Lethocerus) in these striated muscles as well as in chicken skeletal muscle. Antibodies to the repetitive immunoglobulin-like regions and also to the kinase domain of nematode twitchin crossreact with scallop mini-titin and label the A-band of scallop myofibrils. Electron microscopy of single molecules shows that antibodies to twitchin kinase bind to scallop mini-titin near one end of the molecule, suggesting how the scallop structure might be aligned with the sequence of nematode twitchin.

The significance of specific IgM antibody in the diagnosis of rubella employing the immunofluorescence technique

PubMed Central

Iwakata, S.; Rhodes, A. J.; Labzoffsky, N. A.

1972-01-01

The applicability of the immunofluorescence (IF) test to the diagnosis of primary rubella infection was investigated. The test is based on the detection of rubella-specific antibodies in the IgM fraction of immunoglobulins. The results indicate the usefulness of the IF test for the diagnosis of primary rubella infection on a single serum specimen collected at a proper time. The test is also of value in the differentiation of primary infection from reinfection, since in reinfection no rubella-specific antibodies are found in the IgM fraction. The test is also valuable for the detection of fetal infection in utero since the persistence of IgM antibodies in pregnant women is indicative of fetal infection. PMID:4551395

Indirect personality assessment of the violent true believer.

PubMed

Meloy, J Reid

2004-04-01

The violent true believer is an individual committed to an ideology or belief system which advances homicide and suicide as a legitimate means to further a particular goal. The author explores useful sources of evidence for an indirect personality assessment of such individuals. He illustrates both idiographic and nomothetic approaches to indirect personality assessment through comparative analyses of Timothy McVeigh, an American who bombed the federal building in Oklahoma City in 1995, and Mohamed Atta, an Egyptian who led the airplane attacks against the World Trade Center and the Pentagon in 2001. The risks of indirect personality assessment and ethical concerns are identified.

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.

Light sheet microscopy.

PubMed

Weber, Michael; Mickoleit, Michaela; Huisken, Jan

2014-01-01

This chapter introduces the concept of light sheet microscopy along with practical advice on how to design and build such an instrument. Selective plane illumination microscopy is presented as an alternative to confocal microscopy due to several superior features such as high-speed full-frame acquisition, minimal phototoxicity, and multiview sample rotation. Based on our experience over the last 10 years, we summarize the key concepts in light sheet microscopy, typical implementations, and successful applications. In particular, sample mounting for long time-lapse imaging and the resulting challenges in data processing are discussed in detail. © 2014 Elsevier Inc. All rights reserved.

The indirect cost of Patient-Specific Instruments.

PubMed

Thienpont, Emmanuel; Paternostre, Frederic; Van Wymeersch, Charles

2015-09-01

To calculate the indirect costs of Patient Specific Instruments (PSI) based on an opportunity cost, cost of efforts and a supply chain cost model to compare PSI for value with conventional total knee arthroplasty (TKA). In 81 patients the total (direct+indirect) cost of PSI-assisted TKA was compared with conventional TKA. Surgical times and coronal mechanical alignment were measured to evaluate the effectiveness of the PSI system. Indirect costs (459 euro) make up 40% of the total cost that can run up to 1142 euro for a patient operated with PSI guides. No difference in surgical times or coronal alignment was observed in between both groups. Considering the total cost of PSI no value was found for the use of PSI in primary TKA as measured by surgical times or for obtaining a neutral mechanical axis in the coronal plane.

Characterization of specific anti-Candida IgM, IgA and IgE: diagnostic value in deep-seated infections.

PubMed

Aubert, D; Puygauthier-Toubas, D; Leon, P; Pignon, B; Foudrinier, F; Marnef, F; Boulant, J; Pinon, J M

1996-01-01

The proposed serological diagnosis of systemic Candida infections is based on a microplate immunocapture technique detecting IgM, IgA and IgE anti-Candida antibodies. Activity is revealed with a suspension of human erythrocytes sensitized with somatic antigen of Candida albicans, and is quantified on an automated plate reader. The sera were obtained from patients with deep-seated (n = 56) and superficial (n = 193) candidosis. We compared this immunological method with a combination of indirect immunofluorescence and co-immunoelectrodiffusion. The immunocapture method was more sensitive (80.4% vs. 48.2% with indirect immunofluorescence and 58.9% with co-immunoelectrodiffusion), and often provided the diagnosis at an earlier stage, with clear therapeutic advantages. The IgA isotype was a particularly valuable marker of deep-seated Candida infections.

New approaches in renal microscopy: volumetric imaging and superresolution microscopy.

PubMed

Kim, Alfred H J; Suleiman, Hani; Shaw, Andrey S

2016-05-01

Histologic and electron microscopic analysis of the kidney has provided tremendous insight into structures such as the glomerulus and nephron. Recent advances in imaging, such as deep volumetric approaches and superresolution microscopy, have the capacity to dramatically enhance our current understanding of the structure and function of the kidney. Volumetric imaging can generate images millimeters below the surface of the intact kidney. Superresolution microscopy breaks the diffraction barrier inherent in traditional light microscopy, enabling the visualization of fine structures. Here, we describe new approaches to deep volumetric and superresolution microscopy of the kidney. Rapid advances in lasers, microscopic objectives, and tissue preparation have transformed our ability to deep volumetric image the kidney. Innovations in sample preparation have allowed for superresolution imaging with electron microscopy correlation, providing unprecedented insight into the structures within the glomerulus. Technological advances in imaging have revolutionized our capacity to image both large volumes of tissue and the finest structural details of a cell. These new advances have the potential to provide additional profound observations into the normal and pathologic functions of the kidney.

Analysis of replication factories in human cells by super-resolution light microscopy

PubMed Central

2009-01-01

Background DNA replication in human cells is performed in discrete sub-nuclear locations known as replication foci or factories. These factories form in the nucleus during S phase and are sites of DNA synthesis and high local concentrations of enzymes required for chromatin replication. Why these structures are required, and how they are organised internally has yet to be identified. It has been difficult to analyse the structure of these factories as they are small in size and thus below the resolution limit of the standard confocal microscope. We have used stimulated emission depletion (STED) microscopy, which improves on the resolving power of the confocal microscope, to probe the structure of these factories at sub-diffraction limit resolution. Results Using immunofluorescent imaging of PCNA (proliferating cell nuclear antigen) and RPA (replication protein A) we show that factories are smaller in size (approximately 150 nm diameter), and greater in number (up to 1400 in an early S- phase nucleus), than is determined by confocal imaging. The replication inhibitor hydroxyurea caused an approximately 40% reduction in number and a 30% increase in diameter of replication factories, changes that were not clearly identified by standard confocal imaging. Conclusions These measurements for replication factory size now approach the dimensions suggested by electron microscopy. This agreement between these two methods, that use very different sample preparation and imaging conditions, suggests that we have arrived at a true measurement for the size of these structures. The number of individual factories present in a single nucleus that we measure using this system is greater than has been previously reported. This analysis therefore suggests that each replication factory contains fewer active replication forks than previously envisaged. PMID:20015367

[Direct and indirect somatic embryogenesis in Freesia refracta].

PubMed

Wang, L; Duan, X G; Hao, S

1999-06-01

Somatic embryogenesis can be induced in tissue cultures of Freesia refracta either directly from the epidermal cells of explant, or indirectly via intervening callus. In direct pathway, somatic embryos were in contact with maternal tissue in a suspensor-like structure. In indirect pathway, the explants first proliferacted to give rise to calluses before embryoids were induced. The two sorts of calluses were defined to embryogenic callus and non-embryogenic callus according to producing of somatic embryos. An indirect somatic embryo is developed from a pre-embryogenically determined cell. This kind of somatic embryo has no suspensor structure instead of a complex with maternal tissue. Somatic embryos have their own vascular tissues, and can develop new plantlets independently.

Theoretical and testing performance of an innovative indirect evaporative chiller

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

Jiang, Yi; Xie, Xiaoyun

2010-12-15

An indirect evaporative chiller is a device used to produce chilled water at a temperature between the wet bulb temperature and dew point of the outdoor air, which can be used in building HVAC systems. This article presents a theoretical analysis and practical performance of an innovative indirect evaporative chiller. First, the process of the indirect evaporative chiller is introduced; then, the matching characteristics of the process are presented and analyzed. It can be shown that the process that produces cold water by using dry air is a nearly-reversible process, so the ideal produced chilled water temperature of the indirectmore » evaporative chiller can be set close to the dew point temperature of the chiller's inlet air. After the indirect evaporative chiller was designed, simulations were done to analyze the output water temperature, the cooling efficiency relative to the inlet dew point temperature, and the COP that the chiller can performance. The first installation of the indirect evaporative chiller of this kind has been run for 5 years in a building in the city of Shihezi. The tested output water temperature of the chiller is around 14-20 C, which is just in between of the outdoor wet bulb temperature and dew point. The tested COP{sub r,s} of the developed indirect evaporative chiller reaches 9.1. Compared with ordinary air conditioning systems, the indirect evaporative chiller can save more than 40% in energy consumption due to the fact that the only energy consumed is from pumps and fans. An added bonus is that the indirect evaporative chiller uses no CFCs that pollute to the aerosphere. The tested internal parameters, such as the water-air flow rate ratio and heat transfer area for each heat transfer process inside the chiller, were analyzed and compared with designed values. The tested indoor air conditions, with a room temperature of 23-27 C and relative humidity of 50-70%, proved that the developed practical indirect evaporative chiller

Paranodal myelin retraction in relapsing experimental autoimmune encephalomyelitis visualized by coherent anti-Stokes Raman scattering microscopy

NASA Astrophysics Data System (ADS)

Fu, Yan; Frederick, Terra J.; Huff, Terry B.; Goings, Gwendolyn E.; Miller, Stephen D.; Cheng, Ji-Xin

2011-10-01

How demyelination is initiated is a standing question for pathology of multiple sclerosis. By label-free coherent anti-Stokes Raman scattering (CARS) imaging of myelin lipids, we investigate myelin integrity in the lumbar spinal cord tissue isolated from naïve SJL mice, and from mice at the onset, peak acute, and remission stages of relapsing experimental autoimmune encephalomyelitis (EAE). Progressive demyelinating disease is initially characterized by the retraction of paranodal myelin both at the onset of disease and at the borders of acute demyelinating lesions. Myelin retraction is confirmed by elongated distribution of neurofascin proteins visualized by immunofluorescence. The disruption of paranodal myelin subsequently exposes Kv1.2 channels at the juxtaparanodes and lead to the displacement of Kv1.2 channels to the paranodal and nodal domains. Paranodal myelin is partially restored during disease remission, indicating spontaneous myelin regeneration. These findings suggest that paranodal domain injury precedes formation of internodal demyelinating lesions in relapsing EAE. Our results also demonstrate that CARS microscopy is an effective readout of myelin disease burden.

Rapid determination of myosin heavy chain expression in rat, mouse, and human skeletal muscle using multicolor immunofluorescence analysis.

PubMed

Bloemberg, Darin; Quadrilatero, Joe

2012-01-01

Skeletal muscle is a heterogeneous tissue comprised of fibers with different morphological, functional, and metabolic properties. Different muscles contain varying proportions of fiber types; therefore, accurate identification is important. A number of histochemical methods are used to determine muscle fiber type; however, these techniques have several disadvantages. Immunofluorescence analysis is a sensitive method that allows for simultaneous evaluation of multiple MHC isoforms on a large number of fibers on a single cross-section, and offers a more precise means of identifying fiber types. In this investigation we characterized pure and hybrid fiber type distribution in 10 rat and 10 mouse skeletal muscles, as well as human vastus lateralis (VL) using multicolor immunofluorescence analysis. In addition, we determined fiber type-specific cross-sectional area (CSA), succinate dehydrogenase (SDH) activity, and α-glycerophosphate dehydrogenase (GPD) activity. Using this procedure we were able to easily identify pure and hybrid fiber populations in rat, mouse, and human muscle. Hybrid fibers were identified in all species and made up a significant portion of the total population in some rat and mouse muscles. For example, rat mixed gastrocnemius (MG) contained 12.2% hybrid fibers whereas mouse white tibialis anterior (WTA) contained 12.1% hybrid fibers. Collectively, we outline a simple and time-efficient method for determining MHC expression in skeletal muscle of multiple species. In addition, we provide a useful resource of the pure and hybrid fiber type distribution, fiber CSA, and relative fiber type-specific SDH and GPD activity in a number of rat and mouse muscles.

Cold Regions Test of Indirect Fire Weapons Ammunition

DTIC Science & Technology

1983-03-08

COLD REGIONS TEST OF INDIRECT FIRE WEAPONS AMMUNITION Paragraph 1 . SCOPE. 1 2. FACILITIES AND INSTRUMENTATION .......... 3. PREPARATION FOR TEST...A- 1 B. Data Collection Sheets ..... .............. B- 1 C. References ..... .................... ... C- 1 D. Cold-Dry...Uniform .D...... .. ... .. ... 0- 1 1 . SCOPE. The procedures outlined in this TOP are designed to determine the c-h-arac-teristics of indirect artillery

Indirect methods for reference interval determination - review and recommendations.

PubMed

Jones, Graham R D; Haeckel, Rainer; Loh, Tze Ping; Sikaris, Ken; Streichert, Thomas; Katayev, Alex; Barth, Julian H; Ozarda, Yesim

2018-04-19

Reference intervals are a vital part of the information supplied by clinical laboratories to support interpretation of numerical pathology results such as are produced in clinical chemistry and hematology laboratories. The traditional method for establishing reference intervals, known as the direct approach, is based on collecting samples from members of a preselected reference population, making the measurements and then determining the intervals. An alternative approach is to perform analysis of results generated as part of routine pathology testing and using appropriate statistical techniques to determine reference intervals. This is known as the indirect approach. This paper from a working group of the International Federation of Clinical Chemistry (IFCC) Committee on Reference Intervals and Decision Limits (C-RIDL) aims to summarize current thinking on indirect approaches to reference intervals. The indirect approach has some major potential advantages compared with direct methods. The processes are faster, cheaper and do not involve patient inconvenience, discomfort or the risks associated with generating new patient health information. Indirect methods also use the same preanalytical and analytical techniques used for patient management and can provide very large numbers for assessment. Limitations to the indirect methods include possible effects of diseased subpopulations on the derived interval. The IFCC C-RIDL aims to encourage the use of indirect methods to establish and verify reference intervals, to promote publication of such intervals with clear explanation of the process used and also to support the development of improved statistical techniques for these studies.

Preschool Children’s Behavioral Tendency toward Social Indirect Reciprocity

PubMed Central

Kanazawa, Tadahiro; Hinobayashi, Toshihiko

2013-01-01

Social indirect reciprocity seems to be crucial in enabling large-scale cooperative networks among genetically unrelated individuals in humans. However, there are relatively few studies on social indirect reciprocity in children compared to adults. Investigating whether young children have a behavioral tendency toward social indirect reciprocity will help us understand how and when the fundamental ability to form cooperative relationships among adults is acquired. Using naturalistic observation at a nursery school, this study examined whether 5- to 6-year-olds show a behavioral tendency to engage in social indirect reciprocity in response to their peers’ prosocial behavior toward a third party. The results revealed that bystander children tended to display prosocial behavior toward their peers more frequently after observing these peers’ prosocial behavior toward third-party peers, compared with control situations; this suggests that 5- to 6-year-olds may have an essential behavioral tendency to establish social indirect reciprocity when interacting with peers in their daily lives. In addition, bystanders tended to display affiliative behavior after observing focal children’s prosocial behavior. In other words, observing peers’ prosocial behavior toward third-party peers evoked bystanders’ positive emotions toward the helpers. Considering both the present results and previous findings, we speculate that in preschoolers, such positive emotions might mediate the increase in the bystander’s prosocial behavior toward the helper. In addition, an intuitional emotional process plays an important role in the preschooler’s behavioral tendency toward social indirect reciprocity in natural interactions with peers. PMID:23951040

Microscopy and microanalysis 1996

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

Bailey, G.W.; Corbett, J.M.; Dimlich, R.V.W.

1996-12-31

The Proceedings of this Annual Meeting contain paper of members from the three societies. These proceedings emphasizes the common research interests and attempts to eliminate some unwanted overlap. Topics covered are: microscopic analysis of animals with altered gene expression and in-situ gene and antibody localizations, high-resolution elemental mapping of nucleoprofein interactions, plant biology and pathology, quantitative HREM analysis of perfect and defected materials, computational methods for TEM image analysis, high-resolution FESM in materials research, frontiers in polymer microscopy and microanalysis, oxidation and corrosion, micro XRD and XRF, molecular microspectroscopy and spectral imaging, advances in confocal and multidimensional light microscopy, analyticalmore » electron microscopy in biology, correlative microscopy in biological sciences, grain-boundary microengineering, surfaces and interfaces, telepresence microscopy in education and research, MSA educational outreach, quantitative electron probe microanalysis, frontiers of analytical electron microscopy, critical issues in ceramic microstructures, dynamic organization of the cell, pathology, microbiology, high-resolution biological and cryo SEM, and scanning-probe microscopy.« less

Visualization and quantification of GPCR trafficking in mammalian cells by confocal microscopy.

PubMed

Nooh, Mohammed M; Bahouth, Suleiman W

2017-01-01

G protein-coupled receptors (GPCRs) are recognized as one of the most fruitful group of therapeutic targets, accounting for more than 40% of all approved pharmaceuticals on the market. Therefore, the search for selective agents that affect GPCR function is of major interest to the pharmaceutical industry. This chapter describes methods for measuring agonist-promoted GPCR trafficking, which involves the internalization of the GPCR and its subsequent recycling back to the plasma membrane or retention and eventual degradation. These pathways will be analyzed by confocal cellular imaging, using the β 1 -adrenergic receptor (β 1 -AR) as a primary model. A major problem encountered in studying GPCR trafficking is the unavailability of antibodies that would recognize the native receptor in cells or tissues. Therefore, wild-type, point mutants, and β 1 -AR chimeras are generated as epitope-tagged proteins, which are stably- or transiently expressed in mammalian cells. GPCR are labeled with a fluorophore-conjugated antibody directed against the N-terminal epitope tag. The trafficking of the fluorophore-tagged GPCR between divergent trafficking pathways that result in retention and eventual degradation or recycling and reinsertion into the plasma membrane can be followed by confocal immunofluorescence microscopy techniques outlined in this review. © 2017 Elsevier Inc. All rights reserved.

Automobile Industry Retail Price Equivalent and Indirect Cost Multipliers

EPA Science Inventory

This report develops a modified multiplier, referred to as an indirect cost (IC) multiplier, which specifically evaluates the components of indirect costs that are likely to be affected by vehicle modifications associated with environmental regulation. A range of IC multipliers a...

Miniscrew biomechanics: Guidelines for the use of rigid indirect anchorage mechanics.

PubMed

Baumgaertel, Sebastian; Jones, Clark L; Unal, Mustafa

2017-09-01

Indirect anchorage is an established form of anchorage provided by orthodontic miniscrews. Although there are different ways to set up the mechanics, rigid indirect anchorage offers the greatest biomechanical versatility but is more difficult to install than conventional, nonrigid indirect anchorage or direct anchorage. The purpose of this article was to introduce readers to the concept of rigid indirect anchorage and provide guidelines as to its use. Copyright © 2017 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.

Graphene-enabled electron microscopy and correlated super-resolution microscopy of wet cells.

PubMed

Wojcik, Michal; Hauser, Margaret; Li, Wan; Moon, Seonah; Xu, Ke

2015-06-11

The application of electron microscopy to hydrated biological samples has been limited by high-vacuum operating conditions. Traditional methods utilize harsh and laborious sample dehydration procedures, often leading to structural artefacts and creating difficulties for correlating results with high-resolution fluorescence microscopy. Here, we utilize graphene, a single-atom-thick carbon meshwork, as the thinnest possible impermeable and conductive membrane to protect animal cells from vacuum, thus enabling high-resolution electron microscopy of wet and untreated whole cells with exceptional ease. Our approach further allows for facile correlative super-resolution and electron microscopy of wet cells directly on the culturing substrate. In particular, individual cytoskeletal actin filaments are resolved in hydrated samples through electron microscopy and well correlated with super-resolution results.

Lower molar uprighting with miniscrew anchorage: direct and indirect anchorage.

PubMed

Melo, Ana Cláudia Moreira; Duarte da Silva, Ricarda; Shimizu, Roberto Hideo; Campos, Dolores; Andrighetto, Augusto Ricardo

2013-01-01

Lower molar uprighting has been increasingly indicated in clinical orthodontics, mainly because of adult patients with rehabilitation needs. The aim of this study was to determine the success rate of miniscrew use for lower molar uprighting and to compare the use of direct and indirect anchorage. One hundred and eighty-one miniscrews were inserted in 102 rehabilitation patients, with a mean age of 42.24 years. In 71 patients, indirect anchorage was used (116 miniscrews); in the other 31 patients, direct anchorage was employed (65 miniscrews). The choice of direct or indirect anchorage was determined by the orthodontist according to the force system that would be used for the movement and the bone availability. The overall success rate for miniscrews was 90. 05%; 18 miniscrews failed, including 15 that were used as indirect anchorage and 3 that were used as direct anchorage. Considering the results of this study, it can be concluded that both direct and indirect anchorage can be successfully used for molar uprighting.

Inconsistency between direct and indirect comparisons of competing interventions: meta-epidemiological study.

PubMed

Song, Fujian; Xiong, Tengbin; Parekh-Bhurke, Sheetal; Loke, Yoon K; Sutton, Alex J; Eastwood, Alison J; Holland, Richard; Chen, Yen-Fu; Glenny, Anne-Marie; Deeks, Jonathan J; Altman, Doug G

2011-08-16

To investigate the agreement between direct and indirect comparisons of competing healthcare interventions. Meta-epidemiological study based on sample of meta-analyses of randomised controlled trials. Data sources Cochrane Database of Systematic Reviews and PubMed. Inclusion criteria Systematic reviews that provided sufficient data for both direct comparison and independent indirect comparisons of two interventions on the basis of a common comparator and in which the odds ratio could be used as the outcome statistic. Inconsistency measured by the difference in the log odds ratio between the direct and indirect methods. The study included 112 independent trial networks (including 1552 trials with 478,775 patients in total) that allowed both direct and indirect comparison of two interventions. Indirect comparison had already been explicitly done in only 13 of the 85 Cochrane reviews included. The inconsistency between the direct and indirect comparison was statistically significant in 16 cases (14%, 95% confidence interval 9% to 22%). The statistically significant inconsistency was associated with fewer trials, subjectively assessed outcomes, and statistically significant effects of treatment in either direct or indirect comparisons. Owing to considerable inconsistency, many (14/39) of the statistically significant effects by direct comparison became non-significant when the direct and indirect estimates were combined. Significant inconsistency between direct and indirect comparisons may be more prevalent than previously observed. Direct and indirect estimates should be combined in mixed treatment comparisons only after adequate assessment of the consistency of the evidence.

42 CFR 412.322 - Indirect medical education adjustment factor.

Code of Federal Regulations, 2010 CFR

2010-10-01

... Capital-Related Costs § 412.322 Indirect medical education adjustment factor. (a) Basic data. CMS...] Determination of Transition Period Payment Rates for Capital-Related Costs ... 42 Public Health 2 2010-10-01 2010-10-01 false Indirect medical education adjustment factor. 412...

48 CFR 742.770 - Negotiated indirect cost rate agreement.

Code of Federal Regulations, 2011 CFR

2011-10-01

... rate agreement. 742.770 Section 742.770 Federal Acquisition Regulations System AGENCY FOR INTERNATIONAL DEVELOPMENT CONTRACT MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 742.770 Negotiated indirect cost... Agreement is automatically incorporated in each contract between the parties and shall specify: (a) The...

48 CFR 742.770 - Negotiated indirect cost rate agreement.

Code of Federal Regulations, 2010 CFR

2010-10-01

... rate agreement. 742.770 Section 742.770 Federal Acquisition Regulations System AGENCY FOR INTERNATIONAL DEVELOPMENT CONTRACT MANAGEMENT CONTRACT ADMINISTRATION Indirect Cost Rates 742.770 Negotiated indirect cost... Agreement is automatically incorporated in each contract between the parties and shall specify: (a) The...

Superresolution microscopy for microbiology

PubMed Central

Coltharp, Carla; Xiao, Jie

2014-01-01

Summary This review provides a practical introduction to superresolution microscopy from the perspective of microbiological research. Because of the small sizes of bacterial cells, superresolution methods are particularly powerful and suitable for revealing details of cellular structures that are not resolvable under conventional fluorescence light microscopy. Here we describe the methodological concepts behind three major categories of super-resolution light microscopy: photoactivated localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM), structured illumination microscopy (SIM) and stimulated emission-depletion (STED) microscopy. We then present recent applications of each of these techniques to microbial systems, which have revealed novel conformations of cellular structures and described new properties of in vivo protein function and interactions. Finally, we discuss the unique issues related to implementing each of these superresolution techniques with bacterial specimens and suggest avenues for future development. The goal of this review is to provide the necessary technical background for interested microbiologists to choose the appropriate super-resolution method for their biological systems, and to introduce the practical considerations required for designing and analysing superresolution imaging experiments. PMID:22947061

Foodborne Protozoans

USDA-ARS?s Scientific Manuscript database

Identification of the human pathogens Cryptosporidium and Giardia can be grouped into general morphology by microscopy, chemical and immunofluorescent staining methods aiding microscopy, and biochemical and molecular tests. Microscopic observations can be made using brightfield with or without spec...

Immunofluorescent localization of ubiquitin and proteasomes in nucleolar vacuoles of soybean root meristematic cells

PubMed Central

Stępiński, D.

2012-01-01

In this study, using the immunofluorescent method, the immunopositive signals to ubiquitin and proteasomes in nucleoli of root meristematic cells of soybean seedlings have been observed. In fact, those signals were present exclusively in nucleolar vacuoles. No signals were observed in the nucleolar territory out of the nucleolar vacuoles or in the nucleoli without vacuoles. The ubiquitin-proteasome system (UPS) may act within the nucleoli of plants with high metabolic activities and may provide an additional level of regulation of intracellular proteolysis via compartment-specific activities of their components. It is suggested that the presence of the UPS solely in vacuolated nucleoli serves as a mechanism that enhances the speed of ribosome subunit production in very actively transcribing nucleoli. On the other hand, nucleolar vacuoles in a cell/nucleus could play additional roles associated with temporary sequestration or storage of some cellular factors, including components of the ubiquitin-proteasome system. PMID:22688294

Indirect plant defense against insect herbivores: a review.

PubMed

Aljbory, Zainab; Chen, Ming-Shun

2018-02-01

Plants respond to herbivore attack by launching 2 types of defenses: direct defense and indirect defense. Direct defense includes all plant traits that increase the resistance of host plants to insect herbivores by affecting the physiology and/or behavior of the attackers. Indirect defense includes all traits that by themselves do not have significant direct impact on the attacking herbivores, but can attract natural enemies of the herbivores and thus reduce plant loss. When plants recognize herbivore-associated elicitors, they produce and release a blend of volatiles that can attract predators, parasites, and other natural enemies. Known herbivore-associated elicitors include fatty acid-amino acid conjugates, sulfur-containing fatty acids, fragments of cell walls, peptides, esters, and enzymes. Identified plant volatiles include terpenes, nitrogenous compounds, and indoles. In addition, constitive traits including extrafloral nectars, food bodies, and domatia can be further induced to higher levels and attract natural enemies as well as provide food and shelter to carnivores. A better understanding of indirect plant defense at global and componential levels via advanced high throughput technologies may lead to utilization of indirect defense in suppression of herbivore damage to plants. © 2016 Institute of Zoology, Chinese Academy of Sciences.

The Direct-Indirect Technique for Composite Restorations Revisited.

PubMed

Ritter, André V; Fahl, Newton; Vargas, Marcos; Maia, Rodrigo R

2017-06-01

In the direct-indirect composite technique, composite is applied to a nonretentive tooth preparation (eg, a noncarious cervical lesion or a veneer/inlay/onlay preparation) without any bonding agent, sculpted to a primary anatomic form, and light-cured. The partially polymerized restoration is then removed from the preparation and finished and tempered extraorally chairside. The finished inlay is bonded to the preparation using a resin-based luting agent. Advantages of this technique include enhanced physical and mechanical properties afforded by the extraoral chairside tempering process because of increased monomer conversion, and greater operator control over the final marginal adaptation, surface finishing and polishing, and anatomy of the restoration, given that these elements are defined outside of the patient's mouth. The direct-indirect approach also affords enhanced gingival health and patient comfort. This article presents a clinical case in which the direct-indirect composite technique was used to restore three noncarious cervical lesions on the same quadrant on an adult patient. Clinical steps and tips for success are offered. The authors also present scanning electron microscope and atomic force microscope images showing the excellent marginal fit obtained with the direct-indirect composite technique.

Indirect scaling methods for testing quantitative emotion theories.

PubMed

Junge, Martin; Reisenzein, Rainer

2013-01-01

Two studies investigated the utility of indirect scaling methods, based on graded pair comparisons, for the testing of quantitative emotion theories. In Study 1, we measured the intensity of relief and disappointment caused by lottery outcomes, and in Study 2, the intensity of disgust evoked by pictures, using both direct intensity ratings and graded pair comparisons. The stimuli were systematically constructed to reflect variables expected to influence the intensity of the emotions according to theoretical models of relief/disappointment and disgust, respectively. Two probabilistic scaling methods were used to estimate scale values from the pair comparison judgements: Additive functional measurement (AFM) and maximum likelihood difference scaling (MLDS). The emotion models were fitted to the direct and indirect intensity measurements using nonlinear regression (Study 1) and analysis of variance (Study 2). Both studies found substantially improved fits of the emotion models for the indirectly determined emotion intensities, with their advantage being evident particularly at the level of individual participants. The results suggest that indirect scaling methods yield more precise measurements of emotion intensity than rating scales and thereby provide stronger tests of emotion theories in general and quantitative emotion theories in particular.

Inconsistency between direct and indirect comparisons of competing interventions: meta-epidemiological study

PubMed Central

Xiong, Tengbin; Parekh-Bhurke, Sheetal; Loke, Yoon K; Sutton, Alex J; Eastwood, Alison J; Holland, Richard; Chen, Yen-Fu; Glenny, Anne-Marie; Deeks, Jonathan J; Altman, Doug G

2011-01-01

Objective To investigate the agreement between direct and indirect comparisons of competing healthcare interventions. Design Meta-epidemiological study based on sample of meta-analyses of randomised controlled trials. Data sources Cochrane Database of Systematic Reviews and PubMed. Inclusion criteria Systematic reviews that provided sufficient data for both direct comparison and independent indirect comparisons of two interventions on the basis of a common comparator and in which the odds ratio could be used as the outcome statistic. Main outcome measure Inconsistency measured by the difference in the log odds ratio between the direct and indirect methods. Results The study included 112 independent trial networks (including 1552 trials with 478 775 patients in total) that allowed both direct and indirect comparison of two interventions. Indirect comparison had already been explicitly done in only 13 of the 85 Cochrane reviews included. The inconsistency between the direct and indirect comparison was statistically significant in 16 cases (14%, 95% confidence interval 9% to 22%). The statistically significant inconsistency was associated with fewer trials, subjectively assessed outcomes, and statistically significant effects of treatment in either direct or indirect comparisons. Owing to considerable inconsistency, many (14/39) of the statistically significant effects by direct comparison became non-significant when the direct and indirect estimates were combined. Conclusions Significant inconsistency between direct and indirect comparisons may be more prevalent than previously observed. Direct and indirect estimates should be combined in mixed treatment comparisons only after adequate assessment of the consistency of the evidence. PMID:21846695

Manifestations of Indirect Self-Destructiveness and Dimensions of Emotional Intelligence.

PubMed

Tsirigotis, Konstantinos; Łuczak, Joanna

2016-09-01

While indirect self-destructiveness exerts a rather negative influence on the life and psychological and social functioning of the individual, emotional intelligence may have a favourable effect. The aim of this study has been to explore possible relationships between manifestations of indirect self-destructiveness and dimensions of emotional intelligence. A population of 260 individuals (130 females and 130 males) aged 20-30 (mean age of 24.5) was studied by using the Polish version of the Chronic Self-Destructiveness Scale and INTE, i.e., the Polish version of the Assessing Emotions Scale. Manifestations of indirect self-destructiveness show many significant correlations with variables of the INTE, and those correlations are negative. Generally, it can be said that low emotional intelligence is associated with poor psychosocial and social functioning, which, in turn, is associated with indirect self-destructiveness and its manifestations. It seems advisable to use emotional intelligence in the prophylactic and therapeutic work with individuals suffering from various types of disorders, especially the syndrome of indirect self-destructiveness.

Development and performance evaluation of a novel immunofluorescence chromatographic assay for histidine-rich protein 2 of Plasmodium falciparum.

PubMed

Kang, Keren; Dzakah, Emmanuel E; Huang, Yongping; Xie, Mingquan; Luo, Xiaochun; Li, Wenmei; Wang, Jihua

2015-05-30

The low sensitivity and specificity of Plasmodium falciparum diagnostic tests pose a serious health threat to people living in endemic areas. The objective of the study was to develop a rapid assay for the detection of histidine-rich protein 2 (HRP2) of P. falciparum in whole blood by immunofluorescence chromatographic technology. A total of 1163 positive and negative blood samples were screened. The double-antibody sandwich assay was used to establish the kit and its performance was evaluated for sensitivity, specificity, accuracy, precision, stability, and clinical effectiveness. The cut-off level of detection of the kit was 25 parasites/μl. Common interfering substances in human blood specimens, such as bilirubin, triglyceride and cholesterol had no significant effect on HRP2 antigen detection. The precision of the kit was run with different concentration of standard calibrators and the values were less than 10 %. The performance of this diagnostic kit in the detection of the calibrators has shown that a shelf life of about 12 months gives a more reliable result. Among clinical samples tested, the HRP2 test kit and the reference products had good coincidence rate in a parallel experiment and this test kit had a more sensitive detecting level to the target protein than the reference kits used in this study. The specificity and sensitivity for this test were 99.6 % (800/803) and 99.7 % (1160/1163), respectively. A novel HRP2 immunofluorescence detection method was developed in this study. Overall performance evaluation indicated that the kit has a rapid, high sensitivity and on-spot method for detecting P. falciparum.

Direct and indirect pathways for choosing objects and actions.

PubMed

Hikosaka, Okihide; Kim, Hyoung F; Amita, Hidetoshi; Yasuda, Masaharu; Isoda, Masaki; Tachibana, Yoshihisa; Yoshida, Atsushi

2018-02-23

A prominent target of the basal ganglia is the superior colliculus (SC) which controls gaze orientation (saccadic eye movement in primates) to an important object. This 'object choice' is crucial for choosing an action on the object. SC is innervated by the substantia nigra pars reticulata (SNr) which is controlled mainly by the caudate nucleus (CD). This CD-SNr-SC circuit is sensitive to the values of individual objects and facilitates saccades to good objects. The object values are processed differently in two parallel circuits: flexibly by the caudate head (CDh) and stably by the caudate tail (CDt). To choose good objects, we need to reject bad objects. In fact, these contrasting functions are accomplished by the circuit originating from CDt: The direct pathway focuses on good objects and facilitates saccades to them; the indirect pathway focuses on bad objects and suppresses saccades to them. Inactivation of CDt deteriorated the object choice, because saccades to bad objects were no longer suppressed. This suggests that the indirect pathway is important for object choice. However, the direct and indirect pathways for 'object choice', which aim at the same action (i.e., saccade), may not work for 'action choice'. One possibility is that circuits controlling different actions are connected through the indirect pathway. Additional connections of the indirect pathway with brain areas outside the basal ganglia may also provide a wider range of behavioral choice. In conclusion, basal ganglia circuits are composed of the basic direct/indirect pathways and additional connections and thus have acquired multiple functions. © 2018 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Heard it through the grapevine: indirect networks and employee creativity.

PubMed

Hirst, Giles; Van Knippenberg, Daan; Zhou, Jing; Quintane, Eric; Zhu, Cherrie

2015-03-01

Social networks can be important sources of information and insights that may spark employee creativity. The cross-fertilization of ideas depends not just on access to information and insights through one's direct network-the people one actually interacts with--but at least as much on access to the indirect network one's direct ties connect one to (i.e., people one does not interact with directly, but with whom one's direct ties interact). We propose that the reach efficiency of this indirect network--its nonredundancy in terms of interconnections--is positively related to individual creativity. To help specify the boundaries of this positive influence of the indirect network, we also explore how many steps removed the indirect network still adds to creativity. In addition, we propose that the efficiency (nonredundancy) of one's direct network is important here, because more efficient direct networks give one access to indirect networks with greater reach efficiency. Our hypotheses were supported in a multilevel analysis of multisource survey data from 223 sales representatives nested within 11 divisions of a Chinese pharmaceutical company. This analysis also showed that the creative benefits of reach efficiency were evident for 3 and 4 degrees of separation but were greatest for indirect ties that depend only on one's direct ties. PsycINFO Database Record (c) 2015 APA, all rights reserved.

Sleep-wake time perception varies by direct or indirect query.

PubMed

Alameddine, Y; Ellenbogen, J M; Bianchi, M T

2015-01-15

The diagnosis of insomnia rests on self-report of difficulty initiating or maintaining sleep. However, subjective reports may be unreliable, and possibly may vary by the method of inquiry. We investigated this possibility by comparing within-individual response to direct versus indirect time queries after overnight polysomnography. We obtained self-reported sleep-wake times via morning questionnaires in 879 consecutive adult diagnostic polysomnograms. Responses were compared within subjects (direct versus indirect query) and across groups defined by apnea-hypopnea index and by self-reported insomnia symptoms in pre-sleep questionnaires. Direct queries required a time duration response, while indirect queries required clock times from which we calculated time durations. Direct and indirect queries of sleep latency were the same in only 41% of cases, and total sleep time queries matched in only 5.4%. For both latency and total sleep, the most common discrepancy involved the indirect value being larger than the direct response. The discrepancy between direct and indirect queries was not related to objective sleep metrics. The degree of discrepancy was not related to the presence of insomnia symptoms, although patients reporting insomnia symptoms showed underestimation of total sleep duration by direct response. Self-reported sleep latency and total sleep time are often internally inconsistent when comparing direct and indirect survey queries of each measure. These discrepancies represent substantive challenges to effective clinical practice, particularly when diagnosis and management depends on self-reported sleep patterns, as with insomnia. Although self-reported sleep-wake times remains fundamental to clinical practice, objective measures provide clinically relevant adjunctive information. © 2015 American Academy of Sleep Medicine.

Male and Female University Students' Experiences of Indirect Aggression

ERIC Educational Resources Information Center

Leenaars, Lindsey; Rinaldi, Christina M.

2010-01-01

This study examines the role of sex, gender role orientation, social representations of indirect aggression, and indicators of psychosocial adjustment in indirect aggression and victimization in an emerging adult sample. A total of 42 participants (19 men, 23 women) recruited are required to complete the questionnaires, along with 18 participants…

Translocation of an 89-kDa periplasmic protein is associated with Holospora infection

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

Iwatani, Koichi; Dohra, Hideo; Lang, B. Franz

2005-12-02

The symbiotic bacterium Holospora obtusa infects the macronucleus of the ciliate Paramecium caudatum. After ingestion by its host, an infectious form of Holospora with an electron-translucent tip passes through the host digestive vacuole and penetrates the macronuclear envelope with this tip. To investigate the underlying molecular mechanism of this process, we raised a monoclonal antibody against the tip-specific 89-kDa protein, sequenced this partially, and identified the corresponding complete gene. The deduced protein sequence carries two actin-binding motifs. Indirect immunofluorescence microscopy shows that during escape from the host digestive vacuole, the 89-kDa proteins translocates from the inside to the outside ofmore » the tip. When the bacterium invades the macronucleus, the 89-kDa protein is left behind at the entry point of the nuclear envelope. Transmission electron microscopy shows the formation of fine fibrous structures that co-localize with the antibody-labeled regions of the bacterium. Our findings suggest that the 89-kDa protein plays a role in Holospora's escape from the host digestive vacuole, the migration through the host cytoplasm, and the invasion into the macronucleus.« less

Rapid antigen detection test for respiratory syncytial virus diagnosis as a diagnostic tool.

PubMed

Mesquita, Flávio da Silva; Oliveira, Danielle Bruna Leal de; Crema, Daniela; Pinez, Célia Miranda Nunes; Colmanetti, Thaís Cristina; Thomazelli, Luciano Matsumia; Gilio, Alfredo Elias; Vieira, Sandra Elisabeth; Martinez, Marina Baquerizo; Botosso, Viviane Fongaro; Durigon, Edison Luiz

The aim of this study was to evaluate the QuickVue ® RSV Test Kit (QUIDEL Corp, CA, USA) as a screening tool for respiratory syncytial virus in children with acute respiratory disease in comparison with the indirect immunofluorescence assay as gold standard. In Brazil, rapid antigen detection tests for respiratory syncytial virus are not routinely utilized as a diagnostic tool, except for the diagnosis of dengue and influenza. The authors retrospectively analyzed 486 nasopharyngeal aspirate samples from children under age 5 with acute respiratory infection, between December 2013 and August 2014, the samples were analyzed by indirect immunofluorescence assay and QuickVue ® RSV Test kit. Samples with discordant results were analyzed by real time PCR and nucleotide sequencing. From 313 positive samples by immunofluorescence assays, 282 (90%) were also positive by the rapid antigen detection test, two were positive only by rapid antigen detection test, 33 were positive only by immunofluorescence assays, and 171 were positive by both methods. The 35 samples with discordant results were analyzed by real time PCR; the two samples positive only by rapid antigen detection test and the five positive only by immunofluorescence assays were also positive by real time PCR. There was no relation between the negativity by QuickVue ® RSV Test and viral load or specific strain. The QuickVue ® RSV Test showed sensitivity of 90%, specificity of 98.8%, predictive positive value of 99.3%, and negative predictive value of 94.6%, with accuracy of 93.2% and agreement κ index of 0.85 in comparison to immunofluorescence assay. This study demonstrated that the QuickVue ® RSV Test Kit can be effective in early detection of Respiratory syncytial virus in nasopharyngeal aspirate and is reliable for use as a diagnostic tool in pediatrics. Copyright © 2016 Sociedade Brasileira de Pediatria. Published by Elsevier Editora Ltda. All rights reserved.

Application of information-theoretic measures to quantitative analysis of immunofluorescent microscope imaging.

PubMed

Shutin, Dmitriy; Zlobinskaya, Olga

2010-02-01

The goal of this contribution is to apply model-based information-theoretic measures to the quantification of relative differences between immunofluorescent signals. Several models for approximating the empirical fluorescence intensity distributions are considered, namely Gaussian, Gamma, Beta, and kernel densities. As a distance measure the Hellinger distance and the Kullback-Leibler divergence are considered. For the Gaussian, Gamma, and Beta models the closed-form expressions for evaluating the distance as a function of the model parameters are obtained. The advantages of the proposed quantification framework as compared to simple mean-based approaches are analyzed with numerical simulations. Two biological experiments are also considered. The first is the functional analysis of the p8 subunit of the TFIIH complex responsible for a rare hereditary multi-system disorder--trichothiodystrophy group A (TTD-A). In the second experiment the proposed methods are applied to assess the UV-induced DNA lesion repair rate. A good agreement between our in vivo results and those obtained with an alternative in vitro measurement is established. We believe that the computational simplicity and the effectiveness of the proposed quantification procedure will make it very attractive for different analysis tasks in functional proteomics, as well as in high-content screening. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

Indirect carotid cavernous fistula mimicking ocular myasthenia.

PubMed

Leishangthem, Lakshmi; Satti, Sudhakar Reddy

2017-10-19

71-year-old woman with progressive left-sided, monocular diplopia and ptosis. Her symptoms mimicked ocular myasthenia, but she had an indirect carotid cavernous fistula (CCF). She was diagnosed with monocular myasthenia gravis (negative acetylcholinesterase antibody) after a positive ice test and started on Mestinon and underwent a thymectomy complicated by a brachial plexus injury. Months later, she developed left-sided proptosis and ocular bruit. She was urgently referred to neuro-interventional surgery and was diagnosed with an indirect high-flow left CCF, which was treated with Onyx liquid and platinum coil embolisation. Mestinon was discontinued. Her ophthalmic symptoms resolved. However, she was left with a residual left arm and hand hemiparesis and dysmetria secondary to a brachial plexus injury. Indirect CCF usually can present with subtle and progressive symptoms leading to delayed diagnosis or misdiagnosis. It is important for ophthalmologists to consider this differential in a patient with progressive ocular symptoms. © BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Geant4-DNA simulation of DNA damage caused by direct and indirect radiation effects and comparison with biological data.

NASA Astrophysics Data System (ADS)

Villagrasa, Carmen; Meylan, Sylvain; Gonon, Geraldine; Gruel, Gaëtan; Giesen, Ulrich; Bueno, Marta; Rabus, Hans

2017-09-01

In this work we present results obtained in the frame of the BioQuaRT project. The objective of the study was the correlation between the number of radiation-induced double strand breaks (DSB) of the DNA molecule and the probability of detecting nuclear foci after targeted microbeam irradiation of cells with protons and alpha particles of different LET. The former were obtained by simulation with new methods integrated into Geant4-DNA that permit calculating the number of DSB in a DNA target model induced by direct and indirect radiation effects. A particular focus was laid in this work on evaluating the influence of different criteria applied to the simulated results for predicting the formation of a direct SSB. Indeed, these criteria have an important impact on the predicted number of DSB per particle track and its dependence with LET. Among the criteria tested in this work, the case that a direct radiation interaction leads to a strand break if the cumulative energy deposited in the backbone part of one nucleotide exceeds a threshold of 17.5 eV leads to the best agreement with the relative LET dependence of number of radiation induced foci. Further calculations and experimental data are nevertheless needed in order to fix the simulation parameters and to help interpreting the biological experimental data observed by immunofluorescence in terms of the DSB complexity.

Indirect photodegradation of the lampricides TFM and niclosamide.

PubMed

McConville, Megan B; Mezyk, Stephen P; Remucal, Christina K

2017-08-16

3-Trifluromethyl-4-nitrophenol (TFM) and 2',5-dichloro-4'-nitrosalicylanilide (niclosamide) are lampricides used in tributaries of the Great Lakes to kill the invasive parasitic sea lamprey (Petromyzon marinus). Although the lampricides have been applied since the late 1950s, their photochemical behavior in natural environments is still not well understood. This study examines the indirect photodegradation of these two compounds and the resulting yields of organic and inorganic photoproducts in water samples collected from five tributaries of Lake Michigan. The tributaries were selected to span the length of Lake Michigan and its natural carbonate geologic gradient. In the presence of dissolved organic matter (DOM), the niclosamide photodegradation rate triples, while the rate of TFM photodegradation is unchanged. Additionally, the yield of lampricide organic products is influenced by DOM because many of the organic photoproducts themselves are prone to DOM-mediated indirect photodegradation. The indirect photodegradation of niclosamide is primarily mediated by reaction with singlet oxygen, which accounts for more than 50% of the increased photodegradation rate. Additionally, hydroxyl radicals and carbonate radicals (CO 3 - ˙) influence niclosamide indirect photolysis, and their contribution is dependent on the specific river water chemistry. For example, CO 3 - ˙ contribution to niclosamide photodegradation, while small, is greater in southern tributaries where there is higher carbonate alkalinity.

A Nerve Clamp Electrode Design for Indirect Stimulation of Skeletal Muscle

DTIC Science & Technology

2010-10-01

neurons. This device enables stimulation of muscle contraction indirectly as opposed to contraction from direct muscle stimulation. The electrode is able...to stimulate indirect muscle contraction when tested on ex vivo preparations from rodent phrenic nerve-hemidiaphragm muscle in similar fashion to...unsuccessful in stimulating indirect muscle contraction . Therefore, this novel electrode is useful for physiological assessment of nerve agents and

Advances in Urine Microscopy.

PubMed

Becker, Gavin J; Garigali, Giuseppe; Fogazzi, Giovanni B

2016-06-01

Urine microscopy is an important tool for the diagnosis and management of several conditions affecting the kidneys and urinary tract. In this review, we describe the automated instruments, based either on flow cytometry or digitized microscopy, that are currently in use in large clinical laboratories. These tools allow the examination of large numbers of samples in short periods. We also discuss manual urinary microscopy commonly performed by nephrologists, which we encourage. After discussing the advantages of phase contrast microscopy over bright field microscopy, we describe the advancements of urine microscopy in various clinical conditions. These include persistent isolated microscopic hematuria (which can be classified as glomerular or nonglomerular on the basis of urinary erythrocyte morphology), drug- and toxin-related cystalluria (which can be a clue for the diagnosis of acute kidney injury associated with intrarenal crystal precipitation), and some inherited conditions (eg, adenine phosphoribosyltransferase deficiency, which is associated with 2,8-dihydroxyadenine crystalluria, and Fabry disease, which is characterized by unique urinary lamellated fatty particles). Finally, we describe the utility of identifying "decoy cells" and atypical malignant cells, which can be easily done with phase contrast microscopy in unfixed samples. Copyright © 2016 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.

Water Rockets and Indirect Measurement.

ERIC Educational Resources Information Center

Inman, Duane

1997-01-01

Describes an activity that teaches a number of scientific concepts including indirect measurement, Newton's third law of motion, manipulating and controlling variables, and the scientific method of inquiry. Uses process skills such as observation, inference, prediction, mensuration, and communication as well as problem solving and higher-order…

Building Electricity Consumption as an Indicator of Indirect Carbon Dioxide Emissions

NASA Astrophysics Data System (ADS)

Ma’mun, S.; Sukirman; Alel, A. E.; Hasanah, M.

2018-05-01

The global CO2 emissions have continually increased from year to year and reached 32 Gt in 2010. The increased CO2 emissions may lead to a higher temperature and cause climate change on a global scale. Building energy-using equipment in Indonesia continuously increases annually leading to increasing indirect CO2 emissions from the buildings. The objective of this study is to measure the indirect CO2 emissions from the Faculty of Industrial Technology (FIT), Universitas Islam Indonesia (UII) Yogyakarta, Indonesia. The research data were taken from the electricity consumption by reading the electric meter at specified time intervals for 7 weeks from 26 September to 13 November 2016. The amount of electricity consumption indirectly indicates the amount of CO2 emission in the FIT where the FIT has consumed the electricity of 18.6 kWh/day corresponding to the average indirect CO2 emission of 15.9 kg CO2-eq/day. The results obtained would, therefore, give some recommendations to the FIT to take some policy actions related to the indirect CO2 emission by improving energy management system to minimize the indirect CO2 emission in the FIT.

34 CFR 76.564 - Restricted indirect cost rate-formula.

Code of Federal Regulations, 2014 CFR

2014-07-01

... CFR 75.563 that is not a State or local government agency may use— (1) An indirect cost rate computed.... 1221e-3(a)(1), 2831(a), 2974(b), and 3474) [59 FR 59583, Nov. 17, 1994] ... 34 Education 1 2014-07-01 2014-07-01 false Restricted indirect cost rate-formula. 76.564 Section...

Indirect aerosol effect increases CMIP5 models projected Arctic warming

DOE PAGES

Chylek, Petr; Vogelsang, Timothy J.; Klett, James D.; ...

2016-02-20

Phase 5 of the Coupled Model Intercomparison Project (CMIP5) climate models’ projections of the 2014–2100 Arctic warming under radiative forcing from representative concentration pathway 4.5 (RCP4.5) vary from 0.9° to 6.7°C. Climate models with or without a full indirect aerosol effect are both equally successful in reproducing the observed (1900–2014) Arctic warming and its trends. However, the 2014–2100 Arctic warming and the warming trends projected by models that include a full indirect aerosol effect (denoted here as AA models) are significantly higher (mean projected Arctic warming is about 1.5°C higher) than those projected by models without a full indirect aerosolmore » effect (denoted here as NAA models). The suggestion is that, within models including full indirect aerosol effects, those projecting stronger future changes are not necessarily distinguishable historically because any stronger past warming may have been partially offset by stronger historical aerosol cooling. In conclusion, the CMIP5 models that include a full indirect aerosol effect follow an inverse radiative forcing to equilibrium climate sensitivity relationship, while models without it do not.« less

Indirect aerosol effect increases CMIP5 models projected Arctic warming

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

Chylek, Petr; Vogelsang, Timothy J.; Klett, James D.

Phase 5 of the Coupled Model Intercomparison Project (CMIP5) climate models’ projections of the 2014–2100 Arctic warming under radiative forcing from representative concentration pathway 4.5 (RCP4.5) vary from 0.9° to 6.7°C. Climate models with or without a full indirect aerosol effect are both equally successful in reproducing the observed (1900–2014) Arctic warming and its trends. However, the 2014–2100 Arctic warming and the warming trends projected by models that include a full indirect aerosol effect (denoted here as AA models) are significantly higher (mean projected Arctic warming is about 1.5°C higher) than those projected by models without a full indirect aerosolmore » effect (denoted here as NAA models). The suggestion is that, within models including full indirect aerosol effects, those projecting stronger future changes are not necessarily distinguishable historically because any stronger past warming may have been partially offset by stronger historical aerosol cooling. In conclusion, the CMIP5 models that include a full indirect aerosol effect follow an inverse radiative forcing to equilibrium climate sensitivity relationship, while models without it do not.« less

Pregnancy complicated with Alport syndrome: a good obstetric outcome and failure to diagnose an infant born to a mother with Alport syndrome by umbilical cord immunofluorescence staining.

PubMed

Matsubara, Shigeki; Ueda, Yoshihiko; Takahashi, Hisako; Nagai, Takashi; Kuwata, Tomoyuki; Muto, Shigeaki; Yamaguchi, Takehiko; Takizawa, Toshihiro; Suzuki, Mitsuaki

2009-12-01

Alport syndrome is a familial progressive nephritis. The most frequent type is X-linked Alport syndrome, caused by genetic abnormalities in the alpha 5 chain of type IV collagen. Skin biopsy is a useful tool for diagnosing this disease. It is not well known how this syndrome affects pregnancy and how it is affected by pregnancy, or whether the umbilical cord may provide material for detecting this collagen abnormality. We report a primigravida with Alport syndrome with mild proteinuria who gave birth abdominally to a term male infant without deteriorating renal function during pregnancy. The umbilical cord from not only this infant but also from an Alport (-) control infant showed negative immunofluorescence staining for the alpha 5 chain of type IV collagen. Women with Alport syndrome without renal dysfunction may follow an uneventful obstetrical course until term. The cord may not be suitable for diagnosing Alport syndrome with immunofluorescence staining.

Caffeinated and Non-caffeinated Alcohol Use and Indirect Aggression: The Impact of Self-regulation

PubMed Central

Sheehan, Brynn E.; Linden-Carmichael, Ashley N.; Lau-Barraco, Cathy

2016-01-01

Research shows that heavier alcohol use is associated with physical aggression. Scant research has examined the way in which alcohol relates to other forms of aggression, such as indirect aggression (e.g., malicious humor, social exclusion). Given the possible negative consequences of indirect aggression and the limited evidence suggesting alcohol use can elicit indirectly aggressive responses, research is needed to further investigate the association between drinking behavior and indirect aggression. Additionally, specific alcoholic beverages, such as caffeinated alcoholic beverages (CABs; e.g., Red Bull and vodka), may potentiate aggression above the influence of typical use, thus warrant examination with regard to indirect aggression. One factor that may impact the strength of the alcohol-indirect aggression and CAB-indirect aggression relationships is one's level of self-regulation. Consequently, our study examined the relationships between (1) alcohol use and indirect aggression, (2) CAB use and indirect aggression, and (3) self-regulation as a moderator. Participants were 733 (67.6% female) undergraduate students who reported their CAB and alcohol use, self-regulation, and aggressive behaviors. Results revealed that heavier alcohol use was associated with more frequent indirect aggression after controlling for dispositional aggression. Heavier CAB use was related to more frequent indirect aggression after accounting for typical use and dispositional aggression. Self-regulation moderated these associations such that for those with lower self-regulation, greater alcohol and CAB consumption was associated with greater indirect aggression. Our findings suggest that heavier alcohol and CAB consumption may be risk factors for engaging in indirect aggression and this risk is impacted by one's regulatory control. PMID:26905765

New insights into FtsZ rearrangements during the cell division of Escherichia coli from single-molecule localization microscopy of fixed cells.

PubMed

Vedyaykin, Alexey D; Vishnyakov, Innokentii E; Polinovskaya, Vasilisa S; Khodorkovskii, Mikhail A; Sabantsev, Anton V

2016-06-01

FtsZ - a prokaryotic tubulin homolog - is one of the central components of bacterial division machinery. At the early stage of cytokinesis FtsZ forms the so-called Z-ring at mid-cell that guides septum formation. Many approaches were used to resolve the structure of the Z-ring, however, researchers are still far from consensus on this question. We utilized single-molecule localization microscopy (SMLM) in combination with immunofluorescence staining to visualize FtsZ in Esherichia coli fixed cells that were grown under slow and fast growth conditions. This approach allowed us to obtain images of FtsZ structures at different stages of cell division and accurately measure Z-ring dimensions. Analysis of these images demonstrated that Z-ring thickness increases during constriction, starting at about 70 nm at the beginning of division and increasing by approximately 25% half-way through constriction. © 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Diagnostic electron microscopy

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

Dickersin, G.R.

1988-01-01

In this book the author presents a comprehensive reference text on diagnostic electron microscopy. Throughout the book he illustrates how ultrastructural identification can be helpful for the recognition of cell type and the identification of mechanisms of pathogenesis in various diseases. In addition to electron microscopy photographs, there are also numerous light microscopy photographs for comparison. This text presents the classification of neoplasms in the order and arrangement most familiar to the pathologist. Contents: Introduction; Diagram of a Normal Cell; Normal Cell Function; Embryology; Neoplasms; Infectious Agents; Metabolic Diseases; Renal Diseases; Skeletal Muscle and Peripheral Nerve Diseases; Index.

Control methods and systems for indirect evaporative coolers

DOEpatents

Woods, Jason; Kozubal, Erik

2015-09-22

A control method for operating an indirect evaporative cooler to control temperature and humidity. The method includes operating an airflow control device to provide supply air at a flow rate to a liquid desiccant dehumidifier. The supply air flows through the dehumidifier and an indirect evaporative cooler prior to exiting an outlet into a space. The method includes operating a pump to provide liquid desiccant to the liquid desiccant dehumidifier and sensing a temperature of an airstream at the outlet of the indirect evaporative cooler. The method includes comparing the temperature of the airstream at the outlet to a setpoint temperature at the outlet and controlling the pump to set the flow rate of the liquid desiccant. The method includes sensing space temperature, comparing the space temperature with a setpoint temperature, and controlling the airflow control device to set the flow rate of the supply air based on the comparison.

Nuclear protein accumulation in cellular senescence and organismal aging revealed with a novel single-cell resolution fluorescence microscopy assay.

PubMed

De Cecco, Marco; Jeyapalan, Jessie; Zhao, Xiaoai; Tamamori-Adachi, Mimi; Sedivy, John M

2011-10-01

Replicative cellular senescence was discovered some 50 years ago. The phenotypes of senescent cells have been investigated extensively in cell culture, and found to affect essentially all aspects of cellular physiology. The relevance of cellular senescence in the context of age-associated pathologies as well as normal aging is a topic of active and ongoing interest. Considerable effort has been devoted to biomarker discovery to enable the microscopic detection of single senescent cells in tissues. One characteristic of senescent cells documented very early in cell culture studies was an increase in cell size and total protein content, but whether this occurs in vivo is not known. A limiting factor for studies of protein content and localization has been the lack of suitable fluorescence microscopy tools. We have developed an easy and flexible method, based on the merocyanine dye known as NanoOrange, to visualize and quantitatively measure total protein levels by high resolution fluorescence microscopy. NanoOrange staining can be combined with antibody-based immunofluorescence, thus providing both specific target and total protein information in the same specimen. These methods are optimally combined with automated image analysis platforms for high throughput analysis. We document here increasing protein content and density in nuclei of senescent human and mouse fibroblasts in vitro, and in liver nuclei of aged mice in vivo. Additionally, in aged liver nuclei NanoOrange revealed protein-dense foci that colocalize with centromeric heterochromatin.

Nuclear protein accumulation in cellular senescence and organismal aging revealed with a novel single-cell resolution fluorescence microscopy assay

PubMed Central

De Cecco, Marco; Jeyapalan, Jessie; Zhao, Xiaoai; Tamamori-Adachi, Mimi; Sedivy, John M.

2011-01-01

Replicative cellular senescence was discovered some 50 years ago. The phenotypes of senescent cells have been investigated extensively in cell culture, and found to affect essentially all aspects of cellular physiology. The relevance of cellular senescence in the context of age-associated pathologies as well as normal aging is a topic of active and ongoing interest. Considerable effort has been devoted to biomarker discovery to enable the microscopic detection of single senescent cells in tissues. One characteristic of senescent cells documented very early in cell culture studies was an increase in cell size and total protein content, but whether this occurs in vivo is not known. A limiting factor for studies of protein content and localization has been the lack of suitable fluorescence microscopy tools. We have developed an easy and flexible method, based on the merocyanine dye known as NanoOrange, to visualize and quantitatively measure total protein levels by high resolution fluorescence microscopy. NanoOrange staining can be combined with antibody-based immunofluorescence, thus providing both specific target and total protein information in the same specimen. These methods are optimally combined with automated image analysis platforms for high throughput analysis. We document here increasing protein content and density in nuclei of senescent human and mouse fibroblasts in vitro, and in liver nuclei of aged mice in vivo. Additionally, in aged liver nuclei NanoOrange revealed protein-dense foci that colocalize with centromeric heterochromatin. PMID:22006542

Clinical and Biological Heterogeneity in ARDS: Direct versus Indirect Lung Injury

PubMed Central

Shaver, Ciara M.; Bastarache, Julie A.

2014-01-01

Synopsis The acute respiratory distress syndrome (ARDS) is a heterogeneous group of illnesses affecting the pulmonary parenchyma with acute onset bilateral inflammatory pulmonary infiltrates with associated hypoxemia. ARDS occurs after two major types of pulmonary injury: direct lung injury affecting the lung epithelium or indirect lung injury disrupting the vascular endothelium. Greater understanding of the differences between direct and indirect lung injury may refine our classification of patients with ARDS and lead to development of new therapeutics targeted at specific subpopulations of patients with ARDS. In this review, we will summarize the differences between direct and indirect causes of ARDS in human patients and then will review current knowledge of the similarities and differences in ARDS pathogenesis based on experimental animal models of direct and indirect lung injury. While the separation between direct and indirect causes of ARDS may be oversimplified, it is a useful approach to advancing our current understanding of the pathogenesis of this complex and often fatal disease. PMID:25453415

Evaluation of Two rK39 Dipstick Tests, Direct Agglutination Test, and Indirect Fluorescent Antibody Test for Diagnosis of Visceral Leishmaniasis in a New Epidemic Site in Highland Ethiopia

PubMed Central

Cañavate, Carmen; Herrero, Merce; Nieto, Javier; Cruz, Israel; Chicharro, Carmen; Aparicio, Pilar; Mulugeta, Abate; Argaw, Daniel; Blackstock, Anna J.; Alvar, Jorge; Bern, Caryn

2011-01-01

We assessed the performance characteristics of two rK39 immunochromatographic tests, a direct agglutination test (DAT), and an indirect immunofluorescent antibody test (IFAT) in the site of a new epidemic of visceral leishmaniasis (VL) in northwestern Ethiopia. The study population was composed of 179 patients with suspected VL and 67 controls. The sensitivities of Kalazar Detect®, DiaMed-IT Leish®, DAT, and IFAT in 35 polymerase chain reaction–confirmed VL cases were 94.3%, 91.4%, 91.4%, and 100%, respectively, and the specificities were 98.5%, 94%, 98.5%, and 98.5%, respectively. In a Bayesian latent class analysis of all 246 specimens, the estimated sensitivities were 90.5%, 89%, 88.8%, and 96% for Kalazar Detect®, DiaMed-IT Leish®, DAT, and IFAT, respectively; DAT showed the highest estimated specificity (97.4%). Both rK39 immunochromatographic tests perform as well as DAT, and are suitable for VL diagnosis in first-level health centers in this area of Ethiopia. PMID:21212210

Spatially indirect excitons in coupled quantum wells

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

Lai, Chih-Wei Eddy

2004-03-01

Microscopic quantum phenomena such as interference or phase coherence between different quantum states are rarely manifest in macroscopic systems due to a lack of significant correlation between different states. An exciton system is one candidate for observation of possible quantum collective effects. In the dilute limit, excitons in semiconductors behave as bosons and are expected to undergo Bose-Einstein condensation (BEC) at a temperature several orders of magnitude higher than for atomic BEC because of their light mass. Furthermore, well-developed modern semiconductor technologies offer flexible manipulations of an exciton system. Realization of BEC in solid-state systems can thus provide new opportunitiesmore » for macroscopic quantum coherence research. In semiconductor coupled quantum wells (CQW) under across-well static electric field, excitons exist as separately confined electron-hole pairs. These spatially indirect excitons exhibit a radiative recombination time much longer than their thermal relaxation time a unique feature in direct band gap semiconductor based structures. Their mutual repulsive dipole interaction further stabilizes the exciton system at low temperature and screens in-plane disorder more effectively. All these features make indirect excitons in CQW a promising system to search for quantum collective effects. Properties of indirect excitons in CQW have been analyzed and investigated extensively. The experimental results based on time-integrated or time-resolved spatially-resolved photoluminescence (PL) spectroscopy and imaging are reported in two categories. (i) Generic indirect exciton systems: general properties of indirect excitons such as the dependence of exciton energy and lifetime on electric fields and densities were examined. (ii) Quasi-two-dimensional confined exciton systems: highly statistically degenerate exciton systems containing more than tens of thousands of excitons within areas as small as (10 micrometer) 2 were

Indirect Competitive Enzyme-Linked Immunosorbent Assay (ELISA).

PubMed

Kohl, Thomas O; Ascoli, Carl A

2017-07-05

The indirect competitive ELISA (indirect cELISA) pits plate-immobilized antigen against antigens in solution for binding to antigen-specific antibody. The antigens in solution are in the test sample and are first incubated with antigen-specific antibody. These antibody-antigen complexes are then added to microtiter plates whose wells have been coated with purified antigen. The wells are washed to remove unbound antigen-antibody complexes and free antigen. A reporter-labeled secondary antibody is then added followed by the addition of substrate. Substrate hydrolysis yields a signal that is inversely proportional to antigen concentration within the sample. This is because when antigen concentration is high in the test sample, most of the antibody is bound before adding the solution to the plate. Most of the antibody remains in solution (as complexes) and is thus washed away before the addition of the reporter-labeled secondary antibody and substrate. Thus, the higher the antigen concentration in the test sample, the weaker the resultant signal in the detection step. The indirect cELISA is often used for competitive detection and quantification of antibodies against viral diseases in biological samples. © 2017 Cold Spring Harbor Laboratory Press.

Fluorescence (Multiwave) Confocal Microscopy.

PubMed

Welzel, J; Kästle, Raphaela; Sattler, Elke C

2016-10-01

In addition to reflectance confocal microscopy, multiwave confocal microscopes with different laser wavelengths in combination with exogenous fluorophores allow fluorescence mode confocal microscopy in vivo and ex vivo. Fluorescence mode confocal microscopy improves the contrast between the epithelium and the surrounding soft tissue and allows the depiction of certain structures, like epithelial tumors, nerves, and glands. Copyright © 2016 Elsevier Inc. All rights reserved.

Ectoparasitic hematophagous dipters: potential reservoirs of dengue virus?

PubMed

Setién, Álvaro Aguilar; Baltazar, Anahí García; Leyva, Ignacio Olave; Rojas, Mónica Salas; Koldenkova, Vadim Pérez; García, Mariem Pérez-Peña; Ceballos, Nidia Aréchiga; Romero, Guillermo Gálvez; Villegas, Edgar Olivier López; Malacara, Juan Bibiano Morales; Marín, Cenia Almazán

2017-01-01

Recently, the presence of antibodies and dengue virus (DV) RNA in neotropical wild mammals, including Desmodus rotundus, was reported. In a previous study, DV was also found in a high percentage (39.6%) of ectoparasitic hematophagous dipters specifics of these hematophagous bats. In order to verify the susceptibility of these ectoparasites to DV, in this work experimental infections with VD2 of organs explants of Strebla wiedemanni and of Melophagus ovinus were performed using C6/36 cells as control. Viral titers (UFP/mL) were determined at 0, 48 and 96 hrs pi. Infected organs were observed by electron microscopy and under the confocal microscopy indirect immunofluorescence (IIF) using specific conjugates against DV. The infected organs of both species of ectoparasites replicated DV at titers similar to those obtained with the C6/36 cell line (≥10 6 UFP/mL). Electron microscopy and IIF showed DV replication in the digestive tract, tracheoles, reproductive organs of males but not in females, and milk glands (MG) of both species. In the fatty bodies of the MG of M. ovinus, zones with a high affinity for the DV were observed. In this work the susceptibility of S. wiedemanni and M. ovinus to DV was demonstrated and consequently the probable role of this ectoparasites as wild reservoirs of DV. Copyright: © 2017 SecretarÍa de Salud.

Simultaneous visualization of Propionibacterium acnes and Propionibacterium granulosum with immunofluorescence and fluorescence in situ hybridization.

PubMed

Jahns, Anika C; Oprica, Cristina; Vassilaki, Ismini; Golovleva, Irina; Palmer, Ruth H; Alexeyev, Oleg A

2013-10-01

Propionibacterium acnes (P. acnes) and Propionibacterium granulosum (P. granulosum) are common skin colonizers that are implicated as possible contributing factors in acne vulgaris development. We have established direct visualization tools for the simultaneous detection of these closely related species with immunofluorescence assay and fluorescence in situ hybridization (FISH). As proof of principle, we were able to distinguish P. acnes and P. granulosum bacteria in multi-species populations in vitro as well as in a mock skin infection model upon labelling with 16S rRNA probes in combinatorial FISH as well as with antibodies. Furthermore, we report the co-localization of P. acnes and P. granulosum in the stratum corneum and hair follicles from patients with acne vulgaris as well as in healthy individuals. Further studies on the spatial distribution of these bacteria in skin structures in various skin disorders are needed. Copyright © 2013 Elsevier Ltd. All rights reserved.

Immunofluorescence Approach to the Study of the Ecology of Thermoplasma acidophilum In Coal Refuse Material

PubMed Central

Bohlool, B. B.; Brock, T. D.

1974-01-01

Specific immunofluorescence staining was applied to the study of the localization, distribution, and growth of Thermoplasma acidophilum in its natural habitat, the coal refuse pile. Different antigenic groups of T. acidophilum could be isolated from the same refuse pile, and the same antigenic groups were isolated from piles from different geographical areas. No correlation could be established between the antigenic groups and the pH or temperature of the habitats. Brightly fluorescing cells of T. acidophilum were detected on microscope slides buried in contact with the coal refuse material or immersed in the water in the stream draining a refuse pile. T. acidophilum grew when inoculated into either coal refuse material and/or an aqueous extract of coal refuse when incubated at its optimal temperature of 55 C, but not when incubated at room temperature or 37 C. The coal refuse pile appears to be a primary habitat for T. acidophilum. PMID:4602306

Observation of DNA Molecules Using Fluorescence Microscopy and Atomic Force Microscopy

ERIC Educational Resources Information Center

Ito, Takashi

2008-01-01

This article describes experiments for an undergraduate instrumental analysis laboratory that aim to observe individual double-stranded DNA (dsDNA) molecules using fluorescence microscopy and atomic force microscopy (AFM). dsDNA molecules are observed under several different conditions to discuss their chemical and physical properties. In…

48 CFR 227.7005 - Indirect notice of patent infringement claims.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 3 2010-10-01 2010-10-01 false Indirect notice of patent... REGULATIONS SYSTEM, DEPARTMENT OF DEFENSE GENERAL CONTRACTING REQUIREMENTS PATENTS, DATA, AND COPYRIGHTS Infringement Claims, Licenses, and Assignments 227.7005 Indirect notice of patent infringement claims. (a) A...

48 CFR 227.7005 - Indirect notice of patent infringement claims.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 48 Federal Acquisition Regulations System 3 2012-10-01 2012-10-01 false Indirect notice of patent... REGULATIONS SYSTEM, DEPARTMENT OF DEFENSE GENERAL CONTRACTING REQUIREMENTS PATENTS, DATA, AND COPYRIGHTS Infringement Claims, Licenses, and Assignments 227.7005 Indirect notice of patent infringement claims. (a) A...

Intravital microscopy

PubMed Central

Masedunskas, Andrius; Milberg, Oleg; Porat-Shliom, Natalie; Sramkova, Monika; Wigand, Tim; Amornphimoltham, Panomwat; Weigert, Roberto

2012-01-01

Intravital microscopy is an extremely powerful tool that enables imaging several biological processes in live animals. Recently, the ability to image subcellular structures in several organs combined with the development of sophisticated genetic tools has made possible extending this approach to investigate several aspects of cell biology. Here we provide a general overview of intravital microscopy with the goal of highlighting its potential and challenges. Specifically, this review is geared toward researchers that are new to intravital microscopy and focuses on practical aspects of carrying out imaging in live animals. Here we share the know-how that comes from first-hand experience, including topics such as choosing the right imaging platform and modality, surgery and stabilization techniques, anesthesia and temperature control. Moreover, we highlight some of the approaches that facilitate subcellular imaging in live animals by providing numerous examples of imaging selected organelles and the actin cytoskeleton in multiple organs. PMID:22992750

Supporting Risk Assessment: Accounting for Indirect Risk to Ecosystem Components

PubMed Central

Mach, Megan E.; Martone, Rebecca G.; Singh, Gerald G.; O, Miriam; Chan, Kai M. A.

2016-01-01

The multi-scalar complexity of social-ecological systems makes it challenging to quantify impacts from human activities on ecosystems, inspiring risk-based approaches to assessments of potential effects of human activities on valued ecosystem components. Risk assessments do not commonly include the risk from indirect effects as mediated via habitat and prey. In this case study from British Columbia, Canada, we illustrate how such “indirect risks” can be incorporated into risk assessments for seventeen ecosystem components. We ask whether (i) the addition of indirect risk changes the at-risk ranking of the seventeen ecosystem components and if (ii) risk scores correlate with trophic prey and habitat linkages in the food web. Even with conservative assumptions about the transfer of impacts or risks from prey species and habitats, the addition of indirect risks in the cumulative risk score changes the ranking of priorities for management. In particular, resident orca, Steller sea lion, and Pacific herring all increase in relative risk, more closely aligning these species with their “at-risk status” designations. Risk assessments are not a replacement for impact assessments, but—by considering the potential for indirect risks as we demonstrate here—they offer a crucial complementary perspective for the management of ecosystems and the organisms within. PMID:27632287

Direct and indirect effects of body weight on adult wages.

PubMed

Han, Euna; Norton, Edward C; Powell, Lisa M

2011-12-01

Previous estimates of the association between body weight and wages in the literature have been conditional on education and occupation. In addition to the effect of current body weight status (body mass index (BMI) or obesity) on wages, this paper examines the indirect effect of body weight status in the late-teenage years on wages operating through education and occupation choice. Using the National Longitudinal Survey of Youth 1979 data, for women, we find that a one-unit increase in BMI is directly associated with 1.83% lower hourly wages whereas the indirect BMI wage penalty is not statistically significant. Neither a direct nor an indirect BMI wage penalty is found for men. However, results based on clinical weight classification reveal that the indirect wage penalty occurs to a larger extent at the upper tail of the BMI distribution for both men and women via the pathways of education and occupation outcomes. Late-teen obesity is indirectly associated with 3.5% lower hourly wages for both women and men. These results are important because they imply that the total effect of obesity on wages is significantly larger than has been estimated in previous cross-sectional studies. 2011 Elsevier B.V. All rights reserved.

Nonlinear vibrational microscopy

DOEpatents

Holtom, Gary R.; Xie, Xiaoliang Sunney; Zumbusch, Andreas

2000-01-01

The present invention is a method and apparatus for microscopic vibrational imaging using coherent Anti-Stokes Raman Scattering or Sum Frequency Generation. Microscopic imaging with a vibrational spectroscopic contrast is achieved by generating signals in a nonlinear optical process and spatially resolved detection of the signals. The spatial resolution is attained by minimizing the spot size of the optical interrogation beams on the sample. Minimizing the spot size relies upon a. directing at least two substantially co-axial laser beams (interrogation beams) through a microscope objective providing a focal spot on the sample; b. collecting a signal beam together with a residual beam from the at least two co-axial laser beams after passing through the sample; c. removing the residual beam; and d. detecting the signal beam thereby creating said pixel. The method has significantly higher spatial resolution then IR microscopy and higher sensitivity than spontaneous Raman microscopy with much lower average excitation powers. CARS and SFG microscopy does not rely on the presence of fluorophores, but retains the resolution and three-dimensional sectioning capability of confocal and two-photon fluorescence microscopy. Complementary to these techniques, CARS and SFG microscopy provides a contrast mechanism based on vibrational spectroscopy. This vibrational contrast mechanism, combined with an unprecedented high sensitivity at a tolerable laser power level, provides a new approach for microscopic investigations of chemical and biological samples.

Microscopy and Image Analysis.

PubMed

McNamara, George; Difilippantonio, Michael; Ried, Thomas; Bieber, Frederick R

2017-07-11

This unit provides an overview of light microscopy, including objectives, light sources, filters, film, and color photography for fluorescence microscopy and fluorescence in situ hybridization (FISH). We believe there are excellent opportunities for cytogeneticists, pathologists, and other biomedical readers, to take advantage of specimen optical clearing techniques and expansion microscopy-we briefly point to these new opportunities. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

A Dual Microscopy-Based Assay To Assess Listeria monocytogenes Cellular Entry and Vacuolar Escape.

PubMed

Quereda, Juan J; Pizarro-Cerdá, Javier; Balestrino, Damien; Bobard, Alexandre; Danckaert, Anne; Aulner, Nathalie; Shorte, Spencer; Enninga, Jost; Cossart, Pascale

2016-01-01

Listeria monocytogenes is a Gram-positive bacterium and a facultative intracellular pathogen that invades mammalian cells, disrupts its internalization vacuole, and proliferates in the host cell cytoplasm. Here, we describe a novel image-based microscopy assay that allows discrimination between cellular entry and vacuolar escape, enabling high-content screening to identify factors specifically involved in these two steps. We first generated L. monocytogenes and Listeria innocua strains expressing a β-lactamase covalently attached to the bacterial cell wall. These strains were then incubated with HeLa cells containing the Förster resonance energy transfer (FRET) probe CCF4 in their cytoplasm. The CCF4 probe was cleaved by the bacterial surface β-lactamase only in cells inoculated with L. monocytogenes but not those inoculated with L. innocua, thereby demonstrating bacterial access to the host cytoplasm. Subsequently, we performed differential immunofluorescence staining to distinguish extracellular versus total bacterial populations in samples that were also analyzed by the FRET-based assay. With this two-step analysis, bacterial entry can be distinguished from vacuolar rupture in a single experiment. Our novel approach represents a powerful tool for identifying factors that determine the intracellular niche of L. monocytogenes. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Hematopoiesis in 3 dimensions: human and murine bone marrow architecture visualized by confocal microscopy.

PubMed

Takaku, Tomoiku; Malide, Daniela; Chen, Jichun; Calado, Rodrigo T; Kajigaya, Sachiko; Young, Neal S

2010-10-14

In many animals, blood cell production occurs in the bone marrow. Hematopoiesis is complex, requiring self-renewing and pluripotent stem cells, differentiated progenitor and precursor cells, and supportive stroma, adipose tissue, vascular structures, and extracellular matrix. Although imaging is a vital tool in hematology research, the 3-dimensional architecture of the bone marrow tissue in situ remains largely uncharacterized. The major hindrance to imaging the intact marrow is the surrounding bone structures are almost impossible to cut/image through. We have overcome these obstacles and describe a method whereby whole-mounts of bone marrow tissue were immunostained and imaged in 3 dimensions by confocal fluorescence and reflection microscopy. We have successfully mapped by multicolor immunofluorescence the localization pattern of as many as 4 cell features simultaneously over large tiled views and to depths of approximately 150 μm. Three-dimensional images can be assessed qualitatively and quantitatively to appreciate the distribution of cell types and their interrelationships, with minimal perturbations of the tissue. We demonstrate its application to normal mouse and human marrow, to murine models of marrow failure, and to patients with aplastic anemia, myeloid, and lymphoid cell malignancies. The technique should be generally adaptable for basic laboratory investigation and for clinical diagnosis of hematologic diseases.

IDENTIFICATION OF ALGAE WHICH INTERFERE WITH THE DETECTION OF GIARDIA CYSTS AND CRYPTOSPORIDIUM OOCYSTS AND A METHOD FOR ALLEVIATING THIS INTERFERENCE

EPA Science Inventory

Fifty-four algal species were tested for cross-reaction in the American Society for Testing and Materials Giardia/Cryptosporidium indirect immunofluorescence assay, and 24 showed some degree of fluorescence. Two species, Navicula minima and Synechococcus elongatus, exhibited a br...

THE IDENTIFICATION OF ALGAE WHICH INTERFERE WITH THE DETECTION OF GIARDIA AND CRYPTOSPORIDIUM AND A METHOD FOR ALLEVIATING THIS INTERFERENCE

EPA Science Inventory

Fifty-four algal species were tested for cross-reaction in the American Society for Testing and Materials Giardia/Cryptosporidium indirect immunofluorescence assay, and 24 showed some degree of fluorescence. Two species, Navicula minima and Synechococcus elongatus, exhibited a b...

Clinical attachment loss: estimation by direct and indirect methods.

PubMed

Barbosa, Viviane Leal; Angst, Patricia D Melchiors; Finger Stadler, Amanda; Oppermann, Rui V; Gomes, Sabrina Carvalho

2016-06-01

This observational study aimed to compare the estimation of clinical attachment loss (CAL) as measured by direct (CALD ) and indirect (CALI ) methods. Periodontitis patients (n = 75; mean age: 50.9 ± 8.02 years; 72.2% women; 50.6% smokers) received a periodontal examination (six sites/tooth) to determine the presence of visible plaque and calculus, the gingival bleeding index (GBI), periodontal probing depth (PPD), bleeding on probing (BOP), CALD and gingival recession (GR). CALI values resulted from the sum of PPD and GR values. Statistical analysis considered only data from sites with visible GR (e.g. the gingival margin apical to the cemento-enamel junction; n = 4,757 sites) and determined the mean difference between CALI and CALD measurements. Based on the mean difference, univariate and multivariate analyses were also performed. Mean CALD and CALI values were 3.96 ± 2.07 mm and 4.47 ± 2.03 mm, respectively. The indirect method overestimated CAL compared with the direct method (mean difference: 0.51 ± 1.23 mm; P < 0.001). On uni- and multivariate analyses, absence of GBI and BOP, PPD and proximal site location had significant influences on the overestimation of CAL by the indirect method (all P ≤ 0.01). The indirect method increased the CAL value by 0.38 mm for each additional 1 mm in PPD. To decrease the number of probing errors in daily practice it is suggested that direct examination is more appropriate than the indirect method for estimating CAL. © 2016 FDI World Dental Federation.

Photochemical Reactions of Particulate Organic Matter: Deciphering the Role of Direct and Indirect Processes

NASA Astrophysics Data System (ADS)

Carrasquillo, A. J.; Gelfond, C. E.; Kocar, B. D.

2016-12-01

Photochemical reactions of natural organic matter (NOM) represent potentially important pathways for biologically recalcitrant material to be chemically altered in aquatic systems. Irradiation can alter the physical state of organic matter by facilitating the cycling between the particulate (POM) and dissolved (DOM) pools, however, a molecular level understanding of this chemically dynamic system is currently lacking. Photochemical reactions of a target molecule proceed by the direct absorption of a photon, or through reaction with a second photolytically generated species (i.e. the hydroxyl radical, singlet oxygen, excited triplet state NOM, hydrogen peroxide, etc.). Here, we isolate the major direct and indirect photochemical reactions of a lignocellulose-rich POM material (Phragmites australis) to determine their relative importance in changing the the chemical structure of the parent POM, and in the production of DOM. We measured POM molecular structure using a combination of NMR and FTIR for bulk analyses and scanning transmission x-ray microscopy (STXM) for spatially resolved chemistry, while the chemical composition of photo-produced DOM was measured using ultra-high resolution mass spectrometry. Results are discussed in the context of the differences in chemical composition of both NOM pools resulting from the isolated photochemical pathways. All treatments result in an increase in DOM with reaction time, indicating that the larger POM matrix is likely fragmenting into smaller more soluble species. Spectroscopic measurements, on the other hand, point to functionalization reactions which increase the abundance of alcohol, acid, and carbonyl moieties in both carbon pools. This unique dataset provides new insight into how photochemical reactions alter the chemical composition of NOM while highlighting the relative importance of indirect pathways.

Variance in total levels of phospholipase C zeta (PLC-ζ) in human sperm may limit the applicability of quantitative immunofluorescent analysis as a diagnostic indicator of oocyte activation capability.

PubMed

Kashir, Junaid; Jones, Celine; Mounce, Ginny; Ramadan, Walaa M; Lemmon, Bernadette; Heindryckx, Bjorn; de Sutter, Petra; Parrington, John; Turner, Karen; Child, Tim; McVeigh, Enda; Coward, Kevin

2013-01-01

To examine whether similar levels of phospholipase C zeta (PLC-ζ) protein are present in sperm from men whose ejaculates resulted in normal oocyte activation, and to examine whether a predominant pattern of PLC-ζ localization is linked to normal oocyte activation ability. Laboratory study. University laboratory. Control subjects (men with proven oocyte activation capacity; n = 16) and men whose sperm resulted in recurrent intracytoplasmic sperm injection failure (oocyte activation deficient [OAD]; n = 5). Quantitative immunofluorescent analysis of PLC-ζ protein in human sperm. Total levels of PLC-ζ fluorescence, proportions of sperm exhibiting PLC-ζ immunoreactivity, and proportions of PLC-ζ localization patterns in sperm from control and OAD men. Sperm from control subjects presented a significantly higher proportion of sperm exhibiting PLC-ζ immunofluorescence compared with infertile men diagnosed with OAD (82.6% and 27.4%, respectively). Total levels of PLC-ζ in sperm from individual control and OAD patients exhibited significant variance, with sperm from 10 out of 16 (62.5%) exhibiting levels similar to OAD samples. Predominant PLC-ζ localization patterns varied between control and OAD samples with no predictable or consistent pattern. The results indicate that sperm from control men exhibited significant variance in total levels of PLC-ζ protein, as well as significant variance in the predominant localization pattern. Such variance may hinder the diagnostic application of quantitative PLC-ζ immunofluorescent analysis. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Learning and transfer of category knowledge in an indirect categorization task.

PubMed

Helie, Sebastien; Ashby, F Gregory

2012-05-01

Knowledge representations acquired during category learning experiments are 'tuned' to the task goal. A useful paradigm to study category representations is indirect category learning. In the present article, we propose a new indirect categorization task called the "same"-"different" categorization task. The same-different categorization task is a regular same-different task, but the question asked to the participants is about the stimulus category membership instead of stimulus identity. Experiment 1 explores the possibility of indirectly learning rule-based and information-integration category structures using the new paradigm. The results suggest that there is little learning about the category structures resulting from an indirect categorization task unless the categories can be separated by a one-dimensional rule. Experiment 2 explores whether a category representation learned indirectly can be used in a direct classification task (and vice versa). The results suggest that previous categorical knowledge acquired during a direct classification task can be expressed in the same-different categorization task only when the categories can be separated by a rule that is easily verbalized. Implications of these results for categorization research are discussed.

Assessing Mediational Models: Testing and Interval Estimation for Indirect Effects

ERIC Educational Resources Information Center

Biesanz, Jeremy C.; Falk, Carl F.; Savalei, Victoria

2010-01-01

Theoretical models specifying indirect or mediated effects are common in the social sciences. An indirect effect exists when an independent variable's influence on the dependent variable is mediated through an intervening variable. Classic approaches to assessing such mediational hypotheses (Baron & Kenny, 1986; Sobel, 1982) have in recent years…

Powering up with indirect reciprocity in a large-scale field experiment.

PubMed

Yoeli, Erez; Hoffman, Moshe; Rand, David G; Nowak, Martin A

2013-06-18

A defining aspect of human cooperation is the use of sophisticated indirect reciprocity. We observe others, talk about others, and act accordingly. We help those who help others, and we cooperate expecting that others will cooperate in return. Indirect reciprocity is based on reputation, which spreads by communication. A crucial aspect of indirect reciprocity is observability: reputation effects can support cooperation as long as peoples' actions can be observed by others. In evolutionary models of indirect reciprocity, natural selection favors cooperation when observability is sufficiently high. Complimenting this theoretical work are experiments where observability promotes cooperation among small groups playing games in the laboratory. Until now, however, there has been little evidence of observability's power to promote large-scale cooperation in real world settings. Here we provide such evidence using a field study involving 2413 subjects. We collaborated with a utility company to study participation in a program designed to prevent blackouts. We show that observability triples participation in this public goods game. The effect is over four times larger than offering a $25 monetary incentive, the company's previous policy. Furthermore, as predicted by indirect reciprocity, we provide evidence that reputational concerns are driving our observability effect. In sum, we show how indirect reciprocity can be harnessed to increase cooperation in a relevant, real-world public goods game.

Powering up with indirect reciprocity in a large-scale field experiment

PubMed Central

Yoeli, Erez; Hoffman, Moshe; Rand, David G.; Nowak, Martin A.

2013-01-01

A defining aspect of human cooperation is the use of sophisticated indirect reciprocity. We observe others, talk about others, and act accordingly. We help those who help others, and we cooperate expecting that others will cooperate in return. Indirect reciprocity is based on reputation, which spreads by communication. A crucial aspect of indirect reciprocity is observability: reputation effects can support cooperation as long as peoples’ actions can be observed by others. In evolutionary models of indirect reciprocity, natural selection favors cooperation when observability is sufficiently high. Complimenting this theoretical work are experiments where observability promotes cooperation among small groups playing games in the laboratory. Until now, however, there has been little evidence of observability’s power to promote large-scale cooperation in real world settings. Here we provide such evidence using a field study involving 2413 subjects. We collaborated with a utility company to study participation in a program designed to prevent blackouts. We show that observability triples participation in this public goods game. The effect is over four times larger than offering a $25 monetary incentive, the company’s previous policy. Furthermore, as predicted by indirect reciprocity, we provide evidence that reputational concerns are driving our observability effect. In sum, we show how indirect reciprocity can be harnessed to increase cooperation in a relevant, real-world public goods game. PMID:23754399

Silver stain for electron microscopy

NASA Technical Reports Server (NTRS)

Corbett, R. L.

1972-01-01

Ammoniacal silver stain used for light microscopy was adapted advantageously for use with very thin biological sections required for electron microscopy. Silver stain can be performed in short time, has more contrast, and is especially useful for low power electron microscopy.

Direct and indirect urban water footprints of the United States

NASA Astrophysics Data System (ADS)

Chini, Christopher M.; Konar, Megan; Stillwell, Ashlynn S.

2017-01-01

The water footprint of the urban environment is not limited to direct water consumption (i.e., municipal supplies); embedded water in imported resources, or virtual water transfers, provides an additional component of the urban water footprint. Using empirical data, our analysis extends traditional urban water footprinting analysis to quantify both direct and indirect urban resources for the United States. We determine direct water volumes and their embedded energy through open records requests of water utilities. The indirect component of the urban water footprint includes water indirectly consumed through energy and food, relating to the food-energy-water nexus. We comprehensively quantify the indirect water footprint for 74 metropolitan statistical areas through the combination of various databases, including the Commodity Flow Survey of the U.S. Census Bureau, the U.S. Department of Agriculture, the Water Footprint Network, and the Energy Information Administration. We then analyze spatial heterogeneity in both direct and indirect water footprints, determining the average urban water footprint in the United States to be 1.64 million gallons of water per person per year [6200 m3/person/yr or 17,000 L/person/d], dominated by indirect water. Additionally, our study of the urban water cycle extends beyond considering only water resources to include embedded energy and equivalent carbon dioxide emissions. The inclusion of multiple sectors of the urban water cycle and their underlying processes provides important insights to the overall urban environment, the interdependencies of the food-energy-water nexus, and water resource sustainability. Our results provide opportunities for benchmarking the urban energy-water nexus, water footprints, and climate change potential.

Influence of indirect information on interpersonal trust despite direct information.

PubMed

Zarolia, Pareezad; Weisbuch, Max; McRae, Kateri

2017-01-01

Trust is integral to successful relationships. The development of trust stems from how one person treats others, and there are multiple ways to learn about someone's trust-relevant behavior. The present research captures the development of trust to examine if trust-relevant impressions and behavior are influenced by indirect behavioral information (i.e., descriptions of how a person treated another individual)-even in the presence of substantial direct behavioral information (i.e., self-relevant, first-hand experience with a person). Participants had repeated interpersonal exchanges with a partner who was trustworthy or untrustworthy with participants' money. The present studies vary the frequency with which (Studies 1 & 2), the order in which (Study 3) and the number of people for whom (Study 4) indirect information (i.e., brief vignettes describing trustworthy or untrustworthy behavior) were presented. As predicted, across 4 studies, we observed a robust effect of indirect-information despite the presence of substantial direct information. Even after dozens of interactions in which a partner betrayed (or not), a brief behavioral description of a partner influenced participants' willingness to actually trust the partner with money, memory-based estimates of partner-behavior, and impressions of the partner. These effects were observed even though participants were also sensitive to partners' actual trust behavior, and even when indirect behavioral descriptions were only presented a single time. Impressions were identified as a strong candidate mechanism for the effect of indirect-information on behavior. We discuss implications of the persistence of indirect information for impression formation, relationship development, and future studies of trust. (PsycINFO Database Record (c) 2016 APA, all rights reserved).

3D X-ray ultra-microscopy of bone tissue.

PubMed

Langer, M; Peyrin, F

2016-02-01

We review the current X-ray techniques with 3D imaging capability at the nano-scale: transmission X-ray microscopy, ptychography and in-line phase nano-tomography. We further review the different ultra-structural features that have so far been resolved: the lacuno-canalicular network, collagen orientation, nano-scale mineralization and their use as basis for mechanical simulations. X-ray computed tomography at the micro-metric scale is increasingly considered as the reference technique in imaging of bone micro-structure. The trend has been to push towards increasingly higher resolution. Due to the difficulty of realizing optics in the hard X-ray regime, the magnification has mainly been due to the use of visible light optics and indirect detection of the X-rays, which limits the attainable resolution with respect to the wavelength of the visible light used in detection. Recent developments in X-ray optics and instrumentation have allowed to implement several types of methods that achieve imaging that is limited in resolution by the X-ray wavelength, thus enabling computed tomography at the nano-scale. We review here the X-ray techniques with 3D imaging capability at the nano-scale: transmission X-ray microscopy, ptychography and in-line phase nano-tomography. Further, we review the different ultra-structural features that have so far been resolved and the applications that have been reported: imaging of the lacuno-canalicular network, direct analysis of collagen orientation, analysis of mineralization on the nano-scale and use of 3D images at the nano-scale to drive mechanical simulations. Finally, we discuss the issue of going beyond qualitative description to quantification of ultra-structural features.

Assessing the indirect effects due to natural hazards on a mesoscale

NASA Astrophysics Data System (ADS)

Pfurtscheller, C.; Schwarze, R.

2009-04-01

Measuring indirect economic costs and other effects from natural hazards, especially floods in alpine and other mountainous regions, are a necessary part of a comprehensive economic assessment. Their omission seriously affects the relative economic benefits of structural or non structural measures of flood defence. Surpassing controversial, IO-model-based economic estimates, analysing indirect economic effects lead to the key question of identifying and evaluating the drivers of indirect economic effects and resilience to system effects in the regional economy, i.e. at the meso-level. This investigation takes place for the catastrophic floods in summer 2005 in the provinces of Tyrol and Vorarlberg, Austria, which caused an estimated € 670 Mio direct loss on private and public assets and severe interruptions in lifeline services. The paper starts out with differentiating the concept of indirect economic costs from direct costs, examing different temporal (short vs. long-term) and spatial (macro-, meso- vs. microeconomic) system boundaries. It surveys common theories of economic resilience and vulnerability at the regional economy level. Indirect effects at the regional economy level can be defined as interferences of the economic exchange of goods and services triggered by breakdowns of transport lines and critical production inputs. The extent and persistence of indirect effects of natural hazards is not only by parameters of the extreme event, such as duration and amplitude of the flood, but much more by resilience parameters of the regional economy such as size of enterprises, the network structure (linkages) of the regional economy, availability of insurance and relief funds, and the stock of inventory. These effects can only be dissected by means of expert judgement and event studies. This paper presents the results of a survey conducted among business practioneers, members of chamber of commerce, civil protection agencies to identify and scale the drivers of

Types of suggestibility: Relationships among compliance, indirect, and direct suggestibility.

PubMed

Polczyk, Romuald; Pasek, Tomasz

2006-10-01

It is commonly believed that direct suggestibility, referring to overt influence, and indirect suggestibility, in which the intention to influence is hidden, correlate poorly. This study demonstrates that they are substantially related, provided that they tap similar areas of influence. Test results from 103 students, 55 women and 48 men, were entered into regression analyses. Indirect suggestibility, as measured by the Sensory Suggestibility Scale for Groups, and compliance, measured by the Gudjonsson Compliance Scale, were predictors of direct suggestibility, assessed with the Barber Suggestibility Scale. Spectral analyses showed that indirect suggestibility is more related to difficult tasks on the BSS, but compliance is more related to easy tasks on this scale.

Evidence for Limited Indirect Aerosol Forcing in Stratocumulus

NASA Technical Reports Server (NTRS)

Ackerman, Andrew S.; Toon, O. B.; Stevens, D. E.

2003-01-01

Increases in cloud cover and condensed water contribute more than half of the indirect aerosol effect in an ensemble of general circulation model (GCM) simulations estimating the global radiative forcing of anthropogenic aerosols. We use detailed simulations of marine stratocumulus clouds and airborne observations of ship tracks to show that increases in cloud cover and condensed water in reality are far less than represented by the GCM ensemble. Our results offer an explanation for recent simplified inverse climate calculations indicating that indirect aerosol effects are greatly exaggerated in GCMs.

Food and Drug Administration workshop on indirect mechanisms of carcinogenesis.

PubMed

Poirier, L A

1996-01-01

A workshop sponsored by the Food and Drug Administration (FDA) was held on March 4-5, 1996, at the Lister Hill Auditorium of the National Institutes of Health (NIH) Campus in Bethesda, Maryland. The workshop considered both the scientific aspects and the regulatory implications of indirect-acting carcinogens. A wide variety of agents and of prospective mechanisms was discussed. The organizing committee for the workshop consisted of Drs. James Farrelly and Joseph DeGeorge of the Center for Drug Evaluation and Research (CDER), Ronald J. Lorentzen and Sidney Green of the Center for Food Safety and Applied Nutrition (CFSAN), Martin D. Green of the Center for Biologics, Evaluation and Research (CBER), C. Darnell Jackson and Lionel A. Poirier of the National Center for Toxicological Research (NCTR). Rosalie K. Elespuru of the Center for Devices and Radiological Health (CDRH), and David G. Longfellow of the National Cancer Institute (NCI). Following an introduction by Dr. Poirier, who provided a description of indirect carcinogens, the major talks were grouped into three formal sessions: indirect-acting compounds and agents of FDA interest, biological and biochemical endpoints commonly seen with indirect agents, and specific problems associated with the indirect-acting compounds. A panel discussion followed and the concluding remarks were made by Dr. Bernard A. Schwetz, Associate Commissioner for Science, FDA.

Characterizing nanoscale scanning probes using electron microscopy: A novel fixture and a practical guide

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

Jacobs, Tevis D. B., E-mail: tjacobs@pitt.edu; Wabiszewski, Graham E.; Goodman, Alexander J.

2016-01-15

The nanoscale geometry of probe tips used for atomic force microscopy (AFM) measurements determines the lateral resolution, contributes to the strength of the tip-surface interaction, and can be a significant source of uncertainty in the quantitative analysis of results. While inverse imaging of the probe tip has been used successfully to determine probe tip geometry, direct observation of the tip profile using electron microscopy (EM) confers several advantages: it provides direct (rather than indirect) imaging, requires fewer algorithmic parameters, and does not require bringing the tip into contact with a sample. In the past, EM-based observation of the probe tipmore » has been achieved using ad hoc mounting methods that are constrained by low throughput, the risk of contamination, and repeatability issues. We report on a probe fixture designed for use in a commercial transmission electron microscope that enables repeatable mounting of multiple AFM probes as well as a reference grid for beam alignment. This communication describes the design, fabrication, and advantages of this probe fixture, including full technical drawings for machining. Further, best practices are discussed for repeatable, non-destructive probe imaging. Finally, examples of the fixture’s use are described, including characterization of common commercial AFM probes in their out-of-the-box condition.« less

Characterizing nanoscale scanning probes using electron microscopy: A novel fixture and a practical guide

NASA Astrophysics Data System (ADS)

Jacobs, Tevis D. B.; Wabiszewski, Graham E.; Goodman, Alexander J.; Carpick, Robert W.

2016-01-01

The nanoscale geometry of probe tips used for atomic force microscopy (AFM) measurements determines the lateral resolution, contributes to the strength of the tip-surface interaction, and can be a significant source of uncertainty in the quantitative analysis of results. While inverse imaging of the probe tip has been used successfully to determine probe tip geometry, direct observation of the tip profile using electron microscopy (EM) confers several advantages: it provides direct (rather than indirect) imaging, requires fewer algorithmic parameters, and does not require bringing the tip into contact with a sample. In the past, EM-based observation of the probe tip has been achieved using ad hoc mounting methods that are constrained by low throughput, the risk of contamination, and repeatability issues. We report on a probe fixture designed for use in a commercial transmission electron microscope that enables repeatable mounting of multiple AFM probes as well as a reference grid for beam alignment. This communication describes the design, fabrication, and advantages of this probe fixture, including full technical drawings for machining. Further, best practices are discussed for repeatable, non-destructive probe imaging. Finally, examples of the fixture's use are described, including characterization of common commercial AFM probes in their out-of-the-box condition.

Concepts in Light Microscopy of Viruses

PubMed Central

Witte, Robert; Georgi, Fanny

2018-01-01

Viruses threaten humans, livestock, and plants, and are difficult to combat. Imaging of viruses by light microscopy is key to uncover the nature of known and emerging viruses in the quest for finding new ways to treat viral disease and deepening the understanding of virus–host interactions. Here, we provide an overview of recent technology for imaging cells and viruses by light microscopy, in particular fluorescence microscopy in static and live-cell modes. The review lays out guidelines for how novel fluorescent chemical probes and proteins can be used in light microscopy to illuminate cells, and how they can be used to study virus infections. We discuss advantages and opportunities of confocal and multi-photon microscopy, selective plane illumination microscopy, and super-resolution microscopy. We emphasize the prevalent concepts in image processing and data analyses, and provide an outlook into label-free digital holographic microscopy for virus research. PMID:29670029

Concepts in Light Microscopy of Viruses.

PubMed

Witte, Robert; Andriasyan, Vardan; Georgi, Fanny; Yakimovich, Artur; Greber, Urs F

2018-04-18

Viruses threaten humans, livestock, and plants, and are difficult to combat. Imaging of viruses by light microscopy is key to uncover the nature of known and emerging viruses in the quest for finding new ways to treat viral disease and deepening the understanding of virus–host interactions. Here, we provide an overview of recent technology for imaging cells and viruses by light microscopy, in particular fluorescence microscopy in static and live-cell modes. The review lays out guidelines for how novel fluorescent chemical probes and proteins can be used in light microscopy to illuminate cells, and how they can be used to study virus infections. We discuss advantages and opportunities of confocal and multi-photon microscopy, selective plane illumination microscopy, and super-resolution microscopy. We emphasize the prevalent concepts in image processing and data analyses, and provide an outlook into label-free digital holographic microscopy for virus research.

7 CFR 3430.54 - Indirect costs.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 15 2011-01-01 2011-01-01 false Indirect costs. 3430.54 Section 3430.54 Agriculture Regulations of the Department of Agriculture (Continued) NATIONAL INSTITUTE OF FOOD AND AGRICULTURE COMPETITIVE AND NONCOMPETITIVE NON-FORMULA FEDERAL ASSISTANCE PROGRAMS-GENERAL AWARD ADMINISTRATIVE PROVISIONS...

Entrainment, Drizzle, and the Indirect Effect in Stratiform Clouds

NASA Technical Reports Server (NTRS)

Ackerman, Andrew

2005-01-01

Activation of some fraction of increased concentrations of sub-micron soluble aerosol particles lead to enhanced cloud droplet concentrations and hence smaller droplets, increasing their total cross sectional area and thus reflecting solar radiation more efficiently (the Twomey, or first indirect, effect). However, because of competition during condensational growth, droplet distributions tend to broaden as numbers increase, reducing the sensitivity of cloud albedo to droplet concentration on the order of 10%. Also, smaller droplets less effectively produce drizzle through collisions and coalescence, and it is widely expected (and found in large-scale models) that decreased precipitation leads to clouds with more cloud water on average (the so-called cloud lifetime, or second indirect, effect). Much of the uncertainty regarding the overall indirect aerosol effect stems from inadequate understanding of such changes in cloud water. Detailed simulations based on FIRE-I, ASTEX, and DYCOMS-II conditions show that suppression of precipitation from increased droplet concentrations leads to increased cloud water only when sufficient precipitation reaches the surface, a condition favored when the overlying air is-humid or droplet concentrations are very low. Otherwise, aerosol induced suppression of precipitation enhances entrainment of overlying dry air, thereby reducing cloud water and diminishing the indirect climate forcing.

An improved recommendation algorithm via weakening indirect linkage effect

NASA Astrophysics Data System (ADS)

Chen, Guang; Qiu, Tian; Shen, Xiao-Quan

2015-07-01

We propose an indirect-link-weakened mass diffusion method (IMD), by considering the indirect linkage and the source object heterogeneity effect in the mass diffusion (MD) recommendation method. Experimental results on the MovieLens, Netflix, and RYM datasets show that, the IMD method greatly improves both the recommendation accuracy and diversity, compared with a heterogeneity-weakened MD method (HMD), which only considers the source object heterogeneity. Moreover, the recommendation accuracy of the cold objects is also better elevated in the IMD than the HMD method. It suggests that eliminating the redundancy induced by the indirect linkages could have a prominent effect on the recommendation efficiency in the MD method. Project supported by the National Natural Science Foundation of China (Grant No. 11175079) and the Young Scientist Training Project of Jiangxi Province, China (Grant No. 20133BCB23017).

48 CFR 42.707 - Cost-sharing rates and limitations on indirect cost rates.

Code of Federal Regulations, 2011 CFR

2011-10-01

... final indirect cost rate ceiling in a contract. Examples of such circumstances are when the proposed... ACQUISITION REGULATION CONTRACT MANAGEMENT CONTRACT ADMINISTRATION AND AUDIT SERVICES Indirect Cost Rates 42... authorized, may call for the contractor to participate in the costs of the contract by accepting indirect...

Validation of Digital Microscopy Compared With Light Microscopy for the Diagnosis of Canine Cutaneous Tumors.

PubMed

Bertram, Christof A; Gurtner, Corinne; Dettwiler, Martina; Kershaw, Olivia; Dietert, Kristina; Pieper, Laura; Pischon, Hannah; Gruber, Achim D; Klopfleisch, Robert

2018-07-01

Integration of new technologies, such as digital microscopy, into a highly standardized laboratory routine requires the validation of its performance in terms of reliability, specificity, and sensitivity. However, a validation study of digital microscopy is currently lacking in veterinary pathology. The aim of the current study was to validate the usability of digital microscopy in terms of diagnostic accuracy, speed, and confidence for diagnosing and differentiating common canine cutaneous tumor types and to compare it to classical light microscopy. Therefore, 80 histologic sections including 17 different skin tumor types were examined twice as glass slides and twice as digital whole-slide images by 6 pathologists with different levels of experience at 4 time points. Comparison of both methods found digital microscopy to be noninferior for differentiating individual tumor types within the category epithelial and mesenchymal tumors, but diagnostic concordance was slightly lower for differentiating individual round cell tumor types by digital microscopy. In addition, digital microscopy was associated with significantly shorter diagnostic time, but diagnostic confidence was lower and technical quality was considered inferior for whole-slide images compared with glass slides. Of note, diagnostic performance for whole-slide images scanned at 200× magnification was noninferior in diagnostic performance for slides scanned at 400×. In conclusion, digital microscopy differs only minimally from light microscopy in few aspects of diagnostic performance and overall appears adequate for the diagnosis of individual canine cutaneous tumors with minor limitations for differentiating individual round cell tumor types and grading of mast cell tumors.

A novel fluorescence microscopy approach to estimate quality loss of stored fruit fillings as a result of browning.

PubMed

Cropotova, Janna; Tylewicz, Urszula; Cocci, Emiliano; Romani, Santina; Dalla Rosa, Marco

2016-03-01

The aim of the present study was to estimate the quality deterioration of apple fillings during storage. Moreover, a potentiality of novel time-saving and non-invasive method based on fluorescence microscopy for prompt ascertainment of non-enzymatic browning initiation in fruit fillings was investigated. Apple filling samples were obtained by mixing different quantities of fruit and stabilizing agents (inulin, pectin and gellan gum), thermally processed and stored for 6-month. The preservation of antioxidant capacity (determined by DPPH method) in apple fillings was indirectly correlated with decrease in total polyphenols content that varied from 34±22 to 56±17% and concomitant accumulation of 5-hydroxymethylfurfural (HMF), ranging from 3.4±0.1 to 8±1mg/kg in comparison to initial apple puree values. The mean intensity of the fluorescence emission spectra of apple filling samples and initial apple puree was highly correlated (R(2)>0.95) with the HMF content, showing a good potentiality of fluorescence microscopy method to estimate non-enzymatic browning. Copyright © 2015 Elsevier Ltd. All rights reserved.

Indirect-fired gas turbine bottomed with fuel cell

DOEpatents

Micheli, P.L.; Williams, M.C.; Parsons, E.L.

1995-09-12

An indirect-heated gas turbine cycle is bottomed with a fuel cell cycle with the heated air discharged from the gas turbine being directly utilized at the cathode of the fuel cell for the electricity-producing electrochemical reaction occurring within the fuel cell. The hot cathode recycle gases provide a substantial portion of the heat required for the indirect heating of the compressed air used in the gas turbine cycle. A separate combustor provides the balance of the heat needed for the indirect heating of the compressed air used in the gas turbine cycle. Hot gases from the fuel cell are used in the combustor to reduce both the fuel requirements of the combustor and the NOx emissions therefrom. Residual heat remaining in the air-heating gases after completing the heating thereof is used in a steam turbine cycle or in an absorption refrigeration cycle. Some of the hot gases from the cathode can be diverted from the air-heating function and used in the absorption refrigeration cycle or in the steam cycle for steam generating purposes. 1 fig.

Indirect-fired gas turbine bottomed with fuel cell

DOEpatents

Micheli, Paul L.; Williams, Mark C.; Parsons, Edward L.

1995-01-01

An indirect-heated gas turbine cycle is bottomed with a fuel cell cycle with the heated air discharged from the gas turbine being directly utilized at the cathode of the fuel cell for the electricity-producing electrochemical reaction occurring within the fuel cell. The hot cathode recycle gases provide a substantial portion of the heat required for the indirect heating of the compressed air used in the gas turbine cycle. A separate combustor provides the balance of the heat needed for the indirect heating of the compressed air used in the gas turbine cycle. Hot gases from the fuel cell are used in the combustor to reduce both the fuel requirements of the combustor and the NOx emissions therefrom. Residual heat remaining in the air-heating gases after completing the heating thereof is used in a steam turbine cycle or in an absorption refrigeration cycle. Some of the hot gases from the cathode can be diverted from the air-heating function and used in the absorption refrigeration cycle or in the steam cycle for steam generating purposes.

Clinical Predictors of Hospital Mortality Differ Between Direct and Indirect ARDS.

PubMed

Luo, Liang; Shaver, Ciara M; Zhao, Zhiguo; Koyama, Tatsuki; Calfee, Carolyn S; Bastarache, Julie A; Ware, Lorraine B

2017-04-01

Direct (pulmonary) and indirect (extrapulmonary) ARDS are distinct syndromes with important pathophysiologic differences. The goal of this study was to determine whether clinical characteristics and predictors of mortality differ between direct or indirect ARDS. This retrospective observational cohort study included 417 patients with ARDS. Each patient was classified as having direct (pneumonia or aspiration, n = 250) or indirect (nonpulmonary sepsis or pancreatitis, n = 167) ARDS. Patients with direct ARDS had higher lung injury scores (3.0 vs 2.8; P < .001), lower Simplified Acute Physiology Score II scores (51 vs 62; P < .001), lower Acute Physiology and Chronic Health Evaluation II scores (27 vs 30; P < .001), and fewer nonpulmonary organ failures (1 vs 2; P < .001) compared with patients with indirect ARDS. Hospital mortality was similar (28% vs 31%). In patients with direct ARDS, age (OR, 1.29 per 10 years; P = .01; test for interaction, P = .03), lung injury scores (OR, 2.29 per point; P = .001; test for interaction, P = .058), and number of nonpulmonary organ failures (OR, 1.67; P = .01) were independent risk factors for increased hospital mortality. Preexisting diabetes mellitus was an independent risk factor for reduced hospital mortality (OR, 0.47; P = .04; test for interaction, P = .02). In indirect ARDS, only the number of organ failures was an independent predictor of mortality (OR, 2.08; P < .001). Despite lower severity of illness and fewer organ failures, patients with direct ARDS had mortality rates similar to patients with indirect ARDS. Factors previously associated with mortality during ARDS were only associated with mortality in direct ARDS. These findings suggest that direct and indirect ARDS have distinct features that may differentially affect risk prediction and clinical outcomes. Copyright © 2016 American College of Chest Physicians. Published by Elsevier Inc. All rights reserved.

Difference in the color stability of direct and indirect resin composites

PubMed Central

LEE, Yong-Keun; YU, Bin; LIM, Ho-Nam; LIM, Jin Ik

2011-01-01

Indirect resin composites are generally regarded to have better color stability than direct resin composites since they possess higher conversion degree Objective The present study aimed at comparing the changes in color (∆E) and color coordinates (∆L, ∆a and ∆b) of one direct (Estelite Sigma: 16 shades) and 2 indirect resin composites (BelleGlass NG: 16 shades; Sinfony: 26 shades) after thermocycling. Material and Methods Resins were packed into a mold and light cured; post-curing was performed on indirect resins. Changes in color and color coordinates of 1-mm-thick specimens were determined after 5,000 cycles of thermocycling on a spectrophotometer. Results ∆E values were in the range of 0.3 to 1.2 units for direct resins, and 0.3 to 1.5 units for indirect resins, which were clinically acceptable (∆E<3.3). Based on t-test, ∆E values were not significantly different by the type of resins (p>0.05), while ∆L, ∆a and ∆b values were significantly different by the type of resins (p<0.05). For indirect resins, ∆E values were influenced by the brand, shade group and shade designation based on three-way ANOVA (p<0.05). Conclusion Direct and indirect resin composites showed similar color stability after 5,000 cycles of thermocycling; however, their changes in the color coordinates were different. PMID:21552717

18 CFR 367.4572 - Account 457.2, Indirect costs charged to associate companies.

Code of Federal Regulations, 2010 CFR

2010-04-01

..., Indirect costs charged to associate companies. This account must include recovery of those indirect costs... 18 Conservation of Power and Water Resources 1 2010-04-01 2010-04-01 false Account 457.2, Indirect costs charged to associate companies. 367.4572 Section 367.4572 Conservation of Power and Water...

Direct and Indirect Effects of PM on the Cardiovascular System

PubMed Central

Nelin, Timothy D.; Joseph, Allan M.; Gorr, Matthew W.; Wold, Loren E.

2011-01-01

Human exposure to particulate matter (PM) elicits a variety of responses on the cardiovascular system through both direct and indirect pathways. Indirect effects of PM on the cardiovascular system are mediated through the autonomic nervous system, which controls heart rate variability, and inflammatory responses, which augment acute cardiovascular events and atherosclerosis. Recent research demonstrates that PM also affects the cardiovascular system directly by entry into the systemic circulation. This process causes myocardial dysfunction through mechanisms of reactive oxygen species production, calcium ion interference, and vascular dysfunction. In this review, we will present key evidence in both the direct and indirect pathways, suggest clinical applications of the current literature, and recommend directions for future research. PMID:22119171

INDIRECT EFFECT OF X-RADIATION ON BONE GROWTH IN RATS

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

Conard, R.A.

1962-12-21

Effects of 200 to 600 r of x irradiation on tibial bone growth in groups of weanling male rats were studied by in vivo measurement of tibial bone growth in serial radiographs. By comparison of growth rates in shielded with unshielded legs, direct and indirect effects of radiation were demonstrated, both roughly dose dependent, but with the indirect effect being about twice that of the direct effect. Pair-feeding experiments showed that about 70% of the indirect effect was due to radiation-induced lowered food consumption. By partial-body shielding experiments, using pnir-fed controls, it was shown that the abdomen may be themore » site of a non-nutritional abscopal effect. (auth)« less

The indirect costs of ankylosing spondylitis: a systematic review and meta-analysis.

PubMed

Malinowski, Krzysztof Piotr; Kawalec, Paweł

2015-04-01

The aim of this systematic review was to collect and summarize all current data on the indirect costs related to absenteeism and presenteeism associated with ankylosing spondylitis. The search was conducted using Medline, Embase and Centre for Reviews and Dissemination databases. All collected costs were recalculated to average annual cost per patient, expressed in 2013 prices USD using the consumer price index and purchasing power parity. Identified studies were then analyzed to assess their possible inclusion in the meta-analysis. We identified 32 records. The average annual indirect cost per patient varies among all the identified results from US$660.95 to 45,953.87. The mean annual indirect per patient equals US$6454.76. This systematic review summarizes current data related to indirect costs generated by ankylosing spondylitis; it revealed the great economic burden of the disease for society. We observed a great variety of the considered components of indirect costs and their definitions.

A comparative investigation of seven indirect attitude measures.

PubMed

Bar-Anan, Yoav; Nosek, Brian A

2014-09-01

We compared the psychometric qualities of seven indirect attitude measures across three attitude domains (race, politics, and self-esteem) with a large sample (N = 23,413). We compared the measures on internal consistency, sensitivity to known effects, relationships with indirect and direct measures of the same topic, the reliability and validity of single-category attitude measurement, their ability to detect meaningful variance among people with nonextreme attitudes, and their robustness to the exclusion of misbehaving or well-behaving participants. All seven indirect measures correlated with each other and with direct measures of the same topic. These relations were always weak for self-esteem, moderate for race, and strong for politics. This pattern suggests that some of the sources of variation in the reliability and predictive validity of the indirect measures is a function of the concepts rather than the methods. The Implicit Association Test (IAT) and Brief IAT (BIAT) showed the best overall psychometric quality, followed by the Go–No-Go association task, Single-Target IAT (ST-IAT), Affective Misattribution Procedure (AMP), Sorting Paired Features task, and Evaluative Priming. The AMP showed a steep decline in its psychometric qualities when people with extreme attitude scores were removed. Single-category attitude scores computed for the IAT and BIAT showed good relationships with other attitude measures but no evidence of discriminant validity between paired categories. The other measures, especially the AMP and ST-IAT, showed better evidence for discriminant validity. These results inform us on the validity of the measures as attitude assessments, but do not speak to the implicitness of the measured constructs.

Role of Direct and Indirect Violence Exposure on Externalizing Behavior in Children.

PubMed

Fleckman, Julia M; Drury, Stacy S; Taylor, Catherine A; Theall, Katherine P

2016-06-01

The objective of this study was to examine the association between externalizing behaviors and indirect violence exposure, assessed both within the household and at the community level, as well as the interaction effect of indirect and direct violence exposure. A sample of parents of children ages 4-15 who have not been referred or enrolled in child welfare (n = 82) were recruited from the greater New Orleans community. Externalizing behavior was assessed with the Child Behavior Checklist (CBCL). The child's indirect exposure to violence included witnessing community violence, witnessing physical assault, and witnessing fighting or domestic violence at home. Direct exposure to violence included the child experiencing physical aggression from a caregiver. All assessments were based on caregiver reports. To decrease potential for confounding, children were matched for analysis based on age, Hurricane Katrina exposure, and their propensity to be exposed to high indirect violence. Cumulative indirect exposure to violence was significantly positively correlated with CBCL scores. After controlling for key covariates, CBCL externalizing T score increased significantly by approximately 1.25 points for each level increase in indirect violence exposure (β = 1.25, SE = 0.57, p = 0.027). There also was a significant interaction between indirect and direct exposure to violence in the association with CBCL score (β = -0.08, SE = 0.03, p = 0.002). These findings extend previous research by demonstrating that exposure to both direct and cross-contextual indirect violence influences externalizing behaviors in children. Additionally, the findings suggest that community and household social environments are both important targets for interventions designed to decrease externalizing behaviors and improve long-term outcomes for youth at risk of exposure to violence.

27 CFR 6.42 - Indirect inducement through third party arrangements.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2011-04-01 2011-04-01 false Indirect inducement through third party arrangements. 6.42 Section 6.42 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Furnishing Things of Value § 6.42 Indirect inducement...

27 CFR 6.42 - Indirect inducement through third party arrangements.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Indirect inducement through third party arrangements. 6.42 Section 6.42 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS âTIED-HOUSEâ Unlawful Inducements Furnishing Things of Value § 6.42 Indirect inducement...

Bacillus Anthracis Spores of the bclA Mutant Exhibit Increased Adherence to Epithelial Cells, Fibroblasts, and Endothelial Cells but not to Macrophages

DTIC Science & Technology

2007-09-01

immunofluorescence (IFM) and light microscopy. Samples were fixed in forma- lin, stained with immunofluorescent dyes (as described below) or spore stain ( malachite ...BEC. Adherence was assessed by microscopic observation of the infected cells stained with malachite green and counterstaining of the BEC. For enzymatic...this significant difference, BEC infected with spores were stained with malachite green and counter- stained with Wright-Giemsa (Fig. 1B and C). This

Microscopy techniques in flavivirus research.

PubMed

Chong, Mun Keat; Chua, Anthony Jin Shun; Tan, Terence Tze Tong; Tan, Suat Hoon; Ng, Mah Lee

2014-04-01

The Flavivirus genus is composed of many medically important viruses that cause high morbidity and mortality, which include Dengue and West Nile viruses. Various molecular and biochemical techniques have been developed in the endeavour to study flaviviruses. However, microscopy techniques still have irreplaceable roles in the identification of novel virus pathogens and characterization of morphological changes in virus-infected cells. Fluorescence microscopy contributes greatly in understanding the fundamental viral protein localizations and virus-host protein interactions during infection. Electron microscopy remains the gold standard for visualizing ultra-structural features of virus particles and infected cells. New imaging techniques and combinatory applications are continuously being developed to push the limit of resolution and extract more quantitative data. Currently, correlative live cell imaging and high resolution three-dimensional imaging have already been achieved through the tandem use of optical and electron microscopy in analyzing biological specimens. Microscopy techniques are also used to measure protein binding affinities and determine the mobility pattern of proteins in cells. This chapter will consolidate on the applications of various well-established microscopy techniques in flavivirus research, and discuss how recently developed microscopy techniques can potentially help advance our understanding in these membrane viruses. Copyright © 2013 Elsevier Ltd. All rights reserved.

Posttraumatic Stress Disorder and Suicidal Behavior: Indirect Effects of Impaired Social Functioning.

PubMed

Dutton, Courtney E; Rojas, Sasha M; Badour, Christal L; Wanklyn, Sonya G; Feldner, Matthew T

2016-01-01

Social functioning is negatively impacted by the presence of PTSD, while increasing risk of suicidal behavior among individuals with PTSD. However, little research has examined the specific role of social functioning in the association between PTSD and suicidal behavior. Parallel multiple indirect effects analyses were performed to understand the unique indirect effects of four aspects of social functioning. Indirect effects of PTSD on suicidal ideation were significant through three pathways: interpersonal conflict, perceived family support, and interpersonal apprehension. Perceived family support was the only indirect pathway significantly associated with suicide attempt. Findings suggest that social functioning should be assessed and potentially targeted during treatment to help modify the risk for suicidal behavior among individuals with PTSD.

Practical diagnostic testing for human immunodeficiency virus.

PubMed Central

Jackson, J B; Balfour, H H

1988-01-01

Since the discovery of human immunodeficiency virus (HIV) as the causative agent of acquired immunodeficiency syndrome in 1983, there has been a proliferation of diagnostic tests. These assays can be used to detect the presence of HIV antibody, HIV antigen, HIV ribonucleic and deoxyribonucleic acids, and HIV reverse transcriptase. Enzyme-linked immunosorbent assays, Western blot, radioimmunoprecipitation assays, indirect immunofluorescence assays, reverse transcriptase assays, and several molecular hybridization techniques are currently available. Enzyme-linked immunosorbent, Western blot, and indirect immunofluorescence assays for HIV antibody are very sensitive, specific, and adaptable to most laboratories. An enzyme-linked immunosorbent assay for HIV antigen is also readily adaptable to most laboratories and will be commercially available soon. While the other assays are more tedious, they are valuable confirmatory tests and are suitable for reference laboratories. The biohazards of performing HIV testing can be minimized with proper biosafety measures. Images PMID:3060241

Ultrastructural study of electron dense deposits in renal tubular basement membrane: prevalence and relationship to epithelial atrophy.

PubMed

Yong, Jim L C; Killingsworth, Murray C

2014-08-01

This study reports the prevalence of immune deposits associated with the proximal and distal tubules in a series of routine renal biopsies received in our department during a single calendar year. From 87 cases, 65 (74%) were found to have glomerular immune deposits by immunofluorescence. Tubular immune deposits were found in 12 cases (18%), 3 of which had no glomerular deposits. By transmission electron microscopy (EM), 58 cases (66%) were found to have deposits of granular or vesicular material associated with the tubular basement membranes (TBM). Finely granular electron dense deposits appeared to correspond to the immune deposits seen by immunofluorescence microscopy (IF) and may be a sensitive marker of immune deposition.

Ultrastructural Study of Electron Dense Deposits in Renal Tubular Basement Membrane: Prevalence and Relationship to Epithelial Atrophy

PubMed Central

Killingsworth, Murray C.

2014-01-01

This study reports the prevalence of immune deposits associated with the proximal and distal tubules in a series of routine renal biopsies received in our department during a single calendar year. From 87 cases, 65 (74%) were found to have glomerular immune deposits by immunofluorescence. Tubular immune deposits were found in 12 cases (18%), 3 of which had no glomerular deposits. By transmission electron microscopy (EM), 58 cases (66%) were found to have deposits of granular or vesicular material associated with the tubular basement membranes (TBM). Finely granular electron dense deposits appeared to correspond to the immune deposits seen by immunofluorescence microscopy (IF) and may be a sensitive marker of immune deposition. PMID:24933115

Fluorescence confocal microscopy for pathologists.

PubMed

Ragazzi, Moira; Piana, Simonetta; Longo, Caterina; Castagnetti, Fabio; Foroni, Monica; Ferrari, Guglielmo; Gardini, Giorgio; Pellacani, Giovanni

2014-03-01

Confocal microscopy is a non-invasive method of optical imaging that may provide microscopic images of untreated tissue that correspond almost perfectly to hematoxylin- and eosin-stained slides. Nowadays, following two confocal imaging systems are available: (1) reflectance confocal microscopy, based on the natural differences in refractive indices of subcellular structures within the tissues; (2) fluorescence confocal microscopy, based on the use of fluorochromes, such as acridine orange, to increase the contrast epithelium-stroma. In clinical practice to date, confocal microscopy has been used with the goal of obviating the need for excision biopsies, thereby reducing the need for pathological examination. The aim of our study was to test fluorescence confocal microscopy on different types of surgical specimens, specifically breast, lymph node, thyroid, and colon. The confocal images were correlated to the corresponding histological sections in order to provide a morphologic parallel and to highlight current limitations and possible applications of this technology for surgical pathology practice. As a result, neoplastic tissues were easily distinguishable from normal structures and reactive processes such as fibrosis; the use of fluorescence enhanced contrast and image quality in confocal microscopy without compromising final histologic evaluation. Finally, the fluorescence confocal microscopy images of the adipose tissue were as accurate as those of conventional histology and were devoid of the frozen-section-related artefacts that can compromise intraoperative evaluation. Despite some limitations mainly related to black/white images, which require training in imaging interpretation, this study confirms that fluorescence confocal microscopy may represent an alternative to frozen sections in the assessment of margin status in selected settings or when the conservation of the specimen is crucial. This is the first study to employ fluorescent confocal microscopy on

48 CFR 31.203 - Indirect costs.

Code of Federal Regulations, 2010 CFR

2010-10-01

... fiscal year used for financial reporting purposes in accordance with generally accepted accounting... contract or other work, indirect costs are those remaining to be allocated to intermediate or two or more... basis of the benefits accruing to intermediate and final cost objectives. When substantially the same...

Indirect reciprocity can overcome free-rider problems on costly moral assessment.

PubMed

Sasaki, Tatsuya; Okada, Isamu; Nakai, Yutaka

2016-07-01

Indirect reciprocity is one of the major mechanisms of the evolution of cooperation. Because constant monitoring and accurate evaluation in moral assessments tend to be costly, indirect reciprocity can be exploited by cost evaders. A recent study crucially showed that a cooperative state achieved by indirect reciprocators is easily destabilized by cost evaders in the case with no supportive mechanism. Here, we present a simple and widely applicable solution that considers pre-assessment of cost evaders. In the pre-assessment, those who fail to pay for costly assessment systems are assigned a nasty image that leads to them being rejected by discriminators. We demonstrate that considering the pre-assessment can crucially stabilize reciprocal cooperation for a broad range of indirect reciprocity models. In particular for the most leading social norms, we analyse the conditions under which a prosocial state becomes locally stable. © 2016 The Authors.

Popularity and Resource Control Goals as Predictors of Adolescent Indirect Aggression.

PubMed

Dyches, Karmon D; Mayeux, Lara

2015-01-01

Resource Control Theory conceptualizes aggression as a behavior that allows access to, and control of, limited resources (P. H. Hawley, 1999 ). This study investigated the associations of adolescents' indirect aggression with their resource control goals, or goals related to controlling social resources such as dating opportunities and peer status, and with their levels of popularity and social intelligence. Participants were 109 seventh-graders (52% girls) who completed a resource control goals measure, the Tromsø Social Intelligence Scale, and peer nominations of popularity and indirect aggression. Results indicated positive associations between resource control goals and peer-nominated indirect aggression, with popularity further moderating these associations. These findings suggest that the resource control goals of adolescents can be a motivating force to engage in hurtful behaviors. They provide a context from which peer relations researchers can improve their understanding and prevention of adolescents' indirect aggression.

Concurrent activation of striatal direct and indirect pathways during action initiation.

PubMed

Cui, Guohong; Jun, Sang Beom; Jin, Xin; Pham, Michael D; Vogel, Steven S; Lovinger, David M; Costa, Rui M

2013-02-14

The basal ganglia are subcortical nuclei that control voluntary actions, and they are affected by a number of debilitating neurological disorders. The prevailing model of basal ganglia function proposes that two orthogonal projection circuits originating from distinct populations of spiny projection neurons (SPNs) in the striatum--the so-called direct and indirect pathways--have opposing effects on movement: activity of direct-pathway SPNs is thought to facilitate movement, whereas activity of indirect-pathway SPNs is presumed to inhibit movement. This model has been difficult to test owing to the lack of methods to selectively measure the activity of direct- and indirect-pathway SPNs in freely moving animals. Here we develop a novel in vivo method to specifically measure direct- and indirect-pathway SPN activity, using Cre-dependent viral expression of the genetically encoded calcium indicator (GECI) GCaMP3 in the dorsal striatum of D1-Cre (direct-pathway-specific) and A2A-Cre (indirect-pathway-specific) mice. Using fibre optics and time-correlated single-photon counting (TCSPC) in mice performing an operant task, we observed transient increases in neural activity in both direct- and indirect-pathway SPNs when animals initiated actions, but not when they were inactive. Concurrent activation of SPNs from both pathways in one hemisphere preceded the initiation of contraversive movements and predicted the occurrence of specific movements within 500 ms. These observations challenge the classical view of basal ganglia function and may have implications for understanding the origin of motor symptoms in basal ganglia disorders.

Concurrent Activation of Striatal Direct and Indirect Pathways During Action Initiation

PubMed Central

Cui, Guohong; Jun, Sang Beom; Jin, Xin; Pham, Michael D.

2014-01-01

Summary The basal ganglia are subcortical nuclei that control voluntary actions, and are affected by a number of debilitating neurological disorders1–4. The prevailing model of basal ganglia function proposes that two orthogonal projection circuits originating from distinct populations of spiny projection neurons (SPNs) in the striatum5,6 - the so-called direct and indirect pathways - have opposing effects on movement: while activity of direct-pathway SPNs purportedly facilitates movement, activity of indirect-pathway SPNs inhibits movement1,2. This model has been difficult to test due to the lack of methods to selectively measure the activity of direct- and indirect-pathway SPNs in freely moving animals. We developed a novel in-vivo method that allowed us to specifically measure direct- and indirect-pathway SPN activity using Cre-dependent viral expression of the genetically encoded calcium indicator (GECI) GCAMP3 in the dorsal striatum of D1-Cre (direct-pathway specific6,7) and A2A-Cre (indirect-pathway specific8,9) mice10. Using fiber optics and time-correlated single photon counting (TCSPC) in mice performing an operant task, we observed transient increases in neural activity in both direct- and indirect-pathway SPNs when animals initiated actions, but not when they were inactive. Concurrent activation of SPNs from both pathways in one hemisphere preceded the initiation of contraversive movements, and predicted the occurrence of specific movements within 500 ms. These observations challenge the classical view of basal ganglia function, and may have implications for understanding the origin of motor symptoms in basal ganglia disorders. PMID:23354054

Estimation of indirect effect when the mediator is a censored variable.

PubMed

Wang, Jian; Shete, Sanjay

2017-01-01

A mediation model explores the direct and indirect effects of an initial variable ( X) on an outcome variable ( Y) by including a mediator ( M). In many realistic scenarios, investigators observe censored data instead of the complete data. Current research in mediation analysis for censored data focuses mainly on censored outcomes, but not censored mediators. In this study, we proposed a strategy based on the accelerated failure time model and a multiple imputation approach. We adapted a measure of the indirect effect for the mediation model with a censored mediator, which can assess the indirect effect at both the group and individual levels. Based on simulation, we established the bias in the estimations of different paths (i.e. the effects of X on M [ a], of M on Y [ b] and of X on Y given mediator M [ c']) and indirect effects when analyzing the data using the existing approaches, including a naïve approach implemented in software such as Mplus, complete-case analysis, and the Tobit mediation model. We conducted simulation studies to investigate the performance of the proposed strategy compared to that of the existing approaches. The proposed strategy accurately estimates the coefficients of different paths, indirect effects and percentages of the total effects mediated. We applied these mediation approaches to the study of SNPs, age at menopause and fasting glucose levels. Our results indicate that there is no indirect effect of association between SNPs and fasting glucose level that is mediated through the age at menopause.

Confidence Limits for the Indirect Effect: Distribution of the Product and Resampling Methods

ERIC Educational Resources Information Center

MacKinnon, David P.; Lockwood, Chondra M.; Williams, Jason

2004-01-01

The most commonly used method to test an indirect effect is to divide the estimate of the indirect effect by its standard error and compare the resulting z statistic with a critical value from the standard normal distribution. Confidence limits for the indirect effect are also typically based on critical values from the standard normal…

Correlative fluorescence microscopy and scanning transmission electron microscopy of quantum-dot-labeled proteins in whole cells in liquid.

PubMed

Dukes, Madeline J; Peckys, Diana B; de Jonge, Niels

2010-07-27

Correlative fluorescence microscopy and transmission electron microscopy (TEM) is a state-of-the-art microscopy methodology to study cellular function, combining the functionality of light microscopy with the high resolution of electron microscopy. However, this technique involves complex sample preparation procedures due to its need for either thin sections or frozen samples for TEM imaging. Here, we introduce a novel correlative approach capable of imaging whole eukaryotic cells in liquid with fluorescence microscopy and with scanning transmission electron microscopy (STEM); there is no additional sample preparation necessary for the electron microscopy. Quantum dots (QDs) were bound to epidermal growth factor (EGF) receptors of COS7 fibroblast cells. Fixed whole cells in saline water were imaged with fluorescence microscopy and subsequently with STEM. The STEM images were correlated with fluorescence images of the same cellular regions. QDs of dimensions 7x12 nm were visible in a 5 microm thick layer of saline water, consistent with calculations. A spatial resolution of 3 nm was achieved on the QDs.

Indirect detection of neutrino portal dark matter

NASA Astrophysics Data System (ADS)

Batell, Brian; Han, Tao; Shams Es Haghi, Barmak

2018-05-01

We investigate the feasibility of the indirect detection of dark matter in a simple model using the neutrino portal. The model is very economical, with right-handed neutrinos generating neutrino masses through the type-I seesaw mechanism and simultaneously mediating interactions with dark matter. Given the small neutrino Yukawa couplings expected in a type-I seesaw, direct detection and accelerator probes of dark matter in this scenario are challenging. However, dark matter can efficiently annihilate to right-handed neutrinos, which then decay via active-sterile mixing through the weak interactions, leading to a variety of indirect astronomical signatures. We derive the existing constraints on this scenario from Planck cosmic microwave background measurements, Fermi dwarf spheroidal galaxy and Galactic center gamma-ray observations, and AMS-02 antiproton observations, and we also discuss the future prospects of Fermi and the Cherenkov Telescope Array. Thermal annihilation rates are already being probed for dark matter lighter than about 50 GeV, and this can be extended to dark matter masses of 100 GeV and beyond in the future. This scenario can also provide a dark matter interpretation of the Fermi Galactic center gamma-ray excess, and we confront this interpretation with other indirect constraints. Finally we discuss some of the exciting implications of extensions of the minimal model with large neutrino Yukawa couplings and Higgs portal couplings.

Indirect electroreduction as pretreatment to enhance biodegradability of metronidazole.

PubMed

Saidi, I; Soutrel, I; Floner, D; Fourcade, F; Bellakhal, N; Amrane, A; Geneste, F

2014-08-15

The removal of metronidazole, a biorecalcitrant antibiotic, by coupling an electrochemical reduction with a biological treatment was examined. Electroreduction was performed in a home-made flow cell at -1.2V/SCE on graphite felt. After only one pass through the cell, analysis of the electrolyzed solution showed a total degradation of metronidazole. The biodegradability estimated from the BOD5/COD ratio increased from 0.07 to 0.2, namely below the value usually considered as the limit of biodegradability (0.4). In order to improve these results, indirect electrolysis of metronidazole was performed with a titanium complex known to reduce selectively nitro compounds into amine. The catalytic activity of the titanium complex towards electroreduction of metronidazole was shown by cyclic voltammetry analyses. Indirect electrolysis led to an improvement of the biodegradability from 0.07 to 0.42. To confirm the interest of indirect electroreduction to improve the electrochemical pretreatment, biological treatment was then carried out on activated sludge after direct and indirect electrolyses; different parameters were followed during the culture such as pH, TOC and metronidazole concentration. Both electrochemical processes led to a more efficient biodegradation of metronidazole compared with the single biological treatment, leading to an overall mineralization yield for the coupling process of 85%. Copyright © 2014 Elsevier B.V. All rights reserved.

Indirect Positive Evidence in the Acquisition of a Subset Grammar

ERIC Educational Resources Information Center

Schwartz, Misha; Goad, Heather

2017-01-01

This article proposes that second language learners can use indirect positive evidence (IPE) to acquire a phonological grammar that is a subset of their L1 grammar. IPE is evidence from errors in the learner's L1 made by native speakers of the learner's L2. It has been assumed that subset grammars may be acquired using direct or indirect negative…

17 CFR 256.457-2 - Indirect costs charged to associate companies.

Code of Federal Regulations, 2010 CFR

2010-04-01

... charged to associate companies. This account shall include recovery of those indirect costs which cannot... 17 Commodity and Securities Exchanges 3 2010-04-01 2010-04-01 false Indirect costs charged to... COMMISSION (CONTINUED) UNIFORM SYSTEM OF ACCOUNTS FOR MUTUAL SERVICE COMPANIES AND SUBSIDIARY SERVICE...

Engineering and Narrative: Literary Prerequisites as Indirect Communication for Technical Writing

ERIC Educational Resources Information Center

Jeyaraj, Joseph

2014-01-01

While Engineering values direct communication, indirect communication produces a kind of literacy salient for engineers that direct communication may not offer in the way indirect communication does. This article emphasizes the inadequacies of overly emphasizing direct communication for Engineering majors and explains how teaching indirect…

21 CFR 174.5 - General provisions applicable to indirect food additives.

Code of Federal Regulations, 2013 CFR

2013-04-01

... additives. 174.5 Section 174.5 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) INDIRECT FOOD ADDITIVES: GENERAL § 174.5 General provisions applicable to indirect food additives. (a) Regulations prescribing conditions under...