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Sample records for fluorescein diacetate staining

  1. NEW METHOD TO DETERMINE 'GIARDIA' CYST VIABILITY: CORRELATION OF FLUORESCEIN DIACETATE AND PROPIDIUM IODIDE STAINING WITH ANIMAL INFECTIVITY

    EPA Science Inventory

    The viability of Giardia muris cysts was studied using the fluorogenic dyes, fluorescein diacetate (FDA) and propidium iodide (PI). Using the mouse model for giardiasis, FDA or PI stained cysts were inoculated into neonatal mice. Feces were examined at days 3, 5, 8, and 11 post-i...

  2. Limitations in the Use of Fluorescein Diacetate/Propidium Iodide (FDA/PI) and Cell Permeable Nucleic Acid Stains for Viability Measurements of Isolated Islets of Langerhans

    PubMed Central

    Boyd, Vinc; Cholewa, Olivia Maria; Papas, Klearchos K.

    2010-01-01

    Background A review of current literature shows that the combined use of the cell permeable esterase-substrate fluorescein diacetate (FDA) and the cell impermeant nucleic acid stain propidium iodide (PI) to be one of the most common fluorescence-based methods to assess the viability of isolated islets of Langerhans, and it is currently used for islet product release prior to transplantation in humans. However, results from this assay do not correlate with islet viability and function or islet transplantation success in animals or humans (Eckhard et al. 2004; Ricordi et al. 2001). This may be in part attributed to considerable differences as well as discrepancies in the use of these reagents on islets. We critically surveyed the literature and evaluated the impact of a number of variables associated with the use of FDA/PI to determine their reliability in assessing islet cell viability. In addition, we evaluated other fluorescent stains, such as SYTO®13, SYTO®24 and SYBR®14 as possible alternatives to FDA. Results We found that the stability of stains in storage and stock solutions, the number of islets stained, concentration of stains, staining incubation time, the buffer/media used, and the method of examining islets were significant in the final scoring of viability. For archival file photos, the exposure time and camera/software settings can also impact interpretation of viability. Although our results show that FDA does detect intracellular esterase activity and staining with PI does assess cell membrane integrity, the results obtained from using these stains did not correlate directly with expected islet function and viability per transplantation into diabetic athymic nude mice (Papas et al. 2007). In addition, the use of two nucleic acid stains, such as SYTO®13 and PI, for live/dead scoring exhibited staining anomalies which limit their accuracy in assessing islet viability. Conclusions From a review of the literature and from our observations on the impact of reagent handling and various staining and imaging parameters used to visually evaluate islets, consistent interpretation of islet cell membrane integrity and viability is dependent upon a number of factors. We discuss the utility and limitations of these reagents in evaluating islet cell membrane integrity and viability. PMID:20814586

  3. Limitations in the Use of Fluorescein Diacetate/Propidium Iodide (FDA/PI) and Cell Permeable Nucleic Acid Stains for Viability Measurements of Isolated Islets of Langerhans.

    PubMed

    Boyd, Vinc; Cholewa, Olivia Maria; Papas, Klearchos K

    2008-03-01

    BACKGROUND: A review of current literature shows that the combined use of the cell permeable esterase-substrate fluorescein diacetate (FDA) and the cell impermeant nucleic acid stain propidium iodide (PI) to be one of the most common fluorescence-based methods to assess the viability of isolated islets of Langerhans, and it is currently used for islet product release prior to transplantation in humans. However, results from this assay do not correlate with islet viability and function or islet transplantation success in animals or humans (Eckhard et al. 2004; Ricordi et al. 2001). This may be in part attributed to considerable differences as well as discrepancies in the use of these reagents on islets. We critically surveyed the literature and evaluated the impact of a number of variables associated with the use of FDA/PI to determine their reliability in assessing islet cell viability. In addition, we evaluated other fluorescent stains, such as SYTO(R)13, SYTO(R)24 and SYBR(R)14 as possible alternatives to FDA. RESULTS: We found that the stability of stains in storage and stock solutions, the number of islets stained, concentration of stains, staining incubation time, the buffer/media used, and the method of examining islets were significant in the final scoring of viability. For archival file photos, the exposure time and camera/software settings can also impact interpretation of viability. Although our results show that FDA does detect intracellular esterase activity and staining with PI does assess cell membrane integrity, the results obtained from using these stains did not correlate directly with expected islet function and viability per transplantation into diabetic athymic nude mice (Papas et al. 2007). In addition, the use of two nucleic acid stains, such as SYTO(R)13 and PI, for live/dead scoring exhibited staining anomalies which limit their accuracy in assessing islet viability. CONCLUSIONS: From a review of the literature and from our observations on the impact of reagent handling and various staining and imaging parameters used to visually evaluate islets, consistent interpretation of islet cell membrane integrity and viability is dependent upon a number of factors. We discuss the utility and limitations of these reagents in evaluating islet cell membrane integrity and viability. PMID:20814586

  4. Fluorescein eye stain

    MedlinePlus

    ... will be stained by the dye and appear green under the blue light. The provider can determine the location and likely cause of the cornea problem depending on the size, location, and shape of the staining.

  5. Fluorescein isothiocyanate staining and characterization of avian heterophils.

    PubMed

    Rath, N C; Huff, G R; Balog, J M; Huff, W E

    1998-06-30

    Fluorescein isothiocyanate (FITC) was found to stain cytoplasmic granules of avian heterophilgranulocytes. In tissue sections, the fluorescent granulocytes were predominantly distributed adjacent to trabecular bones. The fluorescein stained granulocytes were abundant in synovial fluids of chickens with synovitis. A significant correlation was observed in the percent of fluorescein labeled granulocytes in blood smears and the percent of heterophils determined using an automated counting method, in unstained blood from normal and Escherichia coli-infected turkeys. The fluorescein-binding heterophils purified from chickens showed a time dependent increases in the oxidation of 2',7'-dichlorofluorescin diacetate (DCF-DA) and the reduction of nitroblue tetrazolium (NBT) which were indicative of changes in oxidative burst in response to phorbol 12-myristate 13-acetate (PMA), Salmonella typhimurium lipopolysaccharide (LPS), and zymosan A (ZA). These heterophil-activating agents, also, caused significant degranulation at 16 h post-treatment, as indicated by the loss fluorescence. There were microscopically visible alterations in the cell shapes and a decrease in the density of granules due to treatment with LPS, PMA or ZA. In addition, these cells also showed phagocytic response which was evident at 30 min of incubation with fluorescent latex particles. Both chicken and turkey heterophils produced interleukin-6 in vitro at 24 h in response to LPS but not to PMA, FMLP or ZA. The chicken heterophils showed spontaneous production of matrix metalloproteinases (MMP) which was significantly enhanced by treatment with LPS, PMA, and ZA; however, LPS appeared to be most effective in inducing MMP production. These results demonstrate that the functions of heterophils can be differentially regulated by different activating agents and the fluorescein binding property of these cells may be useful for their histochemical identification. PMID:9656433

  6. Rapid cost-effective analysis of microbial activity in soils using modified fluorescein diacetate method.

    PubMed

    Schumacher, Thomas E; Eynard, Anna; Chintala, Rajesh

    2015-03-01

    Fluorescein diacetate (FDA) is commonly used to determine the hydrolyzing activity of microbial organisms in the soil. However, the costs of chemical reagents and time required to perform routine analysis of large number of samples by soil testing laboratories are limiting. Moreover, existing methods generate significant volumes of hazardous waste. In this context, this study was designed to determine the minimum amount of terminating chemical reagent needed to evaluate microbial hydrolyzing activity. The results showed that 0.2 mL of chloroform was enough to effectively stop the hydrolyzing activity in soil. This proposed terminating chemical reagent (0.2 mL chloroform) was also evaluated by comparing with the 10 mL of chloroform and 5 mL of methanol used in the Adam and Duncan method. PMID:25471725

  7. High Sensitivity Bacillus thuringiensis Cry1Ac Protein Detections Using Fluorescein Diacetate Nanoparticles.

    PubMed

    Liu, Cui; Zhou, Zhen; Zou, Linling; Cao, Yuan-Cheng; Liu, Jun'An; Lin, Yongjun

    2016-03-01

    A highly sensitive transgenic protein analysis method was proposed here based on fluorescein diacetate (FDA). First, FDA was prepared by the ball mill to harvest the nano-sized organic particles. Further examines showed that the FDA size can be controlled by the speed of centrifugation which can obtain FDA in well-distributed size. Cy3 antibody immobilization tests showed that the proteins can attach onto the FDA particles while keep bioactivities. FDA and Cry1Ac antibody immunoassay tests showed that when the FDA particle was in 150 nm, the linear range was 0.01 ng/L-30 μg/mL. And it has the lower detection limitation of 0.01 ng/L, which is 100 times more sensitive than the ELISA methods. These results indicate that the FDA related immunoassays are the promising approach in the transgenic analysis. PMID:26642804

  8. A rapid method for viability and drug sensitivity of Mycobacterium leprae cultured in macrophages and using fluorescein diacetate.

    PubMed

    Bhagria, A; Mahadevan, P R

    1987-01-01

    The ability of viable M. leprae to hydrolyze Fluorescein diacetate and retain fluorescein inside the bacteria was used to identify viable M. leprae inside the cultured in vitro macrophages. The subjective microscopic count of the FDA test was demonstrated as useful routine test by confirming the results obtained therein with a quantitative and non subjective measurement of fluorescence in spectrofluorimeter. Using this method loss of viability of M. leprae in presence of dapsone and rifampicin was demonstrated. Such an assay, was well correlated with another in vitro assay, the Fc receptor test and also the in vivo mouse foot test. The drug resistance of clinical isolates of M. leprae demonstrated by mouse foot pad was also correlated with FDA test system. Thus we have reported a reliable, consistent and rapid in vitro test system for determining viability and drug sensitivity of M. leprae. PMID:2440960

  9. Biochar amendment to lead-contaminated soil: Effects on fluorescein diacetate hydrolytic activity and phytotoxicity to rice.

    PubMed

    Tan, Xiaofei; Liu, Yunguo; Gu, Yanling; Zeng, Guangming; Hu, Xinjiang; Wang, Xin; Hu, Xi; Guo, Yiming; Zeng, Xiaoxia; Sun, Zhichao

    2015-09-01

    The amendment effects of biochar on total microbial activity was measured by fluorescein diacetate (FDA) hydrolytic activity, and phytotoxicity in Pb(II)-contaminated soils was examined by the application of 4 different biochars to soil, with rice as a test plant. The FDA hydrolytic activities of biochar-amended soils were much higher than that of the control. The survival rate of rice in lead-contaminated biochar-amended soils showed significant improvement over the control, especially for bamboo biochar-amended soil (93.3%). In addition, rice grown in lead-contaminated control sediment displayed lower biomass production than that in biochar-amended soil. The immobilization of Pb(II) and the positive effects of biochar amendment on soil microorganisms may account for these effects. The results suggest that biochar may have an excellent ability to mitigate the toxic effects of Pb(II) on soil microorganisms and rice. PMID:25900615

  10. Development of an automated ballast water treatment verification system utilizing fluorescein diacetate hydrolysis as a measure of treatment efficacy.

    PubMed

    Akram, A C; Noman, S; Moniri-Javid, R; Gizicki, J P; Reed, E A; Singh, S B; Basu, A S; Banno, F; Fujimoto, M; Ram, J L

    2015-03-01

    Methods for verifying ballast water treatments in foreign vessels are needed to protect the Great Lakes from the discharge of live non-native organisms or pathogens. A prototype automated viability test system using fluorescein diacetate (FDA), a membrane permeable fluorogen, to differentiate live from dead bacteria and algae is described. The automated fluorescence intensity detection device (AFIDD) captures cultured algae or organisms in Detroit River water (simulated ballast water) on 0.2 μm filters, backwashes them from the filter into a cuvette with buffer and FDA for subsequent fluorescence intensity measurements, and washes the filters with sterile water for serial automated reuse. Preliminary manual versions of these procedures were also tested. Tests of various buffers determined N,N-Bis(2-hydroxyethyl)-2-aminoethanesulfonic acid, N,N-Bis(2-hydroxyethyl)taurine (BES) and 3-(N-morpholino)propanesulfonic acid (MOPS) at pH 7.0 to be the best buffers, causing the least spontaneous FDA breakdown without inhibiting enzymatic activity. Fluorescence in the presence of live organisms increased linearly over time, and the rate of increase was dependent on the sample concentration. Following simulated ballast water treatments with heat or chlorine, the fluorescence produced by Detroit River samples decreased to near control (sterile water) levels. Automated measurements of FDA hydrolysis with a reusable filter backwash system should be applicable to near real-time remote-controlled monitoring of live organisms in ballast water. PMID:25555225

  11. Intraoperative Fluorescein Staining of Cryopreserved Amniotic Membrane Grafts to Improve Visualization During and After Pterygium Surgery: A Novel Technique

    PubMed Central

    Martinez, J. Alberto; Korchak, Michael; Cremers, Sandra L.

    2016-01-01

    Purpose: To describe a new method of enhancing the visualization of amniotic membrane grafts with fluorescein staining during pterygium surgery. Methods: Pterygium excision surgery using intraoperatively stained cryopreserved amniotic membranes was performed on 346 eyes. A sterile 0.6 mg sodium fluorescein strip was placed directly onto the amniotic membrane in the manufacturer's original packaging, and the stained allograft was then transplanted onto the planned site. Staining intensities, at 3, 5, and 10 minutes of dye immersion, were compared. Immediate postoperative pain rating (scale 010), visibility of the fluorescein-stained amniotic membrane graft, and conjunctival autograft and amniotic membrane graft elevation, dehiscence, retraction, or displacement were recorded. The recurrence rate of the study population was compared with that of a previous cohort of 121 patients who underwent pterygium excision with conjunctival autograft without stained amniotic membrane. Results: Direct contact of the fluorescein strip on the amniotic membrane at 3, 5, and 10 minutes showed no differences in subjective staining intensity. Fluorescein-stained amniotic membrane was easily detected on the ocular surface during and 24 hours after pterygium surgery. The average immediate postoperative pain rating was 0.8 1.8. No intraoperative complications or postoperative amniotic membrane graft dehiscence, retraction, or displacement occurred. The recurrence rate using fluorescein-stained amniotic membrane (3 patients, 0.9%, mean follow-up time 31.8 18.6 weeks) did not differ from that of the previous cohort without the stained amniotic membrane (2.5%; ?2(1) = 1.837, P = 0.183). Conclusions: Fluorescein strip staining of the amniotic membrane is a novel and safe intraoperative method to enhance visualization and handling of the graft during and after ocular surgeries. PMID:26751995

  12. Fluorescein Punctate Staining Traced to Superficial Corneal Epithelial Cells by Impression Cytology and Confocal Microscopy

    PubMed Central

    Mokhtarzadeh, Maryam; Casey, Richard

    2011-01-01

    Purpose. The basis of fluorescein-associated superficial punctate staining in dry eyes is controversial. Prior explanations include fluorescein pooling in surface erosive defects, intercellular trapping of fluorescein, and intracellular staining in dead cells. In this study, the hypothesis that punctate erosions are individual cells with enhanced fluorescence was tested. Methods. Ten impression cytology membrane materials were compared, to optimize cellular yield in buccal mucosa and cornea. Clinicocytologic correlation of punctate fluorescent spots was performed in four dry eye patients. Individual punctate spots were localized by fiducial marks in photographs, before and after removal with impression membranes, and were traced in fluorescence microscopy and cytologic staining. Two-way contingency table analysis was used to determine the correlation of punctate spots with cells removed by the membrane. Clinicopathologic correlation of punctate spots was performed in 10 corneas removed in dry eye patients by transplantation for concurrent diseases. Punctate fluorescence was tracked in specimens by fiducial marks and epifluorescence. The distribution of fluorescent spots in specific cell layers of the cornea was determined by confocal microscopy. Results. Cellular yield was greatest with impressions from polytetrafluoroethylene (PTFE [Teflon]; BioPore; Millipore, Billerica, MA) membrane compared with its closest rival (P = 0.019). Punctate fluorescent spots, most of which disappeared after impression cytology (71%), correlated with cells on the membranes (P = 0.009). The punctate spots were more frequent in the superficial cell layers of the cornea (80%) compared with the deepest two layers (0%) (P < 0.00049). Conclusions. Punctate epithelial erosions correspond to enhanced fluorescence in epithelial cells predominantly in superficial layers of the cornea and would be more aptly named fluorescent epithelial cells (FLECs). PMID:21212176

  13. Fluorescent Staining of Tea Pathogenic Fungi in Tea Leaves Using Fluorescein-labeled Lectin

    NASA Astrophysics Data System (ADS)

    Yamada, Kengo; Yoshida, Katsuyuki; Sonoda, Ryoichi

    Fluorochrome-labeled lectin, fluorescein conjugated wheat germ agglutinin (F-WGA) was applied to stain tea pathogenic fungi in tea leaf tissue. Infected leaves were fixed and decolorized with a mixture of ethanol and acetic acid, and cleared with 10% KOH for whole mount before staining with F-WGA. Hyphae of Pestalotiopsis longiseta, Pseudocercospora ocellata, Botrytis cinerea and Colletotrichum theae-sinensis fluoresced brightly in whole mount and sectioned samples of infected leaf tissue. In browned tissue, hyphae did not fluoresce frequently in whole mount sample. Autofluorescence of leaf tissue was strong in browned tissue of sections, it was removed by 10% KOH treatment before staining. Penetration hyphae of C. theae-sinensis in cell wall of trichome and hyphae in basal part of trichome did not fluoresced frequently. In whole mount samples of tea leaf infected with Exobasidium vexans and E. reticulatum, hymenia appeared on leaf surface fluoresced, but hyphae in leaf tissue did not fluoresce. In sectioned samples, hyphae fluoresced brightly when sections were treated with 10% KOH before staining.

  14. Direct staining of mouse T lymphoblasts with fluoresceinated Vicia villosa lectin.

    PubMed Central

    Lang, I; Banga, J P; Varey, A M; Gunn, H; Cooke, A; Roitt, I M

    1982-01-01

    Fluorescein conjugated Vicia villosa (Vv) lectin was used for direct staining of the surface of viable cells of various mouse lymphocyte populations. A varying proportion of polyclonally-activated T cells expressed Vv receptor although less strongly than cells of an influenza virus-specific cytotoxic T lymphocyte (CTL) clone and blasts generated in the mixed lymphocyte reaction. Cyclosporin A (CyA) treatment markedly reduced the expression of Vv receptor following concanavalin A (Con A) stimulation by between 66% and 93% with a concurrent inhibition of blastogenesis and complete abrogation of cytolytic function. Resting mouse lymphocytes and B-cell blasts were always Vv-negative. However, non-specific suppressor factor producing non-cytotoxic T-cell lines also expressed Vv receptor as shown by the weak, but specific, surface fluorescence of EL4 and BW5147 cells stained with Vv. Vv-positive cells were not restricted to a particular Ly phenotype, Vv-positive cells being found among both Lyt 1+ and Lyt2+ MLC stimulated lymphocytes. Our data suggest that the receptor for Vv lectin cannot be regarded as an exclusive differential marker for CTL. Images Figure 1 Figure 2 PMID:6179856

  15. Highly sensitive and simple fluorescence staining of proteins in sodium dodecyl sulfate-polyacrylamide-based gels by using hydrophobic tail-mediated enhancement of fluorescein luminescence.

    PubMed

    Kang, Chulhun; Kim, Hyun Jung; Kang, Donghoon; Jung, Duk Young; Suh, Myungkoo

    2003-10-01

    Fluorescein has an extremely low luminescence intensity in acidic aqueous media. However, when it was bound to proteins, subsequent increase of luminescence intensity took place. Furthermore, when a hydrophobic tail, such as aliphatic hydrocarbons, was introduced to fluorescein, more dramatic increase of luminescence intensity was observed upon binding to proteins. In the present study, by utilizing this luminescence enhancement, three hydrophobic fluorescein dyes (5-dodecanoyl amino fluorescein, 5-hexadecanoyl amino fluorescein, and 5-octadecanoyl amino fluorescein) were examined as noncovalent fluorescent stains of protein bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Effective incorporation of the dyes to proteins in gels was accomplished either simply by adding dyes at the protein fixation step, or by treating gels with a staining solution after the fixation. The sensitivity of this staining method using the fluorescein derivatives was approximately 1 ng/band for most proteins. For some cases, protein bands containing as low as 0.1 ng were successfully visualized. In addition, the detection sensitivity showed much less protein-to-protein variation than silver staining. This new staining method was also successfully applied to two-dimensional electrophoresis of rat brain proteins. Its overall sensitivity was comparable to that of silver staining. PMID:14595675

  16. Proteomic analysis of carbonylated proteins in two-dimensional gel electrophoresis using avidin-fluorescein affinity staining.

    PubMed

    Yoo, Byoung-Sam; Regnier, Fred E

    2004-05-01

    A method for detecting carbonylated proteins in two-dimensional electrophoresis (2-DE) was developed using biotinylation and avidin-fluorescein isothiocyanate (FITC) affinity staining. The method was used to examine oxidatively modified proteins associated with oxidative stress. Carbonyl formation in proteins was first examined in a model system by subjecting bovine serum albumin (BSA) and ribonuclease A (RNase A) to metal-catalyzed oxidation (MCO). Carbonyl group formation was found to occur at multiple sites along with a small amount of polypeptide chain cleavage. In vivo studies were conducted in yeast cell cultures using 5 mM hydrogen peroxide to induce oxidative stress. Biotinylation of yeast protein was accomplished during extraction at 4 degrees C in a lysis buffer containing 5 mM biotin-hydrazide. Biotin-hydrazide forms a Schiff base with a carbonyl group on an oxidized protein that is subsequently reduced before electrophoresis. Proteins were separated by either 2-DE or sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Biotinylated species were detected using avidin-FITC affinity staining. Detection sensitivity with biotinylated proteins was five times higher than achieved by silver staining. The limit of detection with avidin-FITC staining approached 0.64 pmol of protein-associated carbonyls. Twenty carbonylated proteins were identified in the proteome of yeast following oxidative stress with hydrogen peroxide. Matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) analysis of tryptic peptides was used to identify peptides extracted from gels. Aconitase, heat shock protein SSA1 and SSC1, pyruvate decarboxylase isozyme 1, pyruvate kinase 1, enolase 1 and 2, phosphoglycerate kinase, fructose-bisphosphate aldorase, and glyceraldehyde-3-phosphate dehydrogenase were among the major targets of oxidative stress. PMID:15174056

  17. A preservable two color staining procedure to detect toxicant impacts on algae (Selenastrum capricornutum) using flow cytometry

    SciTech Connect

    Faber, M.; Smith, L.; Boermans, H.; Stephenson, G.; Thompson, D.G.

    1995-12-31

    Over the last several years, the use of flow cytometry to assess the impacts of toxicants on algae has increased. Previous studies have tested cell viability using chlorophyll autofluorescence or single stain flow cytometric analysis in fresh algal cultures. A rapid, two-color flow cytometric assay to evaluate viability and cytotoxicity of Selenastrum capricomutum in preserved samples is described. The staining procedure involved fluorescein diacetate, a fluorogenic esterase substrate cleaved in viable cells to fluorescein (green 525 nm) and ethidium homodimer-1 dye which passes through plasma membranes of compromised and dead cells staining DNA (red 620 nm). The auto fluorescence of chlorophyll-a (deep red 675 nm) was used to assess cell viability and examined for use as an internal comparison with the staining procedure. For storage, stained cells were fixed in glutaraldehyde, flash frozen in liquid nitrogen and stored frozen ({minus}20C) for assessment at a later date. Weekly analysis of stored samples showed that the fluorescence of fluorescein diacetate and ethidium homodimer-1 stained cells was stable under these conditions for the 2 months used in this study. A prepared culture of Selenastrum capricomutum containing a 50% (v/v) mixture of live and heat killed cells showed 36.1 % stained live, 13.2% as compromised, 12% unlabeled, and 38.7% dead algal cells. This two-color flow cytometric procedure has proven to be a reliable, sensitive assay to determine viability of Selenastrum capricomutum in preserved samples. The sensitivity of this dual-color assay and the ability to store samples for later analysis are significant improvements over current techniques. This method is being tested for detection of chemical induced algal cytotoxicity and for potential adaptations to field studies.

  18. The photography of fluorescein

    SciTech Connect

    Welch, J.D.

    1982-06-01

    The last few years have seen a number of new flaps described and a renewed interest in the use of fluorescein, but there have been few photographs of the fluorescein effect, because special light sources were required with the filters that were employed. The realization that fluorescein can be excited by electromagnetic radiation in the visible range allows a simplified technique in which an ordinary electronic flash unit may serve as the only light source. The photography of fluorescein is not difficult to perform, and since minimal additional equipment is required, all workers who use fluorescein should begin to document their work more accurately and dramatically.

  19. 21 CFR 582.6197 - Calcium diacetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Calcium diacetate. 582.6197 Section 582.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is...

  20. 21 CFR 182.6197 - Calcium diacetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Calcium diacetate. 182.6197 Section 182.6197 Food... HUMAN CONSUMPTION (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Sequestrants 1 § 182.6197 Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is generally recognized...

  1. 21 CFR 582.6197 - Calcium diacetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Calcium diacetate. 582.6197 Section 582.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is...

  2. 21 CFR 582.6197 - Calcium diacetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Calcium diacetate. 582.6197 Section 582.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is...

  3. 21 CFR 182.6197 - Calcium diacetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Calcium diacetate. 182.6197 Section 182.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Sequestrants 1 § 182.6197 Calcium diacetate. (a) Product. Calcium diacetate....

  4. 21 CFR 182.6197 - Calcium diacetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Calcium diacetate. 182.6197 Section 182.6197 Food... HUMAN CONSUMPTION (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Sequestrants 1 § 182.6197 Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is generally recognized...

  5. 21 CFR 182.6197 - Calcium diacetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Calcium diacetate. 182.6197 Section 182.6197 Food... HUMAN CONSUMPTION (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Sequestrants 1 § 182.6197 Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is generally recognized...

  6. 21 CFR 582.6197 - Calcium diacetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Calcium diacetate. 582.6197 Section 582.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is...

  7. 21 CFR 182.6197 - Calcium diacetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Calcium diacetate. 182.6197 Section 182.6197 Food... HUMAN CONSUMPTION (CONTINUED) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Sequestrants 1 § 182.6197 Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is generally recognized...

  8. 21 CFR 582.6197 - Calcium diacetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Calcium diacetate. 582.6197 Section 582.6197 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Calcium diacetate. (a) Product. Calcium diacetate. (b) Conditions of use. This substance is...

  9. 21 CFR 582.6754 - Sodium diacetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Sodium diacetate. 582.6754 Section 582.6754 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium diacetate. (a) Product. Sodium diacetate. (b) Conditions of use. This substance is...

  10. 21 CFR 582.6754 - Sodium diacetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Sodium diacetate. 582.6754 Section 582.6754 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium diacetate. (a) Product. Sodium diacetate. (b) Conditions of use. This substance is...

  11. 21 CFR 582.6754 - Sodium diacetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Sodium diacetate. 582.6754 Section 582.6754 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium diacetate. (a) Product. Sodium diacetate. (b) Conditions of use. This substance is...

  12. 21 CFR 582.6754 - Sodium diacetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Sodium diacetate. 582.6754 Section 582.6754 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium diacetate. (a) Product. Sodium diacetate. (b) Conditions of use. This substance is...

  13. 21 CFR 582.6754 - Sodium diacetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Sodium diacetate. 582.6754 Section 582.6754 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Sodium diacetate. (a) Product. Sodium diacetate. (b) Conditions of use. This substance is...

  14. Fluorescein Derivatives in Intravital Fluorescence Imaging

    PubMed Central

    Robertson, Thomas A.; Bunel, Florestan; Roberts, Michael S.

    2013-01-01

    Intravital fluorescence microscopy enables the direct imaging of fluorophores in vivo and advanced techniques such as fluorescence lifetime imaging (FLIM) enable the simultaneous detection of multiple fluorophores. Consequently, it is now possible to record distribution and metabolism of a chemical in vivo and to optimise the delivery of fluorophores in vivo. Recent clinical applications with fluorescein and other intravital fluorescent stains have occurred in neurosurgery, dermatology [including photodynamic therapy (PDT)] and endomicroscopy. Potential uses have been identified in periodontal disease, skin graft and cancer surgery. Animal studies have demonstrated that diseased tissue can be specifically stained with fluorophore conjugates. This review focuses on the fluorescein derived fluorophores in common clinical use and provides examples of novel applications from studies in tissue samples. PMID:24709799

  15. 21 CFR 184.1754 - Sodium diacetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Sodium diacetate. 184.1754 Section 184.1754 Food... Specific Substances Affirmed as GRAS § 184.1754 Sodium diacetate. (a) Sodium diacetate (C4H7O4Na·xH2O, CAS Reg. No. 126-96-5) is a molecular compound of acetic acid, sodium acetate, and water of hydration....

  16. 21 CFR 184.1754 - Sodium diacetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sodium diacetate. 184.1754 Section 184.1754 Food... Specific Substances Affirmed as GRAS § 184.1754 Sodium diacetate. (a) Sodium diacetate (C4H7O4Na·xH2O, CAS Reg. No. 126-96-5) is a molecular compound of acetic acid, sodium acetate, and water of hydration....

  17. 21 CFR 184.1754 - Sodium diacetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Sodium diacetate. 184.1754 Section 184.1754 Food... GRAS § 184.1754 Sodium diacetate. (a) Sodium diacetate (C4H7O4Na·xH2O, CAS Reg. No. 126-96-5) is a molecular compound of acetic acid, sodium acetate, and water of hydration. The technical grade is...

  18. 21 CFR 184.1754 - Sodium diacetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Sodium diacetate. 184.1754 Section 184.1754 Food... Specific Substances Affirmed as GRAS § 184.1754 Sodium diacetate. (a) Sodium diacetate (C4H7O4Na·xH2O, CAS Reg. No. 126-96-5) is a molecular compound of acetic acid, sodium acetate, and water of hydration....

  19. 21 CFR 184.1754 - Sodium diacetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Sodium diacetate. 184.1754 Section 184.1754 Food... Specific Substances Affirmed as GRAS § 184.1754 Sodium diacetate. (a) Sodium diacetate (C4H7O4Na·xH2O, CAS Reg. No. 126-96-5) is a molecular compound of acetic acid, sodium acetate, and water of hydration....

  20. Confusion over live/dead stainings for the detection of vital microorganisms in oral biofilms - which stain is suitable?

    PubMed Central

    2014-01-01

    Background There is confusion over the definition of the term “viability state(s)” of microorganisms. “Viability staining” or “vital staining techniques” are used to distinguish live from dead bacteria. These stainings, first established on planctonic bacteria, may have serious shortcomings when applied to multispecies biofilms. Results of staining techniques should be compared with appropriate microbiological data. Discussion Many terms describe “vitality states” of microorganisms, however, several of them are misleading. Authors define “viable” as “capable to grow”. Accordingly, staining methods are substitutes, since no staining can prove viability. The reliability of a commercial “viability” staining assay (Molecular Probes) is discussed based on the corresponding product information sheet: (I) Staining principle; (II) Concentrations of bacteria; (III) Calculation of live/dead proportions in vitro. Results of the “viability” kit are dependent on the stains’ concentration and on their relation to the number of bacteria in the test. Generally this staining system is not suitable for multispecies biofilms, thus incorrect statements have been published by users of this technique. To compare the results of the staining with bacterial parameters appropriate techniques should be selected. The assessment of Colony Forming Units is insufficient, rather the calculation of Plating Efficiency is necessary. Vital fluorescence staining with Fluorescein Diacetate and Ethidium Bromide seems to be the best proven and suitable method in biofilm research. Regarding the mutagenicity of staining components users should be aware that not only Ethidium Bromide might be harmful, but also a variety of other substances of which the toxicity and mutagenicity is not reported. Summary – The nomenclature regarding “viability” and “vitality” should be used carefully. – The manual of the commercial “viability” kit itself points out that the kit is not suitable for natural multispecies biofilm research, as supported by an array of literature. – Results obtained with various stains are influenced by the relationship between bacterial counts and the amount of stain used in the test. Corresponding vitality data are prone to artificial shifting. – As microbiological parameter the Plating Efficiency should be used for comparison. – Ethidium Bromide is mutagenic. Researchers should be aware that alternative staining compounds may also be or even are mutagenic. PMID:24410850

  1. Gram stain

    MedlinePlus

    A Gram stain is a test used to identify bacteria. It is one of the most common ways to ... Urethral discharge Gram stain; Feces Gram stain; Stool Gram stain; Joint fluid Gram stain; Pericardial fluid Gram stain; Gram stain of ...

  2. Intracarotid Fluorescein Angiography

    PubMed Central

    Feindel, William; Yamamoto, Y. Lucas; Hodge, Charles P.

    1967-01-01

    Patterns of blood flow were examined in the surface vessels of the surgically exposed brain by intracarotid injection of 1% fluorescein and rapid serial photographs timed by a photo-cell signal. Matching colour filters were used for black and white or Ektachrome film. As developed in cats and monkeys, and applied in five patients during craniotomy, the technique gave a picture of flow patterns in the pial and cortical vascular bed, demonstrating “water-shed” areas bordering major arterial territories, laminar flow in veins, and, in particular, the details of filling and clearing in the fine pial vessels, the superficial cortical capillary bed and in the vascular beds of tumours. Since these features are rendered in finer detail and sharper contrast than by standard x-ray angiography, the method affords a new means of more adequately examining the epicerebral circulation in man during craniotomy for a variety of lesions. ImagesFig. 1Fig. 2aFig. 2bFig. 3Fig. 4Fig. 5Fig. 6 PMID:4959610

  3. "Geyser" leakage on fluorescein angiography.

    PubMed

    Levy, Jaime; Fagan, Xavier J; Lifshitz, Tova; Schneck, Marina

    2013-01-01

    An 82-year-old patient with diabetes was followed up due to moderate nonproliferative diabetic retinopathy with macular edema in the right eye. Visual acuity was 6/36. Focal macular laser was conducted (A). Three years later, the patient presented with blurry vision in the right eye. Visual acuity was 3/60. Vitreous hemorrhage was observed (B), and neovascularization of the disc was suspected (C). Fluorescein angiography (D, mid venous phase; E-F, recirculation phase) confirmed neovascularization of the disc and depicted a striking vertical leakage. Panretinal photocoagulation was started. Possible explanations for the "geyser" leakage may be either a partial posterior vitreous detachment allowing the fluorescein to track upwards but not elsewhere or a pocket of syneretic vitreous allowing the fluorescein passage in which to diffuse, much like the passage the blood would have taken. PMID:24548789

  4. Method and compositions for detecting of bloodstains using fluorescin-fluorescein reaction

    DOEpatents

    Di Benedetto, John; Kyle, Kevin; Boan, Terry; Marie, Charlene

    2004-02-17

    A method, compositions and kit are set forth for detecting blood stains. A reactant solution includes fluorescin solubilized (reduced) in acetic acid in ethanol. The solution may be buffered to a pH of approximately 9. After spraying the reactant solution on the suspected area an oxidizer is applied to promote the fluorescin to fluorescein reaction with the blood. The reacted fluorescein is then detected through luminescence for capture by photography.

  5. Gram Stain

    MedlinePlus

    ... Gram Stain Related tests: Susceptibility Testing , Bacterial Wound Culture , Blood Culture , Body Fluid Analysis , CSF Analysis , Urine Culture , AFB Testing , Gonorrhea , Stool Culture , Fungal Tests , Sputum ...

  6. Evaluation of intravenous fluorescein in intradermal allergy testing in psittacines.

    PubMed

    Nett, Claudia S; Hosgood, Giselle; Heatley, J Jill; Foil, Carol S; Tully, Thomas N

    2003-12-01

    This study was designed to improve the clinical feasibility of intradermal skin testing of psittacine birds using intravenous fluorescein stain. Twenty-five healthy, anaesthetized Hispaniolan Amazon parrots (Amazona ventralis) were injected intravenously with 10 mg kg-1 fluorescein-sodium 1% followed by intradermal injections of 0.02 mL phosphate-buffered saline, histamine phosphate (1:100,000 w/v) and codeine phosphate (1:100,000 w/v) at the sternal apteria. Wheal diameters of reaction sites were measured grossly and under illumination with a Wood's lamp after 5 and 10 min. Fluorescence-enhanced injection sites were scored between 0 and 2, with 0 equivalent to normal skin and 2 equivalent to a plucked feather follicle. The presence of a fluorescent halo around intradermal injections was also recorded. Under Wood's light illumination at 10 min, histamine and saline were evaluated as positive and negative controls, respectively, based on a positive test having a halo and a score of 2. Sensitivity and specificity were each 76% for halo, 84 and 42% for score and 64 and 77% for combination of score and halo, respectively. Further, mean histamine reactions were significantly larger than codeine phosphate and saline (8.8 +/- 0.4 mm; 7.2 +/- 0.3 mm; 5.9 +/- 0.6 mm); however, this finding was not consistent in individual birds. Wheal size, halo presence and score were affected by site location independent from the injected compound. Intravenous fluorescein improved the readability of avian skin tests; however, the compounds tested raised inconsistent reactions in wheal size, score or halo presence. The compound-independent site effect raises concern on the validity of avian skin testing and warrants investigation of other techniques such as in vitro allergy testing. Based on our findings, intradermal allergy testing in psittacines with or without fluorescein is unreliable and cannot be recommended for practical clinical use. PMID:14678444

  7. 77 FR 23269 - Determination That FUNDUSCEIN-25 (fluorescein sodium injection), 25%, and AK-FLUOR (fluorescein...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-04-18

    ... injection), 25%, and AK-FLUOR (fluorescein sodium injection), 25%, Were Not Withdrawn From Sale for Reasons...%, and AK-FLUOR (fluorescein sodium injection), 25%, were not withdrawn from sale for reasons of safety... include what is now section 505(j)(7) of the Federal Food, Drug, and Cosmetic Act (21 U.S.C....

  8. Oral fluorescein angiography and fluoroscopy: determination of plasma fluorescein levels and clinical application.

    PubMed

    Gómez-Ulla, F; Malvar, A; Parafita, M; Polo, P; Seoane, I

    1992-12-01

    One hundred oral fluorescein angiographic explorations were performed in 85 patients of both sexes, mean age 55.57 years, and range 8 to 77 years. Fluorescein was administered as one 250-mg (15 patients) or 500-mg (70 patients) capsule per 15 kg of body weight. In 15 fasting and 15 postprandial patients administered 500-mg capsules, no significant differences by group were found between fluorescein plasma levels. Fundus fluorescence under slitlamp illumination was poor for patients receiving the lower dosage of fluorescein, whereas at the higher dosage it was visible in all but one patient. The best filters were found to be a Kodak-Wratten 47A for the excitation filter and Kodak-Wratten 8 for the barrier filter. With a conventional fundus camera and fluorescein angiography filters, good photographs were achieved in all cases. The integrity of the blood-retina barriers was explored for various chorioretinal pathologies. PMID:1300524

  9. Advantage of Microscope Integrated for Both Indocyanine Green and Fluorescein Videoangiography on Aneurysmal Surgery: Case Report

    PubMed Central

    YOSHIOKA, Hideyuki; KINOUCHI, Hiroyuki; NISHIYAMA, Yoshihisa; KANEMARU, Kazuya; YAGI, Takashi; HANIHARA, Mitsuto; HORIKOSHI, Toru

    2014-01-01

    Neck clipping of a large middle cerebral artery aneurysm was performed using a newly developed surgical microscope integrated with modules for both indocyanine green (ICG) and fluorescein videoangiography. During surgery, ICG and fluorescein videoangiography by intra-arterial or intravenous injection were safely carried out without interrupting the surgical procedure. Based on the findings obtained from the case, we evaluated the differences between the dyes and the injection routes. With intra-arterial injection, fluorescein offered sharper contrast images and was better at depicting fine arteries than ICG. Patchy staining of vessel walls was observed in intravenous fluorescein videoangiography, while it was not evident in ICG. Intra-arterial injection method had a great advantage in the rapid clearance of the dyes, which allowed us to perform repeated videoangiography within a short period, and was useful in detecting incomplete clipping in this case; however, catheter insertion requires additional work and carries a potential risk. Use of a microscope integrated for both ICG and fluorescein videoangiography would be another method for repeated evaluation. Namely, alternate use of the dyes enables us to perform videoangiography in a short time even via intravenous injection. PMID:24097092

  10. Advantage of microscope integrated for both indocyanine green and fluorescein videoangiography on aneurysmal surgery: case report.

    PubMed

    Yoshioka, Hideyuki; Kinouchi, Hiroyuki; Nishiyama, Yoshihisa; Kanemaru, Kazuya; Yagi, Takashi; Hanihara, Mitsuto; Horikoshi, Toru

    2014-01-01

    Neck clipping of a large middle cerebral artery aneurysm was performed using a newly developed surgical microscope integrated with modules for both indocyanine green (ICG) and fluorescein videoangiography. During surgery, ICG and fluorescein videoangiography by intra-arterial or intravenous injection were safely carried out without interrupting the surgical procedure. Based on the findings obtained from the case, we evaluated the differences between the dyes and the injection routes. With intra-arterial injection, fluorescein offered sharper contrast images and was better at depicting fine arteries than ICG. Patchy staining of vessel walls was observed in intravenous fluorescein videoangiography, while it was not evident in ICG. Intra-arterial injection method had a great advantage in the rapid clearance of the dyes, which allowed us to perform repeated videoangiography within a short period, and was useful in detecting incomplete clipping in this case; however, catheter insertion requires additional work and carries a potential risk. Use of a microscope integrated for both ICG and fluorescein videoangiography would be another method for repeated evaluation. Namely, alternate use of the dyes enables us to perform videoangiography in a short time even via intravenous injection. PMID:24097092

  11. 21 CFR 522.1078 - Gonadorelin diacetate tetrahydrate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Gonadorelin diacetate tetrahydrate. 522.1078 Section 522.1078 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL...

  12. IRRADIATION (GAMMA) OF FINE EMULSION SAUSAGE THAT CONTAINTED SODIUM DIACETATE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Listeria monocytogenes, a psychrotrophic food-borne pathogen, is a frequent post-process contaminant on ready-to-eat meat (RTE) products including frankfurters and bologna. Ionizing radiation can eliminate L. monocytogenes from ready-to-eat meats. Sodium diacetate (SDA) inhibits the growth of L. mo...

  13. 21 CFR 522.1078 - Gonadorelin diacetate tetrahydrate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Gonadorelin diacetate tetrahydrate. 522.1078 Section 522.1078 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL...

  14. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2004-02-01

    This unit describes protocols for detecting protein in a gel by either Coomassie blue, silver or fluorescent staining. Alternate rapid staining procedures are provided for each method and a support protocol describes how to photograph stained gels. Fluorescent staining (e.g., with SYPRO Orange or Red) is described as a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Staining of proteins in SDS-polyacrylamide gels is described, and an alternate protocol details variations in the procedure for proteins in nondenaturing gels. A final support protocol describes the photography of fluorescently stained proteins. PMID:18432935

  15. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2009-01-01

    This unit describes protocols for detecting protein in a gel by Coomassie blue, silver, or fluorescent staining. As a general protein stain, Coomassie is easier and more rapid; however, fluorescent and silver staining methods are considerably more sensitive and thus can be used to detect smaller amounts of protein. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Alternate protocols describe rapid Coomassie and silver staining methods, as well as fluorescent stains that are specific for phosphoproteins and glycoproteins. Staining of proteins in SDS-polyacrylamide gels is described; variations for fluorescent staining of proteins in nondenaturing gels are also included. Support protocols describe photography of stained proteins. PMID:19170026

  16. Port-Wine Stain

    MedlinePlus

    ... and rashes clinical tools newsletter | contact Share | Port-Wine Stain A parent's guide for infants and babies ... a three-month-old infant with a port-wine stain. Overview A port-wine stain is a ...

  17. Electrospun fluorescein/polymer composite nanofibers and their photoluminescent properties

    NASA Astrophysics Data System (ADS)

    Zhang, Zhi-Hua; Long, Yun-Ze; Yin, Hong-Xin; Sun, Bin; Zheng, Jie; Zhang, Hong-Di; Ji, Xin-Ming; Gu, Chang-Zhi

    2012-09-01

    Fluorescein/polyvinyl pyrrolidone (PVP) composite nanofibers with different fluorescein loadings (with a weight concentration of 0-5.0%) are fabricated via electrospinning. Morphologies, structures and photoluminescent (PL) properties of these straight, helical or wavelike fibers are characterized by scanning electron microscopy (SEM), fluorescence microscopy and a spectrophotometer. It is found that the maximum emission of the as-spun fluorescein/PVP fibers occurs at 510 nm. The PL intensity of the composite fiber increases with fluorescein concentration, then fluorescence quenching appears when the concentration reaches 1.67%. The mechanism of fluorescence quenching of fluorescein is discussed. In addition, the composite fibers exhibit a much stronger PL intensity than fluorescein/PVP bulk film owing to larger specific surface area, which makes them promising materials for biomedical applications such as probes and sensors.

  18. Staining proteins in gels.

    PubMed

    Gallagher, Sean; Chakavarti, Deb

    2008-01-01

    Following separation by electrophoretic methods, proteins in a gel can be detected by several staining methods. This unit describes protocols for detecting proteins by four popular methods. Coomassie blue staining is an easy and rapid method. Silver staining, while more time consuming, is considerably more sensitive and can thus be used to detect smaller amounts of protein. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and is often as sensitive as silver staining. Staining of proteins with SYPRO Orange and SYPRO Ruby are also demonstrated here. PMID:19066521

  19. Staining proteins in gels.

    PubMed

    Sasse, Joachim; Gallagher, Sean R

    2003-08-01

    This unit describes protocols for detecting protein in a gel by either Coomassie blue staining or silver staining. The former is easier and more rapid; however, silver staining methods are considerably more sensitive and thus can be used to detect smaller amounts of protein. Alternate rapid staining procedures are provided for each method and a support protocol describes how to photograph stained gels. Fluorescent staining is a popular alternative to traditional staining procedures, mainly because it is more sensitive than Coomassie staining, and often as sensitive as silver staining. Staining of proteins in SDS-polyacrylamide gels is described, and an alternate protocol details variations in the procedure for proteins in nondenaturing gels. A final support protocol describes the photography of fluorescently stained proteins. PMID:18265316

  20. [Leopard-spot pattern in fluorescein angiography].

    PubMed

    Friedel, S; Polak, A

    2013-04-01

    The uveal effusion syndrome is a rare disease characterized by serous choroidal detachment. The pathogenesis of idiopathic uveal effusion syndrome has not yet been conclusively established. One hypothesis is an abnormality of diffusion of extravascular proteins in the choroid leading to decompensation of the pigment epithelium pumping capacity. Fluid then accumulates in the subretinal space leading to retinal detachment which results in loss of visual acuity. It typically affects males and hypermetropia is another risk factor. When looking at the fundus a circular serous detachment of the choroid and choroidal puckering is typical. The fluorescein angiography shows hyperfluorescence in the form of a leopard-spot pattern. Space-occupying lesions have to be excluded with the help of ultrasound or magnetic resonance tomography. The uveal effusion syndrome is a diagnosis by exclusion. Treatment varies because of the different hypotheses for the pathogenesis. An intraocular tamponade in combination with laser coagulation may for example be an effective treatment. PMID:23338531

  1. Protein stains and applications.

    PubMed

    Sundaram, Ranjini K; Balasubramaniyan, Natarajan; Sundaram, Pazhani

    2012-01-01

    Staining of proteins separated on gels provides the basis for determination of the critical properties of these biopolymers, such as their molecular weight and/or charge. Detection of proteins on gels and blots require stains. These stains vary in sensitivity, ease of use, color, stability, versatility, and specificity. This review discusses different stains and applications with details on how to use the advantages and disadvantages of each stain. It also compiles some important points to be considered in imaging and evaluation. Commonly used colorimetric and fluorescent dyes for general protein staining, and posttranslational modification-specific detection methods are also discussed. PMID:22585510

  2. Simplified method for DNA and protein staining of human hematopoietic cell samples. [Cell flow systems

    SciTech Connect

    Crissman, H.A.; Egmond, J.V.; Holdrinet, R.S.; Pennings, A.; Haanen, C.

    1981-01-01

    A rapid reproducible method yielding high resolution analysis of DNA and protein in human hematopoietic cell samples has been developed by modification of the propidium iodide and fluorescein isothiocyanate procedure. Cell staining involves sequential addition of each reagent (RNase, fluorescein isothiocyanate and propidium iodide) to ethanol-fixed cells and requires no centrifugation steps. Stained cells are analyzed in the reagent solutions. Analysis of bone marrow samples from multiple myeloma patients showed mixed normal and aneuploid populations with a major portion of the aneuploid cells having a significantly higher protein content. This approach permitted differential cell cycle analysis of normal and the aneuploid populations.

  3. Fluorescent staining of gels.

    PubMed

    Buxbaum, Engelbert

    2012-01-01

    Certain transition metal complexes show intensive fluorescence when bound to proteins. They can be used to stain gels after electrophoresis with a sensitivity approaching that of silver staining, but in a much simpler and more reproducible procedure. Stains can be prepared easily and at a fraction of the cost of commercially available reagents.Hydrophobic dyes can be used to stain gels without fixing; they do not interfere with later blotting or electro-elution. PMID:22585519

  4. Port-wine stain

    MedlinePlus

    A port-wine stain is a birthmark in which swollen blood vessels create a reddish-purplish discoloration of the skin. ... Port-wine stains occur in about 3 out of 1,000 people. In rare cases, port-wine stains are ...

  5. Fluorescein-labeled glutathione to study protein S-glutathionylation.

    PubMed

    Landino, Lisa M; Brown, Carolyn M; Edson, Carolyn A; Gilbert, Laura J; Grega-Larson, Nathan; Wirth, Anna Jean; Lane, Kelly C

    2010-07-01

    Numerous studies of S-glutathionylation of cysteine thiols indicate that this protein modification plays a key role in redox regulation of proteins. To facilitate the study of protein S-glutathionylation, we developed a synthesis and purification to produce milligram quantities of fluorescein-labeled glutathione. The amino terminus of the glutathione tripeptide reacted with fluorescein isothiocyanate readily in ammonium bicarbonate. Purification by solid phase extraction on C8 and C18 columns separated excess reactants from desired products. Both oxidized and reduced fluorescein-labeled glutathione reacted with a variety of thiol-containing proteins to yield fluorescent proteins. PMID:20156418

  6. Gram stain of urethral discharge

    MedlinePlus

    Urethral discharge Gram stain ... microscope slide. A series of stains called a Gram stain is applied to the specimen. The stained ... culture ) should be performed in addition to the gram stain. More sophisticated diagnostic tests (such as PCR ...

  7. Using carboxyfluorescein diacetate succinimidyl ester to monitor intracellular protein glycation.

    PubMed

    Boucher, Julie; Simard, Élie; Froehlich, Ulrike; D'Orléans-Juste, Pedro; Grandbois, Michel

    2015-06-01

    Protein glycation is a ubiquitous process involved in vascular complications observed in diabetes. Glyoxal (GO), an intracellular reactive oxoaldehyde that is one of the most potent glycation agents, readily reacts with amines present on proteins to produce the lysine-derived adduct carboxymethyllysine, which is a prevalent advanced glycation end-product (AGE). Our group previously showed that cell exposure to GO leads to an alteration in the cell contractile activity that could occur as a result of the glycation of various proteins regulating the cell contractile machinery. Here, we measured the extent of glycation on three functionally distinct proteins known to participate in cell contraction and cytoskeletal organization-Rho-kinase (ROCK), actin, and gelsolin (GSN)-using an assay based on the reaction of the cell membrane-permeable fluorescent probe carboxyfluorescein diacetate succinimidyl ester (CFDA-SE), which reacts with primary amine groups of proteins. By combining CFDA-SE fluorescence and Western blot detection, we observed (following GO incubation) increased glycation of actin and ROCK as well as an increased interaction between actin and GSN as observed by co-immunoprecipitation. Thus, we conclude that the use of the fluorescent probe CFDA-SE offers an interesting alternative to perform a comparative analysis of the extent of intracellular protein glycation in live cells. PMID:25800564

  8. Ultrabright Fluorescein-Labeled Antibodies Near Silver Metallic Surfaces

    PubMed Central

    Lakowicz, Joseph R.; Malicka, Joanna; Huang, Jun; Gryczynski, Zygmunt; Gryczynski, Ignacy

    2009-01-01

    Fluorescein-labeled antibodies are widely used in clinical assays and fluorescence microscopy. The fluorescent signal per labeled antibody is limited by fluorescein self-quenching, which occurs when the antibody is heavily labeled with multiple fluoresceins. We examined immunoglobulin G (IgG) when labeled with 0.7 to about 30 fluoresceins per antibody molecule. The extent of self-quenching was decreased, and the signal increased, when the labeled antibody was in close proximity to metallic silver particles. Time-resolved measurements showed that the intensity increase was due in part to a silver-induced increase in the radiative decay rate. These results suggest the use of labeled antibodies conjugated to silver particles as ultrabright probes for imaging or analytical applications. PMID:15274090

  9. Acute Toxicity of Sodium Fluorescein to Ashy Pebblesnails Fluminicola fuscus

    USGS Publications Warehouse

    Stockton, Kelly A.; Moffitt, Christine M.; Blew, David L.; Farmer, C. Neil

    2011-01-01

    Water resource agencies and groundwater scientists use fluorescein dyes to trace ground water flows that supply surface waters that may contain threatened or endangered mollusk species. Since little is known of the toxicity of sodium fluorescein to mollusks, we tested the toxicity of sodium fluorescein to the ashy pebblesnail Fluminicola fuscus. The pebblesnail was selected as a surrogate test species for the threatened Bliss Rapid snail Taylorcocha serpenticola that is endemic to the Snake River and its tributaries in the Hagerman Valley, Idaho. In laboratory tests, we expose replicated groups of snails to a series of concentrations of fluorescein in a static 24 h exposure at 15 degrees C. Following the exposure, we removed snails, rinsed them, and allowed a 48 h recovery in clean water before recording mortality. We estimated 377 mg/L as the median lethal dose. Mortality to snails occurred at concentrations well above those expected in test wells during the monitoring efforts.

  10. Stain-less staining for computed histopathology

    PubMed Central

    Mayerich, David; Walsh, Michael J.; Kadjacsy-Balla, Andre; Ray, Partha S.; Hewitt, Stephen M.; Bhargava, Rohit

    2015-01-01

    Dyes such as hematoxylin and eosin (H&E) and immunohistochemical stains have been increasingly used to visualize tissue composition in research and clinical practice. We present an alternative approach to obtain the same information using stain-free chemical imaging. Relying on Fourier transform infrared (FT-IR) spectroscopic imaging and computation, stainless computed histopathology can enable a rapid, digital, quantitative and non-perturbing visualization of morphology and multiple molecular epitopes simultaneously in a variety of research and clinical pathology applications. PMID:26029735

  11. Dramatic Stained Glass.

    ERIC Educational Resources Information Center

    Prater, Michael

    2002-01-01

    Describes an art project that is appropriate for students in fifth through twelfth grade in which they create Gothic-style stained-glass windows. Discusses how college students majoring in elementary education created stained-glass windows. Addresses how to adapt this lesson for younger students. (CMK)

  12. Localization of Legionella pneumophila in tissue using FITC-conjugated specific antibody and a background stain

    SciTech Connect

    Lowry, B.S.; Vega, F.G. Jr.; Hedlund, K.W.

    1982-05-01

    Lightly staining formalin-fixed or fresh tissue with Gram's crystal violet obviates interfering nonspecific fluorescence by acting as a metachromatic stain in ultraviolet light. Against the easily recognized background of tissues and cells fluorescein isothiocyanate-tagged Legionella pneumophila antibodies can then identify this bacterium in or on individual cells. This procedure can be run at room temperature in two hours and has the potential for further widespread applicability.

  13. Ayanin diacetate-induced cell death is amplified by TRAIL in human leukemia cells

    SciTech Connect

    Marrero, Maria Teresa; Estevez, Sara; Negrin, Gledy; Quintana, Jose; Leon, Francisco; Estevez, Francisco

    2012-11-09

    Highlights: Black-Right-Pointing-Pointer Ayanin diacetate as apoptotic inducer in leukemia cells. Black-Right-Pointing-Pointer Cell death was prevented by caspase inhibitors and by the overexpression of Bcl-x{sub L}. Black-Right-Pointing-Pointer The intrinsic and the extrinsic pathways are involved in the mechanism of action. Black-Right-Pointing-Pointer Death receptors are up-regulated and TRAIL enhances apoptotic cell death. -- Abstract: Here we demonstrate that the semi-synthetic flavonoid ayanin diacetate induces cell death selectively in leukemia cells without affecting the proliferation of normal lymphocytes. Incubation of human leukemia cells with ayanin diacetate induced G{sub 2}-M phase cell cycle arrest and apoptosis which was prevented by the non-specific caspase inhibitor z-VAD-fmk and reduced by the overexpression of Bcl-x{sub L}. Ayanin diacetate-induced cell death was found to be associated with: (i) loss of inner mitochondrial membrane potential, (ii) the release of cytochrome c, (iii) the activation of multiple caspases, (iv) cleavage of poly(ADP-ribose) polymerase and (v) the up-regulation of death receptors for TRAIL, DR4 and DR5. Moreover, the combined treatment with ayanin diacetate and TRAIL amplified cell death, compared to single treatments. These results provide a basis for further exploring the potential applications of this combination for the treatment of cancer.

  14. Detection of disseminated peritoneal tumors by fluorescein diacrylate in mice

    NASA Astrophysics Data System (ADS)

    Harada, Yoshinori; Furuta, Hirokazu; Murayama, Yasutoshi; Dai, Ping; Fujikawa, Yuta; Urano, Yasuteru; Nagano, Tetsuo; Morishita, Koki; Hasegawa, Akira; Takamatsu, Tetsuro

    2009-02-01

    Tumor invasion to the peritoneum is a poor prognostic factor in cancer patients. Accurate diagnosis of disseminated peritoneal tumors is essential to accurate cancer staging. To date, peritoneal washing cytology during laparotomy has been used for diagnosis of peritoneal dissemination of gastrointestinal cancer, but its sensitivity has not been satisfactory. Thus, a more direct approach is indispensable to detect peritoneal dissemination in vivo. Fluorescein diacrylate (FDAcr) is an esterase-sensitive fluorescent probe derived from fluorescein. In cancer cells, fluorescent fluorescein generated by exogenous application of FDAcr selectively deposits owing to its stronger hydrolytic enzyme activity and its lower leakage rate. We examined whether FDAcr can specifically detect disseminated peritoneal tumors in athymic nude mouse models. Intraperitoneally administered FDAcr revealed disseminated peritoneal microscopic tumors not readily recognized on white-light imaging. These results suggest that FDAcr is a useful probe for detecting disseminated peritoneal tumors.

  15. Effect of 2% fluorescein on Scheimpflug central corneal thickness measurements

    PubMed Central

    Briggs, Stella; Bin Moammar, Marwa

    2016-01-01

    AIM To assess central corneal thickness (CCT) changes measured with Scheimpflug device following instillation of 2% fluorescein in normal subjects. METHODS This was a prospective randomized study of 60 hospital volunteers. After baseline CCT measurements of both eyes of 40 subjects were obtained using Scheimpflug system, a drop of preservative-free 2% fluorescein, was instilled in one eye and in other eye, one drop of normal saline (control). Measurements were repeated after 1, 2, 5, 10, 20, 30, 40, 50 and 60min (continuous assessment group). Twenty subjects had baseline CCT taken, then fluorescein was instilled in one eye and measurements were taken at 1min. Ten eyes had saline rinse after 1min and 10 other eyes did not, measurements were repeated at 2min (eye rinse group). RESULTS The mean baseline CCT for continuous assessment group was 546.2 ±32.1 µm (range, 489.0-606.0), control eyes was 546.6±30.7 µm (range, 489.0-602.0). At 1min after fluorescein instillation, CCT significantly increased by 37.0±34.0 µm (P<0.001), then decreased gradually, reaching baseline at 60min. CCT variations were not significant in control group (P>0.05). For eye rinse group, CCT mean differences between baseline and 2min were 18.2 µm (95 % CI: -54.7 to 18.3) with rinse and 26.5 µm (95% CI: -62.9 to 9.9) without rinse; paired sample tests were not significant (P>0.05). CONCLUSION The presence of fluorescein increased CCT value to a clinically relevant level of 6.8%. Eye rinse did not significantly reduce the effect at 2min post fluorescein timepoint. PMID:26949642

  16. Port-Wine Stains

    MedlinePlus

    ... their own, they can be treated. In fact, laser therapies can make many port-wine stains much ... mark might be. The good news is that lasers (highly concentrated light energy) can make many kids' ...

  17. Candida, fluorescent stain (image)

    MedlinePlus

    This microscopic film shows a fluorescent stain of Candida. Candida is a yeast (fungus) that causes mild disease, but in immunocompromised individuals it may cause life-threatening illness. (Image ...

  18. Apparatus Would Stain Microscope Slides

    NASA Technical Reports Server (NTRS)

    Breeding, James D.

    1993-01-01

    Proposed apparatus meters specific amounts of fluid out of containers at specific times to stain microscope slides. Intended specifically for semiautomated staining of microbiological and hematological samples in microgravity, leakproof apparatus used in other environments in which technicians have little time to allocate to staining procedures and/or exposure to toxic staining agents or to micro-organisms to be stained hazardous. Apparatus adapted to perform almost any staining procedure and accommodates multiple staining reagents, useful for small or remote clinical laboratories.

  19. A Model for Tear Film Thinning With Osmolarity and Fluorescein

    PubMed Central

    Braun, Richard J.; Gewecke, Nicholas R.; Begley, Carolyn G.; King-Smith, P. Ewen; Siddique, Javed I.

    2014-01-01

    Purpose. We developed a mathematical model predicting dynamic changes in fluorescent intensity during tear film thinning in either dilute or quenching regimes and we model concomitant changes in tear film osmolarity. Methods. We solved a mathematical model for the thickness, osmolarity, fluorescein concentration, and fluorescent intensity as a function of time, assuming a flat and spatially uniform tear film. Results. The tear film thins to a steady-state value that depends on the relative importance of the rates of evaporation and osmotic supply, and the resulting increase of osmolarity and fluorescein concentrations are calculated. Depending on the initial thickness, the rate of osmotic supply and the tear film thinning rate, the osmolarity increase may be modest or it may increase by as much as a factor of eight or more from isosmotic levels. Regarding fluorescent intensity, the quenching regime occurs for initial concentrations at or above the critical fluorescein concentration where efficiency dominates, while lower concentrations show little change in fluorescence with tear film thinning. Conclusions. Our model underscores the importance of using fluorescein concentrations at or near the critical concentration clinically so that quenching reflects tear film thinning and breakup. In addition, the model predicts that, depending on tear film and osmotic factors, the osmolarity within the corneal compartment of the tear film may increase markedly during tear film thinning, well above levels that cause marked discomfort. PMID:24458153

  20. Multiwalled carbon nanotubes inhibit fluorescein extrusion and reduce plasma membrane potential in in vitro human glioma cells.

    PubMed

    Xu, Yonghong; Chen, Xiao; Cheng, Yuli; Xing, Yiqiao

    2010-06-01

    In the study on the interactions of carbon nanotubes with living cells, the cell membrane deserves particular attention as it provides the first interface to initiate CNTs-cell interactions. In the present study, the inhibiting effect of multiwalled carbon nanotubes on the extrusion of fluorescein in human glioma cells was demonstrated using two procedures. To provide clues to explanation of this effect, intracellular glutathione content and reactive oxygen species production were determined as fluorescein is a specific substrate of cell membrane multidrug resistance-related protein whose transport activity requires glutathione which can be depleted under oxidative stress. The plasma membrane potential was also probed as the susceptibility of fluorescein efflux to modulation of the plasma membrane potential has been documented. Results showed a remarkable decrease in cellular glutathione level as well as an increase in reactive oxygen species production. Probe staining also indicated decreased plasma membrane potential. The data suggested that multiwalled carbon nanotubes may affect the transport activity of cell membrane multidrug resistance-related protein through reduction of intracellular glutathione content. Hypopolarization of the plasma membrane may also contribute to MWCNTs' effect. Implications of these findings are discussed. PMID:21179943

  1. Italian multicentre study on intrathecal fluorescein for craniosinusal fistulae

    PubMed Central

    Felisati, G; Bianchi, A; Lozza, P; Portaleone, S

    2008-01-01

    Summary Cerebrospinal fluid leak (CSF), clinical sign of a dural lesion of the skull base, is a relatively rare event that can present with a variety of symptoms. Every craniosinus fistula should be considered a serious, potentially life-threatening situation (even those cases with hidden CSF leak). Reports of experience concerning diagnosis and treatment of craniosinus fistulae have appeared in the Literature. In the last few years, the endoscopic nasal approach is proving effective as it makes diagnosis much easier and is the least invasive surgical approach, with the greatest percentage of success. Various classifications are being proposed to improve clinical evaluation of CSF leaks and to simplify the diagnostic and therapeutic approach. The most common parameters of classification are: aetiology (traumatic, iatrogenic, non-traumatic, etc.) site, type of flow (high or low pressure) and, as far as concerns treatment, the type of graft used, all of which have contributed to various diagnostic and therapeutic algorithms being proposed. Therefore, the subject seems to be widely schematized and the therapeutic attitude widely agreed. However, one of the diagnostic and therapeutic approaches is now being questioned. For some, it is the heart of the clinical approach, while for others, it is a useful tool yet too dangerous to be used on account of potential side effects: namely, the fluorescein test. This procedure, consisting of intrathecal injection of a colorant (fluorescein), is well known by the Food and Drug Administration (FDA) which neither explicitly prohibits it, nor allows it, intrathecal administration is, therefore. an off label use. As far as the Authors know, authorization of this procedure has not been forthcoming anywhere in the world although the procedure itself is widely employed. As far as concerns the use of intrathecal fluorescein, many scientific papers have been written, clearly supporting its clinical usefulness. One limit to the use of fluorescein derives from frequent reports of complications, often related to the intrathecal administration; such complications are, however, always due to an incorrect dosage. In order to perform correct monitoring of any complication related to the use of intrathecal fluorescein and to investigate in a strictly scientific fashion, the legal problem related to the off label use (intrathecal administration) of an authorised substance, the Authors coordinated an Italian multicentre study aimed at establishig the tolerability of the lumbar intrathecal administration of fluorescein. Aim of the study was to review the literature focusing on CSF leaks, to set up to date diagnostic and therapeutic indications of fluorescein and to report the preliminary results of the Italian multicentre study. PMID:18939706

  2. Flash Pasteurization inactivation of Listeria innocua on frankfurters that contain potassium lactate and sodium diacetate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Listeria monocytogenes, a psychrotrophic food-borne pathogen, is a recurring post-process contaminant on ready-to-eat meat (RTE) products including frankfurters. Potassium lactate (PL) and sodium diacetate (SDA) are FDA approved antimicrobials that inhibit the growth of L. monocytogenes when incorp...

  3. "Stained Glass" Landscape Windows

    ERIC Educational Resources Information Center

    Vannata, Janine

    2008-01-01

    Both adults and children alike marvel at the grand vivid stained-glass windows created by American artist Louis Comfort Tiffany. Today he is commonly recognized as one of America's most influential designers and artists throughout the last nineteenth and early twentieth century. In the lesson described in this article, students created their own

  4. Stained-Glass Pastels

    ERIC Educational Resources Information Center

    Laird, Shirley

    2009-01-01

    The author has always liked the look of stained-glass windows. Usually the designs are simplified and the shapes are easier for younger students to draw. This technique seemed to be the perfect place for her fifth-graders to try their hand at color mixing. The smaller spaces and simple shapes were just what she needed for this group. Her students…

  5. "Stained Glass" Landscape Windows

    ERIC Educational Resources Information Center

    Vannata, Janine

    2008-01-01

    Both adults and children alike marvel at the grand vivid stained-glass windows created by American artist Louis Comfort Tiffany. Today he is commonly recognized as one of America's most influential designers and artists throughout the last nineteenth and early twentieth century. In the lesson described in this article, students created their own…

  6. Stained-Glass Pastels

    ERIC Educational Resources Information Center

    Laird, Shirley

    2009-01-01

    The author has always liked the look of stained-glass windows. Usually the designs are simplified and the shapes are easier for younger students to draw. This technique seemed to be the perfect place for her fifth-graders to try their hand at color mixing. The smaller spaces and simple shapes were just what she needed for this group. Her students

  7. Joint fluid Gram stain

    MedlinePlus

    Gram stain of joint fluid ... A sample of joint fluid is needed. The fluid sample is sent to a lab where a small drop is placed in a ... on how to prepare for the removal of joint fluid, see joint fluid aspiration .

  8. THE USE OF FLUORESCEIN DYE TO STUDY THE MOVEMENT OF WATER INTO WHEAT KERNELS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The dispersion of fluorescein dye in water was investigated during the tempering process in wheat kernels. Durum and soft wheat kernels were immersed in fluorescein (free acid crystalline) solution (1%, pH7.54) for 12,24 and 48 hours. The fluorescein pattern showed that the dye solution entered th...

  9. Ionic Liquid-Based Fluorescein Colorimetric pH Nanosensors

    PubMed Central

    Das, Susmita; Magut, Paul K. S.; de Rooy, Sergio L.; Hasan, Farhana; Warner, Isiah M.

    2014-01-01

    A novel pH sensitive, colorimetric ionic liquid nanosensor based on phosphonium salts of fluorescein is reported. Herein, fluorescein salts of various stoichiometries were synthesized by use of a trihexyltetradecylphosphonium cation [TTP]+ in combination with dianionic [FL]2? and monoanionic [FL]? fluorescein. Nanomaterials derived from these two compounds yielded contrasting colorimetric responses in neutral and acidic environments. Variations in fluorescence spectra as a function of pH were also observed. Examination of TEM and DLS data revealed significant expansion in the diameter of [TTP]2[FL] nanodroplets in acidic environments of variable pHs. A similar trend was also observed for [TTP][FL] nanoparticles. The pH dependent colorimetric and other optical properties of these nanomaterials are attributed to alterations in molecular orientations and stacking as suggested by measuring the absorption, fluorescence, and zeta potential. Since the pH is an important indicator for many diseases, including cancer, these nanosensors are considered to be potential candidates for biomedical applications. PMID:25264488

  10. Stain length passive dosimeters

    SciTech Connect

    Sefton, M.V.; Kostas, A.V.; Lombardi, C.

    1982-11-01

    Passive dosimeters with instant readout capability have been devised by combining the principles of a gas indicator tube with membrane control of mass transfer. The membrane controls the diffusion of the gas or vapor to the reagent impregnanted support where it reacts to produce a stain. Results with H/sub 2/S and benzene monitors demonstrated that time weighted average concentration of ambient gas or vapor can be measured accurately and precisely by following the movement of the colored stain in the specially prepared and calibrated indicator tube. The 95% confidence interval of such measurements at the TLV (80 ppm-hrs) is +/- 20% for H/sub 2/S and +/- 15% for benzene, well within NIOSH limits of acceptability.

  11. Length of stain dosimeter

    NASA Technical Reports Server (NTRS)

    Lueck, Dale E. (Inventor)

    1994-01-01

    Payload customers for the Space Shuttle have recently expressed concerns about the possibility of their payloads at an adjacent pad being contaminated by plume effluents from a shuttle at an active pad as they await launch on an inactive pad. As part of a study to satisfy such concerns a ring of inexpensive dosimeters was deployed around the active pad at the inter-pad distance. However, following a launch, dosimeters cannot be read for several hours after the exposure. As a consequence factors such as different substrates, solvent systems, and possible volatilization of HCl from the badges were studied. This observation led to the length of stain (LOS) dosimeters of this invention. Commercial passive LOS dosimeters are sensitive only to the extent of being capable of sensing 2 ppm to 20 ppm if the exposure is 8 hours. To map and quantitate the HCl generated by Shuttle launches, and in the atmosphere within a radius of 1.5 miles from the active pad, a sensitivity of 2 ppm HCl in the atmospheric gases on an exposure of 5 minutes is required. A passive length of stain dosimeter has been developed having a sensitivity rendering it capable of detecting a gas in a concentration as low as 2 ppm on an exposure of five minutes.

  12. Blood stain pattern analysis.

    PubMed

    Peschel, O; Kunz, S N; Rothschild, M A; Mützel, E

    2011-09-01

    Bloodstain pattern analysis (BPA) refers to the collection, categorization and interpretation of the shape and distribution of bloodstains connected with a crime. These kinds of stains occur in a considerable proportion of homicide cases. They offer extensive information and are an important part of a functional, medically and scientifically based reconstruction of a crime. The following groups of patterns can essentially be distinguished: dripped and splashed blood, projected blood, impact patterns, cast-off stains, expirated and transferred bloodstains. A highly qualified analysis can help to estimate facts concerning the location, quality and intensity of an external force. A sequence of events may be recognized, and detailed questions connected with the reconstruction of the crime might be answered. In some cases, BPA helps to distinguish between accident, homicide and suicide or to identify bloodstains originating from a perpetrator. BPA is based on systematic training, a visit to the crime scene or alternatively good photographic documentation, and an understanding and knowledge of autopsy findings or statements made by the perpetrator and/or victim. A BPA working group has been established within the German Society of Legal Medicine aiming to put the knowledge and practical applications of this subdiscipline of forensic science on a wider basis. PMID:21069481

  13. Adhesion of a Mycobacterium sp. to cellulose diacetate membranes used in reverse osmosis.

    PubMed Central

    Ridgway, H F; Rigby, M G; Argo, D G

    1984-01-01

    The kinetics of adhesion of a Mycobacterium sp. to cellulose diacetate reverse-osmosis membranes is described. This Mycobacterium sp. (strain BT2-4) was previously implicated in the initial stages of reverse-osmosis membrane biofouling at a wastewater reclamation facility. Adhesion of BT2-4 cells to the cellulose diacetate membrane surfaces occurred within 1 to 2 h at 30 degrees C and exhibited saturation-type kinetics which conformed closely to the Langmuir adsorption isotherm (Pearson r correlation coefficient = 0.977), a mathematical expression describing the partitioning of substances between a solution and solid-liquid interface. This suggests that the cellulose diacetate membrane surfaces may possess a finite number of available binding sites to which the mycobacteria can adhere. Treatment of the attached mycobacteria with different enzymes suggested that cell surface polypeptides, alpha-1, 4- or alpha-1,6-linked glucan polymers, and carboxyl ester bond-containing substances (possibly peptidoglycolipids) may be involved in mycobacterial adhesion. The possible implication of these findings for reverse-osmosis membrane biofouling are discussed. Images PMID:6696424

  14. Microscopic oxygen imaging based on fluorescein bleaching efficiency measurements.

    PubMed

    Beutler, Martin; Heisterkamp, Ines M; Piltz, Bastian; Stief, Peter; De Beer, Dirk

    2014-05-01

    Photobleaching of the fluorophore fluorescein in an aqueous solution is dependent on the oxygen concentration. Therefore, the time-dependent bleaching behavior can be used to measure of dissolved oxygen concentrations. The method can be combined with epi-fluorescence microscopy. The molecular states of the fluorophore can be expressed by a three-state energy model. This leads to a set of differential equations which describe the photobleaching behavior of fluorescein. The numerical solution of these equations shows that in a conventional wide-field fluorescence microscope, the fluorescence of fluorescein will fade out faster at low than at high oxygen concentration. Further simulation showed that a simple ratio function of different time-points during a fluorescence decay recorded during photobleaching could be used to describe oxygen concentrations in an aqueous solution. By careful choice of dye concentration and excitation light intensity the sensitivity in the oxygen concentration range of interest can be optimized. In the simulations, the estimation of oxygen concentration by the ratio function was very little affected by the pH value in the range of pH 6.5-8.5. Filming the fluorescence decay by a charge-coupled-device (ccd) camera mounted on a fluorescence microscope allowed a pixelwise estimation of the ratio function in a microscopic image. Use of a microsensor and oxygen-consuming bacteria in a sample chamber enabled the calibration of the system for quantification of absolute oxygen concentrations. The method was demonstrated on nitrifying biofilms growing on snail and mussel shells, showing clear effects of metabolic activity on oxygen concentrations. PMID:24610786

  15. Digitization of stained glass

    NASA Astrophysics Data System (ADS)

    MacDonald, Lindsay W.

    1997-04-01

    Digital photography was applied to the capture of images of the stained glass windows in the historic parish church in Fairford, Gloucestershire, England. Because of their size, the windows had to be photographed in 45 separate sections in order to capture all the detail present in the painting on the glass. The digital images of each section, approximately 3000 by 2300 pixels, were then mosaiced together in order to construct the very high resolution image needed for the complete window. A special backlight panel was constructed for the purpose, and techniques developed for minimizing the effects of reflected light and for calibrating the color of the images. Improvements in the technology for mounting and positioning the camera were identified as the most significant factors currently preventing the widespread adoption of this technology for virtual heritage applications.

  16. Fluorescein Tri-Aldehyde Promotes the Selective Detection of Homocysteine.

    PubMed

    Barve, Aabha; Lowry, Mark; Escobedo, Jorge O; Thainashmuthu, Josephrajan; Strongin, Robert M

    2016-03-01

    Elevated homocysteine levels are a well-known independent risk factor for cardiovascular disease. To date, relatively few selective fluorescent probes for homocysteine detection have been reported. The lack of sensing reagents and remaining challenges largely derive from issues of sensitivity and/or selectivity. For example, homocysteine is a structural homologue of the more abundant (ca, 20-25 fold) aminothiol cysteine, differing only by an additional methylene group side chain. Fluorescein tri-aldehyde, described herein, has been designed and synthesized as a sensitive and selective fluorophore for the detection of homocysteine in human plasma samples. It responds to analytes selectively via a photoinduced electron transfer (PET) inhibition process that is modulated by predictable analyte-dye product hybridization and ionization states. Mulliken population analysis of fluorescein tri-aldehyde and its reaction products reveals that the characteristic formation of multiple cationic of homocysteine-derived heterocycles leads to enhanced relative negative charge build up on the proximal phenolate oxygen of the fluorophore as a contributing factor to selective emission enhancement. PMID:26780767

  17. Simplified method for DNA and protein staining of human hematopoietic cell samples

    SciTech Connect

    Crissman, H.A.; Egmond, J.V.; Holdrinet, R.S.; Pennings, A.; Haanen, C.

    1980-01-01

    A rapid reproducible method yielding high resolution analysis of DNA and protein in human hematopoietic cell samples was developed by modification of the propidium iodide (PI) and fluorescein isothiocyanate (FITC) procedure. Cell staining involved sequential addition of each reagent (RNase, FITC, and PI) to ethanol-fixed cells and requires no centrifiguation steps. Stained cells are analyzed in the reagent solutions. Analysis of bone marrow samples from multiple myeloma patients revealed mixed 2C DNA and aneuploid populations with the aneuploid cells having a significantly higher protein content. This approach permitted differential cell cycle kinetic analysis of the 2C DNA and the aneuploid population.

  18. Can 1% chlorhexidine diacetate and ethanol stabilize resin-dentin bonds?

    PubMed Central

    Manso, Adriana Pigozzo; Grande, Rosa Helena Miranda; Bedran-Russo, Ana Karina; Reis, Alessandra; Loguercio, Alessandro D.; Pashley, David Henry; Carvalho, Ricardo Marins

    2014-01-01

    Objectives To examine the effects of the combined use of chlorhexidine and ethanol on the durability of resin-dentin bonds. Methods Forty-eight flat dentin surfaces were etched (32% phosphoric acid), rinsed (15 s) and kept wet until bonding procedures. Dentin surfaces were blot-dried with absorbent paper and re-wetted with water (Water, control), 1% chlorhexidine diacetate in water (CHD/Water), 100% ethanol (Ethanol), or 1% chlorhexidine diacetate in ethanol (CHD/Ethanol) solutions for 30 s. They were then bonded with All Bond 3 (AB3, Bisco) or Excite (EX, Ivoclar-Vivadent) using a smooth, continuous rubbing application (10 s), followed by 15 s gentle air stream to evaporate solvents. The adhesives were light-cured (20 s) and resin composite build-ups constructed for the microtensile method. Bonded beams were obtained and tested after 24-hours, 6-months and 15-months of water storage at 37°C. Storage water was changed every month. Effects of treatment and testing periods were analyzed (ANOVA, Holm-Sidak, p<0.05) for each adhesive. Results There were no interactions between factors for both etch-and-rinse adhesives. AB3 was significantly affected only by storage (p = 0.003). Excite was significantly affected only by treatments (p = 0.048). AB3 treated either with ethanol or CHD/ethanol resulted in reduced bond strengths after 15 months. The use of CHD/ethanol resulted in higher bond strengths values for Excite. Conclusions Combined use of ethanol/1% chlorhexidine diacetate did not stabilize bond strengths after 15 months. PMID:24815823

  19. Protein variations in Listeria monocytogenes exposed to sodium lactate, sodium diacetate, and their combination.

    PubMed

    Mbandi, Evelyne; Phinney, Brett S; Whitten, Douglas; Shelef, Leora A

    2007-01-01

    Most studies of the effect of adverse conditions on survival of Listeria monocytogenes have focused on stress caused by acid or sodium chloride. However, no information is available on resistance of this pathogen to stress caused by salts of organic acids. Sodium lactate and sodium diacetate are generally recognized as safe substances and are approved as ingredients for use in foods. We evaluated antilisterial properties of each of these salts and the enhanced inhibition effected by their combination in ready-to-eat meat products at pH 6.3. Changes in proteins found in this pathogen were studied in the presence of the salts in a chemically defined medium at the same pH using a proteomic approach. The total numbers of protein spots obtained from two-dimensional electrophoresis were 198, 150, and 131 for sodium diacetate, sodium lactate, and the control, respectively. Sodium diacetate treatment produced the highest number of unmatched proteins (124 versus 53 in lactate), the greatest increase in expression (20 versus 5 in lactate), and the highest number of novel proteins (90 versus 45 in lactate). The number of repressed proteins was highest in the combination treatment (41 versus -30 in the single salt treatment). Six proteins that increased or decreased by > or = 10-fold were further investigated; oxidoreductase and lipoprotein were upregulated, and DNA-binding protein, alpha amylase, and two SecA proteins were downregulated or completely suppressed by the salt treatment. Identification of all protein spots is essential for comparison with proteins induced or suppressed under other stress conditions. PMID:17265861

  20. Regionally Discrete Aqueous Humor Outflow Quantification Using Fluorescein Canalograms

    PubMed Central

    Roy, Pritha; Schuman, Joel S.; Sigal, Ian A.; Loewen, Nils A.

    2016-01-01

    Purpose To visualize and quantify conventional outflow directly in its anatomic location. Methods We obtained fluorescein canalograms in six porcine whole eyes and six porcine anterior segment cultures. Eyes were perfused with a constant pressure of 15 mmHg using media containing 0.017 mg/ml fluorescein. Flow patterns were visualized using a stereo dissecting microscope equipped for fluorescent imaging. Images were captured every 30 seconds for 20 minutes for time lapse analysis. Anterior chamber cultures were imaged again on day three of culture. Canalograms were first analyzed for filling time per quadrant. We then wrote a program to automatically compute focal flow fits for each macropixel and to detect convergent perilimbal flow patterns with macropixels grouped into 3 equal-radial width rings around the cornea. A generalized additive model was used to determine fluorescence changes of individual macropixels. Results The resulting imaging algorithm deployed 1024 macropixels that were fit to determine maximum intensity and time to fill. These individual fits highlighted the focal flow function. In whole eyes, significantly faster flow was seen in the inferonasal (IN) and superonasal (SN) quadrants compared to the superotemporal (ST) and inferotemporal (IT) ones (p<0.05). In anterior chamber cultures, reduced flow on day 1 increased in all quadrants on day 3 except in IT (p<0.05). Perilimbal ring analysis uncovered convergent perilimbal flow. Conclusions An algorithm was developed that analyzes regional and circumferential outflow patterns. This algorithm found flow patterns that changed over time and differ in whole eyes and anterior segment cultures. PMID:26998833

  1. Comparison of fluorescein angiography with microvascular anatomy of macaque retinas.

    PubMed

    Weinhaus, R S; Burke, J M; Delori, F C; Snodderly, D M

    1995-07-01

    Recent anatomic work has shown that the capillary network of the fovea is multilaminar. We have identified the elements of this network that are visualized by fluorescein angiography and those that are missed. Fluorescein angiograms of monkey retinas (Macaca fascicularis) with good visualization of individual capillaries were obtained by standard clinical techniques. Retinal whole mounts were prepared from the same animals. Anatomic drawings made from the whole mounts were used to identify which parts of the capillary network were visualized angiographically. Angiographic estimates of dimensions of the foveal avascular zone corresponded closely to the anatomy. Capillary visibility declined rapidly from near perfect visualization at the edge of the foveal avascular zone to less than 40% by 900 microns eccentricity. While all the widest capillary segments (diameter 6.1-7.0 microns) were visualized, only 43% of the modal group of capillary segments (diameter 4.1-4.5 microns) were detected. When a relatively homogeneous population of capillaries was analyzed (diameters limited to the narrow range of 4.0-5.0 microns), visualization declined monotonically with depth in the retina. Capillary segments in the nerve fiber plane were visualized more than four times as effectively as segments of comparable diameter in the deepest vascular plane. High quality angiograms accurately delineate the foveal avascular zone, but they visualize only a fraction of the adjacent multilaminar network. Therefore, current techniques may not detect the earliest nonperfusion of capillaries in vaso-occlusive diseases. Capillary visibility is a joint function of diameter and of retinal depth. The decline in visualization with retinal depth implies that light scattering in the retina degrades the angiographic image. PMID:7556462

  2. Heterogeneous catalyst for the production of ethylidene diacetate from acetic anhydride

    DOEpatents

    Ramprasad, Dorai; Waller, Francis Joseph

    1998-01-01

    This invention relates to a process for producing ethylidene diacetate by the reaction of acetic anhydride, acetic acid, hydrogen and carbon monoxide at elevated temperatures and pressures in the presence of an alkyl halide and a heterogeneous, bifunctional catalyst that is stable to hydrogenation and comprises an insoluble polymer having pendant quaternized heteroatoms, some of which heteroatoms are ionically bonded to anionic Group VIII metal complexes, the remainder of the heteroatoms being bonded to iodide. In contrast to prior art processes, no accelerator (promoter) is necessary to achieve the catalytic reaction and the products are easily separated from the catalyst by filtration. The catalyst can be recycled without loss in activity.

  3. Hexaaquacobalt(II) 2,2?-[naphthalene-1,8-diylbis(oxy)]diacetate dihydrate

    PubMed Central

    Shi, Hui Fang; Wu, Tao; Jiang, Peng Gang; Hao, Zhi; Zhang, Miao Miao

    2013-01-01

    In the title compound, [Co(H2O)6](C14H10O6)2H2O, the 2,2?-[naphthalene-1,8-diylbis(oxy)]diacetate dianion L is not coordinated to the CoII ion. The asymmetric unit contains half of the L dianion, half of a [Co(H2O)6]2+ cation (both molecules being completed by inversion symmetry), and one water molecule. The crystal packing features OH?O hydrogen bonding between the carboxylate groups, the aqua ligands and the hydrate water molecules. PMID:23424401

  4. Heterogeneous catalyst for the production of ethylidene diacetate from acetic anhydride

    DOEpatents

    Ramprasad, D.; Waller, F.J.

    1998-06-16

    This invention relates to a process for producing ethylidene diacetate by the reaction of acetic anhydride, acetic acid, hydrogen and carbon monoxide at elevated temperatures and pressures in the presence of an alkyl halide and a heterogeneous, bifunctional catalyst that is stable to hydrogenation and comprises an insoluble polymer having pendant quaternized heteroatoms, some of which heteroatoms are ionically bonded to anionic Group VIII metal complexes, the remainder of the heteroatoms being bonded to iodide. In contrast to prior art processes, no accelerator (promoter) is necessary to achieve the catalytic reaction and the products are easily separated from the catalyst by filtration. The catalyst can be recycled without loss in activity.

  5. Preparation, characterization, and controlled release from coprecipitates of fluorescein and magnesium hydroxide.

    PubMed

    Hickey, A J; Byron, P R

    1986-08-01

    Magnesium hydroxide was precipitated as a lyophobic sol in the presence of various concentrations of fluorescein sodium (3'6'-dihydroxyspiro[isobenzofuran-1(3H),9'-[9H]xanthen++ +]-3-one, disodium salt) ranging in molar equivalents between 0.1 and 2 times that of the hydroxide. Coprecipitates were washed and dried, and release of the dye and magnesium was determined (pH 7.4, 37 degrees C) from rotating disks. Release rates varied depending upon fluorescein content. The rate of dye release was retarded by less than or equal to 10(4) times that of fluorescein sodium alone, implying the existence of some form of solid association between the components of the coprecipitates. The presence of the dye in certain concentrations reduced magnesium hydroxide dissolution rates by a factor of three. Fluorescein dissolution rates, when expressed as percent release, passed through a minimum (coincident with the dye-induced reduction in Mg(OH)2 dissolution). Adsorption experiments revealed evidence for multiaffinity binding of fluorescein at the surface of freshly precipitated Mg(OH)2. Magnesium, fluorescein, and water contents of the coprecipitates were characterized by atomic absorption and UV spectroscopy and by thermogravimetric analysis. Fluorescein content increased in direct proportion to its initial concentration in solution. Controlled, but variable release of this easily assayed dye is possible by employing precipitates with different fluorescein contents. PMID:3772746

  6. 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.

  7. A novel protocol of whole mount electro-immunofluorescence staining

    PubMed Central

    Liu, Hongshan

    2009-01-01

    Purpose To develop a new method of whole mount immunostaining that improves the penetration of staining reagents into the cornea and decreases non-specific binding and background. Methods Adult mouse corneas were fixed overnight in 4% paraformaldehyde or a mixture of 4% paraformaldehyde and 0.2% glutaraldehyde in 0.1 M phosphate buffer, pH 7.4, at 4 °C. After washing with 0.1% Triton X-100, corneas were embedded in 1% solidified agarose in a plastic column and fluorescent staining reagents, e.g., FITC-IgG (Fluorescein isothiocyanate- immunoglobulinG) conjugates in 0.5% solidified agarose was overlaid onto the specimens. The column was directionally immersed in a submarine gel electrophoresis apparatus filled with Tris-glycine buffer (TGB, pH=7.4) and electrophoresed at 4–10 mA for 10–24 h. For comparison, conventional protocols of immune fluorescent staining were also employed. The outcomes were evaluated by confocal microscopy. Results Antibody conjugates recognizing extracellular matrix (ECM) components, integral membrane protein, and intracellular structural proteins were used in whole mount corneas. The images of confocal laser scanning microscopy (CLSM) displayed a uniform distribution pattern of keratocan in corneal stroma, which is similar to that of section-staining. Anti-β-tubulin antibodies bound to microtubes that are distributed within the whole cell body of superficial corneal epithelium cells and stromal keratocytes, but it was found perinuclear of corneal epithelial wing layers and endothelium; integral membrane protein, FAK (focal adhesion kinase), specifically labeled stromal cells of keratectomy corneas that healed for three weeks. In comparison, conventional protocols of immune fluorescent staining using the same antibody conjugates were also employed but did not yield satisfactory results. It was found that IgG conjugates examined did not readily penetrate into stroma and/or intact corneal epithelium. Phalloidin is a small molecule that can readily penetrate into deep tissue and preferentially binds to F-actin. After the whole mount electrofluorescent staining of phalloidin-rhodamine in the mouse cornea, the results were the same as conventional whole mount staining during the healing of epithelial debridement. The cytoplasmic protrusion formed by lamellipodia and filopodia can be clearly demonstrated. Conclusions These results indicate that the whole mount electro-immunofluorescent staining allows the detection of antigens in all layers of cornea, i.e., epithelium, stroma, and endothelium. PMID:19262742

  8. Sensitivity and specificity of intrathecal fluorescein and white light excitation for detecting intraoperative cerebrospinal fluid leak in endoscopic skull base surgery: a prospective study.

    PubMed

    Raza, Shaan M; Banu, Matei A; Donaldson, Angela; Patel, Kunal S; Anand, Vijay K; Schwartz, Theodore H

    2016-03-01

    OBJECT The intraoperative detection of CSF leaks during endonasal endoscopic skull base surgery is critical to preventing postoperative CSF leaks. Intrathecal fluorescein (ITF) has been used at varying doses to aid in the detection of intraoperative CSF leaks. However, the sensitivity and specificity of ITF at certain dosages is unknown. METHODS A prospective database of all endoscopic endonasal procedures was reviewed. All patients received 25 mg ITF diluted in 10 ml CSF and were pretreated with dexamethasone and Benadryl. Immediately after surgery, the operating surgeon prospectively noted if there was an intraoperative CSF leak and fluorescein was identified. The sensitivity, specificity, and positive and negative predictive power of ITF for detecting intraoperative CSF leak were calculated. Factors correlating with postoperative CSF leak were determined. RESULTS Of 419 patients, 35.8% of patients did not show a CSF leak. Fluorescein-tinted CSF (true positive) was noted in 59.7% of patients and 0 false positives were encountered. CSF without fluorescein staining (false negative) was noted in 4.5% of patients. The sensitivity and specificity of ITF were 92.9% and 100%, respectively. The negative and positive predictive values were 88.8% and 100%, respectively. Postoperative CSF leaks only occurred in true positives at a rate of 2.8%. CONCLUSIONS ITF is extremely specific and very sensitive for detecting intraoperative CSF leaks. Although false negatives can occur, these patients do not appear to be at risk for postoperative CSF leak. The use of ITF may help surgeons prevent postoperative CSF leaks by intraoperatively detecting and confirming a watertight repair. PMID:26295912

  9. The congo red stain revisited.

    PubMed

    Elghetany, M T; Saleem, A; Barr, K

    1989-01-01

    The Congo red stain has undergone several modifications since it was first used by Bennhold in 1922 in order to increase the specificity for staining amyloid. Most of the laboratories in the United States use the method of Puchtler which uses alkaline Congo red solution. Some of the variables associated with the procedure were investigated by us. Our results showed the following: (1) amyloid showed green birefringence at all levels between 4 to 12 mu thick sections with better visualization of small deposits with increased thickness. Best results were obtained with 8 mu thick sections; (2) omission of the pretreatment with alkaline alcoholic solution of sodium chloride (NaCl) did not affect the sensitivity of the method; (3) the use of polar mounting media had no effect on amyloid and collagen birefringence; (4) 50 percent saturation of the Congo red staining solution with NaCl caused strong staining of collagen, elastic fibers and eosinophilic granules. In addition, collagen showed green birefringence and dichroism and its differentiation from amyloid became difficult; and (5) using the staining solution fully saturated with NaCl, no positive staining was seen with tissues other than amyloid. Collagen and elastic fibers showed red fluorescence which was of less intensity than amyloid. It is our conclusion that the method of Puchtler for detecting amyloid gives better results if the staining solution is fully saturated with NaCl. The pretreatment step may be deleted without compromising the quality of staining. Improved staining of amyloid enhances the specificity of green birefringence, dichroism, and red fluorescence. PMID:2471435

  10. Fluorescein as a model molecular calculator with reset capability

    NASA Astrophysics Data System (ADS)

    Margulies, David; Melman, Galina; Shanzer, Abraham

    2005-10-01

    The evolution of molecules capable of performing boolean operations has gone a long way since the inception of the first molecular AND logic gate, followed by other logic functions, such as XOR and INHIBIT, and has reached the stage where these tiny processors execute arithmetic calculations. Molecular logic gates that process a variety of chemical inputs can now be loaded with arrays of logic functions, enabling even a single molecular species to execute distinct algebraic operations: addition and subtraction. However, unlike electronic or optical signals, the accumulation of chemical inputs prevents chemical arithmetic systems from resetting. Consequently, a set of solutions is required to complete even the simplest arithmetic cycle. It has been suggested that these limitations can be overcome by washing off the input signals from solid supports. An alternative approach, which does not require solvent exchange or incorporation of bulk surfaces, is to reset the arithmetic system chemically. Ultimately, this is how some biological systems regenerate. Here we report a highly efficient and exceptionally simple molecular arithmetic system based on a plain fluorescein dye, capable of performing a full scale of elementary addition and subtraction algebraic operations. This system can be reset following each separate arithmetic step. The ability to selectively eradicate chemical inputs brings us closer to the realization of chemical computation.

  11. Potentiation of Femtosecond Laser Intratissue Refractive Index Shaping (IRIS) in the Living Cornea with Sodium Fluorescein

    PubMed Central

    Nagy, Lana J.; Ding, Li; Xu, Lisen; Knox, Wayne H.

    2010-01-01

    Purpose. To assess the effectiveness of intratissue refractive index shaping (IRIS) in living corneas and test the hypothesis that it can be enhanced by increasing the two-photon absorption (TPA) of the tissue. Methods. Three corneas were removed from adult cats and cut into six pieces, which were placed in preservative (Optisol-GS; Bausch & Lomb, Inc., Irvine, CA) containing 0%, 0.25%, 1%, 1.5%, or 2.5% sodium fluorescein (Na-Fl). An 800-nm Ti:Sapphire femtosecond laser with a 100-fs pulse duration and 80-MHz repetition rate was used to perform IRIS in each piece, creating several refractive index (RI) modification lines at different speeds (between 0.1 and 5 mm/s). The lines were 1 μm wide, 10 μm apart, and ∼150 μm below the tissue surface. The RI change of each grating was measured using calibrated, differential interference contrast microscopy. TUNEL staining was performed to assess whether IRIS or Na-Fl doping causes cell death. Results. Scanning at 0.1 mm/s changed the RI of undoped, living corneas by 0.005. In doped corneas, RI changes between 0.01 and 0.02 were reliably achieved with higher scanning speeds. The magnitude of RI changes attained was directly proportional to Na-Fl doping concentration and inversely proportional to the scanning speed used to create the gratings. Conclusions. IRIS can be efficiently performed in living corneal tissue. Increasing the TPA of the tissue with Na-Fl increased both the scanning speeds and the magnitude of RI changes in a dose-dependent manner. Ongoing studies are exploring the use of IRIS to alter the optical properties of corneal tissue in situ, over an extended period. PMID:19815735

  12. Comparative Efficacy of Potassium Levulinate with/without Potassium Diacetate and Potassium Propionate vs Potassium Lactate and Sodium Diacetate for Control of Listeria monocytogenes on commercially prepared uncured t.breast

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We evaluated the efficacy of potassium levulinate, potassium diacetate, and potassium propionate to inhibit Listeria monocytogenes on commercially-prepared, uncured turkey breast during refrigerated storage. Whole muscle, uncured turkey breast chubs (ca. 5 kg each) were formulated with or without po...

  13. Influences of acid on molecular forms of fluorescein and photoinduced electron transfer in fluorescein-dispersing sol-gel titania films.

    PubMed

    Nishikiori, Hiromasa; Setiawan, Rudi Agus; Miyashita, Kyohei; Teshima, Katsuya; Fujii, Tsuneo

    2014-01-01

    Fluorescein-dispersing titania gel films were prepared by the acid-catalyzed sol-gel reaction using a titanium alkoxide solution containing fluorescein. The molecular forms of fluorescein in the films, depending on its acid-base equilibria, and the complex formation and photoinduced electron transfer process between the dye and titania surface were investigated by fluorescence and photoelectric measurements. The titanium species were coordinated to the carboxylate and phenolate-like groups of the fluorescein species. The quantum efficiencies of the fluorescence quenching and photoelectric conversion were higher upon excitation of the dianion species interacting with the titania, i.e. the dye-titania complex. This result indicated that the dianion form was the most favorable for formation of the dye-titania complex exhibiting the highest electron transfer efficiency. Using nitric acid as the catalyst, the titania surface bonded to the fluorescein instead of the adsorbed nitrate ion during the steam treatment. The dye-titania complex formation played an important role in the electron injection from the dye to the titania conduction band. PMID:24502447

  14. Effect of ionic crosslinking on the drug release properties of chitosan diacetate matrices.

    PubMed

    Aiedeh, Khaled M; Taha, Mutasem O; Al-Hiari, Yusuf; Bustanji, Yasser; Alkhatib, Hatim S

    2007-01-01

    Chitosan diacetate (CDA) was prepared by alkylating the amino moieties of chitosan with mono-iodoacetic acid. Subsequently, CDA was cross-linked with Al3+, Zn2+, and Ca2+ ions to yield three ionotropically crosslinked polymeric matrices. These composite matrices were characterized employing infrared spectroscopy (IR) and differential scanning calorimetry (DSC). Subsequently, they were loaded with caffeine, as a model drug, and were assessed as sustained release carriers by evaluating their caffeine release profiles. Interestingly, only CDA-Zn2+ complex sustained the release of caffeine effectively in a zero-order manner. The drug release and thermal behavior of the tested matrices agree with the relative strength of the ionic or coordination character of the bonds. This, in turn, depends on the position of the complexing ions on the electrophilic softness/hardness scale. PMID:17031867

  15. Process for the production of ethylidene diacetate from dimethyl ether using a heterogeneous catalyst

    DOEpatents

    Ramprasad, Dorai; Waller, Francis Joseph

    1998-01-01

    This invention relates to a process for producing ethylidene diacetate by the reaction of dimethyl ether, acetic acid, hydrogen and carbon monoxide at elevated temperatures and pressures in the presence of an alkyl halide and a heterogeneous, bifunctional catalyst that is stable to hydrogenation and comprises an insoluble polymer having pendant quaternized heteroatoms, some of which heteroatoms are ionically bonded to anionic Group VIII metal complexes, the remainder of the heteroatoms being bonded to iodide. In contrast to prior art processes, no accelerator (promoter) is necessary to achieve the catalytic reaction and the products are easily separated from the catalyst by filtration. The catalyst can be recycled for 3 consecutive runs without loss in activity.

  16. Process for the production of ethylidene diacetate from dimethyl ether using a heterogeneous catalyst

    DOEpatents

    Ramprasad, D.; Waller, F.J.

    1998-04-28

    This invention relates to a process for producing ethylidene diacetate by the reaction of dimethyl ether, acetic acid, hydrogen and carbon monoxide at elevated temperatures and pressures in the presence of an alkyl halide and a heterogeneous, bifunctional catalyst that is stable to hydrogenation and comprises an insoluble polymer having pendant quaternized heteroatoms, some of which heteroatoms are ionically bonded to anionic Group VIII metal complexes, the remainder of the heteroatoms being bonded to iodide. In contrast to prior art processes, no accelerator (promoter) is necessary to achieve the catalytic reaction and the products are easily separated from the catalyst by filtration. The catalyst can be recycled for 3 consecutive runs without loss in activity.

  17. Supramolecular architecture of betulin diacetate complexes with arabinogalactan from Larix sibirica.

    PubMed

    Mikhailenko, Mikhail A; Shakhtshneider, Tatyana P; Eltsov, Ilia V; Kozlov, Alexander S; Kuznetsova, Svetlana A; Karacharov, Аnton А; Boldyrev, Vladimir V

    2016-03-15

    Supramolecular ensembles of arabinogalactan (AG) and its complexes with betulin diacetate (BDA) were studied in water and dimethyl sulfoxide (DMSO) using ablation, induced by submillimeter radiation from the free electron laser. Solutions of 1wt% AG resulted in formation of aerosol particles with a maximum size of 60-70nm. In contrast, with DMSO as the solvent, the majority of particles were significantly smaller. Nevertheless, the addition of water shifted the particle size distribution to a larger size, suggesting the cross-linking of AG chains due to hydrogen bonding through water molecules. The ensembles of molecules were larger in solutions of the AG-BDA complex as compared to pure AG aqueous solution, and the distribution was narrow. The role of side chain interactions in the formation of AG-BDA complexes in aqueous solutions was confirmed by NMR. PMID:26794731

  18. A tracer test at the Beowawe geothermal field, Nevada, using fluorescein and tinopal CBS

    SciTech Connect

    Rose, P.E.; Adams, M.C.; Benoit, D.

    1995-12-31

    An interwell tracer test using fluorescein and tinopal CBS was performed at the Beowawe geothermal field in north-central Nevada in order to assess the effects of recent changes to the injection strategy. Fluorescein return curves established injection-production flow patterns and verified that produced water is being reinjected into a region of the reservoir that is in excellent communication with the production wells. An analysis of the tinopal CBS return curves indicated that tinopal CBS was apparently strongly adsorbed onto the reservoir rock. The fluorescein return curves were used to estimate the overall (fractures and matrix) reservoir volume.

  19. 76 FR 75886 - Determination That DEMULEN 1/50-28 (Ethinyl Estradiol; Ethynodiol Diacetate) Tablet and Four...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-05

    ...; Ethynodiol Diacetate) Tablet and Four Other Drug Products Were Not Withdrawn From Sale for Reasons of Safety... withdrawn from sale for reasons of safety or effectiveness. This determination means that FDA will not begin..., Drug, and Cosmetic Act (21 U.S.C. 355(j)(7)), which requires FDA to publish a list of all...

  20. Sodium lactate, sodium diacetate and pediocin: effects and interactions on the thermal inactivation of Listeria monocytogenes on bologna

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The effects and interactions of temperature (56.3C-60C), sodium lactate (SL; 0-4.8%), sodium diacetate (SD; 0-0.25%) and pediocin (0-10,000 AU) on Listeria monocytogenes inoculated on bologna were studied and a predictive inactivation model was developed. Bologna was manufactured with different SL ...

  1. Ultraviolet Light (254 nm) Inactivation of Listeria monocytogenes on frankfurters that contain potassium lactate and sodium diacetate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Listeria monocytogenes, a psychrotrophic food-borne pathogen, is an occasional post-process contaminant on ready-to-eat meat (RTE) products including frankfurters. Potassium lactate (PL) and sodium diacetate (SDA) are FDA approved antimicrobials that inhibit the growth of L. monocytogenes when incor...

  2. EFFECTS AND INTERACTIONS OF TEMPERATURE, SODIUM LACTATE, SODIUM DIACETATE AND PEDIOCIN ON THE STARVED CELLS OF LISTERIA MONOCYTOGENES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The effects and interactions of temperature (55-60 degrees C), sodium lactate (SL; 0.0-4.8%), sodium diacetate (SDA; 0.0-0.4%) and pediocin (0.0-10000 AU) on the starved cells of L. monocytogenes inoculated on the surface of the frankfurters were investigated, and a predictive model was developed. C...

  3. Fluorescein angiography and optical coherence tomography in myopic choroidal neovascularization

    PubMed Central

    Chhablani, J; Deepa, M J; Tyagi, M; Narayanan, R; Kozak, I

    2015-01-01

    Purpose To assess intra/inter-observer agreement, and diagnostic capabilities of a color fundus photograph, fundus fluorescein angiography (FFA), and spectral domain optical coherence tomography (SD-OCT) in making a diagnosis of myopic choroidal neovascularization (CNV). Patients and methods Two masked observers evaluated FFA and SD-OCT images to identify the presence of myopic CNV in 80 high-myopic eyes of 57 patients. A third masked observer identified CNV on a color fundus photo. Presence of myopic CNV on a fundus photo was defined as presence of subretinal hemorrhage, thickening of the retina and/or visible membrane at the macula. Presence of myopic CNV on FFA was defined as hyperfluorescence in the early phase with increase in intensity and size in the late phase; presence of a large irregular lesion; and hypofluorsescence due to subretinal hemorrhage. Myopic CNV on SD-OCT was defined as the hyper-reflective lesion with or without intraretinal fluid or subretinal fluid with retinal thickening. Results Intraobserver repeatability on FFA and SD-OCT was 0.54 and 0.44, respectively. Agreement (kappa) between FFA and SD-OCT was 0.38 and 0.3, respectively. Among 34 eyes, which had the presence of CNV on a color fundus photo, CNV was diagnosed in 18 (53%) eyes on FFA and in 20 (58.8%) eyes on SD-OCT. Sensitivity and specificity of FFA was 47 and 80.4%, respectively, and that of SD-OCT was 58.8 and 86.9%, respectively. Conclusion Repeatability and reproducibility for diagnosis of myopic CNV was better with FFA compared with SD-OCT; however, agreement is very poor between FFA and SD-OCT. SD-OCT is comparatively a better tool to rule out presence of myopic CNV. PMID:25613842

  4. Transscleral diffusion of ethacrynic acid and sodium fluorescein

    PubMed Central

    Lin, Cheng-Wen; Wang, Yong; Challa, Pratap; Epstein, David L.

    2007-01-01

    Purpose One of the current limitations in developing novel glaucoma drugs that target the trabecular meshwork (TM) is the induced corneal toxicity from eyedrop formulations. To avoid the corneal toxicity, an alternative approach would be to deliver TM drugs through the sclera. To this end, we quantified ex vivo diffusion coefficient of a potential TM drug, ethacrynic acid (ECA), and investigated mechanisms of ECA transport in the sclera. Methods An Ussing-type diffusion apparatus was built to measure the apparent diffusion coefficient of ECA in fresh porcine sclera at 4 C. To understand mechanisms of ECA transport, we quantified the transscleral transport of a fluorescent tracer, sodium fluorescein (NaF), that has a similar molecular weight but is more hydrophilic compared to ECA. Furthermore, we developed a mathematical model to simulate the transport processes and used it to analyze the experimental data. The model was also used to investigate the dependence of diffusion coefficients on volume fraction of viable cells and the binding of NaF and ECA to scleral tissues. Results The diffusion coefficients of ECA and NaF in the sclera were 48.515.1x10-7 cm2/s (n=9) and 5.231.93x10-7 cm2/s (n=8), respectively. Both diffusion coefficients were insensitive to cell shrinkage caused by ECA during the diffusion experiments and cell damage caused by the storage of tissues ex vivo before the experiments. Binding of ECA to scleral tissues could not be detected. The apparent maximum binding capacity and the apparent equilibrium dissociation constant for NaF were 805 mM and 2.50.5 mM (n=3), respectively. Conclusions These data demonstrated that ECA diffusion was minimally hindered by structures in the sclera, presumably due to the lack of cells and binding sites for ECA in the sclera. PMID:17356511

  5. Objective Area Measurement Technique for Choroidal Neovascularization from Fluorescein Angiography

    PubMed Central

    Guthrie, Micah J.; Osswald, Christian R.; Valio, Nicole L.; Mieler, William F.; Kang-Mieler, Jennifer J.

    2014-01-01

    The purpose of this study was to develop a non-biased method of quantitatively measuring choroidal neovascularization (CNV) areas based on late-phase fluorescein angiography (FA) images. Experimental CNV was induced in Long Evans rats by laser disruption of the Bruch’s membrane. FA was performed weekly for 5 weeks. Multi-Otsu thresholding (MOT) was used to quantify CNV in late-phase FA images from both experimental rodent CNV and wet age-related macular degeneration patients (wAMD). Images were automatically thresholded into three levels based on the image histogram, with the highest level containing CNV. To determine the technique’s ability to quantify CNV areas, rats were given either triamcinolone acetonide or dexamethasone sodium phosphate to treat CNV and compared to untreated rats. The rat CNV lesion areas measured from 5-week histology sections from each treatment group were compared to areas measured from the corresponding FA images. MOT was able to detect statistical decreases in rodent CNV area in the treatment groups versus control from weeks 3 through 5. The ratio of CNV area measured from histology to area measured from FA images was not statistically different between groups. Finally, to determine the usefulness of MOT on pathological morphologies of CNV, MOT was performed on late-phase FA images from patients with classic and diffuse CNV. The technique was able to segment classical CNV in wAMD patients, but performed poorly with diffuse CNV. MOT provides a robust, objective, and quantifiable area measurement of CNV lesion area in both experimentally-induced and pathological CNV. The results indicate that MOT could be a useful research tool in helping evaluate the effects of therapeutics on CNV growth. PMID:24316422

  6. Development of and Clinical Experience with a Simple Device for Performing Intraoperative Fluorescein Fluorescence Cerebral Angiography: Technical Notes

    PubMed Central

    ICHIKAWA, Tsuyoshi; SUZUKI, Kyouichi; WATANABE, Yoichi; SATO, Taku; SAKUMA, Jun; SAITO, Kiyoshi

    2016-01-01

    To perform intraoperative fluorescence angiography (FAG) under a microscope without an integrated FAG function with reasonable cost and sufficient quality for evaluation, we made a small and easy to use device for fluorescein FAG (FAG filter). We investigated the practical use of this FAG filter during aneurysm surgery, revascularization surgery, and brain tumor surgery. The FAG filter consists of two types of filters: an excitatory filter and a barrier filter. The excitatory filter excludes all wavelengths except for blue light and the barrier filter passes long waves except for blue light. By adding this FAG filter to a microscope without an integrated FAG function, light from the microscope illuminating the surgical field becomes blue, which is blocked by the barrier filter. We put the FAG filter on the objective lens of the operating microscope correctly and fluorescein sodium was injected intravenously or intra-arterially. Fluorescence (green light) from vessels in the surgical field and the dyed tumor were clearly observed through the microscope and recorded by a memory device. This method was easy and could be performed in a short time (about 10 seconds). Blood flow of small vessels deep in the surgical field could be observed. Blood flow stagnation could be evaluated. However, images from this method were inferior to those obtained by currently commercially available microscopes with an integrated FAG function. In brain tumor surgery, a stained tumor on the brain surface could be observed using this method. FAG could be performed with a microscope without an integrated FAG function easily with only this FAG filter. PMID:26597335

  7. Development of and Clinical Experience with a Simple Device for Performing Intraoperative Fluorescein Fluorescence Cerebral Angiography: Technical Notes.

    PubMed

    Ichikawa, Tsuyoshi; Suzuki, Kyouichi; Watanabe, Yoichi; Sato, Taku; Sakuma, Jun; Saito, Kiyoshi

    2016-03-15

    To perform intraoperative fluorescence angiography (FAG) under a microscope without an integrated FAG function with reasonable cost and sufficient quality for evaluation, we made a small and easy to use device for fluorescein FAG (FAG filter). We investigated the practical use of this FAG filter during aneurysm surgery, revascularization surgery, and brain tumor surgery. The FAG filter consists of two types of filters: an excitatory filter and a barrier filter. The excitatory filter excludes all wavelengths except for blue light and the barrier filter passes long waves except for blue light. By adding this FAG filter to a microscope without an integrated FAG function, light from the microscope illuminating the surgical field becomes blue, which is blocked by the barrier filter. We put the FAG filter on the objective lens of the operating microscope correctly and fluorescein sodium was injected intravenously or intra-arterially. Fluorescence (green light) from vessels in the surgical field and the dyed tumor were clearly observed through the microscope and recorded by a memory device. This method was easy and could be performed in a short time (about 10 seconds). Blood flow of small vessels deep in the surgical field could be observed. Blood flow stagnation could be evaluated. However, images from this method were inferior to those obtained by currently commercially available microscopes with an integrated FAG function. In brain tumor surgery, a stained tumor on the brain surface could be observed using this method. FAG could be performed with a microscope without an integrated FAG function easily with only this FAG filter. PMID:26597335

  8. Control of Listeria monocytogenes on commercially-produced frankfurters prepared with and without potassium lactate and sodium diacetate and surface .....using the Sprayed Lethality in Container (SLIC®) delivery method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The viability of Listeria monocytogenes was monitored on commercially-produced frankfurters that were formulated with no, low, or high levels of potassium lactate and sodium diacetate (low = 0.68% lactate and 0.097% diacetate and high = 1.36% lactate and 0.19% diacetate) and then treated with 22 or ...

  9. INTER-LABORATORY STUDY OF CELLULAR FLUORESCENCE INTENSITY MEASUREMENTS WITH FLUORESCEIN-LABELED MICROBEAD STANDARDS

    EPA Science Inventory

    To determine the precision of cellular fluorescence intensity (FI) measurements derived from labeled microbead standards, FI results were compared from 43 different flow cytometers in 34 laboratories. ll laboratories analyzed prepared aliquots of fluoresceinated calf thymocyte nu...

  10. Automated single-slide staining system

    NASA Technical Reports Server (NTRS)

    Mills, S. M.; Wilkins, J. R.

    1974-01-01

    Apparatus developed to Gram-stain single slides automatically is flexible enough to accommodate other types of staining procedures. Method frees operator and eliminates necessity for subjective evaluations as to length of staining or decolorizing time.

  11. [Histochemical staining methods for lanthanum].

    PubMed

    Miyagawa, Makoto

    2011-09-01

    In recent years lanthanum compounds have been widely used in the optical and electronic industries. Although release of lanthanum (La) into the environment and exposure to humans are feared, acute or chronic biologic effects of La remain to be elucidated. The present study was undertaken to establish the experimental animal model for La toxicity and histological staining methods for La. After intraperitoneal injections of lanthanum chloride, white precipitates deposited on the surface of the liver. Existence of La in the precipitates was confirmed by a X-ray fluorescent microanalysis. Liver tissues from La treated rats were fixed in paraformaldehyde, embedded in paraffin and cut into thin sections which were used as a La containing standard section. Several reagents for histological stains and spectrophotometry for metals were applied in both test-tube experiments and stainings of tissue sections to test for La. Alizarin complexone (ALC) was found of capable of staining La in tissue. A simple new technique used was described for light microscopic detection of La. PMID:22111301

  12. [Histochemical staining methods for lanthanum].

    TOXLINE Toxicology Bibliographic Information

    Miyagawa M

    2011-09-01

    In recent years lanthanum compounds have been widely used in the optical and electronic industries. Although release of lanthanum (La) into the environment and exposure to humans are feared, acute or chronic biologic effects of La remain to be elucidated. The present study was undertaken to establish the experimental animal model for La toxicity and histological staining methods for La. After intraperitoneal injections of lanthanum chloride, white precipitates deposited on the surface of the liver. Existence of La in the precipitates was confirmed by a X-ray fluorescent microanalysis. Liver tissues from La treated rats were fixed in paraformaldehyde, embedded in paraffin and cut into thin sections which were used as a La containing standard section. Several reagents for histological stains and spectrophotometry for metals were applied in both test-tube experiments and stainings of tissue sections to test for La. Alizarin complexone (ALC) was found of capable of staining La in tissue. A simple new technique used was described for light microscopic detection of La.

  13. Silver staining in clinical cytogenetics.

    PubMed

    Martin, A O

    1985-09-01

    Silver staining of human chromosomes at prometaphase or metaphase identifies variants in the stalk (nucleolar organizing) regions of acrocentric chromosomes (Nos. 13, 14, 15, 21, 22). Variants are defined by size, number, and morphology of silver staining areas. They are heritable polymorphisms and have not been associated with clinical abnormalities. However, these variants are useful in clinical cytogenetics, specifically in studies attempting to determine whether genetic material has been gained or lost in chromosomal rearrangements, the origin of chromosomal aberrations, the origin of cells in tissue culture, the chromosomal location of single genes, clonal origin of tumors, the zygosity of twins, and paternity. Some chromosomal aberrations require silver staining for their definition. Because loss of the stalk regions per se is apparently not deleterious, demonstration that chromosomal breaks occurred within this region without concomitant loss or gain of genetic material essential for normal human development provides basis for a good prognosis for the individual with the chromosomal rearrangement resulting from such breakage. The principle underlying most of the other applications is to determine whether variants being compared are identical or dissimilar, and to make inferences from these results (e.g., variants in monozygotic twins should all be identical, whereas in dizygotic twins they are as similar as in any pair of sibs). Silver staining is a valuable technique for special questions in clinical analysis. PMID:2412316

  14. Whole Blood Cell Staining Device

    NASA Technical Reports Server (NTRS)

    Sams, Clarence F.; Clift, Vaughan L.; McDonald, Kelly E.

    2000-01-01

    An apparatus and method for staining particular cell markers is disclosed. The apparatus includes a flexible tube that is reversibly pinched into compartments with one or more clamps. Each compartment of the tube contains a separate reagent and is in selective fluid communication with adjoining compartments.

  15. Tear Film, Contact Lens, and Patient Factors Associated with Corneal Staining

    PubMed Central

    Sinnott, Loraine T.

    2011-01-01

    Purpose. The purpose of this study was to examine ocular surface and tear film, contact lens, care solution, medical, and patient-related factors that are associated with corneal staining in contact lens wearers. Methods. In this cross-sectional/nested case–control study, in addition to the assessment of corneal staining with fluorescein, a variety of tear film and ocular surface, contact lens, and patient-related factors were examined. Poisson regression models were used to examine the relation between corneal staining and these factors. Results. Data from 413 patients were eligible for the analyses described. The average age was 30.6 ± 11.1 years, and 277 (67.1%) of the patients were women. Several factors were shown to be related to increased corneal staining in multivariate modeling, including increased daily wearing times (P = 0.0006), lower income (P = 0.0008), lissamine green conjunctival staining (P = 0.002), contact lens deposition (P = 0.007), increased tear meniscus height (P = 0.007), and decreased hydrogel nominal water content (P = 0.02). The wearing of silicone hydrogels (as opposed to hydrogels) was protective against corneal staining (P = 0.0004). Notably, neither contact lens care solutions nor disinfectants were associated with corneal staining. Conclusions. Corneal staining in contact lens wearers continues to be a frequent, but not well understood, outcome. These data suggest that contact lens factors (water content, material, wearing time, and deposition) are more generally associated with corneal staining than are contact lens care solutions or other ocular surface and tear film, demographic, or medical factors. PMID:21087960

  16. Degradation of phorbol 12,13-diacetate in aqueous solution by gamma irradiation

    NASA Astrophysics Data System (ADS)

    Kongmany, Santi; Furuta, Masakazu; Matsuura, Hiroto; Okuda, Shuichi; Imamura, Kiyoshi; Maeda, Yasuaki

    2014-12-01

    Phorbol esters (PEs) are highly toxic compounds that cause skin irritation, inflammation, and tumor promotion upon contact with humans or animals. These compounds are naturally present in Jatropha curcas L. To promote the use of J. curcas seed oil in bio-diesel production industries and reduce environmental concerns, it is necessary to find methods of degrading PEs. In this study, the degradation of phorbol 12,13-diacetate (PDA), as a representative PE, in aqueous solution at a concentration of 10 mg/L by 60Co-γ-irradiation was investigated. The results demonstrate that PDA was effectively degraded by this treatment and the degradation efficiency increased with the absorbed dose within the range of 0.5-3 kGy. Complete degradation of PDA was achieved at a dose of 3 kGy. In the presence of radical scavengers (i.e., methanol, tert-butanol, 2-propanol), reactive species from water radiolysis were scavenged, and significant inhibition of PDA degradation was observed at absorbed doses less than 1 kGy. In the presence of nitrous oxide, the generation of hydroxyl radicals (rad OH) was promoted during gamma irradiation and PDA degradation was drastically enhanced.

  17. Encapsulation of T4 bacteriophage in electrospun poly(ethylene oxide)/cellulose diacetate fibers.

    PubMed

    Korehei, Reza; Kadla, John F

    2014-01-16

    Phage therapy is a potentially beneficial approach to food preservation and storage. Sustained delivery of bacteriophage can prevent bacterial growth on contaminated food surfaces. Using coaxial electrospinning bacteriophage can be encapsulated in electrospun fibers with high viability. The resulting bio-based electrospun fibers may have potential as a food packaging material. In the present work, T4 bacteriophage (T4 phage) was incorporated into core/shell electrospun fibers made from poly(ethylene oxide) (PEO), cellulose diacetate (CDA), and their blends. Fibers prepared using PEO as the shell polymer showed an immediate burst release of T4 phage upon submersion in buffer. The blending of CDA with PEO significantly decreased the rate of phage release, with no released T4 phage being detected from the solely CDA fibers. Increasing the PEO molecular weight increased the electrospun fiber diameter and viscosity of the releasing medium, which resulted in a relatively slower T4 phage release profile. SEM analyses of the electrospun fiber morphologies were in good agreement with the T4 phage release profiles. Depending on the PEO/CDA ratio, the post-release electrospun fiber morphologies varied from discontinuous fibers to minimally swollen fibers. From these results it is suggested that the T4 phage release mechanism is through solvent activation/polymer dissolution in the case of the PEO fibers and/or by diffusion control from the PEO/CDA blend fibers. PMID:24188849

  18. Methods for chromosome-specific staining

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    1995-01-01

    Methods and compositions for chromosome-specific staining are provided. Compositions comprise heterogenous mixtures of labeled nucleic acid fragments having substantially complementary base sequences to unique sequence regions of the chromosomal DNA for which their associated staining reagent is specific. Methods include methods for making the chromosome-specific staining compositions of the invention, and methods for applying the staining compositions to chromosomes.

  19. Salt stains from evaporating droplets.

    PubMed

    Shahidzadeh, Noushine; Schut, Marthe F L; Desarnaud, Julie; Prat, Marc; Bonn, Daniel

    2015-01-01

    The study of the behavior of sessile droplets on solid substrates is not only associated with common everyday phenomena, such as the coffee stain effect, limescale deposits on our bathroom walls , but also very important in many applications such as purification of pharmaceuticals, de-icing of airplanes, inkjet printing and coating applications. In many of these processes, a phase change happens within the drop because of solvent evaporation, temperature changes or chemical reactions, which consequently lead to liquid to solid transitions in the droplets. Here we show that crystallization patterns of evaporating of water drops containing dissolved salts are different from the stains reported for evaporating colloidal suspensions. This happens because during the solvent evaporation, the salts crystallize and grow during the drying. Our results show that the patterns of the resulting salt crystal stains are mainly governed by wetting properties of the emerging crystal as well as the pathway of nucleation and growth, and are independent of the evaporation rate and thermal conductivity of the substrates. PMID:26012481

  20. Salt stains from evaporating droplets

    PubMed Central

    Shahidzadeh, Noushine; Schut, Marthe F. L.; Desarnaud, Julie; Prat, Marc; Bonn, Daniel

    2015-01-01

    The study of the behavior of sessile droplets on solid substrates is not only associated with common everyday phenomena, such as the coffee stain effect, limescale deposits on our bathroom walls , but also very important in many applications such as purification of pharmaceuticals, de-icing of airplanes, inkjet printing and coating applications. In many of these processes, a phase change happens within the drop because of solvent evaporation, temperature changes or chemical reactions, which consequently lead to liquid to solid transitions in the droplets. Here we show that crystallization patterns of evaporating of water drops containing dissolved salts are different from the stains reported for evaporating colloidal suspensions. This happens because during the solvent evaporation, the salts crystallize and grow during the drying. Our results show that the patterns of the resulting salt crystal stains are mainly governed by wetting properties of the emerging crystal as well as the pathway of nucleation and growth, and are independent of the evaporation rate and thermal conductivity of the substrates. PMID:26012481

  1. 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.

  2. Fluorogenic Substrate Detection of Viable Intracellular and Extracellular Pathogenic Protozoa

    NASA Astrophysics Data System (ADS)

    Jackson, Peter R.; Pappas, Michael G.; Hansen, Brian D.

    1985-01-01

    Viable Leishmania promastigotes and amastigotes were detected by epifluorescence microscopy with fluorescein diacetate being used to mark living parasites and the nucleic acid-binding compound ethidium bromide to stain dead cells. This procedure is superior to other assays because it is faster and detects viable intracellular as well as extracellular Leishmania. Furthermore, destruction of intracellular pathogens by macrophages is more accurately determined with fluorescein diacetate than with other stains. The procedure may have applications in programs to develop drugs and vaccines against protozoa responsible for human and animal disease.

  3. [Preparation and vitality detection of protoplast in Salvia miltiorrhiza Bunge].

    PubMed

    Zhu, Nan; Liu, Jun; Zhang, Xinyu; Dong, Juan'e

    2014-10-01

    We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts. PMID:25726586

  4. The use of fluorescein sodium in the biopsy and gross-total resection of a tectal plate glioma.

    PubMed

    Ung, Timothy H; Kellner, Christopher; Neira, Justin A; Wang, Shih-Hsiu J; D'Amico, Randy; Faust, Phyllis L; Canoll, Peter; Feldstein, Neil A; Bruce, Jeffrey N

    2015-12-01

    Intravenous administration of fluorescein sodium fluoresces glioma burden tissue and can be visualized using the surgical microscope with a specialized filter. Intraoperative guidance afforded through the use of fluorescein may enhance the fidelity of tissue sampling, and increase the ability to accomplish complete resection of tectal lesions. In this report the authors present the case of a 19-year-old man with a tectal anaplastic pilocytic astrocytoma in which the use of fluorescein sodium and a Zeiss Pentero surgical microscope equipped with a yellow 560 filter enabled safe complete resection. In conjunction with neurosurgical navigation, added intraoperative guidance provided by fluorescein may be beneficial in the resection of brainstem gliomas. PMID:26407010

  5. Inactivation of Listeria innocua on frankfurters that contain potassium lactate and sodium diacetate by flash pasteurization.

    PubMed

    Sommers, C H; Geveke, D J; Fan, X

    2008-03-01

    Listeria monocytogenes, a psychrotrophic foodborne pathogen, is a recurring postprocess contaminant on ready-to-eat meat (RTE) products, including frankfurters. Potassium lactate (PL) and sodium diacetate (SDA) are FDA-approved antimicrobials that inhibit the growth of L. monocytogenes when incorporated into the formulation of fine emulsion sausage. Flash (steam) pasteurization (FP) has been shown to reduce levels of L. monocytogenes, and its surrogate L. innocua, on frankfurter surfaces. The ability of FP to inactivate and prevent the growth of the L. monocytogenes surrogate L. innocua in a pilot plant setting was investigated. FP treatment (1.5 s, 121 degrees C) of single layers of frankfurters that were surface-inoculated with either 5, 4, or 3 log CFU/g of L. innocua immediately before FP (1.5 s, 121 degrees C) resulted in log reductions of 1.97 (+/- 0.11), 2.03 (+/- 0.10), or 2.07 (+/- 0.14), respectively. Inoculum level had no effect on the inactivation of L. innocua. Following 8 wk of refrigerated storage (4 degrees C), L. innocua levels decreased by 0.5 log in non-FP-treated frankfurter packs, while the 2 log reduction of L. innocua was maintained for FP-treated frankfurters. FP (1.5 s, 121 degrees C) had no effect on frankfurter color or texture. Because the numbers of L. monocytogenes associated with contaminations of ready-to-eat meats are typically very low, the use of FP in combination with PL and SDA has the potential to reduce the number of frankfurter recalls and foodborne illness outbreaks. PMID:18298739

  6. Improved measurements of intracellular nitric oxide in intact microvessels using 4,5-diaminofluorescein diacetate

    PubMed Central

    Zhou, Xueping

    2011-01-01

    4,5-Diaminofluorescein diacetate (DAF-2 DA) has been widely used for the measurement of nitric oxide (NO) in living cells and tissues. We previously established a method that demonstrated platelet activating factor (PAF)-induced endothelial NO production in intact venules using DAF-2 DA. In previous applications, the loading dye was removed from the extracellular space before NO measurements. However, in high permeability vessels, endothelial cells quickly released the accumulated intracellular DAF-2 after the washout, which compromises the NO measurement. The objective of this study was to investigate if the presence of DAF-2 DA during NO measurements could overcome the dye retention problem and enhance the sensitivity of NO detection. Experiments were conducted in individually perfused rat venules, and endothelial NO was measured using fluorescence imaging under basal and stimulated conditions with continuous perfusion of DAF-2 DA. Continuous dye perfusion was found to promote a relatively constant endothelial dye concentration in both normal and high permeability vessels throughout the experiment. With the use of this method, the basal and stimulated NO was quantified after endothelial DAF-2 concentrations reached a steady state. Our results showed enhanced sensitivity of detecting PAF-stimulated NO compared with a previous method. We also found that the hydrolyzed intracellular DAF-2, the precursor of DAF-2 triazole, contributed significantly to the measured fluorescence and that an appropriate subtraction of non-NO-dependent intracellular DAF-2 fluorescence is critical for the assessment of NO in living tissues. This method overcame the dye leakage problem, enhanced the sensitivity of NO detection, and improved NO quantification, demonstrating significant advantages over existing methodologies using DAF-2. PMID:21536843

  7. Early Diagnosis of Subclinical Interferon Alpha-Associated Optic Neuropathy Using Fluorescein Angiography.

    PubMed

    Cestari, Dean M; Lessell, Simmons; Mantopoulos, Dimosthenis

    2015-09-01

    We report a case of a 57-year-old man who presented with decreased visual acuity in the left eye secondary to nonarteritic anterior ischemic optic neuropathy (NAION) while on therapy with interferon-α for hepatitis C. Fundus fluorescein angiography revealed late leakage of both optic discs, consistent with bilateral disease. One week later, the patient developed clinical signs and symptoms consistent with NAION in the fellow eye. Fluorescein angiography may play an important role in identifying subclinical NAION in patients taking interferon-α. PMID:25793363

  8. Safer staining method for acid fast bacilli.

    PubMed

    Ellis, R C; Zabrowarny, L A

    1993-06-01

    To develop a method for staining acid fast bacilli which excluded highly toxic phenol from the staining solution. A lipophilic agent, a liquid organic detergent, LOC High Studs, distributed by Amway, was substituted. The acid fast bacilli stained red; nuclei, cytoplasm, and cytoplasmic elements stained blue on a clear background. These results compare very favourably with acid fast bacilli stained by the traditional method. Detergents are efficient lipophilic agents and safer to handle than phenol. The method described here stains acid fast bacilli as efficiently as traditional carbol fuchsin methods. LOC High Suds is considerably cheaper than phenol. PMID:7687254

  9. Safer staining method for acid fast bacilli.

    PubMed Central

    Ellis, R C; Zabrowarny, L A

    1993-01-01

    To develop a method for staining acid fast bacilli which excluded highly toxic phenol from the staining solution. A lipophilic agent, a liquid organic detergent, LOC High Studs, distributed by Amway, was substituted. The acid fast bacilli stained red; nuclei, cytoplasm, and cytoplasmic elements stained blue on a clear background. These results compare very favourably with acid fast bacilli stained by the traditional method. Detergents are efficient lipophilic agents and safer to handle than phenol. The method described here stains acid fast bacilli as efficiently as traditional carbol fuchsin methods. LOC High Suds is considerably cheaper than phenol. Images PMID:7687254

  10. A double fluorescence staining protocol to determine the cross-sectional area of myofibers using image analysis

    NASA Technical Reports Server (NTRS)

    Mozdziak, P. E.; Fassel, T. A.; Schultz, E.; Greaser, M. L.; Cassens, R. G.

    1996-01-01

    A double fluorescence staining protocol was developed to facilitate computer based image analysis. Myofibers from experimentally treated (irradiated) and control growing turkey skeletal muscle were labeled with the anti-myosin antibody MF-20 and detected using fluorescein-5-isothiocyanate (FITC). Extracellular material was stained with concanavalin A (ConA)-Texas red. The cross-sectional area of the myofibers was determined by calculating the number of pixels (0.83 mu m(2)) overlying each myofiber after subtracting the ConA-Texas red image from the MF-20-FITC image for each region of interest. As expected, myofibers in the irradiated muscle were smaller (P < 0.05) than those in the non-irradiated muscle. This double fluorescence staining protocol combined with image analysis is accurate and less labor-intensive than classical procedures for determining the cross-sectional area of myofibers.

  11. A Quantitative and Standardized Method for the Evaluation of Choroidal Neovascularization Using MICRON III Fluorescein Angiograms in Rats

    PubMed Central

    Wigg, Jonathan P.; Zhang, Hong; Yang, Dong

    2015-01-01

    Introduction In-vivo imaging of choroidal neovascularization (CNV) has been increasingly recognized as a valuable tool in the investigation of age-related macular degeneration (AMD) in both clinical and basic research applications. Arguably the most widely utilised model replicating AMD is laser generated CNV by rupture of Bruch’s membrane in rodents. Heretofore CNV evaluation via in-vivo imaging techniques has been hamstrung by a lack of appropriate rodent fundus camera and a non-standardised analysis method. The aim of this study was to establish a simple, quantifiable method of fluorescein fundus angiogram (FFA) image analysis for CNV lesions. Methods Laser was applied to 32 Brown Norway Rats; FFA images were taken using a rodent specific fundus camera (Micron III, Phoenix Laboratories) over 3 weeks and compared to conventional ex-vivo CNV assessment. FFA images acquired with fluorescein administered by intraperitoneal injection and intravenous injection were compared and shown to greatly influence lesion properties. Utilising commonly used software packages, FFA images were assessed for CNV and chorioretinal burns lesion area by manually outlining the maximum border of each lesion and normalising against the optic nerve head. Net fluorescence above background and derived value of area corrected lesion intensity were calculated. Results CNV lesions of rats treated with anti-VEGF antibody were significantly smaller in normalised lesion area (p<0.001) and fluorescent intensity (p<0.001) than the PBS treated control two weeks post laser. The calculated area corrected lesion intensity was significantly smaller (p<0.001) in anti-VEGF treated animals at 2 and 3 weeks post laser. The results obtained using FFA correlated with, and were confirmed by conventional lesion area measurements from isolectin stained choroidal flatmounts, where lesions of anti-VEGF treated rats were significantly smaller at 2 weeks (p = 0.049) and 3 weeks (p<0.001) post laser. Conclusion The presented method of in-vivo FFA quantification of CNV, including acquisition variable corrections, using the Micron III system and common use software establishes a reliable method for detecting and quantifying CNV enabling longitudinal studies and represents an important alternative to conventional CNV quantification methods. PMID:26024231

  12. Investigation on interaction and sonodynamic damage of fluorescein derivants to bovine serum albumin (BSA) under ultrasonic irradiation

    NASA Astrophysics Data System (ADS)

    Zou, Mingming; Zhang, Lei; Wang, Jun; Wang, Qi; Gao, Jingqun; Fan, Ping

    2013-06-01

    The fluorescein derivants (Fluorescein: (2-(6-Hydroxy-3-oxo-(3H)-xanthen-9-yl) benzoic acid), Fluorescein-DA: (Bis [N,N-bis (carboxymethyl) aminomethyl] fluorescein) and Fluorescein-DAsbnd Fe(III): (Bis [N,N-bis (carboxymethyl) aminomethyl] fluoresceinsbnd Ferrous(III)) with a tricyclic plane structure were used to study the interaction and sonodynamic damage to bovine serum albumin (BSA) under ultrasonic irradiation through fluorospectrometry and UV-vis spectrophotometry. Besides, because of the existence of Fe(III) ion in Fluorescein-DAsbnd Fe(III), under ultrasonic irradiation the sonocatalytic activity in the damage of BSA molecules was also found. Three-dimensional fluorescence spectra and three-dimensional fluorescence contour profile spectra were mentioned to determine the fluorescence quenching and the conformation change of BSA in the absence and presence of these fluorescein derivants. As judged from the experimental results, the fluorescence quenching of BSA in aqueous solution caused by these fluorescein derivants were all attributed to static quenching process. The damage degree and mode were related to some factors such as ultrasonic irradiation time, fluorescein derivant concentration and ionic strength. Finally, several quenchers were used to determine the amount and kind of generated reactive oxygen species (ROS) during sonodynamic and sonocatalytic reaction processes. It suggests that these fluorescein derivants induce protein damage via various ROS, at least, including singlet oxygen (1O2) and hydroxyl radicals (rad OH). Perhaps, this paper may offer some important subjects for broadening the application of these fluorescein derivants in sonodynamic therapy (SDT) and sonocatalytic therapy (SCT) technologies for tumor treatment.

  13. Methods for chromosome-specific staining

    DOEpatents

    Gray, J.W.; Pinkel, D.

    1995-09-05

    Methods and compositions for chromosome-specific staining are provided. Compositions comprise heterogeneous mixtures of labeled nucleic acid fragments having substantially complementary base sequences to unique sequence regions of the chromosomal DNA for which their associated staining reagent is specific. Methods include ways for making the chromosome-specific staining compositions of the invention, and methods for applying the staining compositions to chromosomes. 3 figs.

  14. Golgi-Cox Staining Step by Step

    PubMed Central

    Zaqout, Sami; Kaindl, Angela M.

    2016-01-01

    Golgi staining remains a key method to study neuronal morphology in vivo. Since most protocols delineating modifications of the original staining method lack details on critical steps, establishing this method in a laboratory can be time-consuming and frustrating. Here, we describe the Golgi-Cox staining in such detail that should turn the staining into an easily feasible method for all scientists working in the neuroscience field. PMID:27065817

  15. Golgi-Cox Staining Step by Step.

    PubMed

    Zaqout, Sami; Kaindl, Angela M

    2016-01-01

    Golgi staining remains a key method to study neuronal morphology in vivo. Since most protocols delineating modifications of the original staining method lack details on critical steps, establishing this method in a laboratory can be time-consuming and frustrating. Here, we describe the Golgi-Cox staining in such detail that should turn the staining into an easily feasible method for all scientists working in the neuroscience field. PMID:27065817

  16. Determination of torasemide by fluorescence quenching method with some dihalogenated fluorescein dyes as probes

    NASA Astrophysics Data System (ADS)

    Cui, Zhiping; Liu, Shaopu; Liu, Zhongfang; Li, Yuanfang; Hu, Xiaoli; Tian, Jing

    2013-10-01

    A novel fluorescence quenching method for the determination of torasemide (TOR) with some dihalogenated fluorescein dyes as fluorescence probes was developed. In acidulous medium, TOR could interact with some dihalogenated fluorescein dyes such as dichlorofluorescein (DCF), dibromofluorescein (DBF) and diiodofluorescein (DIF) to form binary complexes, which could lead to fluorescence quenching of above dihalogenated fluorescein dyes. The maximum fluorescence emission wavelengths were located at 532 nm (TOR-DCF), 535 nm (TOR-DBF) and 554 nm (TOR-DIF). The relative fluorescence intensities (ΔF = F0 - F) were proportional to the concentration of TOR in certain ranges. The detection limits were 4.8 ng mL-1 for TOR-DCF system, 9.8 ng mL-1 for TOR-DBF system and 35.1 ng mL-1 for TOR-DIF system. The optimum reaction conditions, influencing factors were studied; and the effect of coexisting substances was investigated owing to the highest sensitivity of TOR-DCF system. In addition, the reaction mechanism, composition and structure of the complex were discussed by quantum chemical calculation and Job's method. The fluorescence quenching of dihalogenated fluorescein dyes by TOR was a static quenching process judging from the effect of temperature and the Stern-Volmer plots. The method was satisfactorily applied to the determination of TOR in tablets and human urine samples.

  17. Fluoride-induced modulation of ionic transport in asymmetric nanopores functionalized with ``caged'' fluorescein moieties

    NASA Astrophysics Data System (ADS)

    Ali, Mubarak; Ahmed, Ishtiaq; Ramirez, Patricio; Nasir, Saima; Cervera, Javier; Niemeyer, Christof M.; Ensinger, Wolfgang

    2016-04-01

    We demonstrate experimentally and theoretically a nanofluidic fluoride sensing device based on a single conical pore functionalized with ``caged'' fluorescein moieties. The nanopore functionalization is based on an amine-terminated fluorescein whose phenolic hydroxyl groups are protected with tert-butyldiphenylsilyl (TBDPS) moieties. The protected fluorescein (Fcn-TBDPS-NH2) molecules are then immobilized on the nanopore surface via carbodiimide coupling chemistry. Exposure to fluoride ions removes the uncharged TBDPS moieties due to the fluoride-promoted cleavage of the silicon-oxygen bond, leading to the generation of negatively charged groups on the fluorescein moieties immobilized onto the pore surface. The asymmetrical distribution of these groups along the conical nanopore leads to the electrical rectification observed in the current-voltage (I-V) curve. On the contrary, other halides and anions are not able to induce any significant ionic rectification in the asymmetric pore. In each case, the success of the chemical functionalization and deprotection reactions is monitored through the changes observed in the I-V curves before and after the specified reaction step. The theoretical results based on the Nernst-Planck and Poisson equations further demonstrate the validity of an experimental approach to fluoride-induced modulation of nanopore current rectification behaviour.

  18. Fluoride-induced modulation of ionic transport in asymmetric nanopores functionalized with "caged" fluorescein moieties.

    PubMed

    Ali, Mubarak; Ahmed, Ishtiaq; Ramirez, Patricio; Nasir, Saima; Cervera, Javier; Niemeyer, Christof M; Ensinger, Wolfgang

    2016-04-28

    We demonstrate experimentally and theoretically a nanofluidic fluoride sensing device based on a single conical pore functionalized with "caged" fluorescein moieties. The nanopore functionalization is based on an amine-terminated fluorescein whose phenolic hydroxyl groups are protected with tert-butyldiphenylsilyl (TBDPS) moieties. The protected fluorescein (Fcn-TBDPS-NH2) molecules are then immobilized on the nanopore surface via carbodiimide coupling chemistry. Exposure to fluoride ions removes the uncharged TBDPS moieties due to the fluoride-promoted cleavage of the silicon-oxygen bond, leading to the generation of negatively charged groups on the fluorescein moieties immobilized onto the pore surface. The asymmetrical distribution of these groups along the conical nanopore leads to the electrical rectification observed in the current-voltage (I-V) curve. On the contrary, other halides and anions are not able to induce any significant ionic rectification in the asymmetric pore. In each case, the success of the chemical functionalization and deprotection reactions is monitored through the changes observed in the I-V curves before and after the specified reaction step. The theoretical results based on the Nernst-Planck and Poisson equations further demonstrate the validity of an experimental approach to fluoride-induced modulation of nanopore current rectification behaviour. PMID:27050623

  19. Spectroscopic properties and amplified spontaneous emission of fluorescein laser dye in ionic liquids as green media

    NASA Astrophysics Data System (ADS)

    AL-Aqmar, Dalal M.; Abdelkader, H. I.; Abou Kana, Maram T. H.

    2015-09-01

    The use of ionic liquids (ILs) as milieu materials for laser dyes is a promising field and quite competitive with volatile organic solvents and solid state-dye laser systems. This paper investigates some photo-physical parameters of fluorescein dye incorporated into ionic liquids; 1-Butyl-3-methylimidazolium chloride (BMIM Cl), 1-Butyl-3-methylimidazolium tetrachloroaluminate (BMIM AlCl4) and 1-Butyl-3-methylimidazolium tetrafluoroborate (BMIM BF4) as promising host matrix in addition to ethanol as reference. These parameters are: absorption and emission cross-sections, fluorescence lifetime and quantum yield, in addition to the transition dipole moment, the attenuation length and oscillator strength were also investigated. Lasing characteristics such as amplified spontaneous emission (ASE), the gain, and the photostability of fluorescein laser dye dissolved in different host materials were assessed. The composition and properties of the matrix of ILs were found that it has great interest in optimizing the laser performance and photostability of the investigated laser dye. Under transverse pumping of fluorescein dye by blue laser diode (450 nm) of (400 mW), the initial ASE for dye dissolved in BMIM AlCl4 and ethanol were decreased to 39% and 36% respectively as time progressed 132 min. Relatively high efficiency and high fluorescence quantum yield (11.8% and 0.82% respectively) were obtained with good photostability in case of fluorescein in BMIM BF4 that was decreased to ∼56% of the initial ASE after continuously pumping with 400 mW for 132 min.

  20. A method for immunofluorescent demonstration of three coexisting neurotransmitters in rat brain and spinal cord, using the fluorophores fluorescein, lissamine rhodamine, and 7-amino-4-methylcoumarin-3-acetic acid.

    PubMed

    Wessendorf, M W; Appel, N M; Molitor, T W; Elde, R P

    1990-12-01

    Coexistence of neurotransmitters within single nerve fibers or terminals can be convincingly demonstrated by the use of multicolor immunofluorescence. The present study examined whether three-color immunocytochemical localization of coexisting neurotransmitters can be performed using the blue fluorophore AMCA. Spectrofluorometric examination of secondary antibodies conjugated with AMCA, fluorescein, and lissamine rhodamine showed that the peaks of excitation and emission were well separated and that dots of AMCA-conjugated IgG dried on slides were not visible when viewed using microscope filters for rhodamine and fluorescein. These findings suggest that AMCA might be suitable for three-color immunofluorescence. The usefulness of AMCA for triple labeling was tested directly by staining sections of rat brainstem and spinal cord for serotonin (5HT), substance P (SP), and either enkephalin (ENK) or prepro-thyrotropin-releasing hormone 160-169 (ppT), a marker peptide for thyrotropin-releasing hormone. Triple labeling for 5HT, SP, and ppT was observed in both brainstem and spinal cord but was only very rarely observed for 5HT,SP, and ENK. No evidence was found for artifactual triple labeling, although false negatives appeared to be possible in some circumstances. We conclude that AMCA can be combined with fluorescein and lissamine rhodamine for three-color immunofluorescent studies of coexisting neurotransmitters. In addition, the coexistence of 5HT with ENK appears to be much less common than the coexistence of 5HT with either SP or ppT. PMID:1701460

  1. Quantitative Spatial and Temporal Analysis of Fluorescein Angiography Dynamics in the Eye

    PubMed Central

    Hui, Flora; Nguyen, Christine T. O.; Bedggood, Phillip A.; He, Zheng; Fish, Rebecca L.; Gurrell, Rachel; Vingrys, Algis J.; Bui, Bang V.

    2014-01-01

    Purpose We describe a novel approach to analyze fluorescein angiography to investigate fluorescein flow dynamics in the rat posterior retina as well as identify abnormal areas following laser photocoagulation. Methods Experiments were undertaken in adult Long Evans rats. Using a rodent retinal camera, videos were acquired at 30 frames per second for 30 seconds following intravenous introduction of sodium fluorescein in a group of control animals (n?=?14). Videos were image registered and analyzed using principle components analysis across all pixels in the field. This returns fluorescence intensity profiles from which, the half-rise (time to 50% brightness), half-fall (time for 50% decay) back to an offset (plateau level of fluorescence). We applied this analysis to video fluorescein angiography data collected 30 minutes following laser photocoagulation in a separate group of rats (n?=?7). Results Pixel-by-pixel analysis of video angiography clearly delineates differences in the temporal profiles of arteries, veins and capillaries in the posterior retina. We find no difference in half-rise, half-fall or offset amongst the four quadrants (inferior, nasal, superior, temporal). We also found little difference with eccentricity. By expressing the parameters at each pixel as a function of the number of standard deviation from the average of the entire field, we could clearly identify the spatial extent of the laser injury. Conclusions This simple registration and analysis provides a way to monitor the size of vascular injury, to highlight areas of subtle vascular leakage and to quantify vascular dynamics not possible using current fluorescein angiography approaches. This can be applied in both laboratory and clinical settings for in vivo dynamic fluorescent imaging of vasculature. PMID:25365578

  2. A novel biosensor for Escherichia coli O157:H7 based on fluorescein-releasable biolabels.

    PubMed

    Hu, Rong-Rong; Yin, Zheng-Zhi; Zeng, Yan-Bo; Zhang, Jian; Liu, Hai-Qing; Shao, Yong; Ren, Shi-Bin; Li, Lei

    2016-04-15

    New techniques are required for the rapid and sensitive detection of Escherichia coli O157:H7 (E. coli O157:H7), a pathogenic bacterium responsible for serious and sometimes life-threatening diseases in humans. In this study, we developed a highly sensitive and efficient biosensor for the quantitative detection of E. coli O157:H7 by integrating fluorescein-releasable biolabels with a magnetism-separable probe. Hollow silica nanospheres with a diameter of approximately 350 nm were synthesized, enriched with fluorescein, and surface-protected with macromolecule layers of poly (acrylic acid) and poly (dimethyldiallylammonium chloride). These fluorescein-enriched hollow silica nanospheres were characterized using scanning electron microscopy, transmission electron microscopy, and Fourier transform infrared spectroscopy. They were further functionalized as immune labels of E. coli O157:H7 for a sandwich-type immune reaction between this bacterium and magnetic nanoparticles (Fe3O4@SiO2). Next, the E. coli O157:H7 cells were captured, magnetically separated, and quantified based on the fluorescence intensity of the fluorescein released from the biolabels of the fluorescein-enriched hollow silica nanospheres. This analytic process can be completed within 75 min, and the biosensor showed a linear relationship ranging from 4 to 4.0 × 10(8)cfu/mL with a detection limit of 3 cfu/mL. These results show that the developed fluorescent sensor has excellent specificity, and good reproducibility and stability. This study used real spiked samples for detection, indicating that this technique has a wide range of potential applications and may be readily adapted for detecting other pathogens. PMID:26584080

  3. Silver staining techniques of polyacrylamide gels.

    PubMed

    Bartsch, Holger; Arndt, Claudia; Koristka, Stefanie; Cartellieri, Marc; Bachmann, Michael

    2012-01-01

    Although the main application for polyacrylamide gels is the separation and subsequent blotting of proteins for immunodetection, there are tasks that need staining of proteins in the polyacrylamide gel. Several different staining techniques exist for protein staining in SDS gels that differ in their sensitivity, their expenditure of time, and other aspects. Still, silver staining is the most sensitive and reliable staining technique. Because this technique was developed in the 1970s, a huge number of variations exist. Therefore, we will provide herein three methods, which are robust and easy to perform. PMID:22585513

  4. Histological stain evaluation for machine learning applications

    PubMed Central

    Azar, Jimmy C.; Busch, Christer; Carlbom, Ingrid B.

    2013-01-01

    Aims: A methodology for quantitative comparison of histological stains based on their classification and clustering performance, which may facilitate the choice of histological stains for automatic pattern and image analysis. Background: Machine learning and image analysis are becoming increasingly important in pathology applications for automatic analysis of histological tissue samples. Pathologists rely on multiple, contrasting stains to analyze tissue samples, but histological stains are developed for visual analysis and are not always ideal for automatic analysis. Materials and Methods: Thirteen different histological stains were used to stain adjacent prostate tissue sections from radical prostatectomies. We evaluate the stains for both supervised and unsupervised classification of stain/tissue combinations. For supervised classification we measure the error rate of nonlinear support vector machines, and for unsupervised classification we use the Rand index and the F-measure to assess the clustering results of a Gaussian mixture model based on expectation–maximization. Finally, we investigate class separability measures based on scatter criteria. Results: A methodology for quantitative evaluation of histological stains in terms of their classification and clustering efficacy that aims at improving segmentation and color decomposition. We demonstrate that for a specific tissue type, certain stains perform consistently better than others according to objective error criteria. Conclusions: The choice of histological stain for automatic analysis must be based on its classification and clustering performance, which are indicators of the performance of automatic segmentation of tissue into morphological components, which in turn may be the basis for diagnosis. PMID:23766933

  5. Comparison of various staining methods for the detection of Cryptosporidium in cell-free culture.

    PubMed

    Boxell, Annika; Hijjawi, Nawal; Monis, Paul; Ryan, Una

    2008-09-01

    The complete development of Cryptosporidium in host cell-free medium first described in 2004, represented a significant advance that can facilitate many aspects of Cryptosporidium research. A current limitation of host cell-free cultivation is the difficulty involved in visualising the life-cycle stages as they are very small in size, morphologically difficult to identify and dispersed throughout the media. This is in contrast to conventional cell culture methods for Cryptosporidium, where it is possible to focus on the host cells and view the foci of infection on the host cells. In the present study, we compared three specific and three non-specific techniques for visualising Cryptosporidium parvum life-cycle stages in cell-free culture; antibody staining using anti-sporozoite and anti-oocyst wall antibodies (Sporo-Glo and Crypto Cel), fluorescent in-situ hybridization (FISH) using a Cryptosporidium specific rRNA oligonucleotide probe and the non-specific dyes; Texas Red, carboxyfluorescein diacetate succinimidyl ester (CFSE) and 4,6' diamino-2-phenylindole dihydrochloride (DAPI). Results revealed that a combination of Sporo-Glo and Crypto Cel staining resulted in easy and reliable identification of all life-cycle stages. PMID:18547565

  6. "Coffee Ring Effect" in Ophthalmology: "Anionic Dye Deposition" Hypothesis Explaining Normal Lid Margin Staining.

    PubMed

    Rajabi, Mohammad Taher; Sharifzadeh, Morteza

    2016-04-01

    The process of formation of Marx line is studied in this article. Various theories have been proposed previously, in order to explain the mechanisms which lead to the development of Marx line. These theories are based on the characteristics of stained area and do not pay attention to the behavior of dye solution itself on the surface. The aim of this study is to investigate the latter behavior and introduce a new theory based on it, in order to explain the process of the Marx line formation.This study also introduces "Coffee Ring Effect" and its possible applications in explaining some ophthalmological phenomena.The effect of dye solution's behavior on the beneath surface is adopted in order to propose a novel theory. This new hypothesis is called "Anionic Dye Deposition" which was based on "Coffee Ring Effect" phenomenon. For evaluation of this theory, Evaporation pattern of Rose Bengal and fluorescein were analyzed on different surfaces. Furthermore, the effect of tear meniscus alteration on lid margin staining is studied.During the evaporation process of dye solutions, it was observed that almost all of the solute was deposited at the edge of the drop on hydrophilic surfaces. Furthermore, in the study of lid margin staining, it is observed that tear meniscus alteration during gaze affects staining pattern. This observation invalidates former hypotheses which only focus on stained surface characteristics.According to the observations in this study, it is proposed that Marx line staining occurs as a result of "anionic dye deposition" due to evaporation. PMID:27057835

  7. Rapid, one step staining procedures for analysis of cellular DNA and protein by single and dual laser flow cytometry

    SciTech Connect

    Crissman, H.A.; Steinkamp, J.A.

    1982-01-01

    Detailed, simplified techniques are described for simultaneous staining and analysis of DNA and protein in a number of mammalian cell types. Cell staining involves the addition of appropriate dye mixtures to unfixed or ethanol-fixed cells and subsequent analysis of cell populations in the staining reagents generally within 10 to 20 minutes. The approach is novel in that no centrifugation steps are involved during the staining procedure, thus, eliminating cell clumping and cell loss and making the procedures appropriate for samples containing limited numbers of cells. For single wavelength analysis, staining of DNA and protein in ethanol-fixed cells was accomplished with a dye solution containing propidium iodide, fluorescein isothimocyanate and RNase. After 20 minutes at room temperature cells were analyzed using the 488 nanometer (nm) laser excitation line. For dual laser analysis the following dye combinations were employed without RNase: mithramycin-rhodamine 640, mithramycin-substituted rhodamine isothiocyanate, Hoechst 33342-rhodamine 640 and Hoechst 33342-rhodamine isothiocyanate. Unfixed cells were also stained with the Hoechst 33342-rhodamine 640 dye combination. Mithramycin was excited at 457.9 nm, Hoechst 33342 at 333-363 nm, and rhodamine dyes at 568 nm. Cell types analyzed included Chinese hamster ovary cells, cultured mouse colon 26 cells, mouse embryo forelimb bud cells, and rat cell obtained by lung lavage.

  8. Quantitative studies of immunofluorescent staining

    PubMed Central

    Wick, G.; Beutner, E. H.

    1970-01-01

    The antiperinuclear factor (APF) is found in a high percentage of sera from patients with rheumatoid arthritis. It can be demonstrated by direct immunofluorescence using the keratohyaline granules of human buccal mucosa as antigenic substrate. Mixing of some normal goat sera with an APF positive serum from a patient with rheumatoid arthritis resulted in an inhibition of the APF titre of the patient's serum. However, there was no clear cut correlation between the APF-positivity of normal goat sera and their inhibitory effect on the APF-reactivity of a human rheumatoid arthritis patient's serum. In reciprocal screening tests the human rheumatoid arthritis serum blocked only one of the APF-reactive goat sera. The reciprocal blocking activity of this goat serum and the patient's serum could be more exactly evaluated by the use of chessboard titrations in an indirect immunofluorescence blocking test. This test consisted of mixing equal volumes of serial dilutions of a goat serum and the patient's serum and subsequent examination of the mixtures for APF using an anti-human IgG conjugate and an anti-goat immunoglobulin conjugate, respectively. The results point to an antibody nature for the APF in preimmune, normal goat sera and to the value of chessboard titrations of this type in demonstrating the identity, non-identity, partial identity (or very close proximity of antigenic determinants) of the antibodies in different antisera which cannot be distinguished by their immunofluorescent staining patterns. ImagesFIG. 1FIG. 2 PMID:4913803

  9. Antimicrobial activity of lactic acid bacteria against Listeria monocytogenes on frankfurters formulated with and without lactate/diacetate.

    PubMed

    Koo, Ok-Kyung; Eggleton, Mallory; O'Bryan, Corliss A; Crandall, Philip G; Ricke, Steven C

    2012-12-01

    Contamination by Listeria monocytogenes has been a constant public health threat for the ready-to-eat (RTE) meat industry due to the potential for high mortalities from listeriosis. Lactic acid bacteria (LAB) have shown protective action against various pathogenic bacteria. The aim of this study was to evaluate the antilisterial activity of a combination of three LAB strains (Lactiguard) on L. monocytogenes. The combination of the LAB was inhibitory to L. monocytogenes inoculated onto frankfurters not containing lactate/diacetate after 8weeks of refrigerated storage (0.6 log reduction compared to L. monocytogenes only control), and when a cell free extract (CFS) of the LAB was added with LAB even more inhibition was obtained (1.2 log reduction compared with L. monocytogenes only). In frankfurters containing lactate/diacetate the LAB and the LAB plus CFS were more effective in reducing growth of L. monocytogenes after 8 weeks of refrigerated storage (2 and 3.3 log reductions respectively). PMID:22704134

  10. The effect of methionine, thiouracil, dienestrol diacetate and thyroprotein on the development and prevention of fatty liver in pullets.

    PubMed

    Roberson, R H; Trujillo, T

    1975-05-01

    The effect of two levels each of methionine (0.0 and 0.07 percent), thiouracil (0.0 and 0.05 percent), dienestrol diacetate (0.0 and 0.007 percent), and thyroactive casein (0.0 and 0.0125 percent) on the performancy, organ changes, and liver composition in 640 pullets of two strains was studied in a 24 factorial arrangement of treatments. Egg production, egg characteristics, feed conversion, organ weights, and liver composition were parameters measured. Supplemental methionine increased the phosphorus content of liver fat in strain A, but other parameters in the two strains were mot affected by the increase in dietary methionine. The thiouracil increased weight grains, gram of fat per total liver, percent of liver fat, thyroid weight, and heart weight but decreased the phosphorus content of liver fat. Nine typical cases of fatty liver syndrome with large liver hematomas occurred in the thiouracil treated birds and one case occurred in an untreated pullet. Dienestrol diacetate did not affect egg production, egg characteristics, organ weights, and liver composition in the two strains. Thyroprotein decreased weight gain, abdominal fat, liver weight. liver fat, thyroid weight, and percent red cells, but decreased percent blood sports in eggs and adjusted weights of the kidney and heart in both strains. PMID:1153373

  11. The effect of tricaine on use of the fluorescein test for detecting skin and corneal ulcers in fish.

    PubMed

    Davis, Michael W; Stephenson, Jeana; Noga, Edward J

    2008-06-01

    Fluorescein has been used for rapid and sensitive detection of fish skin and corneal ulceration. Effective use of the fluorescein test requires knowledge of conditions that might cause misleading interpretations or otherwise interfere with test reliability. Examination of fish health and the clinical workup often require tricaine as one of the most commonly used anesthetics. However, tricaine may interfere with correct interpretation of the fluorescein test and might also cause significant fish injury. The effects of tricaine exposure sequence on the fidelity of the fluorescein test was studied in Pacific halibut Hippoglossus stenolepis, walleye pollock Theragra chalcogramma, and northern rock soles Lepidopsetta polyxystra by examining the fluorescence of experimentally induced epidermal wounding. Tricaine can quench fluorescence that is emitted by fluorescein retained in skin ulcers, causing a false-negative reaction. Thus, for the fluorescein test to work properly, it is important to avoid the exposure of fluorescein-treated and rinsed ulcers to tricaine. The effects of exposure to buffered versus unbuffered tricaine on epidermal and corneal integrity were studied in Nile tilapia Oreochromis niloticus and channel catfish Ictalurus punctatus subjected to the fluorescein test and histological examination. Fluorescein could detect not only ulcers but also areas with only a partial loss of epithelium (i.e., erosion). The use of unbuffered tricaine to anesthetize these fish caused serious epidermal and corneal damage. If fish are euthanized with unbuffered tricaine for clinical workup, this severe epidermal or corneal damage could be misinterpreted as an antemortem lesion, leading to misdiagnosis. Even in water with alkalinity exceeding 50 mg/L as CaCO3, it would seem prudent to always buffer tricaine with sodium bicarbonate to prevent a pH change that might lead to iatrogenic effects from unbuffered tricaine. Thus, current general recommendations suggesting that tricaine does not need to be buffered in waters with alkalinity greater than 50 mg/L might need to be modified. PMID:18783129

  12. Analysis of chemical equilibrium of silicon-substituted fluorescein and its application to develop a scaffold for red fluorescent probes.

    PubMed

    Hirabayashi, Kazuhisa; Hanaoka, Kenjiro; Takayanagi, Toshio; Toki, Yuko; Egawa, Takahiro; Kamiya, Mako; Komatsu, Toru; Ueno, Tasuku; Terai, Takuya; Yoshida, Kengo; Uchiyama, Masanobu; Nagano, Tetsuo; Urano, Yasuteru

    2015-09-01

    Fluorescein is a representative green fluorophore that has been widely used as a scaffold of practically useful green fluorescent probes. Here, we report synthesis and characterization of a silicon-substituted fluorescein, i.e., 2-COOH TokyoMagenta (2-COOH TM), which is a fluorescein analogue in which the O atom at the 10' position of the xanthene moiety of fluorescein is replaced with a Si atom. This fluorescein analogue forms a spirolactone ring via intramolecular nucleophilic attack of the carboxylic group in a pH-dependent manner. Consequently, 2-COOH TM exhibits characteristic large pH-dependent absorption and fluorescence spectral changes: (1) 2-COOH TM is colorless at acidic pH, whereas fluorescein retains observable absorption and fluorescence even at acidic pH, and the absorption maximum is also shifted; (2) the absorption spectral change occurs above pH 7.0 for 2-COOH TM and below pH 7.0 for fluorescein; (3) 2-COOH TM shows a much sharper pH response than fluorescein because of its pKa inversion, i.e., pKa1 > pKa2. These features are also different from those of a compound without the carboxylic group, 2-Me TokyoMagenta (2-Me TM). Analysis of the chemical equilibrium between pH 3.0 and 11.0 disclosed that 2-COOH TM favors the colorless and nonfluorescent lactone form, compared with fluorescein. Substitution of Cl atoms at the 4' and 5' positions of the xanthene moiety of 2-COOH TM to obtain 2-COOH DCTM shifted the equilibrium so that the new derivative exists predominantly in the strongly fluorescent open form at physiological pH (pH 7.4). To demonstrate the practical utility of 2-COOH DCTM as a novel scaffold for red fluorescent probes, we employed it to develop a probe for β-galactosidase. PMID:26237524

  13. Effect of liposomal fluidity on skin permeation of sodium fluorescein entrapped in liposomes

    PubMed Central

    Subongkot, Thirapit; Ngawhirunpat, Tanasait

    2015-01-01

    The purpose of this study was to investigate the effect of ultradeformable liposome components, Tween 20 and terpenes, on vesicle fluidity. The fluidity was evaluated by electron spin resonance spectroscopy using 5-doxyl stearic acid and 16-doxyl stearic acid as spin labels for phospholipid bilayer fluidity at the C5 atom of the acyl chain near the polar head group (hydrophilic region) and the C16 atom of the acyl chain (lipophilic region), respectively. The electron spin resonance study revealed that Tween 20 increased the fluidity at the C5 atom of the acyl chain, whereas terpenes increased the fluidity at the C16 atom of the acyl chain of the phospholipid bilayer. The increase in liposomal fluidity resulted in the increased skin penetration of sodium fluorescein. Confocal laser scanning microscopy showed that ultradeformable liposomes with terpenes increase the skin penetration of sodium fluorescein by enhancing hair follicle penetration. PMID:26229462

  14. Methods to measure the reactivity of peroxynitrite-derived oxidants toward reduced fluoresceins and rhodamines.

    PubMed

    Wardman, Peter

    2008-01-01

    The commonest probes for "reactive oxygen and nitrogen species" are reduced fluorescein and rhodamine dyes that fluoresce when oxidized. The reduced dyes are reactive toward peroxynitrite, although probably not directly but via free radical oxidants derived from it: hydroxyl, carbonate, and nitrogen dioxide free radicals. The reaction with peroxynitrite can be monitored by rapid mixing and stopped-flow spectrophotometry, but reliable measurement of reactivity of the peroxynitrite-derived radicals requires specialized techniques such as flash photolysis or pulse radiolysis to monitor the fast reactions in real time. A key feature of oxidation by radicals is that the reaction produces an intermediate fluorescein or rhodamine radical, which normally is oxidized further by oxygen to yield the fluorescent, stable product. Susceptibility of the yield of fluorescence to interference by antioxidants can be assessed from kinetic parameters, which reflect reactivity. This chapter outlines methods for estimation of key rate constants involving peroxynitrite-derived oxidants. PMID:18554539

  15. Detection of the halogenating activity of heme peroxidases in leukocytes by aminophenyl fluorescein.

    PubMed

    Flemmig, J; Remmler, J; Zschaler, J; Arnhold, J

    2015-06-01

    The formation of hypochlorous and hypobromous acids by heme peroxidases is a key property of certain immune cells. These products are not only involved in defense against pathogenic microorganisms and in regulation of inflammatory processes, but contribute also to tissue damage in certain pathologies. After a short introduction about experimental approaches for the assessment of the halogenating activity in vitro and in cell suspensions, we are focusing on novel applications of fluorescent dye systems to detect the formation of hypochlorous acid (HOCl) in leukocytes. Special attention is directed to properties and applications of the non-fluorescent dye aminophenyl fluorescein that is converted by HOCl, HOBr, and other strong oxidants to fluorescein. This dye allows the detection of the halogenating activity in samples containing free myeloperoxidase and eosinophil peroxidase as well as in intact granulocytes using fluorescence spectroscopy and flow cytometry, respectively. PMID:25536418

  16. Investigation of Fluorescence Resonance Energy Transfer between Fluorescein and Rhodamine 6G

    NASA Astrophysics Data System (ADS)

    Saha, Jaba; Datta Roy, Arpan; Dey, Dibyendu; Chakraborty, Santanu; Bhattacharjee, D.; Paul, P. K.; Hussain, Syed Arshad

    2015-10-01

    Fluorescence Resonance Energy Transfer between two organic dyes Fluorescein and Rhodamine 6G was investigated in aqueous solution in presence and absence of synthetic clay laponite. Spectroscopic studies suggest that both the dyes were present mainly as monomer in solution. Fluorescence Resonance Energy Transfer occurred from Fluorescein to Rhodamine 6G in solutions. Energy transfer efficiency increases in presence of laponite and the maximum efficiency was 72.00% in aqueous laponite dispersion. Energy transfer efficiency was found to be pH sensitive. It has been demonstrated that with proper calibration it is possible to use the present system under investigation to sense pH over a wide range from 1.5 to 8.0.

  17. Influence of ionization states of antigen on anti-fluorescein antibodies

    NASA Astrophysics Data System (ADS)

    Fukunishi, Hiroaki

    2012-10-01

    Ratios of anion and di-anion states of fluorescein (FLU(-1) and FLU(-2)) are 21.2% and 78.8%, respectively, in the neutral pH. We investigated the influence of ionization states of antigen on anti-fluorescein antibodies. For this purpose, steered molecular dynamics (SMD) simulations were performed. Potential of mean forces (PMF) based on Jarzynski equality showed that wild-type (4-4-20) more strongly binds to FLU(-1) than FLU(-2), whereas its femtomolar-affinity mutant (4M5.3) more strongly binds to FLU(-2) than FLU(-1). It was speculated that the environment or the process of in vivo antibody production had been different from those of the protein engineering.

  18. Development of poly-l-lysine-coated calcium-alginate microspheres encapsulating fluorescein-labeled dextrans

    NASA Astrophysics Data System (ADS)

    Charron, Luc; Harmer, Andrea; Lilge, Lothar

    2005-09-01

    A technique to produce fluorescent cell phantom standards based on calcium alginate microspheres with encapsulated fluorescein-labeled dextrans is presented. An electrostatic ionotropic gelation method is used to create the microspheres which are then exposed to an encapsulation method using poly-l-lysine to trap the dextrans inside. Both procedures were examined in detail to find the optimal parameters producing cell phantoms meeting our requirements. Size distributions favoring 10-20 microns microspheres were obtained by varying the high voltage and needle size parameters. Typical size distributions of the samples were centered at 150 μm diameter. Neither the molecular weight nor the charge of the dextrans had a significant effect on their retention in the microspheres, though anionic dextrans were chosen to help in future capillary electrophoresis work. Increasing the exposure time of the microspheres to the poly-l-lysine solution decreased the leakage rates of fluorescein-labeled dextrans.

  19. Silver staining of proteins in polyacrylamide gels.

    PubMed

    Chevallet, Mireille; Luche, Sylvie; Rabilloud, Thierry

    2006-01-01

    Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It combines excellent sensitivity (in the low nanogram range) with the use of very simple and cheap equipment and chemicals. It is compatible with downstream processing, such as mass spectrometry analysis after protein digestion. The sequential phases of silver staining are protein fixation, then sensitization, then silver impregnation and finally image development. Several variants of silver staining are described here, which can be completed in a time range from 2 h to 1 d after the end of the electrophoretic separation. Once completed, the stain is stable for several weeks. PMID:17487168

  20. Bodian's Silver Method Stains Neurofilament Polypeptides

    NASA Astrophysics Data System (ADS)

    Gambetti, P.; Autilio-Gambetti, L.; Papasozomenos, S. Ch.

    1981-09-01

    Bodian's silver method was used to stain polypeptides of rat spinal cord or peripheral nerve separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The bands corresponding to the three polypeptide subunits of the neurofilaments were intensely impregnated. Two other polypeptides were stained inconsistently and less intensely. The tubulin band was stained weakly or not at all; other polypeptides, including glial fibrillary acidic protein, actin, and vimentin, remained unstained. This novel application of Bodian's method provides indirect proof that neurofilaments are the neuronal subcellular structure stained by the technique.

  1. Orcein-picroindigocarmine--a new multiple stain.

    PubMed

    Steven, P; Paulsen, F; Tillmann, B

    2000-10-01

    A new "orcein-picroindigocarmine staining", a colour combination of orcein, indigo carmine, and picric acid, was developed for histological applications. The new technique was tested on different human tissues. Colours ranging from red to brown, yellow, green and blue were observed in paraffine sections of tissues stained by this method. Nuclear structures in all tissues were stained dark brown to dark blue. Squamous epithelium was stained light brown with varying shades of blue in upper horny layers, whereas the ciliated epithelium was tinged blue grey. When connective tissue was stained, collagen fibrils appeared strongly blue next to elastic fibres, which took on a rust brown tinge; cellular components were all coloured brown. The matrix of hyaline cartilage was stained in different shades of blue, with the chondrocytes rust brown. Sections of bone components appeared dark blue to dark green. Skeletal muscle cells were coloured yellow and green with blue collagenous septa. The new staining is useful for distinguishing connective tissue components such as elastic fibres and collagen fibrils. It also demonstrates chondrocytes in favourable contrast to the cartilage matrix. The technique produces aesthetic staining colouring that could supplement histological investigations and provide an alternative to other staining materials. PMID:11073070

  2. The analysis of an anti-fluorescein response at limit dilution: the effect of lymphokine preparations on the apparent specificity.

    PubMed Central

    Skinner, M A; Prestidge, R; Marbrook, J

    1985-01-01

    The specificity of cytotoxic T cells in an anti-fluorescein response has been analysed at limit dilution. According to the ability to discriminate between fluoresceinated and non-fluoresceinated targets, the apparent specificity of CTL depends on the quality and quantity of the lymphokine preparations used to support the response. An acid-labile component of the factor preparations contributes to the generation of non-specific CTL. No cytotoxicity against NK-sensitive target cells was detected in the 'non-specific' component of the responses. PMID:3926636

  3. Third order nonlinear optical susceptibility of fluorescein-containing polymers determined by electro-absorption spectroscopy

    NASA Astrophysics Data System (ADS)

    Gomez-Sosa, Gustavo; Beristain, Miriam F.; Ortega, Alejandra; Martínez-Viramontes, Jaquelin; Ogawa, Takeshi; Fernández-Hernández, Roberto C.; Tamayo-Rivera, Lis; Reyes-Esqueda, Jorge-Alejandro; Isoshima, Takashi; Hara, Masahiko

    2012-03-01

    Novel polymers containing xanthene groups with high dye concentrations were prepared, and their third order nonlinear optical properties were studied by electroabsorption spectroscopy technique. The polymers were amorphous with refractive indices above 1.6 in the non-resonant region. The UV-Visible absorption spectra indicate the fluoresceins molecules in the polymers are H-aggregated. They showed third order nonlinear susceptibility, χ(3) (-ω:ω, 0, 0), of 2.5-3.5 × 10-12 esu.

  4. 2-D Registration and 3-D Shape Inference of the Retinal Fundus from Fluorescein Images

    PubMed Central

    Choe, Tae Eun; Medioni, Gerard; Cohen, Isaac; Walsh, Alexander C.; Sadda, SriniVas R.

    2008-01-01

    This study presents methods to 2-D registration of retinal image sequences and 3-D shape inference from fluorescein images. The Y-feature is a robust geometric entity that is largely invariant across modalities as well as across the temporal grey level variations induced by the propagation of the dye in the vessels. We first present a Y-feature extraction method that finds a set of Y-feature candidates using local image gradient information. A gradient-based approach is then used to align an articulated model of the Y-feature to the candidates more accurately while optimizing a cost function. Using mutual information, fitted Y-features are subsequently matched across images, including colors and fluorescein angiographic frames, for registration. To reconstruct the retinal fundus in 3-D, the extracted Y-features are used to estimate the epipolar geometry with a plane-and-parallax approach. The proposed solution provides a robust estimation of the fundamental matrix suitable for plane-like surfaces, such as the retinal fundus. The mutual information criterion is used to accurately estimate the dense disparity map, while the Y-features are used to estimate the bounds of the range space. Our experimental results validate the proposed method on a set of difficult fluorescein image pairs. PMID:18060827

  5. A selectively fluorescein-based colorimetric probe for detecting copper(II) ion.

    PubMed

    Zhang, Li; Zhang, Xianhong

    2014-12-10

    A novel fluorescein derivative 3-bromo-5-methylsalicylaldehyde fluorescein hydrazone (BMSFH) has been synthesized by reacting fluorescein hydrazide with 3-bromo-5-methylsalicylaldehyde and was developed as a new colorimetric probe for detection of Cu(2+). In the presence of Cu(2+) the BMSFH exhibits a rapid color change from colorless to yellow together with an obvious new band appeared at 502nm in the UV-vis absorption spectra. However, other common alkali-, alkaline earth-, transition- and rare earth metal ions induced no or minimal spectral changes. This change is attributed to BMSFH via coordination with Cu(2+) in a 1:1 stoichiometry and this binding to Cu(2+) is reversible, as indicated by the bleaching of the color when the Cu(2+) is extracted with EDTA. Experimental results indicate that the BMSFH can provide a rapid, selective and sensitive response to Cu(2+) with a linear dynamic range 3.0-330μmol/L and can be used as a potential Cu(2+) colorimetric probe in aqueous solution. PMID:24929315

  6. Fluorescein thiocarbamyl amino acids as internal standards for migration time correction in capillary sieving electrophoresis.

    PubMed

    Pugsley, Haley R; Swearingen, Kristian E; Dovichi, Norman J

    2009-04-10

    A number of algorithms have been developed to correct for migration time drift in capillary electrophoresis. Those algorithms require identification of common components in each run. However, not all components may be present or resolved in separations of complex samples, which can confound attempts for alignment. This paper reports the use of fluorescein thiocarbamyl derivatives of amino acids as internal standards for alignment of 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ)-labeled proteins in capillary sieving electrophoresis. The fluorescein thiocarbamyl derivative of aspartic acid migrates before FQ-labeled proteins and the fluorescein thiocarbamyl derivative of arginine migrates after the FQ-labeled proteins. These compounds were used as internal standards to correct for variations in migration time over a two-week period in the separation of a cellular homogenate. The experimental conditions were deliberately manipulated by varying electric field and sample preparation conditions. Three components of the homogenate were used to evaluate the alignment efficiency. Before alignment, the average relative standard deviation in migration time for these components was 13.3%. After alignment, the average relative standard deviation in migration time for these components was reduced to 0.5%. PMID:19249052

  7. A selectively fluorescein-based colorimetric probe for detecting copper(II) ion

    NASA Astrophysics Data System (ADS)

    Zhang, Li; Zhang, Xianhong

    2014-12-01

    A novel fluorescein derivative 3-bromo-5-methylsalicylaldehyde fluorescein hydrazone (BMSFH) has been synthesized by reacting fluorescein hydrazide with 3-bromo-5-methylsalicylaldehyde and was developed as a new colorimetric probe for detection of Cu2+. In the presence of Cu2+ the BMSFH exhibits a rapid color change from colorless to yellow together with an obvious new band appeared at 502 nm in the UV-vis absorption spectra. However, other common alkali-, alkaline earth-, transition- and rare earth metal ions induced no or minimal spectral changes. This change is attributed to BMSFH via coordination with Cu2+ in a 1:1 stoichiometry and this binding to Cu2+ is reversible, as indicated by the bleaching of the color when the Cu2+ is extracted with EDTA. Experimental results indicate that the BMSFH can provide a rapid, selective and sensitive response to Cu2+ with a linear dynamic range 3.0-330 μmol/L and can be used as a potential Cu2+ colorimetric probe in aqueous solution.

  8. Special stains in cytology: a technique using one half of a previously stained slide.

    PubMed

    Dhurandhar, N R; Zorrilla, O

    1988-01-01

    We describe a technique for performing special stains on one half of a slide previously stained by the Papanicolaou or Diff-Quick method. The technique varies depending on whether incubation is required during the staining procedure. For stains that do not require incubation, the portion of the slide to be preserved is covered with liquid paraffin. For stains that do require incubation, the area to be preserved is coverslipped with mounting medium and dried overnight prior to staining. The technique is easy, rapid, inexpensive, and enhances diagnostic accuracy. PMID:2474423

  9. Specificity of Immunofluorescent Staining for Study of Aspergillus flavus in Soil

    PubMed Central

    Schmidt, E. L.; Bankole, R. O.

    1965-01-01

    Fluorescein-labeled antiserum prepared with Aspergillus flavus strain CS was tested for specificity by staining fungi grown in soil in the vicinity of buried slides. All 14 strains of A. flavus fluoresced as intensely or nearly as intensely as the antigen control. Among 21 isolates of species of Aspergillus other than A. flavus, 17 reacted with moderate to low fluorescence at intensities readily distinguishable from that of A. flavus. The fluorescence of the remaining four cultures, and particularly A. sydowi, was indistinguishable from that of A. flavus. Fungi other than aspergilli were generally nonreactive. Interfering cross-reactions were encountered for one strain of Spicaria and one strain of Stemphylium; three isolates could not be evaluated because of interfering autofluorescence. An additional 22 isolates were either wholly negative or had a low order of fluorescence. Agglutination tests between each of the fungi and A. flavus CS serum revealed close agreement between agglutination titer and fluorescent-staining reaction. Unknown fungi freshly isolated from soil were checked for reaction to the A. flavus labeled antiserum; only one isolate gave a pronounced staining reaction, and that one proved to be a strain of A. flavus. In a simplified ecological model, the fluorescent-antibody technique was used to follow the development of A. flavus in mixed culture in soil with five other soil fungi. Images Fig. 1 PMID:5325934

  10. Fluorescein angiography

    MedlinePlus

    ... bar to keep your head still during the test. The health care provider will take pictures of the inside of ... to stop taking medicines that could affect the test results. Tell your health care provide about any allergies, particularly reactions to iodine. ...

  11. Effect of antibiotics on viability staining of Escherichia coli in solid phase cytometry.

    PubMed

    D'Haese, E; Nelis, H J

    2000-11-01

    Solid phase cytometry (SPC) has been investigated as a tool to assess the effect of antibiotics on the viability of Escherichia coli. After exposure of the cells to the antibiotic, they are retained on a polyester membrane filter and labelled using a fluorescein derivative as a substrate for intracellular esterases. The number of fluorescent bacteria is automatically counted in an Ar laser scanning device. In the presence of nutrients, all antibiotics tested in concentrations exceeding the MIC inhibited the multiplication of cells but not the labelling per se. However, when no nutrients were added, the cells did not multiply, and inhibition of the fluorescent staining was only observed for membrane permeabilizing antibiotics, even at sub-MIC concentrations. The selective detection by SPC of membrane-permeabilizing antibiotics corroborates the requirement of membrane integrity for viability labelling of bacteria. This selectivity has been exploited to develop a method for the detection of colistin residues in milk. PMID:11119151

  12. Mineral Stains at the No Name Prospect

    USGS scientist Art Bookstrom looks at greenish copper stain and pale pink cobalt bloom on limonite-stained meta-siltite and meta-argillite at the No Name prospect, near Iron Creek, in the southeastern part of the Idaho cobalt belt, in east-central Idaho....

  13. Efficiency of staining hair with indocyanine green

    NASA Astrophysics Data System (ADS)

    Kulyabina, Tatyana V.; Kochubey, Vyacheslav I.

    2005-06-01

    The efficiency of staining hair with indocyanine green (ICG) solution depending on type of hair, natural color, staining time and other parameters was investigated. Bonding ICG with hair material occurs due to interaction between ICG molecules and keratinocyte albumin. The penetration of ICG dye into hair meets with difficulties owing to surface protective layer.

  14. DETERMINATION OF 'GIARDIA MURIS' CYST VIABILITY BY DIFFERENTIAL INTERFERENCE CONTRAST, PHASE, OR BRIGHTFIELD MICROSCOPY

    EPA Science Inventory

    Recent experiments have demonstrated that fluorogenic substrates are taken up by Giardia cysts and that an excellent correlation exists between animal infectivity and vital staining with fluorescein diacetate (FDA) for viable cysts and propidium iodide (PI) for non-viable cysts. ...

  15. DETERMINATION OF ALIPHATIC AMINES IN WATER USING DERIVATIZATION WITH FLUORESCEIN ISOTHIOCYANATE AND CAPILLARY ELECTROPHORESIS/LASER-INDUCED FLUORESCENCE DETECTION.

    EPA Science Inventory

    Detection-oriented derivatization of aliphatic amines and amine functional groups in coumpounds of environmental interest was studied using fluorescein isothiocyanate (FITC) with separation/determination by capillary electrophoresis/laser-induced fluorescence. Determinative level...

  16. Porous cellulose diacetate-SiO2 composite coating on polyethylene separator for high-performance lithium-ion battery.

    PubMed

    Chen, Wenju; Shi, Liyi; Wang, Zhuyi; Zhu, Jiefang; Yang, Haijun; Mao, Xufeng; Chi, Mingming; Sun, Lining; Yuan, Shuai

    2016-08-20

    The developments of high-performance lithium ion battery are eager to the separators with high ionic conductivity and thermal stability. In this work, a new way to adjust the comprehensive properties of inorganic-organic composite separator was investigated. The cellulose diacetate (CDA)-SiO2 composite coating is beneficial for improving the electrolyte wettability and the thermal stability of separators. Interestingly, the pore structure of composite coating can be regulated by the weight ratio of SiO2 precursor tetraethoxysilane (TEOS) in the coating solution. The electronic performance of lithium ion batteries assembled with modified separators are improved compared with the pristine PE separator. When weight ratio of TEOS in the coating solution was 9.4%, the composite separator shows the best comprehensive performance. Compared with the pristine PE separator, its meltdown temperature and the break-elongation at elevated temperature increased. More importantly, the discharge capacity and the capacity retention improved significantly. PMID:27178959

  17. Effects and interactions of sodium lactate, sodium diacetate, and pediocin on the thermal inactivation of starved cells of Listeria monocytogenes on the surface of bologna

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The effects and interactions of temperature (56.3-60C) sodium lactate (SL; 0-4.8%), sodium diacetate (SDA; 0-2.5%), and pediocin (0-10,000 AU) on starved Listeria monocytogenes on bologna were investigated. Bologna slices containing SL and SDA in the formulation were dipped in pediocin, surface inoc...

  18. RADIATION (GAMMA) RESISTANCE AND POST-IRRADIATION GROWTH OF LISTERIA MONOCTYTOGENES SUSPENDED IN BEEF BOLOGNA THAT CONTAINED SODIUM DIACETATE AND POTASSIUM LACTATE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Listeria monocytogenes (Lm), a psychrotrophic food-borne pathogen, is a frequent post-process contaminant on ready-to-eat meat (RTE) products including bologna. Ionizing radiation can eliminate Lm from ready-to-eat meats. Sodium diacetate (SDA) and potassium lactate (PL) mixtures inhibit the growt...

  19. EFFECTS AND INTERACTIONS OF SODIUM LACTATE, SODIUM DIACETATE, AND PEDIOCIN ON THE THERMAL INACTIVATION OF STARVED CELLS OF LISTERIA MONOCYTOGENES ON THE SURFACE OF BOLOGNA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The effects and interactions of sodium lactate, sodium diacetate, and pediocin on the thermal inactivation of starved cells of Listeria monocytogenes on the surface of bologna were investigated. The heating temperatures used in the study were 56.3 to 60 degrees C and the antimicrobials were: sodium ...

  20. PREDICTIVE MODEL FOR THE COMBINED EFFECT OF TEMPERATURE, SODIUM LACTATE, AND SODIUM DIACETATE ON THE HEAT RESISTANCE OF LISTERIA MONOCYTOGENES IN BEEF

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Listeria monocytogenes continues to be one of the most important foodborne psychrotrophic pathogens of public health significance and a major concern to the food industry and regulatory agencies. Sodium lactate (NaL) and sodium diacetate (SDA) are generally regarded as safe and are used in meat prod...

  1. Modeling the Lag Phase and Growth Rate of Listeria monocytogenes in Ground Ham Containing Sodium Lactate and Sodium Diacetate at Various Storage Temperatures

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Refrigerated ready-to-eat (RTE) meats contaminated with L. monocytogenes were implicated in several listeriosis outbreaks. Lactate and diacetate have been shown to control L. monocytogenes in RTE meats. The objective of this study was to examine and model the effect of lactate (1.0-4.2%) and diace...

  2. Survival and growth of Listeria monocytogenes in broth as a function of temperature, pH, and potassium lactate and sodium diacetate concentrations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to determine the antimicrobial effect of a combination of potassium lactate and sodium diacetate (PURASAL P Opti.Form 4TM, 60% solution) on the survival and growth of Listeria monocytogenes Scott A in pH adjusted broth (5.5, 6.0, 6.5 and 7.0) stored at 4, 10, 17, 24, ...

  3. 2, 2'-(phenylazanediyl) diacetic acid modified Fe3O4@PEI for selective removal of cadmium ions from blood

    NASA Astrophysics Data System (ADS)

    Jin, Jun; Yang, Fang; Zhang, Fengwei; Hu, Wuquan; Sun, Shao-Bo; Ma, Jiantai

    2012-01-01

    A water-dispersible and supermagnetic nanocomposite (PAD-PEG-Fe3O4@PEI) has been successfully synthesized using polyethylenimine (PEI, Mol MW = 10000) coated supermagnetic Fe3O4-NH2 which was modified with 2, 2'-(phenylazanediyl) diacetic acid (PAD) through the bridge of poly(ethylene glycol) (PEG, Mol MW = 2000). The average particle size of PAD-PEG-Fe3O4@PEI was determined by TEM, and was about 50 nm. From magnetic hysteresis cycles for PAD-PEG-Fe3O4@PEI at room temperature, the saturation magnetization (Ms) was shown to be 58.14 emu g-1. Inductively coupled plasma spectrometry (ICP) analysis showed that the designed magnetic nanocomposite can remove 98% and 80% of Cd2+ from water and blood, respectively.A water-dispersible and supermagnetic nanocomposite (PAD-PEG-Fe3O4@PEI) has been successfully synthesized using polyethylenimine (PEI, Mol MW = 10000) coated supermagnetic Fe3O4-NH2 which was modified with 2, 2'-(phenylazanediyl) diacetic acid (PAD) through the bridge of poly(ethylene glycol) (PEG, Mol MW = 2000). The average particle size of PAD-PEG-Fe3O4@PEI was determined by TEM, and was about 50 nm. From magnetic hysteresis cycles for PAD-PEG-Fe3O4@PEI at room temperature, the saturation magnetization (Ms) was shown to be 58.14 emu g-1. Inductively coupled plasma spectrometry (ICP) analysis showed that the designed magnetic nanocomposite can remove 98% and 80% of Cd2+ from water and blood, respectively. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr11481j

  4. Standard specimens for stain calibration: application to Romanowsky-Giemsa staining.

    PubMed

    Turner, J N; Weir, B; Collins, D N

    1990-01-01

    Standardized specimens with reproducible staining properties were fabricated from extracts of biological objects (bovine liver, nucleoprotamine and defatted muscle). The standard specimens were stained with two formulations of the Romanowsky-Giemsa stain (RG), using the same azure B and eosin Y. One formulation used methanol and Sorensen's buffer and the other DMSO and Hepes buffer as solvents. The standard specimens were stained either in the composite stain or in the individual dyes dissolved in the same solvents and at the same concentration as the composite stain. Solution spectroscopy demonstrated different spectra for the two formulations with some wavelength regions varying by more than an order of magnitude. The RG spectra were also very different from those of the individual dyes dissolved at the RG concentration in the respective solvents. The stained standard specimens were analyzed by microspectrophotometry and were found to have spectra similar to those of cell smears. Furthermore, the standard specimens were shown to be a repeatable substrate for stain uptake. The transmitted light intensity from random fields of the same standardized specimen varied +/- 5%. When specimens were stained at the same time, the specimen-to-specimen variation depended on preparation conditions and the measurement wavelength, but was as good as +/- 5% for some conditions. The quantitative stain performance of both formulations was studied and compared. The standardized specimens provide a tool for the quantitative study of staining processes and specimen preparation procedures and for stain calibration. PMID:1694314

  5. Tooth staining effects of an alexidine mouthwash.

    PubMed

    Formicola, A J; Deasy, M J; Johnson, D H; Howe, E E

    1979-04-01

    The primary purpose of this study was to determine the amount of tooth staining produced by an alexidine mouthrinse. One hundred and eighty subjects rinsed twice daily for 1 month with either 15 ml of alexidine (0.035%) or a placebo solution. Prior to the study, the subjects were classified according to their smoking, coffee and tea drinking habits and these factors were subsequently considered in the analysis of the stain scores. Additionally, the effects on staining of a prior prophylaxis and the use of a fluoridated toothpaste during the study were determined. Upon termination of the study, subjects utilizing the active mouthrinse manifested a greater degree of staining than placebo users. The amount and intensity of the stain due to alexidine were not influenced (increased) by smoking, tea or coffee drinking habits. A prior prophylaxis did not reduce the staining propensity of alexidine users. The method of scoring developed can be used to assess the degree of tooth staining induced by antiplaque agents. PMID:374706

  6. Compact, Automated Centrifugal Slide-Staining System

    NASA Technical Reports Server (NTRS)

    Feeback, Daniel L.; Clarke, Mark S. F.

    2004-01-01

    The Directional Acceleration Vector-Driven Displacement of Fluids (DAVD-DOF) system, under development at the time of reporting the information for this article, would be a relatively compact, automated, centrifugally actuated system for staining blood smears and other microbiological samples on glass microscope slides in either a microgravitational or a normal Earth gravitational environment. The DAVD-DOF concept is a successor to the centrifuge-operated slide stainer (COSS) concept, which was reported in Slide-Staining System for Microgravity or Gravity (MSC-22949), NASA Tech Briefs, Vol. 25, No. 1 (January, 2001), page 64. The COSS includes reservoirs and a staining chamber that contains a microscope slide to which a biological sample is affixed. The staining chamber is sequentially filled with and drained of staining and related liquids from the reservoirs by use of a weighted plunger to force liquid from one reservoir to another at a constant level of hypergravity maintained in a standard swing-bucket centrifuge. In the DAVD-DOF system, a staining chamber containing a sample would also be sequentially filled and emptied, but with important differences. Instead of a simple microscope slide, one would use a special microscope slide on which would be fabricated a network of very small reservoirs and narrow channels connected to a staining chamber (see figure). Unlike in the COSS, displacement of liquid would be effected by use of the weight of the liquid itself, rather than the weight of a plunger.

  7. De-staining and re-staining mucins in formalin fixed paraffin sections.

    PubMed

    Smith, A A; Glickfield, I

    2011-04-01

    Re-staining of formalin fixed paraffin sections sometimes is required and this requires prior de-staining. Some simple and effective protocols for de-staining are described. Mucihematoxylin and mucicarmine can be removed with acid alcohol. Zirconyl hematoxylin can be removed with periodic acid or Sinha's fixative. Alcian blue can be removed with 5% trifluoroacetic acid in dichloromethane. Colloidal iron can be bleached in 1% household bleach in alcohol. PAS can be removed with hydrogen peroxide or ammonium hydroxide. With few exceptions, de-stained sections can be re-stained with mucihematoxylin, PAS or Gabe's trichrome. PMID:20001228

  8. Vessel extraction from non-fluorescein fundus images using orientation-aware detector.

    PubMed

    Yin, Benjun; Li, Huating; Sheng, Bin; Hou, Xuhong; Chen, Yan; Wu, Wen; Li, Ping; Shen, Ruimin; Bao, Yuqian; Jia, Weiping

    2015-12-01

    The automatic extraction of blood vessels in non-fluorescein eye fundus images is a tough task in applications such as diabetic retinopathy screening. However, vessel shapes have complex variations, and accurate modeling of retinal vascular structures is challenging. We have therefore developed a new approach to accurately extract blood vessels in non-fluorescein fundus images using an orientation-aware detector (OAD). The detector was designed according to the intrinsic property of vessels being locally oriented and having linearly elongated structures. We employ the OAD to extract vessel shapes with no assumptions on parametric orientations of vessel shapes. The orientations of vessels can be efficiently modeled by the energy distribution of Fourier transformation. Accordingly, both wide and thin vessels can be extracted with two-scale segmentation in which line operators are applied in large scale and the Gabor filter bank is applied in small scale. A post-processing technique, based on the path opening operation, is applied to eliminate false responses to nonvascular areas, such as retinal structures (optic disc and macula) and pathologies (exudates, hemorrhages,and microaneurysms). This makes the detector robust and structure-aware. By achieving a competitive CAL measurement of 80.82% for the DRIVE database and 68.94% for the STARE, the experimental results demonstrated that the OAD approach outperforms existing segmentation methods. Furthermore, the proposed approach effectively works with non-fluorescein fundus images and proves highly accurate and robust in complicated regions such as the central reflex, close vessels, and crossover points, despite a high level of illumination noise in the original data. PMID:26474120

  9. The Effect of Fluorescein Angiography on Full-Field Electroretinography Parameters

    PubMed Central

    Azarmina, Mohsen; Moradian, Siamak; Azarmina, Hossein

    2012-01-01

    Purpose To investigate the effect of simultaneously performed fluorescein angiography (FA) on full-field electroretinography (ffERG) parameters. Method Scotopic and photopic ffERG were performed immediately and 60 minutes after conventional FA in patients with retinal photoreceptor disorders; a-and b-wave amplitudes were compared between recordings obtained at the two time intervals in each patient. Results Ten eyes of five (3 male and 2 female) patients with mean age of 19.6±3.8 (range, 15-25) years were studied. Intravenous fluorescein administration caused an immediate reduction in ERG waves which was most prominent in rod and maximal combined responses. Mean a-wave amplitude in maximal combined response, rod response and cone response ERGs was 46.0±18.8, 8.0±7.0 and 5.1±2.0 mv immediately after FA which was increased to 79.0±30.0, 21.5± 22.5 and 6.5±2.4 mv 60 minutes afterwards, respectively (P<0.005 for all comparisons). Mean b-wave amplitude in the same order was 91.0±17.5, 47.7±17.2 and 17.3±14.7mv which was increased to 145.0±54.3, 91.8±48.1 and 20.0±17.7 mv respectively, 60 minutes after FA (P<0.005 for all comparisons). Conclusion The amplitude of ERG a- and b-waves under scotopic and photopic conditions increased significantly one hour after FA. These changes may be explained by disappearance of phototoxic and bleaching effects of strong light exposure from the light source of the angiography machine and fluorescein molecule on retinal photoreceptors. PMID:23503687

  10. Classification of Human Retinal Microaneurysms Using Adaptive Optics Scanning Light Ophthalmoscope Fluorescein Angiography

    PubMed Central

    Dubow, Michael; Pinhas, Alexander; Shah, Nishit; Cooper, Robert F.; Gan, Alexander; Gentile, Ronald C.; Hendrix, Vernon; Sulai, Yusufu N.; Carroll, Joseph; Chui, Toco Y. P.; Walsh, Joseph B.; Weitz, Rishard; Dubra, Alfredo; Rosen, Richard B.

    2014-01-01

    Purpose. Microaneurysms (MAs) are considered a hallmark of retinal vascular disease, yet what little is known about them is mostly based upon histology, not clinical observation. Here, we use the recently developed adaptive optics scanning light ophthalmoscope (AOSLO) fluorescein angiography (FA) to image human MAs in vivo and to expand on previously described MA morphologic classification schemes. Methods. Patients with vascular retinopathies (diabetic, hypertensive, and branch and central retinal vein occlusion) were imaged with reflectance AOSLO and AOSLO FA. Ninety-three MAs, from 14 eyes, were imaged and classified according to appearance into six morphologic groups: focal bulge, saccular, fusiform, mixed, pedunculated, and irregular. The MA perimeter, area, and feret maximum and minimum were correlated to morphology and retinal pathology. Select MAs were imaged longitudinally in two eyes. Results. Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging revealed microscopic features of MAs not appreciated on conventional images. Saccular MAs were most prevalent (47%). No association was found between the type of retinal pathology and MA morphology (P = 0.44). Pedunculated and irregular MAs were among the largest MAs with average areas of 4188 and 4116 μm2, respectively. Focal hypofluorescent regions were noted in 30% of MAs and were more likely to be associated with larger MAs (3086 vs. 1448 μm2, P = 0.0001). Conclusions. Retinal MAs can be classified in vivo into six different morphologic types, according to the geometry of their two-dimensional (2D) en face view. Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging of MAs offers the possibility of studying microvascular change on a histologic scale, which may help our understanding of disease progression and treatment response. PMID:24425852

  11. Indicating pressure and environmental effects by means of the spectral shift with rhodamine B and fluorescein

    NASA Astrophysics Data System (ADS)

    Johann, R. M.

    2015-07-01

    Fluorescence absorption and emission wavelengths can be influenced by environmental conditions, such as pressure, temperature and concentration. Here those effects are explored with an emphasis on determining the potential of rhodamine B and fluorescein as high-pressure indicators. The red shift of the emission peak maxima of rhodamine B and fluorescein are investigated in dependence of pressure up to 200 MPa using as the solvents water, ethanol and poly(dimethylsiloxane) (PDMS) with rhodamine B and water, polystyrene beads and melamine resin beads with fluorescein. Emission spectra recording and peak fitting is done automatically at time intervals of down to a second and with 0.3 nm wavelength resolution. The wavenumber-pressure relation for rhodamine B reveals increasing divergence from linear behavior in the sequence of the solvents water, ethanol and silicone rubber. Graphical correlation of the data diverging only slightly from linearity with a selection of polarity functions is enabled using the concept of `deviation from linearity (DL)' plots. Using the example of rhodamine B dissolved in PDMS elastomer it is shown that there is a temperature induced irreversible molecular reordering, when scanning between 3 and ˜50°C, and a polarity change in the proximity of the embedded dye molecule. Swelling studies are performed with PDMS containing rhodamine B, where the elastomer is first put in water, then in ethanol and again in water. There a complex solvent exchange process is revealed in the elastomer demonstrating the feasibility of fluorescence spectroscopy, when observing variations in wavelength, to indicate and enlighten molecular rearrangements and swelling dynamics in the polymer, and polarity changes and solvent exchange processes in the dye solvation shell.

  12. Comparison of Special Stains for Keratin with Routine Hematoxylin and Eosin Stain

    PubMed Central

    Rao, Roopa S; Patil, Shankargouda; Majumdar, Barnali; Oswal, Rakesh G

    2015-01-01

    Background: Keratins are the most abundant proteins and are characteristic findings in many epithelial pathologies, making it a diagnostically important marker, both histopathologically and immunohistochemically. Since, immunohistochemistry is an expensive diagnostic tool, special stains to detect the degree of keratinization could serve as a faster and economic option. The aim of the present study was to compare the efficacy of special stains for keratin with standard hematoxylin and eosin stain (H and E). Objectives include: (i) To subject the diagnosed cases of keratin disorders to the selected special stains: Ayoub-shklar method, Dane-Herman method, Alcian blue –periodic acid Schiff ’s (PAS), rapid papanicolaou (PAP) and Gram’s stain. (ii) To compare the staining specificity and staining intensity of special stains with respect to routine hematoxylin and eosin (H and E) stain. (iii) To compare the efficacy of special stains to routine H and E stain in identification of the type of keratin present in the selected cases. Materials and Methods: A total of 80 cases of known pathology for keratin were retrieved from the department archive, which included 10 each of normal gingiva, hyperkeratosis, squamous papilloma, verrucous hyperplasia, verrucous carcinoma, well-differentiated squamous cell carcinoma, orthokeratinized odontogenic cyst and keratocystic odontogenic tumors. Six sections of 4 µ each from the paraffin blocks were made, stained with H and E and the special stains and these were evaluated by 2 pathologists based on the modified scoring criteria from Rahma Al-Maaini and Philip Bryant 2008. Results: The results were tabulated using Chi square and kappa statistics. The statistical values for identification of the type of keratinization was insignificant showing that ortho and parakeratinized epithelia could be correctly identified by both H and E as well as all the special stains. Furthermore, all the special stains showed a positive result and statistical significance (P < 0.001) with respect to the staining of keratin. Conclusion: To conclude, though the special stains distinctly stained the keratin with a higher intensity, H and E proves to be overall better stain with respect to specificity PMID:25878469

  13. Cochlear Microdialysis for Quantification of Dexamethasone and Fluorescein Entry into Scala Tympani During Round Window Administration

    PubMed Central

    Hahn, Hartmut; Kammerer, Bernd; DiMauro, Andre; Salt, Alec N.; Plontke, Stefan K.

    2006-01-01

    Before new drugs for the treatment of inner ear disorders can be studied in controlled clinical trials, it is important that their pharmacokinetics be established in inner ear fluids. Microdialysis allows drug levels to be measured in perilymph without the volume disturbances and potential cerebrospinal fluid contamination associated with fluid sampling. The aims of this study were to show: (i) that despite low recovery rates from miniature dialysis probes, significant amounts of drug are removed from small fluid compartments, (ii) that dialysis sampling artifacts can be accounted for using computer simulations and (iii) that microdialysis allows quantification of the entry rates through the round window membrane (RWM) into scala tympani (ST). Initial experiments used microdialysis probes in small compartments in vitro containing sodium fluorescein. Stable concentrations were observed in large compartments (1000 μl) but significant concentration declines were observed in smaller compartments (100, 10 and 5.6 μl) comparable to the size of the inner ear. Computer simulations of these experiments closely approximated the experimental data. In in vivo experiments, sodium fluorescein 10 mg/ml and dexamethasone-dihydrogen-phosphate disodium salt 8 mg/ml were simultaneously applied to the RWM of guinea pigs. Perilymph concentration in the basal turn of ST was monitored using microdialysis. The fluorescein concentration reached after 200 min application (585 ± 527 μg/ml) was approximately twice that of dexamethasone phosphate (291 ± 369 μg/ml). Substantial variation in concentrations was found between animals by approximately a factor of 34 for fluorescein and at least 41 for dexamethasone phosphate. This is, to a large extent, thought to be the result of the RWM permeability varying in different animals. It was not caused by substance analysis variations, because two different analytic methods were used and the concentration ratio between the two substances remained nearly constant across the experiments and because differences were apparent for the repeated samples obtained in each animal. Interpretation of the results using computer simulations allowed RWM permeability to be quantified. It also demonstrated, however, that cochlear clearance values could not be reliably obtained with microdialysis because of the significant contribution of dialysis to clearance. The observed interanimal variation, e.g., in RWM permeability, is likely to be clinically relevant to the local application of drugs in patients. PMID:16442251

  14. Gram staining apparatus for space station applications

    NASA Technical Reports Server (NTRS)

    Molina, T. C.; Brown, H. D.; Irbe, R. M.; Pierson, D. L.

    1990-01-01

    A self-contained, portable Gram staining apparatus (GSA) has been developed for use in the microgravity environment on board the Space Station Freedom. Accuracy and reproducibility of this apparatus compared with the conventional Gram staining method were evaluated by using gram-negative and gram-positive controls and different species of bacteria grown in pure cultures. A subsequent study was designed to assess the performance of the GSA with actual specimens. A set of 60 human and environmental specimens was evaluated with the GSA and the conventional Gram staining procedure. Data obtained from these studies indicated that the GSA will provide the Gram staining capability needed for the microgravity environment of space.

  15. Silver staining of 2D electrophoresis gels.

    PubMed

    Rabilloud, Thierry

    2012-01-01

    Silver staining is used to detect proteins after electrophoretic separation on polyacrylamide gels. It -combines excellent sensitivity (in the low nanogram range) with the use of very simple and cheap equipment and chemicals. For its use in proteomics, two important additional features must be considered, compatibility with mass spectrometry and quantitative response. Both features are discussed in this chapter, and optimized silver staining protocols are proposed. PMID:22665294

  16. New Grocott Stain without Using Chromic Acid.

    PubMed

    Shiogama, Kazuya; Kitazawa, Kayo; Mizutani, Yasuyoshi; Onouchi, Takanori; Inada, Ken-Ichi; Tsutsumi, Yutaka

    2015-01-01

    We established a new "ecological" Grocott stain for demonstrating fungi, based upon a 4R principle of refusal, reduction, reuse, and recycle of waste management. Conventional Grocott stain employs environmentally harsh 5% chromic acid for oxidization. Initially, we succeeded in reducing the concentration of chromic acid from 5% to 1% by incubating the solution at 60°C and using five-fold diluted chromic acid solution at which point it was reusable. Eventually, we reached the refusal level where 1% periodic acid oxidization was efficient enough, when combined with preheating of sections in the electric jar, microwave oven, or pressure pan. For convenience sake, we recommend pressure pan heating in tap water for 10 min. Stainability of fungi in candidiasis and aspergillosis was comparable with conventional Grocott stain, while Mucor hyphae showed enhanced staining. The modified sequence was further applicable to detecting a variety of mycotic pathogens in paraffin sections. Our environmentally-friendly Grocott stain also has the advantage of avoiding risk of human exposure to hexavalent chromium solution in the histopathology laboratory. The simple stain sequence is can be easily applied worldwide. PMID:25861133

  17. Tailoring of optical properties of fluorescein using green synthesized gold nanoparticles.

    PubMed

    John, Jisha; Thomas, Lincy; George, Nibu A; Kurian, Achamma; George, Sajan D

    2015-06-28

    Dye-nanoparticle mixtures hold great promise in biological as well as photonics applications due to their capability to tailor the emission behavior of dye by tuning the nanoparticles parameters. However, as compared to the well-defined dye-nanoparticle distance, studies lack the understanding of homogenous mixtures of dye and nanoparticles. In this work, we investigate the influence of shape and concentration of gold nanoparticles prepared via green synthesis on the optical properties of fluorescein dye in a dye-nanoparticle mixture. We have investigated the radiative path of deexcitation using steady state fluorescence and the non-radiative path is probed using a laser based dual-beam thermal lens technique. The energy transfer efficiency as well as dye-nanoparticle distance is studied using both techniques. Furthermore, we have explored the influence of nanoparticles parameters on the fluorescence quantum yield of fluorescein using the thermal lens technique. The studies indicate that spherical nanoparticles are efficient quenchers while star shaped nanoparticles can probe larger dye-NP distances. The tailoring of dye properties by tuning nanoparticle parameters can be utilized in diverse areas including bioimaging, solar cells, and sensors. PMID:26017461

  18. Wide-Field Fluorescein Angiography in Wet Age-Related Macular Degeneration

    PubMed Central

    Beare, Nicholas

    2014-01-01

    Purpose. The aim of our study was to investigate if peripheral retinal ischaemia contributed to the pathogenesis of neovascular AMD (NvAMD), using wide-field fluorescein angiography (WFFA). Methods. This prospective study included 30 consecutive patients with newly diagnosed NvAMD in the index eye. Wide-field colour fundus images and fluorescein angiograms were obtained using P200C optomap FA and analysed using a grid with three concentric circles of 50°, 100°, and 200° centred on the fovea to define zones Z1, Z2, and Z3. Results. Areas of peripheral retinal nonperfusion were seen in 2 (7%) eyes, peripheral vascular leakage in 5 (17%) eyes, and diffuse dye leakage close to the ora in 5 (17%) eyes. A total of one-third of the study eyes showed changes on WFFA in Z2 and Z3. On comparing index eyes to nonindex eyes in these patients, the presence of NvAMD was associated with peripheral FA changes (P = 0.009, Fisher's test). Conclusion. Frank peripheral retinal non-perfusion does not appear to be associated with NvAMD. In some patients with active NvAMD there is degradation of the peripheral blood-retina barrier. Smoking was also found to be associated with the above-mentioned abnormalities. PMID:25379537

  19. Membrane carbohydrate characterization of Acanthamoeba astronyxis, A. castellanii and Naegleria fowleri by fluorescein-conjugated lectins.

    PubMed

    Bose, K; Ghosh, D K; Bhattacharya, A

    1989-11-01

    A comparative study of membrane carbohydrate characteristics of pathogenic and non-pathogenic trophozoites and cysts of free-living Acanthamoeba castellanii, Naegleria fowleri and A. astronyxis, respectively from sewage sludge in India was carried out by means of fluorescein-conjugated lectin binding using eight lectins. Two lectins, viz. Concanavalin A and Phytohaemagglutinin P, could bind all free-living amoebae at different concentrations. The most notable feature of the study is that peanut agglutinin (PNA) and wheatgerm agglutinin (WGA) can differentiate between the pathogenic A. castellanii and non-pathogenic A. astronyxis strain, respectively. However, Ulex agglutinin I (UEA I) was the only lectin positive to both pathogenic A. castellanii and N. fowleri. During in vitro conversion from trophozoites to cysts, A. castellanii and N. fowleri cysts gained WGA-specific saccharide whereas A. castellanii; A. astronyxis and N. fowleri lost or reduced Dolichos biflorus agglutinin, PNA; WGA and ConA, and UEA I-specific saccharides, respectively. Neuraminidase could not alter the fluorescein-lectin binding to WGA and PNA. These demonstrated that only two lectins can recognize the factors giving Acanthamoeba their pathogenic (PNA-specific) and non-pathogenic (WGA-specific) status. More interestingly, UEA I can only differentiate between pathogenic and non-pathogenic amoebae. It is also suggested that during stage conversion the surface of the organism exhibited replacement of saccharides. PMID:2592141

  20. Digital stain separation for histological images.

    PubMed

    Tadrous, P J

    2010-11-01

    It is often desirable to perform digital image analyses on sections prepared for human interpretation, e.g. nuclear chromatin texture analysis or three-dimensional reconstructions using sections requiring human delineation of structures of interest. Unfortunately such analyses are often more effective using stains with less complex contrast. Here an automated selective 'de-staining' method for digital images is presented. The method separates an image into its red, green and blue and hue, saturation and intensity components. A mask of stained tissue is prepared by automatic percentile thresholding. A single weighted inverted colour channel is then added to each of the three primary colour channels separately by an iterative algorithm that adjusts the weights to give minimum variance within the mask. The modified red, green and blue channels are then recombined. This method is automatic requiring no pre-definition of stain colours or special hardware. The method is demonstrated to 'de-stain' nuclei in haematoxylin and eosin (H&E) sections (and a separate haematoxylin image can be derived from this). An image of isolated brown reaction product is produced with immunoperoxidase preparations counterstained with haematoxylin. Furthermore trichrome (haematoxylin van Gieson, picrosirius red) and other common stains may be separated into their components with modifications of the same algorithm. Although other methods for colour separation do exist (e.g. spectral pathology and colour deconvolution) these require special apparatus or precise calibration and foreknowledge of pure dye colour spectra. The present method of digital stain separation is fully automatic with no such prerequisites. PMID:20946383

  1. A dual-mode signaling response of a AuNP-fluorescein based probe for specific detection of thiourea.

    PubMed

    Chen, Chuanxia; Zhao, Dan; Sun, Jian; Yang, Xiurong

    2016-04-21

    By employing fluorescein and AuNPs as energy donors and acceptors, respectively, a novel fluorescence resonance energy transfer (FRET)-based dual-mode sensor for selective recognition and quantitative detection of thiourea was designed and constructed in this study for the first time. Herein, it is demonstrated that fluorescein could be adsorbed on the surface of AuNPs and induce fluorescence quenching through the well-known FRET process. The preferential introduction of thiourea would reduce the overall surface negative charges of AuNPs by replacing the original citrate groups, leading to the aggregation of AuNPs. Meanwhile, thiourea could prevent the binding between fluorescein and AuNPs, reduce the as-formed FRET effect, and then lead to fluorescence recovery of the fluorescein. Therefore, a dual-mode sensor with AuNP-related colorimetric and fluorescein-based fluorescence readout was rationally developed. Under the optimum conditions, the detection limits were calculated to be 10 nM and 23 nM for fluorescent and colorimetric sensors, individually, and a limit of 0.4 μM was detected by the naked eye. Finally, such a simple, convenient, cost-effective, highly selective and sensitive sensing assay was successfully applied in the detection of thiourea in tap water and fruit juice samples. PMID:27031921

  2. Compositions for chromosome-specific staining

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    1998-01-01

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods are provided to disable the hybridization capacity of shared, high copy repetitive sequences and/or remove such sequences to provide for useful contrast. Still further methods are provided to produce chromosome-specific staining reagents which are made specific to the targeted chromosomal material, which can be one or more whole chromosomes, one or more regions on one or more chromosomes, subsets of chromosomes and/or the entire genome. Probes and test kits are provided for use in tumor cytogenetics, in the detection of disease related loci, in analysis of structural abnormalities, such as translocations, and for biological dosimetry. Further, methods and prenatal test kits are provided to stain targeted chromosomal material of fetal cells, including fetal cells obtained from maternal blood. Still further, the invention provides for automated means to detect and analyse chromosomal abnormalities.

  3. Compositions for chromosome-specific staining

    DOEpatents

    Gray, J.W.; Pinkel, D.

    1998-05-26

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. The methods produce staining patterns that can be tailored for specific cytogenetic analyses. The probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods are provided to disable the hybridization capacity of shared, high copy repetitive sequences and/or remove such sequences to provide for useful contrast. Still further methods are provided to produce chromosome-specific staining reagents which are made specific to the targeted chromosomal material, which can be one or more whole chromosomes, one or more regions on one or more chromosomes, subsets of chromosomes and/or the entire genome. Probes and test kits are provided for use in tumor cytogenetics, in the detection of disease related loci, in analysis of structural abnormalities, such as translocations, and for biological dosimetry. Methods and prenatal test kits are provided to stain targeted chromosomal material of fetal cells, including fetal cells obtained from maternal blood. The invention provides for automated means to detect and analyze chromosomal abnormalities. 17 figs.

  4. Effects of Hoechst33342 staining on the viability and flow cytometric sex-sorting of frozen-thawed ram sperm.

    PubMed

    Quan, Guo Bo; Ma, Yuan; Li, Jian; Wu, Guo Quan; Li, Dong Jiang; Ni, Yi Na; Lv, Chun Rong; Zhu, Lan; Hong, Qiong Hua

    2015-02-01

    Cytometric sorting of frozen-thawed sperm can overcome difficulties caused by the unavailability of sorting facilities on farms where semen is collected from male livestock. In order to optimize the cytometric sex-sorting procedure, effects of Hoechst33342 staining on the viability and cytometric sorting efficiency of frozen-thawed ram sperm were evaluated. The frozen-thawed sperm were stained with Hoechst33342 at various dye concentrations (80 μM, 120 μM, 160 μM, 200 μM, 240 μM, or 320 μM) for 45 min to evaluate effects of dye dose. The frozen-thawed sperm were stained with 160 μM Hoechst33342 for various durations (0 min, 15 min, 30 min, 45 min, 60 min, 75 min, or 90 min) to evaluate effects of staining duration. Sperm motility and moving velocity were analyzed using a computer-assisted sperm analysis system (CASAS). Acrosome status, membrane integrity, and distribution of phosphatidylserine (PS) in Hoechst33342-stained sperm were analyzed using flow cytometry after staining with fluorescein isothiocyanate-labeled lectin from pisum sativum (FITC-PSA), Annexin V, or propidium iodide (PI). The fertility of Hoechst33342-stained sperm was analyzed by in vitro fertilization (IVF). A high-speed cell sorter was used to evaluate effects of Hoechst33342 staining on cytometric sex-sorting of frozen-thawed sperm. The motility, moving velocity, membrane integrity, and PS distribution of Hoechst33342-stained sperm were significantly different from that of immediately thawed sperm (P<0.05). However, there is no significant difference existing among the Hoechst33342-stained groups with respect to the above evaluated parameters. Additionally, along with the staining durations, the adverse effects of the staining procedure on sperm showed a steady increase. However, Hoechst33342 staining did not damage acrosome and in vitro fertilizing capability of frozen-thawed ram sperm. Results of cytometric sorting indicated that frozen-thawed sperm can be efficiently sorted into two sperm populations with X and Y chromosome when the Hoechst33342 concentration was 160 μM. Moreover, when the staining duration was equal to or longer than 45 min, the frozen-thawed sperm can be successfully sorted in the presence of 160μM Hoechst33342. In conclusion, Hoechst33342 staining can detrimentally influence viability of frozen-thawed ram sperm except acrosome and in vitro fertilizing capability. Accordingly, the minimum values of Hoechst33342 concentration and staining duration can be set at 160 μM and 45 min respectively. However, the maximum values of Hoechst33342 concentration and staining duration were not determined based on the current study. Further research on how to reduce injuries caused by freezing, thawing, and Hoechst33342 staining on frozen-thawed ram sperm is needed. PMID:25481668

  5. Rapid coomassie blue staining of protein gels.

    PubMed

    Simpson, Richard J

    2010-04-01

    Coomassie brilliant blue R250 (CBR-250) and silver staining are the most widely used methods for the routine visualization of proteins separated by SDS-PAGE. CBR-250 is an organic dye that complexes with basic amino acids, such as arginine, lysine, and histidine, as well as tyrosine. Conventional CBR-250 staining is capable of detecting as little as 30-100 ng of protein, but sensitivity can be improved by performing the staining and destaining at elevated temperatures. The method described in this protocol, which is a modified version of the conventional Coomassie protocol, speeds up the destaining process for faster results with increased sensitivity and is compatible with mass-spectrometry-based methods for identifying proteins. PMID:20360367

  6. Effect of alumina on triethylene glycol diacetate-2-propenoic acid butyl ester composite polymer electrolytes for flexible lithium ion batteries

    NASA Astrophysics Data System (ADS)

    Wang, Qiujun; Song, Wei-Li; Fan, Li-Zhen; Shi, Qiao

    2015-04-01

    Triethylene glycol diacetate-2-propenoic acid butyl ester (TEGDA-BA) based composite polymer electrolytes (CPE) are fabricated by incorporating alumina (Al2O3) nanoparticles (average particle size 10-20 nm) as inorganic filler via in situ polymerization. Effects of Al2O3 concentration on ionic conductivities, Li+ transfer numbers and charge/discharge properties are studied in details. Due to the uniformly dispersed Al2O3 nanoparticles, significant improvements in the mechanical flexibility and bendability are presented in the resulting polymer electrolytes. The CPE with 5 wt% Al2O3 nanoparticles exhibits the highest ionic conductivity up to 6.02 × 10-3 S cm-1 at 25 °C and the highest Li+ transference number (0.675), coupled with the most stable electrochemical window (>4.5 V vs. Li/Li+). With the presence of Al2O3, the growth of interface resistance is retarded, which increases the interface stability. The Li|CPE|Li4Ti5O12 and Li|CPE|LiFePO4 cells demonstrate remarkably stable charge/discharge performance and excellent capacity retention during cycling test. The results suggest that the CPE holds great application potential in flexible lithium ion batteries.

  7. PREDICTIVE MODEL FOR THE COMBINED EFFECT OF TEMPERATURE, SODIUM LACTATE, AND SODIUM DIACETATE ON THE HEAT RESISTANCE OF LISTERIA MONOCYTOGENES IN BEEF

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The effects and interactions of heating temperature (60 - 73.9C), sodium lactate (NaL; 0.0 - 4.8%, w/w) and/or sodium diacetate SDA; 0.0 - 0.25%, w/w) on the heat resistance of a five-strain mixture of Listeria monocytogenes in 75% lean ground beef were examined. Thermal death times were determined...

  8. Automated single-slide staining device

    NASA Technical Reports Server (NTRS)

    Wilkins, J. R.; Mills, S. M. (Inventor)

    1977-01-01

    A simple apparatus and method is disclosed for making individual single Gram stains on bacteria inoculated slides to assist in classifying bacteria in the laboratory as Gram-positive or Gram-negative. The apparatus involves positioning a single inoculated slide in a stationary position and thereafter automatically and sequentially flooding the slide with increments of a primary stain, a mordant, a decolorizer, a counterstain and a wash solution in a sequential manner without the individual lab technician touching the slide and with minimum danger of contamination thereof from other slides.

  9. Detection Of Concrete Deterioration By Staining

    DOEpatents

    Guthrie, Jr., George D.; Carey, J. William

    1999-09-21

    A method using concentrated aqueous solutions of sodium cobaltinitrite and a rhodamine dye is described which can be used to identify concrete that contains gels formed by the alkali-silica reaction (ASR), and to identify degraded concrete which results in a porous or semi-permeable paste due to carbonation or leaching. These solutions present little health or environmental risk, are readily applied, and rapidly discriminate between two chemically distinct gels; K-rich, Na--K--Ca--Si gels are identified by yellow staining, and alkali-poor, Ca--Si gels are identified by pink staining.

  10. Laser treatment of port-wine stains.

    PubMed

    Brightman, Lori A; Geronemus, Roy G; Reddy, Kavitha K

    2015-01-01

    Port-wine stains are a type of capillary malformation affecting 0.3% to 0.5% of the population. Port-wine stains present at birth as pink to erythematous patches on the skin and/or mucosa. Without treatment, the patches typically darken with age and may eventually develop nodular thickening or associated pyogenic granuloma. Laser and light treatments provide improvement through selective destruction of vasculature. A variety of vascular-selective lasers may be employed, with the pulsed dye laser being the most common and well studied. Early treatment produces more optimal results. Advances in imaging and laser treatment technologies demonstrate potential to further improve clinical outcomes. PMID:25624768

  11. Crystalline lens capsule staining with trypan blue.

    PubMed

    Laureano, Joseph San; Coroneo, Minas T

    2004-10-01

    A 1-step method for staining the anterior lens capsule with trypan blue is described. The dye is instilled via a paracentesis port at the start of cataract extraction. As aqueous humor is allowed to exit the anterior chamber (AC), which consequently shallows, the resulting pupil block confines the dye to the AC. An ophthalmic viscosurgical device (OVD) is used to flush dye-stained aqueous from the AC, circumventing the need for AC washout. Although the OVD may be tinged with dye, this does not impede performing capsulorhexis. This method does not add to the surgical time, requires no additional instruments or materials, and is safe. PMID:15474812

  12. Laser treatment of port-wine stains

    PubMed Central

    Brightman, Lori A; Geronemus, Roy G; Reddy, Kavitha K

    2015-01-01

    Port-wine stains are a type of capillary malformation affecting 0.3% to 0.5% of the population. Port-wine stains present at birth as pink to erythematous patches on the skin and/or mucosa. Without treatment, the patches typically darken with age and may eventually develop nodular thickening or associated pyogenic granuloma. Laser and light treatments provide improvement through selective destruction of vasculature. A variety of vascular-selective lasers may be employed, with the pulsed dye laser being the most common and well studied. Early treatment produces more optimal results. Advances in imaging and laser treatment technologies demonstrate potential to further improve clinical outcomes. PMID:25624768

  13. Immunofluorescence staining of paraffin sections: creating DAB staining like virtual digital images using CMYK color conversion.

    PubMed

    Buchynska, L; Kashuba, E; Szekely, L

    2008-12-01

    Crystal violet treatment of formalin fixed paraffin embedded tissue slides greatly reduces the endogenous autofluorescence, and allows immunofluorescence (IF) staining with FITC or Alexa488 conjugated antibodies. Using cold CCD camera to capture the fluorescence images makes this staining method very sensitive. Here we show that combination of IF with the simultaneous recording of crystal violet induced red and Hoechst 33258 induced blue fluorescence permits the localization of the IF signal over a cytoplasmic: nuclear red:blue stain that visualizes the microscopic anatomy of the underlying tissue. To make the visual interpretation of the IF staining easier for microscopists, who are used to DAB staining over weak hematoxilin-eosin background, we created a simple color conversion procedure that turns the captured three-color fluorescence RGB (red, green, blue) images over a black background into four color CMYK (cyan, magenta, yellow, key color (black)) images. PMID:19112433

  14. Automated Detection of Leakage in Fluorescein Angiography Images with Application to Malarial Retinopathy

    PubMed Central

    Zhao, Yitian; J. C. MacCormick, Ian; G. Parry, David; Leach, Sophie; A. V. Beare, Nicholas; P. Harding, Simon; Zheng, Yalin

    2015-01-01

    The detection and assessment of leakage in retinal fluorescein angiogram images is important for the management of a wide range of retinal diseases. We have developed a framework that can automatically detect three types of leakage (large focal, punctate focal, and vessel segment leakage) and validated it on images from patients with malarial retinopathy. This framework comprises three steps: vessel segmentation, saliency feature generation and leakage detection. We tested the effectiveness of this framework by applying it to images from 20 patients with large focal leak, 10 patients with punctate focal leak, and 5,846 vessel segments from 10 patients with vessel leakage. The sensitivity in detecting large focal, punctate focal and vessel segment leakage are 95%, 82% and 81%, respectively, when compared to manual annotation by expert human observers. Our framework has the potential to become a powerful new tool for studying malarial retinopathy, and other conditions involving retinal leakage. PMID:26030010

  15. Early events in herpes simplex virus type 1 infection: photosensitivity of fluorescein isothiocyanate-treated virions

    SciTech Connect

    DeLuca, N.; Bzik, D.; Person, S.; Snipes, W.

    1981-02-01

    Herpes simplex virus type 1 is photosensitized by treatment with fluorescein isothiocyanate (FITC). The inactivation of FITC-treated virions upon subsequent exposure to light is inhibited by the presence of sodium azide, suggesting the involvement of singlet oxygen in the process. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis revealed that treatment with FITC plus light induces crosslinks in viral envelope glycoproteins. Treatment of virions with high concentrations of FITC (50 ..mu..g/ml) plus light causes a reduction in the adsorption of the virus to monolayers of human embryonic lung cells. For lower concentrations of FITC (10 ..mu..g/ml) plus light, treated virions adsorb to the host cells, but remain sensitive to light until entry occurs. The loss of light sensitivity coincides with the development of resistance to antibodies. These results are most consistent with a mechanism of entry for herpes simplex virus involving fusion of the viral membrane with the plasma membrane of the host cell.

  16. Fully Automatic Segmentation of Fluorescein Leakage in Subjects With Diabetic Macular Edema

    PubMed Central

    Rabbani, Hossein; Allingham, Michael J.; Mettu, Priyatham S.; Cousins, Scott W.; Farsiu, Sina

    2015-01-01

    Purpose. To create and validate software to automatically segment leakage area in real-world clinical fluorescein angiography (FA) images of subjects with diabetic macular edema (DME). Methods. Fluorescein angiography images obtained from 24 eyes of 24 subjects with DME were retrospectively analyzed. Both video and still-frame images were obtained using a Heidelberg Spectralis 6-mode HRA/OCT unit. We aligned early and late FA frames in the video by a two-step nonrigid registration method. To remove background artifacts, we subtracted early and late FA frames. Finally, after postprocessing steps, including detection and inpainting of the vessels, a robust active contour method was utilized to obtain leakage area in a 1500-μm-radius circular region centered at the fovea. Images were captured at different fields of view (FOVs) and were often contaminated with outliers, as is the case in real-world clinical imaging. Our algorithm was applied to these images with no manual input. Separately, all images were manually segmented by two retina specialists. The sensitivity, specificity, and accuracy of manual interobserver, manual intraobserver, and automatic methods were calculated. Results. The mean accuracy was 0.86 ± 0.08 for automatic versus manual, 0.83 ± 0.16 for manual interobserver, and 0.90 ± 0.08 for manual intraobserver segmentation methods. Conclusions. Our fully automated algorithm can reproducibly and accurately quantify the area of leakage of clinical-grade FA video and is congruent with expert manual segmentation. The performance was reliable for different DME subtypes. This approach has the potential to reduce time and labor costs and may yield objective and reproducible quantitative measurements of DME imaging biomarkers. PMID:25634978

  17. Validation of an automated fluorescein method for determining bromide in water

    USGS Publications Warehouse

    Fishman, M. J.; Schroder, L.J.; Friedman, L.C.

    1985-01-01

    Surface, atmospheric precipitation and deionized water samples were spiked with ??g l-1 concentrations of bromide, and the solutions stored in polyethylene and polytetrafluoroethylene bottles. Bromide was determined periodically for 30 days. Automated fluorescein and ion chromatography methods were used to determine bromide in these prepared samples. Analysis of the data by the paired t-test indicates that the two methods are not significantly different at a probability of 95% for samples containing from 0.015 to 0.5 mg l-1 of bromide. The correlation coefficient for the same sets of paired data is 0.9987. Recovery data, except for the surface water samples to which 0.005 mg l-1 of bromide was added, range from 89 to 112%. There appears to be no loss of bromide from solution in either type of container.Surface, atmospheric precipitation and deionized water samples were spiked with mu g l** minus **1 concentrations of bromide, and the solutions stored in polyethylene and polytetrafluoroethylene bottles. Bromide was determined periodically for 30 days. Automated fluorescein and ion chromatography methods were used to determine bromide in these prepared samples. Analysis of the data by the paired t-test indicates that the two methods are not significantly different at a probability of 95% for samples containing from 0. 015 to 0. 5 mg l** minus **1 of bromide. The correlation coefficient for the same sets of paired data is 0. 9987. Recovery data, except for the surface water samples to which 0. 005 mg l** minus **1 of bromide was added, range from 89 to 112%. Refs.

  18. Vascular and avascular retinae in mammals. A funduscopic and fluorescein angiographic study.

    PubMed

    Buttery, R G; Haight, J R; Bell, K

    1990-01-01

    Intraretinal blood vessels are present in some and absent in other vertebrate species, including the mammals. Among the marsupials, both vascular and avascular retinae are seen. We determined the funduscopic appearance of the eye, investigated the functional aspects of ocular blood flow in both types of retina in marsupials and compared our results with known patterns in placental mammals. The Australian polyprotodont marsupials, the Tasmanian devil, Sarcophilus harrisii, and the quoll, Dasyurus viverrinus, together with an American polyprotodont, the Virginia opossum, Didelphis virginiana, demonstrate variable degrees of tapetal differentiation, pigmentation and a very close parallel course of their intraretinal arteries and veins over considerable distances. Using the technique of fluorescein angiography, we found that retinal blood flow in the 3 vascular Australian species commenced with arterial filling. Early venous was seen next, followed by the capillary blush. This unusual sequence of vascular flow differs from that of the arterial-capillary-venous filling seen in placental mammals. This difference is most likely a consequence of the known looped, end artery organisation found within marsupial nervous systems, of which the retinae are a part. The 2 diprotodont marsupials examined, the brushtail possum, Trichosurus vulpecula, and the sugar glider, Petaurus breviceps, possess avascular retinae. Only a small residual tuft of fluorescein-impermeable vessels projects from the optic disc into the vitreous. Interestingly, the structural complexity of the central visual system in diprotodonts all of whom possess avascular retinae) is commonly accepted as being greater than that of the stem polyprotodont line (which possess vascular retinae). If retinal function matches this internal complexity, then retinal avascularity may, as in birds, be associated with superior vision. However, as the retinae of these mammals clearly lack any nutritive mechanisms directly analogous to those in the retinae of, say, birds or the megachiropteran bats, their retinal nutritive pathways remain enigmatic. PMID:2375974

  19. The Language of Stained-Glass Windows

    ERIC Educational Resources Information Center

    Brew, Charl Anne

    2010-01-01

    The splendor and beauty of stained glass punctuates any room. In this article, the author describes a cross-curriculum project which incorporated the French classes' research and written study of France in the Middle Ages. For the project the author suggested Sainte-Chapelle which is considered a reliquary and was built by Louis IX to house the…

  20. Protein Stains to Detect Antigen on Membranes.

    PubMed

    Dsouza, Anil; Scofield, R Hal

    2015-01-01

    Western blotting (protein blotting/electroblotting) is the gold standard in the analysis of complex protein mixtures. Electroblotting drives protein molecules from a polyacrylamide (or less commonly, of an agarose) gel to the surface of a binding membrane, thereby facilitating an increased availability of the sites with affinity for both general and specific protein reagents. The analysis of these complex protein mixtures is achieved by the detection of specific protein bands on a membrane, which in turn is made possible by the visualization of protein bands either by chemical staining or by reaction with an antibody of a conjugated ligand. Chemical methods employ staining with organic dyes, metal chelates, autoradiography, fluorescent dyes, complexing with silver, or prelabeling with fluorophores. All of these methods have differing sensitivities and quantitative determinations vary significantly. This review will describe the various protein staining methods applied to membranes after western blotting. "Detection" precedes and is a prerequisite to obtaining qualitative and quantitative data on the proteins in a sample, as much as to comparing the protein composition of different samples. "Detection" is often synonymous to staining, i.e., the reversible or irreversible binding by the proteins of a colored organic or inorganic chemical. PMID:26139252

  1. Asbestos identification by dispersion staining microscopy

    SciTech Connect

    Ganotes, J.T.; Tan, H.T.

    1980-01-01

    Asbestos can be detected and identified by an optical microscope procedure known as dispersion staining. This procedure can be carried out with most phase contrast equipped microscopes. The primary application is for material samples. Distinction between tremolite and anthophyllite asbestos requires examination between crossed polarizers.

  2. The Language of Stained-Glass Windows

    ERIC Educational Resources Information Center

    Brew, Charl Anne

    2010-01-01

    The splendor and beauty of stained glass punctuates any room. In this article, the author describes a cross-curriculum project which incorporated the French classes' research and written study of France in the Middle Ages. For the project the author suggested Sainte-Chapelle which is considered a reliquary and was built by Louis IX to house the

  3. Histopathological confirmation of similar intramucosal distribution of fluorescein in both intravenous administration and local mucosal application for probe-based confocal laser endomicroscopy of the normal stomach

    PubMed Central

    Nonaka, Kouichi; Ohata, Ken; Ban, Shinichi; Ichihara, Shin; Takasugi, Rumi; Minato, Yohei; Tashima, Tomoaki; Matsuyama, Yasushi; Takita, Maiko; Matsuhashi, Nobuyuki; Neumann, Helmut

    2015-01-01

    Probe-based confocal laser endomicroscopy (pCLE) is capable of acquiring in vivo magnified cross-section images of the gastric mucosa. Intravenous injection of fluorescein sodium is used for confocal imaging. However, it is still under debate if local administration of the dye to the mucosa is also effective for confocal imaging as it is not yet clear if topical application also reveals the intramucosal distribution of fluorescein. The objective of this study was to evaluate the intramucosal distribution of fluorescein sodium after topical application and to compare the distribution to the conventional intravenous injection used for confocal imaging. pCLE of the stomach uninfected with Helicobacter pylori was performed in a healthy male employing intravenous administration and local mucosal application of fluorescein. The mucosa of the lower gastric body was biopsied 1 min and 5 min after intravenous administration or local mucosal application of fluorescein, and the distribution of fluorescein in the biopsy samples was examined histologically. Green fluorescence was already observed in the cytoplasm of fundic glandular cells in the biopsied deep mucosa 1 min after local mucosal application of fluorescein. It was also observed in the foveolar lumen and inter-foveolar lamina propria, although it was noted at only a few sites. In the tissue biopsied 5 min after the local mucosal application of fluorescein, green fluorescence was more frequently noted in the cytoplasm of fundic glandular cells than in that 1 min after the local mucosal application of fluorescein, although obvious green fluorescence was not identified in the foveolar lumen or inter-foveolar lamina propria. The distribution of intravenously administered fluorescein in the cytoplasm of fundic glandular cells was also clearly observed similarly to that after local mucosal application of fluorescein. Green fluorescence in more cells was observed in many cells 5 min after intravenous administration compared with that after 1 min. The presence of fluorescein in the mucosa was observed within a short time after local mucosal application of fluorescein, suggesting that pCLE images similarly to those after intravenous fluorescein administration can be acquired by local mucosal application of fluorescein. PMID:26677449

  4. Enhancing absorption of fluorescein and LHRH across hindgut epithelia in a marsupial, the common brushtail possum Trichosurus vulpecula.

    TOXLINE Toxicology Bibliographic Information

    Wen JY; McLeod BJ; Tucker IG; Davies NM; Ledger R; Butt AG

    2007-09-01

    We have identified differences in transport properties of intestinal epithelia in the marsupial brushtail possum, compared to eutherian mammals. To determine whether differences in its permeability to hydrophilic compounds also occur, the absorption of sodium fluorescein and luteinizing hormone releasing hormone (LHRH) was assessed in vitro and the ability of chemical enhancers and a metabolic inhibitor to promote their absorption investigated. The apparent permeability of colonic and caecal tissues to fluorescein and LHRH and transepithelial resistance (Rt) in the absence or presence of ethylenediamine tetra-acetic acid (EDTA), sodium deoxycholic acid (SDA), dithiothreitol (DTT), polyacrylic acids (PAA), or the inhibitor bacitracin were determined. The effects of SDA and/or DTT on adherent mucus and the release of lactate dehydrogenase (LDH) were also assessed. In the absence of treatment, both tissues had comparable amounts of adherent mucus, Rt and low permeabilities to fluorescein and LHRH. All chemical enhancers increased fluorescein permeability, but SDA at concentrations >0.5 mM also induced LDH release. DTT alone and in combination with SDA reduced the amount of adherent mucus. Bacitracin inhibited LHRH metabolism and increased LHRH permeability. These data indicate that the possum hindgut epithelium represents a significant barrier to the uptake of hydrophilic compounds, similar to that in eutherians.

  5. Lighting up Protons with MorphFl, a Fluorescein-Morpholine Dyad: An Experiment for the Organic Laboratory

    ERIC Educational Resources Information Center

    Miller, Tyson A.; Spangler, Michael; Burdette, Shawn C.

    2011-01-01

    A two-period organic laboratory experiment that includes fluorescence sensing is presented. The pH-sensitive sensor MorphFl is prepared using a Mannich reaction between a fluorescein derivative and the iminium ion of morpholine. During the first laboratory, students prepare MorphFl. The second session begins with characterizing the sensor using…

  6. Lighting up Protons with MorphFl, a Fluorescein-Morpholine Dyad: An Experiment for the Organic Laboratory

    ERIC Educational Resources Information Center

    Miller, Tyson A.; Spangler, Michael; Burdette, Shawn C.

    2011-01-01

    A two-period organic laboratory experiment that includes fluorescence sensing is presented. The pH-sensitive sensor MorphFl is prepared using a Mannich reaction between a fluorescein derivative and the iminium ion of morpholine. During the first laboratory, students prepare MorphFl. The second session begins with characterizing the sensor using

  7. Improved Whole-Blood-Staining Device

    NASA Technical Reports Server (NTRS)

    Sams, Clarence F.; Crucian, Brian; Paul, Bonnie; Melton, Shannon; Guess, Terry

    2012-01-01

    Dramatic improvements have been made in NASA s Whole Blood Staining Device (WBSD) since it was last described in "Whole-Blood-Staining Device," NASA Tech Briefs, Vol. 23, No. 10 (October 1999), page 64. The new system has a longer shelf life, a simpler and more effective operational procedure, improved interface with instrumentation, and shorter processing time. More specifically, the improvements have targeted bag and locking clip materials, sampling ports, and air pocket prevention. The WBSD stains whole blood collected during spaceflight for subsequent flow cytometric analysis. In short, the main device stains white blood cells by use of monoclonal antibodies conjugated to various fluorochromes, followed by lysing and fixing of the cells by use of a commercial reagent that has been diluted according to NASA safety standards. This system is compact, robust, and does not require electric power, precise mixing, or precise incubation times. Figure 1 depicts the present improved version for staining applications, which is a poly(tetrafluoroethylene) bag with a Luer-lock port and plastic locking clips. An InterLink (or equivalent) intravenous- injection port screws into the Luer-lock port. The inflatable/collapsible nature of the bag facilitates loading and helps to minimize the amount of air trapped in the fully loaded bag. Some additional uses have been identified for the device beyond whole blood staining. The WBSD has been configured for functional assays that require culture of live cells by housing sterile culture media, mitogens, and fixatives prior to use [Figure 2(a)]. Simple injection of whole blood allows cell-stimulation culture to be performed in reduced gravity conditions, and product stabilization prior to storage, while protecting astronauts from liquid biohazardous materials. Also, the improved WBSD has reconstituted powdered injectable antibiotics by mixing them with diluent liquids [Figure 2(b)]. Although such mixing can readily be performed on Earth by shaking in glass vials, it cannot readily be performed this way in outer space without entraining air bubbles. The present device can be preloaded with the powder and diluent(s) in separate compartments. The powder and diluent( s) can be mixed, without introducing air bubbles, by removing the clip(s), then shaking. This use of the device could also be advantageous in terrestrial applications because it maintains the isolation of the constituents until the time of use.

  8. A Bifunctional Converter: Fluorescein Quenching scFv/Fluorogen Activating Protein for Photostability and Improved Signal to Noise in Fluorescence Experiments

    PubMed Central

    2015-01-01

    Monoclonal antibodies are one of the most useful and ubiquitous affinity reagents used in the biological sciences. Immunostaining of fixed and live cells for microscopy or cytometry measurements frequently employs fluorescently labeled antibodies, in particular fluorescein-labeled antibodies. This dye emits light at a wavelength overlapping with cellular autofluorescence, making it difficult to measure antibody binding to proteins of relatively low copy number or in cells of high green autofluorescence. A number of high affinity fluorescein binding antibodies and antibody domains have been developed that quench the dye’s fluorescence. Using a fluorescein-binding recombinant antibody domain genetically fused to a fluorogen activating protein (FAP), we demonstrate a molecular converter capable of binding and quenching fluorescein, while binding and activating a fluorogenic triarylmethane dye. This reagent converts fluorescein conjugates to far-red fluorescent probes, where cellular autofluorescence is low, improving signal-to-background of cell-based antibody binding measurements by ∼7-fold. Microscopy experiments show colocalization of both fluorescein and MG fluorescence. This dual affinity fluorescein-quenching-FAP can also be used to convert fluorescein to the red fluorescing MG fluorogen on biological molecules other than antibodies. PMID:25072845

  9. Methods and compositions for chromosome-specific staining

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    2003-07-22

    Methods and compositions for chromosome-specific staining are provided. Compositions comprise heterogenous mixtures of labeled nucleic acid fragments having substantially complementary base sequences to unique sequence regions of the chromosomal DNA for which their associated staining reagent is specific. Methods include methods for making the chromosome-specific staining compositions of the invention, and methods for applying the staining compositions to chromosomes.

  10. 7 CFR 28.441 - Strict Middling Yellow Stained Color.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Strict Middling Yellow Stained Color. 28.441 Section... CONTAINER REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.441 Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper...

  11. 7 CFR 28.441 - Strict Middling Yellow Stained Color.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 2 2012-01-01 2012-01-01 false Strict Middling Yellow Stained Color. 28.441 Section... CONTAINER REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.441 Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper...

  12. 7 CFR 28.442 - Middling Yellow Stained Color.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 2 2012-01-01 2012-01-01 false Middling Yellow Stained Color. 28.442 Section 28.442... REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.442 Middling Yellow Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper...

  13. 7 CFR 28.442 - Middling Yellow Stained Color.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 2 2011-01-01 2011-01-01 false Middling Yellow Stained Color. 28.442 Section 28.442... REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.442 Middling Yellow Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper...

  14. 7 CFR 28.442 - Middling Yellow Stained Color.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 2 2013-01-01 2013-01-01 false Middling Yellow Stained Color. 28.442 Section 28.442... REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.442 Middling Yellow Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper...

  15. 7 CFR 28.441 - Strict Middling Yellow Stained Color.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 2 2011-01-01 2011-01-01 false Strict Middling Yellow Stained Color. 28.441 Section... CONTAINER REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.441 Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper...

  16. 7 CFR 28.442 - Middling Yellow Stained Color.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Middling Yellow Stained Color. 28.442 Section 28.442... REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.442 Middling Yellow Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper...

  17. 7 CFR 28.441 - Strict Middling Yellow Stained Color.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 2 2013-01-01 2013-01-01 false Strict Middling Yellow Stained Color. 28.441 Section... CONTAINER REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.441 Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper...

  18. 7 CFR 28.441 - Strict Middling Yellow Stained Color.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Strict Middling Yellow Stained Color. 28.441 Section... CONTAINER REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.441 Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper...

  19. 7 CFR 28.442 - Middling Yellow Stained Color.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 2 2014-01-01 2014-01-01 false Middling Yellow Stained Color. 28.442 Section 28.442... REGULATIONS COTTON CLASSING, TESTING, AND STANDARDS Standards Yellow Stained Cotton § 28.442 Middling Yellow Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper...

  20. 21 CFR 864.1850 - Dye and chemical solution stains.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Dye and chemical solution stains. 864.1850 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Biological Stains § 864.1850 Dye and chemical solution stains. (a) Identification. Dye and chemical solution stains for medical purposes are mixtures...

  1. 21 CFR 864.1850 - Dye and chemical solution stains.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Dye and chemical solution stains. 864.1850 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Biological Stains § 864.1850 Dye and chemical solution stains. (a) Identification. Dye and chemical solution stains for medical purposes are mixtures...

  2. 21 CFR 864.1850 - Dye and chemical solution stains.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Dye and chemical solution stains. 864.1850 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Biological Stains § 864.1850 Dye and chemical solution stains. (a) Identification. Dye and chemical solution stains for medical purposes are mixtures...

  3. 21 CFR 864.1850 - Dye and chemical solution stains.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Dye and chemical solution stains. 864.1850 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Biological Stains § 864.1850 Dye and chemical solution stains. (a) Identification. Dye and chemical solution stains for medical purposes are mixtures...

  4. 21 CFR 864.1850 - Dye and chemical solution stains.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Dye and chemical solution stains. 864.1850 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Biological Stains § 864.1850 Dye and chemical solution stains. (a) Identification. Dye and chemical solution stains for medical purposes are mixtures...

  5. A rapid method for staining proteins in acrylamide gels.

    PubMed

    LeBlanc, G A; Cochrane, B J

    1987-02-15

    A negative staining procedure for the rapid visualization of proteins in acrylamide gels is described. In the absence of proteins, staining of the gel occurs through the reduction of nitroblue tetrazolium by reduced glutathione. No staining occurs in the presence of proteins. The procedure can be completed in 20 min and is at least as sensitive as Coomassie brilliant blue staining. PMID:2437824

  6. Cell-trappable quinoline-derivatized fluoresceins for selective and reversible biological Zn(II) detection.

    PubMed

    McQuade, Lindsey E; Lippard, Stephen J

    2010-10-18

    The synthesis and spectroscopic characterization of two new, cell-trappable fluorescent probes for Zn(II) are presented. These probes, 2-(4,5-bis(((6-(2-ethoxy-2-oxoethoxy)quinolin-8-yl)amino)methyl)-6-hydroxy-3-oxo-3H-8 xanthen-9-yl)benzoic acid (QZ2E) and 2,2'-((8,8'-(((9-(2-carboxyphenyl)-6-hydroxy-3-oxo-3H-xanthene-4,5-diyl)bis(methylene))bis(azanediyl))bis(quinoline-8,6-diyl))bis(oxy))diacetic acid (QZ2A), are poorly emissive in the off-state but exhibit dramatic increases in fluorescence upon Zn(II) binding (120 ± 10-fold for QZ2E, 30 ± 7-fold for QZ2A). This binding is selective for Zn(II) over other biologically relevant metal cations, toxic heavy metals, and most first-row transition metals and is of appropriate affinity (K(d1)(QZ2E) = 150 ± 100 μM, K(d2)(QZ2E) = 3.5 ± 0.1 mM, K(d1)(QZ2A) = 220 ± 30 μM, K(d2)(QZ2A) = 160 ± 80 μM, K(d3)(QZ2A) = 9 ± 6 μM) to reversibly bind Zn(II) at physiological levels. In live cells, QZ2E localizes to the Gogli apparatus where it can detect Zn(II). It is cell-membrane-permeable until cleavage of its ester groups by intracellular esterases produces QZ2A, a negatively charged acid form that cannot cross the cell membrane. PMID:20849126

  7. Double staining immunofluorescence procedure for enumeration of cells containing cytoplasmic immunoglobulin in human bone marrow: 40 selected cases.

    PubMed Central

    Ayliffe, M J

    1985-01-01

    A double staining immunofluorescence method was developed that allowed the reliable differential counting of cells containing cytoplasmic immunoglobulin in human bone marrow. These cells were stained with a polyspecific antihuman immunoglobulin serum conjugated with rhodamine. In separate preparations subpopulations of cells containing each heavy chain class and light chain type were identified by prior staining with isotype specific fluorescein conjugated antihuman immunoglobulin sera. A standardised counting procedure was adopted to indicate the degree of plasma cell infiltration and to establish the percentage of cells containing each type of immunoglobulin. A series of 122 patients requiring haematological investigation of the bone marrow for various disorders but without evidence of paraproteinaemia was tested to provide results for a nonmyelomatous reference group. These results were compared with those from 40 patients with paraproteins of various classes in their serum. The procedure described here clearly differentiated the two groups. When combined with serum paraprotein assays and conventional haematological cytology of bone marrow these counts provided useful additional information in cases of diagnostic difficulty and in the assessment of individual patients before, during, and after treatment. PMID:3930576

  8. Photodynamic therapy for port wine stains

    NASA Astrophysics Data System (ADS)

    Li, Junheng

    1998-11-01

    Previous therapies for port wine stains usually cause unacceptable scarring or obtain poor effect. Because port wine is a congenital vasculopathy consisting of an abnormal network of capillaries in the upper dermis with an overlying normal epidermis and the researchers found the tumor blood vessels were occluded accompanying the necrosis of the tumor after PDT. The author and his colleagues started a series of animal and clinical studies since 1991 about photodynamic therapy for port wine stain an they established the method of PDT for PWS. The clinical studies of over 1500 cases proved that PWS can be cured by PDT without scar formation because there is no thermal effect involved. No relapse was found within a maximum follow-up of six years.

  9. Lanthanum as an electron microscopic stain.

    PubMed

    Shaklai, M; Tavassoli, M

    1982-12-01

    Applications of lanthanum as an electron microscopic tracer have been reviewed. This electron-dense trivalent cation, which binds avidly to calcium binding sites, can be used as tracer for delineating extracellular spaces and intercellular junctions. It has served as a basis for classification of junctional structures. It can also be used as a calcium probe, a tracer in studying the permeability of barriers, as an intracellular marker and as an electron microscopic stain for such membrane components as surface glycoprotein. Each of these applications may require a different methodology. Thus methodological considerations in the use of this tracer have also been reviewed. The recent recognition that lanthanum is more than a passive tracer and that by reacting with different cell components may serve as a true stain, will extend the use of lanthanum in electron microscope histochemistry. PMID:6185564

  10. Gold staining in cellulose acetate membranes.

    PubMed

    Righetti, P G; Casero, P; Del Campo, G B

    1986-06-15

    A novel method for revealing proteins after electrophoresis on cellulose acetate is described, based on adsorption of mixed gold-Tween 20 micelles onto TCA-fixed protein bands. The sensitivity is 2-300 times higher than conventional Coomassie Blue staining and ca. 10 fold higher than silver coloring, detecting barely 1 ng protein/mm2 gel. Acidic and ammoniacal silver dyeing, based on ionic silver, perform very poorly on cellulose matrices, while a new variant, colloidal silver, stains protein bands on a clear background, but with a sensitivity ca. 10 times lower than micellar gold. Urines, cerebrospinal fluid and other biological liquids can be revealed without resorting to a concentration step prior to electrophoresis. The method is simple to perform and does not require any destaining step. PMID:2424644

  11. Evaluation of lanthanide salts as alternative stains to uranyl acetate.

    PubMed

    Hosogi, Naoki; Nishioka, Hideo; Nakakoshi, Masamichi

    2015-12-01

    Uranyl acetate (UAc) has been generally used not only as a superb staining reagent for ultrathin sections of plastic-embedded biological materials, but also as high-contrast negative stains for biological macromolecules such as particles of protein or virus. However, the use and purchase of radioactive UAc have been restricted. In this study, we determine the performance of ytterbium triacetate, lutetium triacetate, samarium triacetate and gadolinium triacetate as new staining reagents for biological electron microscopy. We observed chemically fixed spinach (Spinacia oleracea) leaves stained with these reagents. Ultrathin sections were stained with these reagents. Some of them were counterstained with lead citrate. The transmission electron microscopy contrast of spinach organelles was evaluated in sections exposed to the conventional stain and new stains. We show acetate salts of samarium, gadolinium, ytterbium and lutetium could be excellent substitutes for UAc for thin section staining and for negative staining. In addition, each reagent showed appreciable negative-staining effects. PMID:26374081

  12. Laser Treatment of Port Wine Stains

    NASA Astrophysics Data System (ADS)

    Majaron, Boris; Nelson, J. Stuart

    Port wine stain (PWS), also called nevus flammeus, is a congenital, cutaneous vascular malformation involving post-capillary venules which produce a light pink to red to dark-red-violet discoloration of human skin [1]. PWS occurs in an estimated 3 children per 1000 live births, affecting males and females and all racial groups equally [2]. There appears to be no hereditary predilection for PWS within families. There are no known risk factors or ways to prevent PWS.

  13. Photodynamic therapy for port wine stains

    NASA Astrophysics Data System (ADS)

    Li, Junheng

    1998-08-01

    Therapies for port wine stains including conventional laser irradiation usually cause unacceptable scarring or obtain poor effect. Pulsed dye laser has better approach, but only few patients obtain complete fading after multiple laser treatment. Because port wine stain is a congenital vasculopathy consisting of an abnormal network of capillaries in the upper dermis with an overlying normal epidermis and the researchers found that tumor blood vessels were occluded accompanying the necrosis of the tumor after PDT. It is though to be the effect primarily by thrombus formation in vessels and shut down of the blood supply to the tumor as well as direct tumor cells kill. The author and his colleagues started a series of animal and clinical studies since 1991 about photodynamic therapy for port wine stains and they established the method of PDT for PWS. An experimental study showed that Hpd appeared rapidly within the human vascular endothelial cells in culture fluid. Animal study using chicken combs as PWS models treated by PDT revealed the possibility of selective destruction of the malformative vasculature in PWS. The clinical studies of over 1700 cases proved that PWS can be cured without scar formation by PDT because there is no thermal effect involved. No relapse was found within a maximum follow-up of seven years. The differences and mechanism between the treatments of PDT and conventional lasers are discussed.

  14. Coffee Stain Effect with Liquid Droplets

    NASA Astrophysics Data System (ADS)

    Mitra, Sushanta; Das, Siddhartha

    2012-11-01

    We discuss the dynamics of immiscible bidispersed oil droplets that are suspended in an evaporating water sessile drop. Therefore, in contrast to classical coffee stain problem, the depositing ``particles'' are replaced by microscopic oil droplets - hence, we discuss a liquid-droplet coffee stain phenomenon. We show experimentally that unlike colloidal particles in a classical coffee stain problem, liquid oil droplets cannot reach the three phase contact line (TPCL) due to the aversion of the oil droplets to form finite oil-air interface in water medium. Therefore, the oil droplets get positioned at a finite distance from the TPCL. We call this distance the ``enclosure'' distance, which being a function of the droplet size, triggers a spontaneous size-based oil droplet separation. In addition, the ``enclosure'' effect is a function of the surface energies of the oil droplet and the rate of evaporation. We develop a theory to describe this effect, and the results show excellent agreement with the experimental findings. NSERC Banting Postdoctoral Fellowship for S. Das.

  15. Double staining of coomassie blue-stained polyacrylamide gels by imidazole-sodium dodecyl sulfate-zinc reverse staining: sensitive detection of coomassie blue-undetected proteins.

    PubMed

    Fernandez-Patron, C; Hardy, E; Sosa, A; Seoane, J; Castellanos, L

    1995-01-01

    The sensitivity, simplicity, and relative rapidity of Coomassie blue staining have made this technique the method of choice for routine detection and quantitative analysis of gel electrophoresis-separated protein bands in many applications. To extend the usefulness of this technique, we have developed a new double-staining method for visualizing SDS-PAGE-separated protein bands that were undetected by Coomassie blue staining of the gel. Coomassie blue-stained gels are washed in distilled water (15 min, two times) and then subjected to imidazole-zinc reverse staining. As a result of the method, a homogeneous white-stained background is generated and two types of protein bands can be observed: (a) typical Coomassie blue-stained bands, which appear superposed on larger transparent bands; and (b) reverse-stained (transparent) bands, which were previously undetected by the Coomassie blue staining. The method is rapid, simple, and reproducible and double-staining gels can be kept in distilled water for months without loss of the protein pattern. The overall sensitivity is high (e.g., 1.6 ng for recombinant streptokinase, 47 kDa) over a wide range of protein molecular weights (10 to 100 kDa) and independent of the degree of Coomassie blue destaining of the gel. Furthermore, a mechanism offering a consistent explanation for the role of imidazole, SDS, and zinc in the reverse staining of gels, particularly after Coomassie blue staining is proposed. PMID:7535985

  16. In Vivo Ocular Fluorophotometry: Delivery of Fluoresceinated Dextrans via Transscleral Diffusion in Rabbits

    PubMed Central

    Berezovsky, Damian E.; Patel, Samirkumar R.; McCarey, Bernard E.

    2011-01-01

    Purpose. To evaluate the transscleral delivery of fluoresceinated dextrans (FITC-D) with molecular mass up to 70 kDa to the rabbit posterior segment using sub-Tenon injections. Methods. Eighteen NZW rabbits received a unilateral 200-μL injection of 2 mg/mL sodium fluorescein (NaF), 25 mg/mL 40-kDa FITC-D, or 25 mg/mL 70-kDa FITC-D, with (n = 9) or without (n = 9) immediate euthanatization. In live animals, fluorescence was measured in the retina/choroid and mid-vitreous by fluorophotometry, immediately after injection and after 4, 24, 48, and 72 hours. Euthanatized animals were examined hourly through 5 or 6 hours. Results. In live animals, the average peak NaF concentration in the retina/choroid was 310.2 ng/mL, measured 3 hours after injection. Average 40- and 70-kDa FITC-D concentrations in the retina/choroid peaked at 5409.6 and 2375.6 ng/mL, respectively, 24 hours after injection. Fluorescence returned to baseline levels 6 hours after NaF injection, and 48 and 72 hours after 40- and 70-kDa FITC-D injections, respectively. Rabbits that received NaF followed by euthanatization exhibited a continuous increase in retina/choroid and mid-vitreous fluorescence, beginning 1 hour after injection, whereas FITC-D-injected eyes did not show elevated retina/choroid or mid-vitreous fluorescence through 6 hours. Conclusions. FITC-D weighing up to 70-kDa, as well as NaF, reached the posterior retina/choroid after sub-Tenon injections in live rabbits. NaF and 40-kDa FITC-D reached higher peak concentrations and were cleared from the eye more rapidly than was 70-kDa FITC-D. There was minimal penetration of NaF and FITC-D into the mid-vitreous in the in vivo experiments. PMID:21791594

  17. Segmentation of blood vessels from red-free and fluorescein retinal images.

    PubMed

    Martinez-Perez, M Elena; Hughes, Alun D; Thom, Simon A; Bharath, Anil A; Parker, Kim H

    2007-02-01

    The morphology of the retinal blood vessels can be an important indicator for diseases like diabetes, hypertension and retinopathy of prematurity (ROP). Thus, the measurement of changes in morphology of arterioles and venules can be of diagnostic value. Here we present a method to automatically segment retinal blood vessels based upon multiscale feature extraction. This method overcomes the problem of variations in contrast inherent in these images by using the first and second spatial derivatives of the intensity image that gives information about vessel topology. This approach also enables the detection of blood vessels of different widths, lengths and orientations. The local maxima over scales of the magnitude of the gradient and the maximum principal curvature of the Hessian tensor are used in a multiple pass region growing procedure. The growth progressively segments the blood vessels using feature information together with spatial information. The algorithm is tested on red-free and fluorescein retinal images, taken from two local and two public databases. Comparison with first public database yields values of 75.05% true positive rate (TPR) and 4.38% false positive rate (FPR). Second database values are of 72.46% TPR and 3.45% FPR. Our results on both public databases were comparable in performance with other authors. However, we conclude that these values are not sensitive enough so as to evaluate the performance of vessel geometry detection. Therefore we propose a new approach that uses measurements of vessel diameters and branching angles as a validation criterion to compare our segmented images with those hand segmented from public databases. Comparisons made between both hand segmented images from public databases showed a large inter-subject variability on geometric values. A last evaluation was made comparing vessel geometric values obtained from our segmented images between red-free and fluorescein paired images with the latter as the "ground truth". Our results demonstrated that borders found by our method are less biased and follow more consistently the border of the vessel and therefore they yield more confident geometric values. PMID:17204445

  18. Whole-mount silver staining of Arabidopsis and maize: a new method for staining secondary wall thickenings in tracheary elements.

    PubMed

    Kitajima, Sakihito; Hasegawa, Yoko; Sugimura, Yukio

    2004-12-01

    Secondary wall thickenings in tracheary elements were specifically stained by incubation of Arabidopsis and maize in Silver Stain Plus (Bio-Rad) staining solution, after pretreatment with SDS and ethanol solution. Scanning electron microscopic analysis of sections of celery revealed that silver particles were deposited on the secondary wall thickenings, indicating that the staining was due to the deposition of silver through the interaction of the stain with lignin. This method is more sensitive than the acidified phloroglucinol method. PMID:15618638

  19. The Transcriptional Response of Listeria monocytogenes during Adaptation to Growth on Lactate and Diacetate Includes Synergistic Changes That Increase Fermentative Acetoin Production▿†

    PubMed Central

    Stasiewicz, Matthew J.; Wiedmann, Martin; Bergholz, Teresa M.

    2011-01-01

    The organic acids lactate and diacetate are commonly used in combination in ready-to-eat foods because they show synergistic ability to inhibit the growth of Listeria monocytogenes. Full-genome microarrays were used to investigate the synergistic transcriptomic responses of two L. monocytogenes strains, H7858 (serotype 4b) and F6854 (serotype 1/2a), to these two organic acids under conditions representing osmotic and cold stress encountered in foods. Strains were exposed to brain heart infusion (BHI) broth at 7°C with 4.65% water-phase (w.p.) NaCl at pH 6.1 with (i) 2% w.p. potassium lactate, (ii) 0.14% w.p. sodium diacetate, (iii) the combination of both at the same levels, or (iv) no organic acids as a control. RNA was extracted 8 h after exposure, during lag phase, to capture gene transcription changes during adaptation to the organic acid stress. Significant differential transcription of 1,041 genes in H7858 and 640 genes in F6854 was observed in at least one pair of the 4 different treatments. The effects of combined treatment with lactate and diacetate included (i) synergistic transcription differences for 474 and 209 genes in H7858 and F6854, respectively, (ii) differential transcription of genes encoding cation transporters and ABC transporters of metals, and (iii) altered metabolism, including induction of a nutrient-limiting stress response, reduction of menaquinone biosynthesis, and a shift from fermentative production of acetate and lactate to energetically less favorable, neutral acetoin. These data suggest that additional treatments that interfere with cellular energy generation processes could more efficiently inhibit the growth of L. monocytogenes. PMID:21666015

  20. Lateral diffusion and conductance properties of a fluorescein-labelled alamethicin in planar lipid bilayers.

    PubMed

    Helluin, O; Dugast, J Y; Molle, G; Mackie, A R; Ladha, S; Duclohier, H

    1997-12-01

    In order to follow alamethicin diffusion within membranes under conditions of pore-formation, a fluorescein isothiocyanate (FITC) analogue was synthesized. To test the influence of the fluorescent probe addition on the pore-forming activity of the new analogue, macroscopic and single-channel experiments into planar lipid bilayers were performed. Although the apparent mean number of monomers per conducting aggregate was equivalent, the voltage-dependence of the new analogue was slightly reduced and hysteresses were broader, in agreement with the much longer duration of the open single-channels. Thus, the conducting aggregates seem to be stabilized by the introduction of the probe, presumably through the interaction of the conjugated cycles with the lipid headgroups, while the added steric hindrance may account for the slightly higher conductances of the open substates. Lateral diffusion of the labelled peptide associated with the bilayer was then investigated by the fluorescence recovery after photobleaching technique. Under applied voltage, associated with high conductance, D, the lateral diffusion coefficient, was reduced by 50% when compared to peptide at rest. These results provide new independent experimental evidence for a voltage-driven insertion of the highly mobile surface-associated peptide into the bilayer as a prominent step in pore formation. PMID:9408182

  1. Correlation between Fluorescein Angiographic Findings and Visual Acuity in Behçet Retinal Vasculitis

    PubMed Central

    Kim, Min; Kwon, Hee Jung; Choi, Eun Young; Kim, Sung Soo; Koh, Hyoung Jun

    2015-01-01

    Purpose To identify significant fluorescein angiographic (FA) characteristics associated with visual acuity (VA) in Behçet retinal vasculitis. Materials and Methods Retrospective review of 86 eyes of 48 patients (age: 35.6±10.2 years) with Behçet retinal vasculitis were performed. VA and FA findings as well as correlation between them were assessed. Results The mean initial VA of eyes with posterior pole-involved vasculitis (63 eyes; 73.3%) was significantly worse than that of those with peripheral vasculitis (23 eye; 26.7%) (logarithm of the minimum angle of resolution VA: 0.554±0.572 vs. 0.078±0.148; p<0.0001). Subgroup analysis revealed a more severe and diffuse pattern of vascular leakage in posterior pole-involved vasculitis compared to peripheral vasculitis (p<0.0001). Retinal vascular leakage (β=0.345; p<0.0001), optic disc hyperfluorescence (β=0.147; p=0.032), and macular leakage (β=0.107; p=0.047) were significantly associated with worse initial VA. During the follow up (mean: 33.3±17.9 months), the change of leakage showed no significant correlation with change of VA in posterior pole-involved vasculitis (τ=0.199, p=0.092). Conclusion Posterior pole involvement, the degree of retinal vascular leakage, optic disc hyperfluorescence, and macular leakage are significantly associated with VA in Behçet retinal vasculitis. PMID:26069134

  2. Groundwater dating using radiocarbon in fulvic acid in groundwater containing fluorescein

    NASA Astrophysics Data System (ADS)

    Nakata, Kotaro; Kodama, Hiroki; Hasegawa, Takuma; Hama, Katsuhiro; Iwatsuki, Teruki; Miyajima, Tohru

    2013-05-01

    Natural DO14C is recognized as one of the most useful tracers in the estimation of groundwater age. Fluorescent dye is commonly used as an indicator of drilling fluid contamination during borehole investigation. Fluorescein (FS) is one of the most frequently used fluorescent dyes, yet as it contains little radiocarbon it may affect DO14C age when it is mixed with natural DOC. In this study, fulvic acid (FA) was isolated from groundwater containing FS and DO14C value of isolated FA was measured. Separation methods were proposed by using the difference between sorption and desorption behavior of FA and FS onto synthetic adsorbent resin. DO14C measurement on FA from a mixture of FA and FS is estimated by removing FS from the mixture and correcting for the amount of C derived from FS. Furthermore, DO14C age is compared with the groundwater age estimated by He. The results show that the values of DO14C age for separated FA estimated by the two methods had good agreement with those that corresponded to groundwater age estimated by He. This result indicates that DO14C is a useful indicator of groundwater age even for groundwater contaminated with FS.

  3. Acute retinal pigment epitheliitis: spectral domain optical coherence tomography, fluorescein angiography, and autofluorescence findings.

    PubMed

    Aydoğan, Tuğba; Güney, Esra; Akçay, Betül İlkay Sezgin; Bozkurt, Tahir Kansu; Ünlü, Cihan; Ergin, Ahmet

    2015-01-01

    A 17-year-old presented with central and paracentral scotomas in his right eye for one week. There was no remarkable medical or ocular history. Blood analyses were within normal range. At presentation both eyes' best-corrected visual acuities were 20/20. Slit-lamp examination result was normal. Fundus examination revealed yellow-white hypopigmented areas in the macula. Fluorescein angiography (FA) showed hypofluorescence surrounded by ring of hyperfluorescence. Fundus autofluorescence (FAF) was slightly increased. Spectral domain optical coherence tomography (SD-OCT) showed disruption of IS/OS junction with expansion of abnormal hyperreflectivity from retinal pigment epithelium to the outer nuclear layer (ONL). One month later fundus examination showed disappearance of the lesions. FA revealed transmission hyperfluorescence. FAF showed increased autofluorescence and pigment clumping. Hyperreflective band in SD-OCT disappeared. Loss of photoreceptor segment layers was observed in some of the macular lesions. The diagnosis of acute retinal pigment epitheliitis can be challenging after disappearance of fundus findings. FA, FAF, and SD-OCT are important tests for diagnosis after resolution of the disease. PMID:25767511

  4. Volatilization of fluorescein mercuric acetate by marine bacterial from Minamata Bay

    SciTech Connect

    Nakamura, Kunihiko )

    1989-05-01

    Some bacteria that live in a mercury-polluted environment are resistant to mercury compounds. A majority of these mercury-resistant bacterial have been found to volatilize organic as well as inorganic mercury compounds into elemental mercury vapor by means of their enzymes. One compound, fluorescein mercuric acetate (FMA) has long been in use as a disinfectant in hospitals; yet, there has been little definitive information on bacterial resistance to this compound. Minamata Bay has been heavily polluted by mercury, which has caused methylmercury poisoning in humans, called Minamata disease. Sediments from the Bay still contain high concentrations of mercury. The percentage of mercury-resistant bacteria in the total bacterial count is higher in these sediments than in those of other marine environments. FMA-pollution, however, has not been reported. Research into the mechanism of bacterial resistance to FMA will not only add to our general understanding of the ability of certain bacteria to resist mercury, but will also help in defining the role bacteria play in the mercury cycle of a mercury-polluted environment. The purpose of the present study is to determine the mechanism of resistance to FMA of the FMA-resistant bacteria living in the Bay.

  5. Dibutyl Maleate and Dibutyl Fumarate Enhance Contact Sensitization to Fluorescein Isothiocyanate in Mice.

    PubMed

    Matsuoka, Takeshi; Kurohane, Kohta; Suzuki, Wakana; Ogawa, Erina; Kobayashi, Kamiyu; Imai, Yasuyuki

    2016-02-01

    Di-n-butyl phthalate (DBP), a phthalate ester, has been shown to have an adjuvant effect on fluorescein isothiocyanate (FITC)-induced contact hypersensitivity (CHS) mouse models. Di-n-butyl maleate (DBM), widely used as a plasticizer for industrial application, has been reported to cause dermatitis in humans. DBM is a butyl alcohol ester of di-carboxylic acid that represents a part of the DBP structure, while di-n-butyl fumarate (DBF) is a trans isomer of DBM. We examined whether DBM or DBF exhibits an adjuvant effect like DBP does. When BALB/c mice were epicutaneously sensitized with FITC in the presence of DBM or DBF, the FITC-specific CHS response was enhanced, as we have observed for DBP. As to underlying mechanisms, DBM and DBF facilitated the trafficking of FITC-presenting CD11c(+) dendritic cells (DCs) from skin to draining lymph nodes and increased the cytokine production by draining lymph nodes. In conclusion, DBM and DBF may have an effect that aggravates contact dermatitis through a skin sensitization process. PMID:26632200

  6. Extending applicability of the oxygen radical absorbance capacity (ORAC-fluorescein) assay.

    PubMed

    Dvalos, Alberto; Gmez-Cordovs, Carmen; Bartolom, Begoa

    2004-01-14

    The ORAC-fluorescein (ORAC-FL) method recently validated using automatic liquid handling systems has now been adapted to manual handling and using a conventional fluorescence microplate reader. As calculated for Trolox, the precision of the method was <3.0, expressed as percent coefficient of variation. The accuracy of the method was <2.3, expressed as percent variation of the mean. The detection and quantification limits were those corresponding to 0.5- and 1-microM Trolox standard solutions, respectively. The method has been applied to 10 pure compounds (benzoic and cinnamic acids and aldehydes, flavonoids, and butylated hydroxyanisole), to 30 white, rose, and bottled- and oak-aged red wines, and to 7 commercial dietary antioxidant supplements. All samples exhibited a good linear response with concentration. As seen by other methodologies, the chemical structure of a compound determines its antioxidant activity (ORAC-FL value). Of particular interest were the results with oak-aged red wines from different vintages (1989-2002) that confirm influence of vintage, but not origin of the oak, in the antioxidant activity of wines from the same variety. Dietary antioxidant supplements presented a great variability (170-fold difference) in their antioxidant potency. This work proves applicability of the ORAC-FL assay in evaluating the antioxidant activity of diverse food samples. PMID:14709012

  7. Automated Detection of Vessel Abnormalities on Fluorescein Angiogram in Malarial Retinopathy

    PubMed Central

    Zhao, Yitian; MacCormick, Ian J. C.; Parry, David G.; Beare, Nicholas A. V.; Harding, Simon P.; Zheng, Yalin

    2015-01-01

    The detection and assessment of intravascular filling defects is important, because they may represent a process central to cerebral malaria pathogenesis: neurovascular sequestration. We have developed and validated a framework that can automatically detect intravascular filling defects in fluorescein angiogram images. It first employs a state-of-the-art segmentation approach to extract the vessels from images and then divide them into individual segments by geometrical analysis. A feature vector based on the intensity and shape of saliency maps is generated to represent the level of abnormality of each vessel segment. An AdaBoost classifier with weighted cost coefficient is trained to classify the vessel segments into normal and abnormal categories. To demonstrate its effectiveness, we apply this framework to 6,358 vessel segments in images from 10 patients with malarial retinopathy. The test sensitivity, specificity, accuracy, and area under curve (AUC) are 74.7%, 73.5%, 74.1% and 74.2% respectively when compared to the reference standard of human expert manual annotations. This performance is comparable to the agreement that we find between human observers of intravascular filling defects. Our method will be a powerful new tool for studying malarial retinopathy. PMID:26053690

  8. Comparison of adaptive optics scanning light ophthalmoscopic fluorescein angiography and offset pinhole imaging.

    PubMed

    Chui, Toco Y P; Dubow, Michael; Pinhas, Alexander; Shah, Nishit; Gan, Alexander; Weitz, Rishard; Sulai, Yusufu N; Dubra, Alfredo; Rosen, Richard B

    2014-04-01

    Recent advances to the adaptive optics scanning light ophthalmoscope (AOSLO) have enabled finer in vivo assessment of the human retinal microvasculature. AOSLO confocal reflectance imaging has been coupled with oral fluorescein angiography (FA), enabling simultaneous acquisition of structural and perfusion images. AOSLO offset pinhole (OP) imaging combined with motion contrast post-processing techniques, are able to create a similar set of structural and perfusion images without the use of exogenous contrast agent. In this study, we evaluate the similarities and differences of the structural and perfusion images obtained by either method, in healthy control subjects and in patients with retinal vasculopathy including hypertensive retinopathy, diabetic retinopathy, and retinal vein occlusion. Our results show that AOSLO OP motion contrast provides perfusion maps comparable to those obtained with AOSLO FA, while AOSLO OP reflectance images provide additional information such as vessel wall fine structure not as readily visible in AOSLO confocal reflectance images. AOSLO OP offers a non-invasive alternative to AOSLO FA without the need for any exogenous contrast agent. PMID:24761299

  9. Saturable CD14-dependent binding of fluorescein-labeled lipopolysaccharide to human monocytes.

    PubMed Central

    Troelstra, A; Antal-Szalmas, P; de Graaf-Miltenburg, L A; Weersink, A J; Verhoef, J; Van Kessel, K P; Van Strijp, J A

    1997-01-01

    We used rough lipopolysaccharide (ReLPS) to construct a fluorescein-labeled LPS (FITC-LPS) with a very high labeling efficiency that bound to isolated human monocytes in a CD14-dependent fashion and that in this respect behaved indistinctively from native LPS. The CD14-dependent binding could be inhibited either by a 1,000-fold excess of unlabeled LPS or by polymyxin B, bactericidal/permeability-increasing protein, cationic protein 18, or soluble CD14. Although this FITC-LPS preparation no longer possessed the ability to prime neutrophils for the production of reactive oxygen species or to stimulate human monocytes to produce tumor necrosis factor, activation of the Limulus amoebocyte lysate cascade was comparable to activation by native LPS. Binding to monocytes was enhanced by human pooled serum (HPS) or LPS-binding protein (LBP) for LPS concentrations up to 100 ng/ml and was completely CD14 dependent. For LPS concentrations exceeding 100 ng/ml, binding was still partially CD14 dependent, but not HPS or LBP dependent. CD14-dependent association of LPS with monocytes was shown to be totally saturable. In conclusion, we found an HPS- or LBP-dependent binding of FITC-LPS to monocytes that was CD14 dependent at up to 100 ng of LPS per ml, and saturation of binding was shown. PMID:9169763

  10. New Parametric Imaging Method with Fluorescein Angiograms for Detecting Areas of Capillary Nonperfusion

    PubMed Central

    Kim, Young Jae; Jeong, Chang Bu; Hwang, Jeong-Min; Yang, Hee Kyung; Lee, Seung Hyun

    2014-01-01

    Objectives Fluorescein angiography (FAG) is currently the most useful diagnostic modality for examining retinal circulation, and it is frequently used for the evaluation of patients with diabetic retinopathy, occlusive diseases, such as retinal venous and arterial occlusions, and wet macular degeneration. This paper presents a method for objectively evaluating retinal circulation by quantifying circulation-related parameters. Methods This method allows the semiautomatic preprocessing and registering of FAG images. The arterial input function is estimated from the registered set of FAG images using gamma-variate fitting. Then, the parameters can be computed by deconvolution on the basis of truncated singular value decomposition, and they can finally be presented as parametric color images in a combination of three colors, red, green, and blue. Results After the estimation of arterial input function, the parameters of relative blood flow and mean transit time were computed using deconvolution analysis based on truncated singular value decomposition. Conclusions The parametric color image is helpful to interpret the status of retinal blood circulation and provides quantitative data on retina ischemia without interobserver variability. This system easily provides the status of retinal blood circulation both qualitatively and quantitatively. It also helps to standardize FAG interpretation and may contribute to network-based telemedicine systems in the future. PMID:25152832

  11. Inhibition of tonoplast ATPase from etiolated mung bean seedlings by fluorescein 5'-isothiocyanate.

    PubMed Central

    Tzeng, C M; Hsu, L H; Pan, R L

    1992-01-01

    Fluorescein 5'-isothiocyanate (FITC) was used to modify the lysine residue in the active site of tonoplast H(+)-ATPase from etiolated mung-bean (Vigna radiata L.) seedlings. FITC caused marked inactivation of the enzyme activities of both membrane-bound and soluble ATPase and its associated H+ translocation. The SDS/PAGE pattern revealed that the FITC-binding site was in the large (A) subunit of ATPase. Inhibition could be substantially prevented by its physiological substrate ATP, pyrophosphate and nucleotides in the decreasing order: ATP greater than pyrophosphate greater than ADP greater than AMP greater than GTP greater than CTP greater than UTP. The mode of inhibition by FITC was competitive with respect to ATP. Loss of ATPase activity followed pseudo-first-order kinetics with a Ki of 0.33 mM, a minimum inactivation half-time of 110 s, and a first-order rate constant of 0.244 s-1. A double-logarithmic plot of apparent rate constant versus FITC concentration gave a slope of 0.913, indicating that inactivation results from reaction of at least one lysine residue at the catalytic site of the large subunit. Labelling studies indicated that the incorporation of approx. 1 mol of FITC/mol of ATPase is sufficient to inhibit ATPase completely. The enhancement and blue shift of emission maxima of FITC after modification of ATPase indicated that the labelled lysine residue was located in a relatively hydrophobic domain. PMID:1386733

  12. Estimating retinal vascular permeability using the adiabatic approximation to the tissue homogeneity model with fluorescein videoangiography

    NASA Astrophysics Data System (ADS)

    Tichauer, Kenneth M.; Osswald, Christian R.; Dosmar, Emily; Guthrie, Micah J.; Hones, Logan; Sinha, Lagnojita; Xu, Xiaochun; Mieler, William F.; St. Lawrence, Keith; Kang-Mieler, Jennifer J.

    2015-06-01

    Clinical symptoms of diabetic retinopathy are not detectable until damage to the retina reaches an irreversible stage, at least by today's treatment standards. As a result, there is a push to develop new, "sub-clinical" methods of predicting the onset of diabetic retinopathy before the onset of irreversible damage. With diabetic retinopathy being associated with the accumulation of long-term mild damage to the retinal vasculature, retinal blood vessel permeability has been proposed as a key parameter for detecting preclinical stages of retinopathy. In this study, a kinetic modeling approach used to quantify vascular permeability in dynamic contrast-enhanced medical imaging was evaluated in noise simulations and then applied to retinal videoangiography data in a diabetic rat for the first time to determine the potential for this approach to be employed clinically as an early indicator of diabetic retinopathy. Experimental levels of noise were found to introduce errors of less than 15% in estimates of blood flow and extraction fraction (a marker of vascular permeability), and fitting of rat retinal fluorescein angiography data provided stable maps of both parameters.

  13. Reproducibility of morphometric measurements of amyloid after various staining methods.

    PubMed

    Kosma, V M; Collan, Y; Kulju, T; Aalto, M L; Jantunen, E; Karhunen, J; Selkäinaho, K

    1985-12-01

    Four histologic staining methods used for detecting amyloid (Congo red, viewed in both normal and polarized light, Sirius red, Crystal violet and Thioflavine T) were applied to heart muscle autopsy samples from 19 patients who suffered from amyloidosis. The amount of amyloid present was evaluated with morphometry (point counting) by five pathologists, and the interobserver reproducibility and variation of point counting in these staining methods were analyzed. The Sirius red method showed the least variation and was the most suitable stain for demonstrating amyloid with respect to reproducibility. Thioflavine T showed the greatest variation and was the least suitable stain with respect to reproducibility. The range of variation was considerable in all staining methods. The results show that stains differ in their specificity and sensitivity in staining amyloid, observers differ in their interpretation of staining results and certain stains result in more uniform interpretations than do others. PMID:2418854

  14. Histological Stains: A Literature Review and Case Study.

    PubMed

    Alturkistani, Hani A; Tashkandi, Faris M; Mohammedsaleh, Zuhair M

    2016-03-01

    The history of histology indicates that there have been significant changes in the techniques used for histological staining through chemical, molecular biology assays and immunological techniques, collectively referred to as histochemistry. Early histologists used the readily available chemicals to prepare tissues for microscopic studies; these laboratory chemicals were potassium dichromate, alcohol and the mercuric chloride to harden cellular tissues. Staining techniques used were carmine, silver nitrate, Giemsa, Trichrome Stains, Gram Stain and Hematoxylin among others. The purpose of this research was to assess past and current literature reviews, as well as case studies, with the aim of informing ways in which histological stains have been improved in the modern age. Results from the literature review has indicated that there has been an improvement in histopathology and histotechnology in stains used. There has been a rising need for efficient, accurate and less complex staining procedures. Many stain procedures are still in use today, and many others have been replaced with new immunostaining, molecular, non-culture and other advanced staining techniques. Some staining methods have been abandoned because the chemicals required have been medically proven to be toxic. The case studies indicated that in modern histology a combination of different stain techniques are used to enhance the effectiveness of the staining process. Currently, improved histological stains, have been modified and combined with other stains to improve their effectiveness. PMID:26493433

  15. Histological Stains: A Literature Review and Case Study

    PubMed Central

    Alturkistani, Hani A; Tashkandi, Faris M; Mohammedsaleh, Zuhair M

    2016-01-01

    The history of histology indicates that there have been significant changes in the techniques used for histological staining through chemical, molecular biology assays and immunological techniques, collectively referred to as histochemistry. Early histologists used the readily available chemicals to prepare tissues for microscopic studies; these laboratory chemicals were potassium dichromate, alcohol and the mercuric chloride to harden cellular tissues. Staining techniques used were carmine, silver nitrate, Giemsa, Trichrome Stains, Gram Stain and Hematoxylin among others. The purpose of this research was to assess past and current literature reviews, as well as case studies, with the aim of informing ways in which histological stains have been improved in the modern age. Results from the literature review has indicated that there has been an improvement in histopathology and histotechnology in stains used. There has been a rising need for efficient, accurate and less complex staining procedures. Many stain procedures are still in use today, and many others have been replaced with new immunostaining, molecular, non-culture and other advanced staining techniques. Some staining methods have been abandoned because the chemicals required have been medically proven to be toxic. The case studies indicated that in modern histology a combination of different stain techniques are used to enhance the effectiveness of the staining process. Currently, improved histological stains, have been modified and combined with other stains to improve their effectiveness. PMID:26493433

  16. Silver(I) 2,2'-(1,2-Phenylenedisulfanediyl)diacetic Acid as a Molecular Building Block for a Silver(I)-Cadmium(II) Coordination Polymer.

    PubMed

    Grosu, Ioana Georgeta; Berghof, Christiane; Lönnecke, Peter; Silaghi-Dumitrescu, Luminita; Hey-Hawkins, Evamarie

    2015-01-01

    Starting from heterotopic multidentate ligand 2,2'-(1,2-phenylenedisulfanediyl)diacetic acid, (RS,RS,RS,RS/SS,SS,SS,SS)-[Ag{1,2-C6H4(SCH2COOH)2-κ2S,S'}2]BF4 (1) was prepared and further used as a building block for the synthesis of heterobimetallic Ag-Cd coordination polymer [Ag2Cd2{1,2-(OOCCH2S)2C6H4}3 (H2O)3·5H2O]n (2). Both complexes were characterized by X-ray structure analysis and conventional spectroscopic techniques. PMID:25946559

  17. Bleaching of fluorosis stains using sodium hypochlorite.

    PubMed

    Penumatsa, Narendra Varma; Sharanesha, Rajashekhara Bhari

    2015-08-01

    Fluorosis staining is commonly considered an esthetic problem because of the psychological impact of unesthetic maxillary anterior teeth. Numerous treatment approaches have been proposed, ranging from bleaching to enamel reduction to restorative techniques. Bleaching of hypomineralized enamel lesions, using 5% sodium hypochlorite, has been useful clinically. The technique described, in this case, appears to have advantages over other methods for improving the appearance of fluorotic lesions. It is simple, low cost, noninvasive, so the enamel keeps its structure, relatively rapid, and safe; it requires no special materials, and it can be used with safety on young permanent teeth. PMID:26538964

  18. Bleaching of fluorosis stains using sodium hypochlorite

    PubMed Central

    Penumatsa, Narendra Varma; Sharanesha, Rajashekhara Bhari

    2015-01-01

    Fluorosis staining is commonly considered an esthetic problem because of the psychological impact of unesthetic maxillary anterior teeth. Numerous treatment approaches have been proposed, ranging from bleaching to enamel reduction to restorative techniques. Bleaching of hypomineralized enamel lesions, using 5% sodium hypochlorite, has been useful clinically. The technique described, in this case, appears to have advantages over other methods for improving the appearance of fluorotic lesions. It is simple, low cost, noninvasive, so the enamel keeps its structure, relatively rapid, and safe; it requires no special materials, and it can be used with safety on young permanent teeth. PMID:26538964

  19. In vivo labelling of the spleen and mesenteric lymph nodes with fluorescein isothiocyanate for lymphocyte migration studies.

    PubMed Central

    Pabst, R; Binns, R M

    1981-01-01

    Lymphocytes in normal young pigs were labelled in vivo with fluorescein isothiocyanate in the spleen using an extracorporeal perfusion system and in mesenteric lymph nodes by direct injection into the nodes. Labelled lymphocytes leave the spleen at a high rate via the splenic vein and migrate to different lymphoid organs. Emigrants from mesenteric lymph nodes left the nodes more slowly and revealed a different homing pattern. Evidence is presented that a considerable number of lymphocytes from the parenchyma leave the nodes via the vein and not by the classical route of recirculating lymphocytes via the efferent lymphatics. Fluorescein labelling of lymphocytes in their normal micro-environment is a suitable method for lymphocyte migration studies. Images Figure 1 Figure 2 PMID:6795108

  20. Treatment of port-wine stains: analysis

    SciTech Connect

    van Gemert, M.J.; Welch, A.J.

    1987-08-01

    Port-wine stains (PWS) are bluish red skin stains that are caused by enlarged, ectatic blood vessels in the dermis. Laser treatment of PWS is analyzed from computation of the spatial distribution of heat production by direct absorption of the laser light and subsequent heat conduction. The absorption and scattering caused by oxyhemoglobin, epidermis, and dermis as a function of wavelength are utilized in this analysis. Ideal treatment is defined as coagulating the ectatic blood vessels without irreversible damage to the epidermis and dermis. The analysis shows that a millisecond pulsed, yellow dye laser at 577 nm (one of the large absorption bands in blood) is the laser of choice to treat PWS, offering as close to the ''ideal treatment'' as possible. The blue-green argon laser, which is currently the most frequently used laser for this purpose, is strongly recommended with irradiation times in milliseconds. Other lasers that are in clinical use, such as the red ruby and near-infrared Nd-YAG lasers, can provide selective treatment only when the epidermis is cooled concurrently. The CO/sub 2/ laser, on the other hand, can coagulate the blood vessels only through heat conduction from the hot epidermis; hence, it has neither the treatment selectivity nor any other physical option to force this selectivity.

  1. Comparison of conventional staining methods and monoclonal antibody-based methods for Cryptosporidium oocyst detection.

    PubMed

    Arrowood, M J; Sterling, C R

    1989-07-01

    The sensitivity and specificity of seven microscopy-based Cryptosporidium oocyst detection methods were compared after application to unconcentrated fecal smears. The seven methods were as follows: (i) a commercial acid-fast (AF) stain (VOLU-SOL) method, (ii) Truant auramine-rhodamine (AR) stain method, (iii) fluorescein-conjugated C1B3 monoclonal antibody (MAb) direct fluorescence method, (iv) OW3 MAb indirect fluorescence method, (v) biotinylated OW3 indirect fluorescence method, (vi) biotinylated OW3-indirect diaminobenzidine (DAB) method, and (vii) biotinylated OW3-aminoethylcarbazole (AEC) method. A total of 281 randomly collected Formalin-fixed fecal samples (submitted to the Maricopa County Health Department, Phoenix, Ariz.) and 30 known positives (Formalin-fixed and K2Cr2O7-preserved stools from our laboratory) were examined in a blind test; 32 of 311 samples (10.3%) were confirmed positive. Of the confirmed positives, 40.6% were identified by the AF method, 93.8% were identified by the AR method, 93.8% were identified by the C1B3 method, 81.3% were identified by the OW3-DAB method, 71.9% were identified by the OW3-AEC method, 100% were identified by the OW3 indirect fluorescence method, and 100% were identified by the biotinylated OW3 indirect fluorescence method. False-positives were encountered by the AF and AR methods (52.0 and 85.7% specificity, respectively), while no false-positives encountered by the MAb-based methods. Oocysts in infected tissue sections were easily detected by the MAb-based methods. PMID:2475523

  2. pH-sensitive intracellular photoluminescence of carbon nanotube-fluorescein conjugates in human ovarian cancer cells

    NASA Astrophysics Data System (ADS)

    Chen, M. T.; Gomez, L. M.; Ishikawa, F. N.; Vernier, P. T.; Zhou, C.; Gundersen, M. A.

    2009-07-01

    To add to the understanding of the properties of functionalized carbon nanotubes in biological applications, we report a monotonic pH sensitivity of the intracellular fluorescence emission of single-walled carbon nanotube-fluorescein carbazide (SWCNT-FC) conjugates in human ovarian cancer cells. Light-stimulated intracellular hydrolysis of the amide linkage and localized intracellular pH changes are proposed as mechanisms. SWCNT-FC conjugates may serve as intracellular pH sensors.

  3. No intermediate channelling in stepwise hydrolysis of fluorescein di-beta-D-galactoside by beta-galactosidase.

    PubMed

    Fieldler, F; Hinz, H

    1994-05-15

    For the hydrolysis of the two glycosidic bonds of fluorescein di-beta-D-galactoside (FDG) by beta-galactosidase from Escherichia coli, small [Hofmann, J. & Sernetz, M. (1983) Anal. Biochem. 131, 180-186] to dramatic [Huang, Z. (1991) Biochemistry 30, 8535-8540] deviations from simple stepwise substrate-intermediate-product kinetics have been reported. Intermediate channelling, a preferred hydrolysis of the intermediate fluorescein mono-beta-D-galactoside (FMG) formed from FDG at the active site and thus in a favourable position for further reaction, has been postulated. As there were reasons to doubt the previous findings and conclusions, the hydrolysis experiments have been repeated at initial FDG concentrations of 7-200 microM, following the concentrations of FDG, FMG and fluorescein with a reliable method, quantitative HPLC, to completion of the reaction. The transient appearance of substantial amounts of the intermediate FMG also in experiments with 200 microM FDG already rules out the existence of the most efficient intermediate channelling deduced by Huang (1991) from measurements of the initially developing fluorescence, incorrectly ascribed to fluorescein. Redetermination of the Michaelis constants for FDG and FMG led to much higher values than those reported previously. Fitting the progress curves by means of nonlinear regression combined with numerical integration of the rate equations resulted in good fits of the normal stepwise substrate-intermediate-product mechanism, without any necessity of assuming a more complex course of the reaction. So one of the rare examples of the hydrolysis of two bonds at a single enzyme-substrate encounter has been invalidated. PMID:8200355

  4. Polyvinyl pyrrolidone capped fluorescent anthracene nanoparticles for sensing fluorescein sodium in aqueous solution and analytical application for ophthalmic samples.

    PubMed

    Bhopate, Dhanaji P; Mahajan, Prasad G; Garadkar, Kalyanrao M; Kolekar, Govind B; Patil, Shivajirao R

    2015-11-01

    Based on the known complexation ability between polyvinyl pyrrolidone (PVP) and fluorescein sodium (FL Na(+)), fluorescent PVP capped anthracene nanoparticles (PVP-ANPs) were prepared using a reprecipitation method for detection of fluorescein in aqueous solution using the fluorescence resonance energy transfer (FRET) approach. A dynamic light scattering histogram of PVP-ANPs showed narrower particle size distribution and the average particle size was 15 nm. The aggregation-induced enhanced emission (AIEE) of PVP-ANPs was red shifted from its monomer by 1087.22 cm(-1). The maximum emission was seen to occur at 420 nm. The presence of FL Na(+) in the vicinity of PVP-ANPs quenched the fluorescence of PVP-ANPs because of its adsorption on the surface of PVP-ANPs in aqueous suspension. The FL Na(+) and PVP-ANPs were brought close enough, typically to 7.89 nm, which was less than the distance of 10 nm that is required between the energy donor-acceptor molecule for efficient FRET. The quenching results fit into the Stern-Volmer relationship even at temperatures greater than ambient temperatures. The thermodynamic parameters determined from FRET results helped to propose binding mechanisms involving hydrophobic and electrostatic molecular interaction. The fluorescence quenching results were used further to develop an analytical method for estimation of fluorescein sodium from ophthalmic samples available commercially in the market. PMID:25736374

  5. Flow cytometric analysis of fluorescein-labeled nerve growth factor binding to A875 human melanoma cells.

    PubMed

    Kasaian, M T; Jacobberger, J W; Neet, K E

    1994-01-01

    The interaction of fluorescein-labeled nerve growth factor (NGF) with human melanoma cells (A875) has been studied in order to assess better methodology for rigorous NGF binding studies. The NGF was modified at a single carboxyl group with iodoacetamidofluorescein after reaction with carbodiimide and cystamine. The modified NGF showed full binding competence in competition with radiolabeled NGF and full biological activity in neurite outgrowth assays compared to native NGF. Binding to unfixed, viable cells was assayed using flow cytometry. This method offers the advantage that unbound ligand need not be separated from that which is cell-associated, thus avoiding perturbation of the binding equilibrium, and accurate, extensive statistical analysis is possible. Binding of fluorescein-NGF was mainly specific and saturable, with analysis by three methods of data treatment indicating a Kd of 0.8 to 3 nM at 4 degrees C. Time-based data acquisition allowed a continuous time course for binding to be generated. Binding reached a steady-state level within 5 min of exposure of the cells to the ligand. Kinetic and steady-state results obtained using fluorescein-NGF agree well with previous data produced by 125I-NGF binding studies. The main limitation of the flow cytometric method in the NGF system is the relative lack of sensitivity compared to the binding of radiolabeled NGF, partially due to unusual quenching of the fluorophore bound to NGF. PMID:8270000

  6. Port wine stain on a child's face (image)

    MedlinePlus

    Port wine stains are always present at birth. In an infant, they are flat, pink, vascular lesions. Common locations ... may be present anywhere on the body. Port wine stains may appear in association with other syndromes.

  7. Laser therapy in plastic surgery: decolorization in port wine stains

    NASA Astrophysics Data System (ADS)

    Peszynski-Drews, Cezary; Wolf, Leszek

    1996-03-01

    For the first time laserotherapy is described as a method of port wine stain decolorization in plastic surgery. The authors present their 20-year experience in the treatment of port wine stains with the argon laser and dye laser.

  8. Spectroscopic characterization of the binding mechanism of fluorescein and carboxyfluorescein in human serum albumin

    NASA Astrophysics Data System (ADS)

    Sulaiman, Saba A. J.; Kulathunga, H. Udani; Abou-Zied, Osama K.

    2015-03-01

    Fluorescein (FL) and some of its precursors have proven to be effective fluorescent tracers in pharmaceutical and medical applications owing to their high quantum yield of fluorescence in physiological conditions and their high membrane permeability. In order to protect FL from metabolic effects during the process of its delivery, human serum albumin (HSA) has been used as a carrier because of its compatibility with the human body. In the present work, we used spectroscopic methods to characterize the binding mechanisms of FL and one of its derivatives, 5(6)- carboxyfluorescein (CFL), in the HSA protein. The absorbance change of the two ligands (FL and CFL) was quantified as a function of the HSA concentration and the results indicate a moderate binding strength for the two ligands inside HSA (1.00 +/- 0.12 x 104 M-1). The quenching effect of FL(CFL) on the fluorescence intensity of W214 (the sole tryptophan in HSA) indicates that FL and CFL occupy Site I in the protein which is known to bind several hydrophobic drugs. By performing site-competitive experiments, the location of the ligands is determined to be similar to that of the anticoagulant drug warfarin. At higher ratios of [ligand]/[HSA], we observed an upward curvature in the Stern-Volmer plots which indicates that the ligands occupy more pockets in Site I, close to W214. Our results indicate that both ligands bind in HSA with a moderate strength that should not affect their release when used as fluorescent reporters. The chemical and physical identities of the two ligands are also preserved inside the HSA binding sites.

  9. Detection of retinal capillary nonperfusion in fundus fluorescein angiogram of diabetic retinopathy

    PubMed Central

    Rasta, Seyed Hossein; Nikfarjam, Shima; Javadzadeh, Alireza

    2015-01-01

    Introduction: Retinal capillary nonperfusion (CNP) is one of the retinal vascular diseases in diabetic retinopathy (DR) patients. As there is no comprehensive detection technique to recognize CNP areas, we proposed a different method for computing detection of ischemic retina, non-perfused (NP) regions, in fundus fluorescein angiogram (FFA) images. Methods: Whilst major vessels appear as ridges, non-perfused areas are usually observed as ponds that are surrounded by healthy capillaries in FFA images. A new technique using homomorphic filtering to correct light illumination and detect the ponds surrounded in healthy capillaries on FFA images was designed and applied on DR fundus images. These images were acquired from the diabetic patients who had referred to the Nikookari hospital and were diagnosed for diabetic retinopathy during one year. Our strategy was screening the whole image with a fixed window size, which is small enough to enclose areas with identified topographic characteristics. To discard false nominees, we also performed a thresholding operation on the screen and marked images. To validate its performance we applied our detection algorithm on 41 FFA diabetic retinopathy fundus images in which the CNP areas were manually delineated by three clinical experts. Results: Lesions were found as smooth regions with very high uniformity, low entropy, and small intensity variations in FFA images. The results of automated detection method were compared with manually marked CNP areas so achieved sensitivity of 81%, specificity of 78%, and accuracy of 91%.The result was present as a Receiver operating character (ROC) curve, which has an area under the curve (AUC) of 0.796 with 95% confidence intervals. Conclusion: This technique introduced a new automated detection algorithm to recognize non-perfusion lesions on FFA. This has potential to assist detecting and managing of ischemic retina and may be incorporated into automated grading diabetic retinopathy structures. PMID:26929922

  10. A fluorescein tracer release experiment in the hydrothermally active crater of Vailulu'u volcano, Samoa

    NASA Astrophysics Data System (ADS)

    Hart, S. R.; Staudigel, H.; Workman, R.; Koppers, A. A. P.; Girard, A. P.

    2003-08-01

    On 3 April 2001, a 20 kg point source of fluorescein dye was released 30 m above the bottom of the active summit caldera of Vailulu'u submarine volcano, Samoa. Vailulu'u crater is 2000 m wide and at water depths of 600-1000 m, with the bottom 200 m completely enclosed; it thus provides an ideal site to study the hydrodynamics of an active hydrothermal system. The magmatically driven hydrothermal system in the crater is currently exporting massive amounts of particulates, manganese, and helium. The dispersal of the dye was tracked for 4 days with a fluorimeter in tow-yo mode from the U.S. Coast Guard icebreaker Polar Sea. Lateral dispersion of the dye ranged from 80 to 500 m d-1; vertical dispersion had two components: a diapycnal diffusivity component averaging 21 cm2 s-1, and an advective component averaging 0.025 cm s-1. These measurements constrain the mass export of water from the crater during this period to be 8-1.3+4.6 × 107 m3 d-1, which leads to a "turnover" time for water in the crater of ˜3.2 days. Coupled with temperature data from CTD profiles and Mn analyses of water samples, the power output from the crater is 610-100+350 MW, and the manganese export flux is ˜240 kg d-1. The Mn/Heat ratio of 4.7 ng J-1 is significantly lower than ratios characteristic of hot smokers and diffuse hydrothermal flows on mid-ocean ridges and points to phase separation processes in this relatively shallow hydrothermal system.

  11. Role of alkoxyl radicals on the fluorescein-based ORAC (Oxygen Radical Absorbance Capacity) assay.

    PubMed

    Dorta, E; Atala, E; Aspee, A; Speisky, H; Lissi, E; Lopez-Alarcon, C

    2014-10-01

    During the last decades the ORAC (Oxygen Radical Absorbance Capacity) assay has been widely employed to evaluate the in vitro antioxidant capacity of polyphenol-rich fruits, vegetables and beverages. The method employs fluorescein (FLH) as target molecule and AAPH (2,2'-azo-bis(2-amidinopropane)dihydrochloride) as the source of peroxyl radicals (ROO•). The protection of FLH, afforded by antioxidants (XH), is often characterized by kinetic profiles with clear lag times (LT), which are directly associated with the stoichiometry (n) of the XH-ROO• reaction. However, even for simple phenolic compounds, the LT measured imply large n values (defined as the number of ROO• moles trapped by each antioxidant molecule) which cannot be explained by a simple reaction mechanism. Nonetheless, they can be explained when considering the formation of alkoxyl radicals (RO•) from the recombination of two AAPH-derived ROO•. In the present work, we provide kinetic data showing that, in the zero order kinetic limit of FLH consumption, there is a low reaction rate incompatible with total trapping of ROO•. Thus, the consumption of FLH should be mostly related to its reaction with RO•. In addition, we present data regarding the assumption that in competitive measurements, the LT is due to efficient trapping of the ROO• by the added phenols, leading to high n values (1.7 to 23) for mono and polyphenols. These values are not in agreement with kinetic studies of the antioxidant consumption mediated by the presence of AAPH carried out by HPLC-DAD technique, which imply a competition by RO•. The results suggest that the use of FLH as probe at low concentrations give, for several antioxidants, ORAC values mainly related to their reaction towards RO• radicals instead of primary ROO•radicals. PMID:26461359

  12. A selective reaction of fructose bisphosphate aldolase with fluorescein isothiocyanate in chicken muscle extracts.

    PubMed

    Gehring, Andrew G; Ezzell, John L; Lebherz, Herbert G

    2008-01-01

    The present work describes the selective covalent modification of fructose bisphosphate aldolase in crude extracts of chicken breast muscle by fluorescein 5'-isothiocyanate (5'-FITC) at pH 7.0 and 35 degrees C. The modification was observed after 1 min while no other major soluble protein was labeled even after 30 min. We calculated that ca. one 5'-FITC molecule was incorporated into each aldolase tetramer after a 30 min reaction which resulted in a minimal loss of enzyme activity. The "native" structure of aldolase was required for the selective modification by 5'-FITC since high pH, high temperature, and ionic detergents either inhibited or prevented the reaction of 5'-FITC with aldolase. Certain metabolites (ATP, ADP, CTP, GTP, FBP) and erythrosin B also inhibited the 5'-FITC modification of aldolase. In contrast, F-6-P, AMP, NADH, and NAD(+) as well as free lysine and most importantly, the 6'-isomer of FITC exhibited no competition with 5'-FITC for the labeling of aldolase. Alone, the 6'-isomer of FITC did not exhibit preferential reaction when combined with aldolase. 5'-FITC-labeled and -unlabeled aldolases were not distinguished by their ability to bind to muscle myofibrils (MFs) or by their abilities to refold following reversible denaturation in urea. Structural analysis revealed that 5'-FITC-labeled a tryptic peptide corresponding to residues 112-134 in the primary structure of aldolase, a peptide that does not contain lysine, the amino acid believed to be the primary target of this reagent. Unlike chicken and rabbit muscle aldolases, chicken brain and liver aldolase isoforms along with several other aldolases derived from diverse biological sources did not exhibit this highly selective modification by 5'-FITC. PMID:18438970

  13. Several staining techniques to enhance the visibility of Acanthamoeba cysts.

    PubMed

    El-Sayed, Nagwa Mostafa; Hikal, Wafaa Mohamed

    2015-03-01

    Acanthamoeba is one of the most common free-living amoebae. It is widespread in the environment and can infect humans causing keratitis. Delayed diagnosis or misdiagnosis leads to extensive corneal inflammation and profound visual loss. Therefore, accurate and rapid diagnosis of Acanthamoeba keratitis is essential for successful treatment and good prognosis. This study was designed to use different staining techniques to facilitate the identification of Acanthamoeba cysts. Acanthamoeba cysts were isolated by cultivation of either corneal scraping specimens or tap water samples onto non-nutrient agar plates seeded with Escherichia coli. Subcultures were done from positive cultures until unique cysts were isolated. Acanthamoeba cysts were stained temporarily using iodine, eosin, methylene blue, and calcofluor white (CFW) stains and as permanent slides after processing for mounting using modified trichrome, Gimenez and Giemsa staining. These stains were compared on the basis of staining quality including clarity of morphological details, differentiation between cytoplasm and nuclei, color and contrast, and also other characteristics of the staining techniques, including ease of handling, time taken for the procedure, and cost effectiveness. The cysts of Acanthamoeba were recognized in the form of double-walled cysts: the outer wall (ectocyst) that was being differentiated from the variably stained surrounding background and the inner wall (endocyst) that was sometimes stellated, polygonal, round, or oval and visualized as separate from the spherical, sometimes irregular, outline of the ectocyst. Regarding the temporary stains, it was found that they were efficient for visualizing the morphological details of Acanthamoeba cysts. In CFW staining, Acanthamoeba cysts appeared as bluish-white or turquoise oval halos although the internal detail was not evident. On the other hand, the results of permanent-stained slides showed the most consistent stain for identification of Acanthamoeba cysts was modified trichrome followed by Gimenez stain and lastly Giemsa stain that gave poor visibility of Acanthamoeba cysts due to the intense staining background and monochrome staining of parasite. In the present study, multi-attribute ranking of the used staining techniques showed the highest rank for iodine stain (92 %) followed by eosin stain (84 %), Gimenez stain (76 %), methylene blue (72 %), CFW (64 %), modified trichrome (56 %), and the least was Giemsa stain (44 %). In conclusion, the staining techniques enhance the overall visibility of Acanthamoeba cysts. PMID:25346196

  14. Staining Protocols for Human Pancreatic Islets

    PubMed Central

    Campbell-Thompson, Martha L.; Heiple, Tiffany; Montgomery, Emily; Zhang, Li; Schneider, Lynda

    2012-01-01

    Estimates of islet area and numbers and endocrine cell composition in the adult human pancreas vary from several hundred thousand to several million and beta mass ranges from 500 to 1500 mg 1-3. With this known heterogeneity, a standard processing and staining procedure was developed so that pancreatic regions were clearly defined and islets characterized using rigorous histopathology and immunolocalization examinations. Standardized procedures for processing human pancreas recovered from organ donors are described in part 1 of this series. The pancreas is processed into 3 main regions (head, body, tail) followed by transverse sections. Transverse sections from the pancreas head are further divided, as indicated based on size, and numbered alphabetically to denote subsections. This standardization allows for a complete cross sectional analysis of the head region including the uncinate region which contains islets composed primarily of pancreatic polypeptide cells to the tail region. The current report comprises part 2 of this series and describes the procedures used for serial sectioning and histopathological characterization of the pancreatic paraffin sections with an emphasis on islet endocrine cells, replication, and T-cell infiltrates. Pathology of pancreatic sections is intended to characterize both exocrine, ductular, and endocrine components. The exocrine compartment is evaluated for the presence of pancreatitis (active or chronic), atrophy, fibrosis, and fat, as well as the duct system, particularly in relationship to the presence of pancreatic intraductal neoplasia4. Islets are evaluated for morphology, size, and density, endocrine cells, inflammation, fibrosis, amyloid, and the presence of replicating or apoptotic cells using H&E and IHC stains. The final component described in part 2 is the provision of the stained slides as digitized whole slide images. The digitized slides are organized by case and pancreas region in an online pathology database creating a virtual biobank. Access to this online collection is currently provided to over 200 clinicians and scientists involved in type 1 diabetes research. The online database provides a means for rapid and complete data sharing and for investigators to select blocks for paraffin or frozen serial sections. PMID:22665223

  15. Ultrafast tissue staining with chemical tags

    PubMed Central

    Kohl, Johannes; Ng, Julian; Cachero, Sebastian; Ciabatti, Ernesto; Dolan, Michael-John; Sutcliffe, Ben; Tozer, Adam; Ruehle, Sabine; Krueger, Daniel; Frechter, Shahar; Branco, Tiago; Tripodi, Marco; Jefferis, Gregory S. X. E.

    2014-01-01

    Genetically encoded fluorescent proteins and immunostaining are widely used to detect cellular and subcellular structures in fixed biological samples. However, for thick or whole-mount tissue, each approach suffers from limitations, including limited spectral flexibility and lower signal or slow speed, poor penetration, and high background labeling, respectively. We have overcome these limitations by using transgenically expressed chemical tags for rapid, even, high-signal and low-background labeling of thick biological tissues. We first construct a platform of widely applicable transgenic Drosophila reporter lines, demonstrating that chemical labeling can accelerate staining of whole-mount fly brains by a factor of 100. Using viral vectors to deliver chemical tags into the mouse brain, we then demonstrate that this labeling strategy works well in mice. Thus this tag-based approach drastically improves the speed and specificity of labeling genetically marked cells in intact and/or thick biological samples. PMID:25157152

  16. Red food coloring stain: new, safer procedures for staining nematodes in roots and egg masses on root surfaces.

    PubMed

    Thies, Judy A; Merrill, Sharon B; Corley, E Luther

    2002-06-01

    Acid fuchsin and phloxine B are commonly used to stain plant-parasitic nematodes in roots and egg masses on root surfaces, respectively. Both stains can be harmful to both the user and the environment and require costly waste disposal procedures. We developed safer methods to replace both stains using McCormick Schilling red food color. Eggs, juveniles, and adults of Meloidogyne incognita stained in roots with red food color were equally as visible as those stained with acid fuchsin. Egg masses stained with red food color appeared as bright-red spheres on the root surfaces and were highly visible even without magnification. Replacement of acid fuchsin and phloxine B with red food color for staining nematodes is safer for the user and the environment, and eliminates costly waste disposal of used stain solutions. PMID:19265929

  17. Centrifuge-operated specimen staining method and apparatus

    NASA Technical Reports Server (NTRS)

    Clarke, Mark S. F. (Inventor); Feeback, Daniel L. (Inventor)

    1999-01-01

    A method of staining preselected, mounted specimens of either biological or nonbiological material enclosed within a staining chamber where the liquid staining reagents are applied and removed from the staining chamber using hypergravity as the propelling force. In the preferred embodiment, a spacecraft-operated centrifuge and method of diagnosing biological specimens while in orbit, characterized by hermetically sealing a shell assembly. The assembly contains slide stain apparatus with computer control therefor, the operative effect of which is to overcome microgravity, for example on board an International Space Station.

  18. Automated single-slide staining device. [in clinical bacteriology

    NASA Technical Reports Server (NTRS)

    Wilkins, J. R.; Mills, S. M.

    1975-01-01

    An automatic single-slide Gram staining device is described. A timer-actuated solenoid controls the dispensing of gentian violet, Gram iodine solution, decolorizer, and 1% aqueous safranin in proper sequence and for the time required for optimum staining. The amount of stain or reagent delivered is controlled by means of stopcocks below each solenoid. Used stains and reagents can be flushed automatically or manually. Smears Gram stained automatically are equal in quality to those prepared manually. The time to complete one Gram cycle is 4.80 min.

  19. Three-dimensional structure, binding, and spectroscopic characteristics of the monoclonal antibody 43.1 directed to the carboxyphenyl moiety of fluorescein.

    PubMed

    Gayda, Susan; Longenecker, Kenton L; Judge, Russell A; Swift, Kerry M; Manoj, Sharmila; Linthicum, D Scott; Tetin, Sergey Y

    2016-04-01

    Unlike other known anti-fluorescein antibodies, the monoclonal antibody 43.1 is directed toward the fluorescein's carboxyl phenyl moiety. It demonstrates a very high affinity (KD ∼ 70 pM) and a fast association rate (kon ∼ 2 × 10(7) M(-1 ) s(-1) ). The three-dimensional structure of the Fab 43.1-fluorescein complex was resolved at 2.4 Å resolution. The antibody binding site is exclusively assembled by the CDR loops. It is comprised of a 14 Å groove-shaped entrance leading to a 9 Å by 7 Å binding pocket. The highly polar binding pocket complementary encloses the fluorescein's carboxyphenyl moiety and tightly fixes it by multiple hydrogen bonds. The fluorescein's xanthene ring is embedded in the more hydrophobic groove and stacked between the side chains of Tyr37L and of Arg99H providing conditions for an excited state electron transfer process. In comparison to fluorescein, the absorption spectrum of the complex in the visible region is shifted to the "red" by 23 nm. The complex demonstrates a very weak fluorescence (Φc  = 0.0018) with two short lifetime components: 0.03 ns (47%) and 0.8 ns (24%), which reflects a 99.8% fluorescein emission quenching effect upon complex formation. The antibody 43.1 binds fluorescein with remarkable affinity, fast association rate, and strongly quenches its emission. Therefore, it may present a practical interest in applications such as molecular sensors and switches. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 234-243, 2016. PMID:26756394

  20. Protein gel staining methods: an introduction and overview.

    PubMed

    Steinberg, Thomas H

    2009-01-01

    Laboratory scientists who encounter protein biochemistry in many of its myriad forms must often ask: is my protein pure? The most frequent response: run a denaturing SDS polyacrylamide gel. Running this gel raises another series of considerations regarding detection, quantitation, and characterization and so the next questions invariably center on suitable protein gel staining and detection methods. A total protein profile can be determined with the colorimetric methods embodied in Coomassie Blue and silver staining methods, or increasingly, with fluorescent stains. Protein quantitation can be done following staining, with fluorescence- and instrumentation-based methods offering the greatest sensitivity and linear dynamic range. Protein posttranslational modifications such as phosphorylation and glycosylation can be reliably determined with several fluorescence-based protocols. Staining and detection with two or more different stains can be done in series to establish relative profiles of modified versus total protein or to assess purity at two levels of quantitative sensitivity. The choice of staining method and protocol depends on the required rigor of detection and quantitation combined with available instrumentation and documentation capabilities. Other considerations for staining methods include intended downstream analytical procedures such as mass spectrometry or peptide sequencing, which preclude some methods. Nonfixative staining methods allow western blotting after gel staining. Laboratory custom and budget or intellectual curiosity may be the ultimate determinate of the chosen gel staining protocol. PMID:19892191

  1. Factors relating to dental stain formation in the rat.

    PubMed

    McDonald, J L; Schemehorn, B R; Stookey, G K

    1985-05-01

    A series of studies was conducted to investigate the use of the rat as an in vivo model for studies of dental stain and to identify dietary factors which influence stain formation in this model. It was determined that appreciable amounts of stain formed on the molar teeth of rats provided a synthetic diet containing lactalbumin, and the amount of stain increased throughout a four-week test period. Stain formation was also observed when rats were provided their diet by gastric intubation. Topical applications of chlorhexidine generally resulted in an increase in stain formation, as did the presence of tea in the drinking water. These studies support the use of the rat for investigations of dental stain. PMID:3858301

  2. Eosin Y staining of proteins in polyacrylamide gels.

    PubMed

    Lin, F; Fan, W; Wise, G E

    1991-08-01

    A staining method is described in which various proteins in polyacrylamide gels can be stained by using eosin Y. After a brief incubation of a polyacrylamide gel in an acidic solution of 1% eosin Y, various proteins, including human erythrocyte membrane sialoglycoproteins which are not detectable by Coomassie blue R-250 (CB), can be detected with a sensitivity of 10 ng protein. This is far more sensitive than CB staining and is comparable to the sensitivity of silver staining. In a Western blot, the antigenicity of an eosin Y stained protein is retained. In addition, proteins on an immunoblot sheet can be detected by eosin Y staining. The method described is rapid, sensitive, and reproducible with various proteins in polyacrylamide gels and has the added advantage of also staining sialoglycoproteins. PMID:1723249

  3. Fundus fluorescein angiographic findings in patients who underwent ventricular assist device implantation.

    PubMed

    Ozturk, Taylan; Nalcaci, Serhad; Ozturk, Pelin; Engin, Cagatay; Yagdi, Tahir; Akkin, Cezmi; Ozbaran, Mustafa

    2013-09-01

    Disruption of microcirculation in various tissues as a result of deformed blood rheology due to ventricular assist device (VAD) implantation causes novel arteriovenous malformations. Capillary disturbances and related vascular leakage in the retina and choroidea may also be seen in patients supported by VADs. We aimed to evaluate retinal vasculature deteriorations after VAD implantation. The charts of 17 patients who underwent VAD implantation surgery for the treatment of end-stage heart failure were retrospectively reviewed. Eight cases (47.1%) underwent pulsatile pump implantation (Berlin Heart EXCOR, Berlin Heart Mediprodukt GmbH, Berlin, Germany); however, nine cases (52.9%) had continuous-flow pump using centrifugal design (HeartWare, HeartWare Inc., Miramar, FL, USA). Study participants were selected among the patients who had survived with a VAD for at least 6 months, and results of detailed ophthalmologic examinations including optic coherence tomography (OCT) and fundus fluorescein angiography (FA) were documented. All of the 17 patients were male, with a mean age of 48.5 ± 14.8 years (15-67 years). Detailed ophthalmologic examinations including the evaluation of retinal vascular deteriorations via FA were performed at a mean of 11.8 ± 3.7 months of follow-up (6-18 months). Mean best-corrected visual acuity and intraocular pressure were found as logMAR 0.02 ± 0.08 and 14.6 ± 1.9 mm Hg, respectively in the study population. Dilated fundoscopy revealed severe focal arteriolar narrowing in two patients (11.8%), and arteriovenous crossing changes in four patients (23.5%); however, no pathological alteration was present in macular OCT scans. In patients with continuous-flow blood pumps, mean arm-retina circulation time (ARCT) and arteriovenous transit time (AVTT) were found to be 16.8 ± 3.0 and 12.4 ± 6.2 s, respectively; whereas those with pulsatile-flow blood pumps were found to be 17.4 ± 3.6 and 14.0 ± 2.1 s in patients (P=0.526 and P=0.356, respectively). FA also revealed a tendency for increased frequency of dye leakage from the optic disc in our study population. Except for remarkable delays in both ARCT and AVTT as well as a tendency for increased frequency of dye leakage from the optic disc, ophthalmologic evaluations revealed no other significant pathology or vascular deterioration in the retina that could be attributed to artificial heart systems. PMID:23826834

  4. Pre-embedding immunogold labeling of TUNEL stain enables evaluation of DNA strand breaks and ultrastructural alterations in individual cells of neuronal tissue.

    PubMed

    Barth, Martin; Oulmi, Yasmina; Ehrenreich, Hannelore; Schilling, Lothar

    2002-12-01

    In the brain apoptosis may occur as a physiological phenomenon during periods of programmed cell death as well as under pathological conditions such as ischemia, trauma, tumor, and degenerative diseases. While the definition of apoptotic cell death was originally based on ultrastructural alterations, the detection of DNA double-strand breaks has become an important feature in studies of apoptosis. Currently, the terminal transferase-mediated dUTP nick-end labeling (TUNEL) procedure is widely used for detection of apoptotic cell death. However, there is a growing body of evidence to suggest that the TUNEL staining does not label apoptotic alterations exclusively. Therefore, a new staining procedure was developed combining TUNEL methodology with pre-embedding nanogold labeling to detect DNA double-strand breaks in individual cells by electron microscopy and assess the accompanying ultrastructural alterations. In vitro DNAse-treated vibratome sections (thickness, 20 micro m) from normal adult rat brains were used to develop the staining procedure consisting of the following steps: (i) TUNEL staining of free-floating vibratome sections using fluorescein isothiocyanate (FITC)-labeled UTP, (ii) conversion of the fluorescence signal into an electron-dense signal using an anti-FITC antibody coupled with ultrasmall (diameter, 0.8 nm) gold particles followed by silver enhancement, and (iii) osmification, embedding in Spurr resin and cutting of ultrathin sections. Early postnatal brain tissue was used to study physiologically occurring apoptotic cell death. Under these conditions different patterns of gold staining were observed probably representing different states of cellular decay along the apoptotic avenue. Severe focal brain ischemia was studied as a pathological situation in which intense TUNEL staining occurs. Under these conditions TUNEL labeling of cells was regularly observed in conjunction with ultrastructural alterations indicative of necrosis. These results suggest that under pathological conditions apoptosis and necrosis are not mutually exclusive mechanisms but rather may occur concurrently along a continuum in which cell death occurs. PMID:12410384

  5. Copper staining: a five-minute protein stain for sodium dodecyl sulfate-polyacrylamide gels.

    PubMed

    Lee, C; Levin, A; Branton, D

    1987-11-01

    We present a new method for visualizing proteins electrophoresed in sodium dodecyl sulfate-polyacrylamide gels. After electrophoresis, gels are incubated in CuCl2 to produce a negative image of colorless protein bands against a semiopaque background. Gels are stained completely within 5 min, do not require destaining, and can be stored indefinitely without loss of the image. Because proteins are not permanently fixed within the gel, they can be quantitatively eluted after chelation of Cu with EDTA. The sensitivity of the CuCl2 stain falls between that of Coomassie blue and silver. We anticipate that CuCl2 will be useful in the rapid analysis of proteins by polyacrylamide gel electrophoresis and in the preparation of purified polypeptides by elution from gel slices. PMID:2449094

  6. Electrostatic control of the coffee stain effect

    NASA Astrophysics Data System (ADS)

    Wray, Alex; Papageorgiou, Demetrios; Sefiane, Khellil; Matar, Omar

    2013-11-01

    The ``coffee stain effect,'' as first explained by Deegan et al. 1997, has received a great deal of attention amongst modellers and experimentalists in recent years, perhaps due in part to its obvious casual familiarity. However, it maintains interest because of its intriguing reliance on an interplay of a trio of effects: contact line pinning, inhomogeneous mass flux, and resulting capillarity-driven flow. What is more, the effect, and especially its suppression or reversal, find applications in fields as diverse as sample recovery, mass spectroscopy and the printing of Organic LEDs. We examine the motion a nanoparticle-laden droplet deposited on a precursor film, incorporating the effects of capillarity, concentration-dependent rheology, together with a heated substrate and resultant mass flux and Marangoni effects. We allow the substrate to act as an electrode and incorporate a second electrode above the droplet. The potential difference together with a disparity in electrical properties between the two regions results in electrical (Maxwell) stresses at the interface. We show via lubrication theory and via direct numerical simulations that the ring effect typically observed may be suppressed or augmented via appropriate use of electric fields. EPSRC DTG

  7. Protein staining methods in quantitative cytochemistry.

    PubMed

    Tas, J; van der Ploeg, M; Mitchell, J P; Cohn, N S

    1980-08-01

    The chemical action and practical application of the Naphthol Yellow S, Alkaline Fast Green, Coomassie Brilliant Blue, Dinitrofluorobenzene and some lesser known protein staining methods have been surveyed with respect to their potentialities for quantitative cytochemical analyses. None of the dyes can be said to bind to any specific protein or group of proteins, but each may be used to analyse the presence of one or more particular amino acid residues. For the cytophotometric measurement of the 'total protein content' of individual cells and cell organelles the covalent binding Dinitrofluorobenzene and the electrostatic binding Naphthol Yellow S can properly be used. Fast Green FCF, applied at alkaline pH, binds electrostatically to the basic amino acid side chains of strongly basic proteins only but not in a quantitative (stoichiometrical) way. Coomassie Brilliant Blue, recently introduced to protein cytochemistry, may be useful for quantitative purposes. The combined Feulgen-Pararosaniline(SO2)/Naphthol Yellow S and Dinitrofluorobenzene/Feulgen-Pararosaniline(SO2) methods enable the simultaneous cytophotometric analysis at two different wavelengths for protein and DNA within the same microscopical preparation. PMID:6157816

  8. Standard specimens for stain calibration and their application to the Papanicolaou stain.

    PubMed

    Turner, J N; Collins, D N

    1987-12-01

    Standardized specimens composed of extracts of biologic objects (nucleoprotamine and bovine liver) were developed as tools for the quantitative evaluation of stain performance on biologic substrates. The specimens are mixtures of proteins and nucleic acids and thus mimic the staining characteristics of cytologic smears. The concentration of each mixture and the specimen thickness can be precisely controlled, ensuring the production of a large number of samples with a nearly identical capability for dye binding. The transmitted light spectra of the standardized specimens varied depending on the extract and the preparation conditions. Spectra similar to those reported from the nuclei and cytoplasm of cell types in Papanicolaou-stained cervicovaginal smears were observed. Light transmission was uniform to +/- 5% across each specimen and from specimen to specimen. The specimen thickness was uniform within +/- 2%. Studies with these standardized samples could reveal the much-needed correlations between the chemical and optical characteristics of dyes and dye solutions and the performance of the dyes on biologic substrates. PMID:2449228

  9. Multi-class stain separation using independent component analysis

    NASA Astrophysics Data System (ADS)

    Trahearn, Nicholas; Snead, David; Cree, Ian; Rajpoot, Nasir

    2015-03-01

    Stain separation is the process whereby a full colour histology section image is transformed into a series of single channel images, each corresponding to a given stain's expression. Many algorithms in the field of digital pathology are concerned with the expression of a single stain, thus stain separation is a key preprocessing step in these situations. We present a new versatile method of stain separation. The method uses Independent Component Analysis (ICA) to determine a set of statistically independent vectors, corresponding to the individual stain expressions. In comparison to other popular approaches, such as PCA and NNMF, we found that ICA gives a superior projection of the data with respect to each stain. In addition, we introduce a correction step to improve the initial results provided by the ICA coefficients. Many existing approaches only consider separation of two stains, with primary emphasis on Haematoxylin and Eosin. We show that our method is capable of making a good separation when there are more than two stains present. We also demonstrate our method's ability to achieve good separation on a variety of different stain types.

  10. IgG Subclass Staining in Routine Renal Biopsy Material.

    PubMed

    Hemminger, Jessica; Nadasdy, Gyongyi; Satoskar, Anjali; Brodsky, Sergey V; Nadasdy, Tibor

    2016-05-01

    Immunofluorescence staining plays a vital role in nephropathology, but the panel of antibodies used has not changed for decades. Further classification of immunoglobulin (Ig)G-containing immune-type deposits with IgG subclass staining (IgG1, IgG2, IgG3, and IgG4) has been shown to be of diagnostic utility in glomerular diseases, but their value in the evaluation of renal biopsies has not been addressed systematically in large renal biopsy material. Between January 2007 and June 2014, using direct immunofluorescence, we stained every renal biopsy for the IgG subclasses if there was moderate to prominent glomerular IgG staining and/or IgG-predominant or IgG-codominant glomerular staining. The total number of biopsies stained was 1084, which included 367 cases of membranous glomerulonephritis, 307 cases of lupus nephritis, 74 cases of fibrillary glomerulonephritis, 53 cases of proliferative glomerulonephritis with monoclonal IgG deposits, and 25 cases of antiglomerular basement membrane disease, among others. We found that monoclonality of IgG deposits cannot always be reliably determined on the basis of kappa and lambda light chain staining alone, particularly if concomitant (frequently nonspecific) IgM staining is present. In IgG heavy and heavy and light chain deposition disease (3 cases), subclass staining is very helpful, and in proliferative glomerulonephritis with monoclonal IgG deposits subclass staining is necessary. IgG subclass staining is useful in differentiating primary from secondary membranous glomerulonephritis. In proliferative glomerulonephritis with polyclonal IgG deposition, IgG1 dominance/codominance with concomitant IgG3 and IgG2 but weak or absent IgG4 staining favors an underlying autoimmune disease. IgG subclass staining is a very useful diagnostic method in a selected cohort of renal biopsies, particularly in biopsies with glomerulonephritis with monoclonal IgG deposits. PMID:26848798

  11. Cadmium-glutathione complex formation in human t-cell and b-cell lymphocytes after their incubation with organo-cadmium diacetate.

    PubMed

    Ullah, Hashmat; Khan, Muhammad Farid; Jan, Syed Umer; Hashmat, Farwa

    2015-11-01

    Cadmium intake is associated with oxidative stress that causes depletion of intracellular as well as extra cellular reduced glutathione. There is strong evidence indicating that reactive oxygen species and reactive nitrogen species generated in the presence of cadmium could be responsible for its toxic effects in many cells and tissues. Depletion of reduced glutathione in various cells, especially in T and B-lymphocytes, causes extreme damage to the antioxidant defense system of body. The aim of this research work was to investigate the metabolic changes that occur in T and B lymphocytes after their incubation with organ cadmium diacetate by using Ellman's spectrophotometric method of thiol quantification. The results of the present study indicate that cadmium depleted T and B lymphocytes GSH to a harmful extent. It is proposed that this depletion is due to the bivalent cadmium glutathione complex formation, oxidation of reduced glutathione (GSH) to its oxidized form, or both. PMID:26639500

  12. Effect of Sodium Fluorescein and Plating Medium on Recovery of Irradiated Escherichia coli and Serratia marcescens from Aerosols

    PubMed Central

    Dorsey, Emerson L.; Berendt, Richard F.; Neff, Everett L.

    1970-01-01

    Irradiation of aerosols of either Escherichia coli or Serratia marcescens with simulated solar (xenon) radiation caused a significant decrease in viability. When sodium fluorescein was employed to determine the physical loss of organisms from the aerosol, an additional adverse effect upon survival was noted. The decay curves indicated that at least two mechanisms of inactivation were operative, one due to aerosolization, the other to irradiation. After collection from aerosols, both species of microorganisms grew better on blood agar base than on Casitone agar, but this finding did not appear to be related to the effect of irradiation. PMID:4922085

  13. Asymmetric incorporation of (/sup 14/C)cyanate and of fluorescein isothiocyanate in mamillary body of conditioned rats

    SciTech Connect

    Burgal, M.; Montes, F.; Grisolia, S.

    1988-05-01

    A marked decrease in overall learning capacity has been observed in rats injected with cyanate. Therefore it was of interest to test whether learning influenced carbamylation of brain proteins. Incorporation of (/sup 14/C)cyanate into proteins of the mamillary body was selectively modified following operant conditioning of the rat, so that trained rats showed an asymmetric image with higher levels of incorporation in the right side than in the left side, as compared to control rats. These results were confirmed using fluorescein isothiocyanate. The asymmetry persisted once the learning had been well established.

  14. Synthesis and Purification of a Hammerhead Ribozyme and a Fluorescein-Labeled RNA Substrate. A Biochemistry Laboratory: Part 1

    NASA Astrophysics Data System (ADS)

    Chow, Christine S.; Somne, Smita

    1999-05-01

    The applications of in vitro transcription and chemical synthesis of RNA are discussed. This laboratory describes the in vitro synthesis of a 38-nucleotide hammerhead ribozyme and the synthesis of a 17-nucleotide fluorescein-labeled RNA substrate by using standard phosphoramidite methodologies, two widely used methods in modern RNA research. The synthesis and purification procedures outlined allow students to develop an understanding of RNA handling procedures, synthesis of modified nucleic acids, gel electrophoresis, visualization of RNA by nonradioactive techniques, and quantitation of nucleic acids. The RNAs that are synthesized have applications in biotechnology and medicine; thus the students gain access to current problems in chemical and clinical research.

  15. The blood-brain barrier penetration and distribution of PEGylated fluorescein-doped magnetic silica nanoparticles in rat brain

    SciTech Connect

    Ku, Shuting; Yan, Feng; Wang, Ying; Sun, Yilin; Yang, Nan; Ye, Ling

    2010-04-16

    PEGylated PAMAM conjugated fluorescein-doped magnetic silica nanoparticles (PEGylated PFMSNs) have been synthesized for evaluating their ability across the blood-brain barrier (BBB) and distribution in rat brain. The obtained nanoparticles were characterized by transmission electron microscopy (TEM), thermal gravimetry analyses (TGA), zeta potential ({zeta}-potential) titration, and X-ray photoelectron spectroscopy (XPS). The BBB penetration and distribution of PEGylated PFMSNs and FMSNs in rat brain were investigated not only at the cellular level with Confocal laser scanning microscopy (CLSM), but also at the subcellular level with transmission electron microscopy (TEM). The results provide direct evidents that PEGylated PFMSNs could penetrate the BBB and spread into the brain parenchyma.

  16. Use of thermography and fluorescein angiography in the management of a Chilean flamingo with avascular necrosis of the wing.

    PubMed

    Hurley-Sanders, Jennifer L; Bowman, Karl F; Wolfe, Barbara A; Nutter, Felicia B; Sladky, Kurt K; Stoskopf, Michael K

    2012-12-01

    A Chilean flamingo (Phoenicopterus chilensis) was presented to the veterinary clinic at the North Carolina Zoological Park for evaluation of acute weakness of the right wing. Results of a physical examination revealed a lack of a palpable pulse in the radial artery, which suggested occlusion or obstruction of the vessel. Radiography, thermography, and fluorescein angiography confirmed right wing injury and vascular compromise. Based on the poor prognosis for return to function associated with irreversible vascular damage, the wing was amputated. After a period of observation and treatment, the bird was returned to public exhibit. PMID:23409438

  17. Use of carboxyfluorescein diacetate succinimidyl ester (CFSE) dye and fluorescent imaging as an in situ method to visualize lymphoid tissues in egg-layer chickens challenged with Salmonella enterica serovar Enteritidis (SE)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Carboxyfluorescein diacetate succinimidyl ester (CFSE) vital dye has been used in leukocyte studies involving mice, rats, sheep, heifers, nonhuman primates, teleost fish and avian embryos. Mice and sheep appear to be the only animals that have received intravenous (IV) CFSE administration, and the ...

  18. Identifying different types of chromatin using Giemsa staining.

    PubMed

    Stockert, Juan C; Blázquez-Castro, Alfonso; Horobin, Richard W

    2014-01-01

    Mixtures of polychrome methylene blue-eosin Y (i.e., Giemsa stain) are widely used in biological staining. They induce a striking purple coloration of chromatin DNA (the Romanowsky-Giemsa effect), which contrasts with the blue-stained RNA-containing cytoplasm and nucleoli. After specific prestaining treatments that induce chromatin disorganization (giving banded or harlequin chromosomes), Giemsa staining produces a differential coloration, with C- and G-bands appearing in purple whereas remaining chromosome regions are blue. Unsubstituted (TT) and bromo-substituted (BT) DNAs also appear purple and blue, respectively. The same occurs in the case of BT and BB chromatids.In addition to discussing the use of Giemsa stain as a suitable method to reveal specific features of chromosome structure, some molecular processes and models are also described to explain Giemsa staining mechanisms of chromatin. PMID:24162977

  19. Manual hematoxylin and eosin staining of mouse tissue sections.

    PubMed

    Cardiff, Robert D; Miller, Claramae H; Munn, Robert J

    2014-06-01

    The hematoxylin and eosin (H&E) stain is the standard used for microscopic examination of tissues that have been fixed, processed, embedded, and sectioned. It can be performed manually or by automation. For economic reasons, the manual technique is generally the method of choice for facilities with a low sample volume. This protocol describes manual H&E staining of fixed, processed, paraffin-embedded, and sectioned mouse tissues. In H&E-stained tissues, the nucleic acids stain dark blue and the proteins stain red to pink or orange. For accurate phenotyping and delineation of tissue detail, the protocol must be adhered to rigorously. This includes frequent reagent changes as well as the use of "in-date" reagents. Appropriate color in a good H&E stain allows for identification of many tissue subtleties that are necessary for accurate diagnosis. PMID:24890205

  20. Factors influencing extract of Hibiscus sabdariffa staining of rat testes.

    PubMed

    Bassey, R B; Bakare, A A; Peter, A I; Oremosu, A A; Osinubi, A A

    2012-08-01

    Some plant extracts can be used in biology and medicine to reveal or identify cellular components and tissues. We investigated the effects of time and concentration on staining of histological sections of rat testes by an acidified extract of Hibiscus sabdariffa. An ethanolic extract of H. sabdariffa was diluted using 1% acetic acid in 70% ethanol to stain histological sections of testes at concentrations of 0.2, 0.1 and 0.05 g/ml for 5, 10, 15, 30, 45 and 60 min. The sections of testes were stained deep red. The staining efficiency of H. sabdariffa was greater at a high concentration and required less time to achieve optimal staining. H. sabdariffa is a strongly basic dye that can be used for various diagnostic purposes. Staining time and concentration must be considered to achieve optimal results. PMID:22548304

  1. Visible luminescence from silicon wafers subjected to stain etches

    NASA Technical Reports Server (NTRS)

    Fathauer, R. W.; George, T.; Ksendzov, A.; Vasquez, R. P.

    1992-01-01

    Etching of Si in a variety of solutions is known to cause staining. These stain layers consist of porous material similar to that produced by anodic etching of Si in HF solutions. In this work, photoluminescence peaked in the red from stain-etched Si wafers of different dopant types, concentrations, and orientations produced in solutions of HF:HNO3:H2O was observed. Luminescence is also observed in stain films produced in solutions of NaNO2 in HF, but not in stain films produced in solutions of CrO3 in HF. The luminescence spectra are similar to those reported recently for porous Si films produced by anodic etching in HF solutions. However, stain films are much easier to produce, requiring no special equipment.

  2. Silver staining of proteins in 2DE gels.

    PubMed

    Lelong, Cécile; Chevallet, Mireille; Luche, Sylvie; Rabilloud, Thierry

    2009-01-01

    Silver staining detects proteins after electrophoretic separation on polyacrylamide gels. Its main positive features are its excellent sensitivity (in the low nanogram range) and the use of very simple and cheap equipment and chemicals. The sequential phases of silver staining are protein fixation, then sensitization, then silver impregnation, and finally image development. Several variants of silver staining are described here, which can be completed in a time range from 2 h to 1 day after the end of the electrophoretic separation. Once completed, the stain is stable for several weeks. PMID:19381593

  3. Morphological changes in the lungs of meconium-stained piglets.

    PubMed

    Castro-Nájera, José A; Martínez-Burnes, Julio; Mota-Rojas, Daniel; Cuevas-Reyes, Humberto; López, Alfonso; Ramírez-Necoechea, Ramiro; Gallegos-Sagredo, Ramón; Alonso-Spilsbury, María

    2006-11-01

    Meconium staining of the skin is a common event associated with fetal hypoxia, stillbirths, weak-born piglets, and neonatal mortality. Aspiration of meconium leads to meconium aspiration syndrome (MAS). This study was undertaken to assess the relationship between the degree of meconium staining of the skin at birth, meconium aspiration, and pulmonary changes in porcine neonates. A total of 353 farrowing sows and 3,693 born piglets were monitored during parturition and for 15 days after delivery. Umbilical cords were classified as normal or ruptured. Meconium staining in the skin was graded as nonstained, mildly, moderately, and severely stained. Mortality from birth to 15 days of age was 8.4%. The lungs from 60 meconium-stained piglets and 60 lungs from nonstained piglets were collected and microscopically examined for meconium aspiration and inflammation. Rupture of the umbilical cord was significantly higher (P < 0.01) in meconium-stained piglets. Microscopically, 32% and 40% of the lungs had evidence of meconium for the stained and nonstained groups, respectively. The microscopic grade of meconium aspiration and inflammatory cells was not different between nonstained and meconium-stained piglets. Aspiration of meconium induced a granulomatous response in the lungs. It was concluded that the grade of meconium staining is a good indicator of fetal hypoxia, but not a good predictor for meconium aspiration and MAS in piglets. PMID:17121098

  4. A Method for Staining Nematode Secretions and Structures

    PubMed Central

    Premachandran, D.; Von Mende, N.; Hussey, R. S.; McClure, M. A.

    1988-01-01

    Secretions from amphids, phasmids, and excretory system were stained by incubating nematodes in 0.1% coomassie brilliant blue G-250 in 40% aqueous methanol containing 10% acetic acid on slides with coverslips sealed with nail polish or Zut. Nematodes incubated in this staining solution usually produced copious amounts of secretions from their amphids and excretory pore. Phasmids also stained dark blue, enabling them to be easily observed. Other biological dyes stained these secretions or were useful for differentiating specific morphological features of nematodes. PMID:19290186

  5. Eosin Y: a reversible stain for detecting electrophoretically resolved protein.

    PubMed

    Selsted, M E; Becker, H W

    1986-06-01

    A method for the rapid staining of polypeptides in low-pH, urea-containing polyacrylamide gels is described. After 30 s of immersion in an alkaline solution of eosin Y, a variety of proteins stained with intensities similar to those seen with Coomassie R-250. By subjecting eosin-stained bands to electrophoretic elution, the eosin-protein complex was dissociated. This allowed for recovery of milligram quantities of electrophoretically resolved, stain-free protein. High recoveries of total protein as well as enzymatic activity were achieved using hen egg white lysozyme as a model protein. PMID:2425660

  6. Functional expression of the 11 human Organic Anion Transporting Polypeptides in insect cells reveals that sodium fluorescein is a general OATP substrate.

    PubMed

    Patik, Izabel; Kovacsics, Daniella; Német, Orsolya; Gera, Melinda; Várady, György; Stieger, Bruno; Hagenbuch, Bruno; Szakács, Gergely; Özvegy-Laczka, Csilla

    2015-12-15

    Organic Anion Transporting Polypeptides (OATPs), encoded by genes of the Solute Carrier Organic Anion (SLCO) family, are transmembrane proteins involved in the uptake of various compounds of endogenous or exogenous origin. In addition to their physiological roles, OATPs influence the pharmacokinetics and drug-drug interactions of several clinically relevant compounds. To examine the function and molecular interactions of human OATPs, including several poorly characterized family members, we expressed all 11 human OATPs at high levels in the baculovirus-Sf9 cell system. We measured the temperature- and inhibitor-sensitive cellular accumulation of sodium fluorescein and fluorescein-methotrexate, two fluorescent substrates of the OATPs, OATP1B1 and 1B3. OATP1B1 and 1B3 were functional in Sf9 cells, showing rapid uptake (t1/2(fluorescein-methotrexate) 2.64 and 4.16 min, and t1/2(fluorescein) 6.71 and 5.58 min for OATP1B1 and 1B3, respectively) and high-affinity transport (Km(fluorescein-methotrexate) 0.23 and 0.53 μM, and Km(fluorescein) 25.73 and 38.55 μM for OATP1B1 and 1B3, respectively) of both substrates. We found that sodium fluorescein is a general substrate of all human OATPs: 1A2, 1B1, 1B3, 1C1, 2A1, 2B1, 3A1, 4A1, 4C1, 5A1 and 6A1, while fluorescein-methotrexate is only transported by 1B1, 1B3, 1A2 and 2B1. Acidic extracellular pH greatly facilitated fluorescein uptake by all OATPs, and new molecular interactions were detected (between OATP2B1 and Imatinib, OATP3A1, 5A1 and 6A1 and estradiol 17-β-d-glucuronide, and OATP1C1 and 4C1 and prostaglandin E2). These studies demonstrate, for the first time, that the insect cell system is suitable for the functional analysis of the entire human OATP family, and for drug-OATP interaction screening. PMID:26415544

  7. Comparison of Ultrafast Papanicolaou Stain with the Standard Papanicolaou Stain in Body Fluids and Fine Needle Aspiration Specimens

    PubMed Central

    Alwahaibi, Nasar Yousuf; Alsubhi, Mariam Said; Aldairi, Najat; Alshukaili, Amna; Bai, Usha Rani

    2016-01-01

    Introduction: Most cytology laboratories in all Gulf countries including Oman, use the standard papanicolaou (PAP) method to stain various cytological specimens. The aim of this study was to investigate the possible application of ultrafast PAP (UF-PAP) method in cytology laboratory. Materials and Methods: Samples from 46 patients containing 26 body fluids and 20 fine needle aspirations (FNAs) (9 thyroids and 11 breasts) were collected. Two air dried and two wet smears from each sample were prepared and stained by UF-PAP and the standard PAP stains, respectively. Background, nuclear staining, cell morphology, and overall staining were independently reviewed by two cytoscreeners. Results: In all cases of FNA, UF-PAP stain gave a good score for the background, nuclear staining, cell morphology, and overall staining when compared with the standard PAP method. Although the correct diagnosis was made in all cases of body cavity fluids cases except in one case, UF-PAP stain gave a fewer score in the assessment of body cavity fluid samples. Conclusion: The findings of this study support the use of UF-PAP method in cytology laboratory with a high emphasis on FNA samples. PMID:27013808

  8. Comparison of tetrachromic VOF stain to other histochemical staining techniques for characterizing stromal soft and hard tissue components.

    PubMed

    Belaldavar, C; Hallikerimath, S; Angadi, P V; Kale, A D

    2014-11-01

    The components of hard tissues including dentin, enamel, cementum, bone and other calcified deposits, and mature and immature collagen pose problems for identification in routine hematoxylin and eosin (H & E) stained sections. Use of combinations of stains can demonstrate the components of hard tissues and soft tissues distinctly. We assessed the efficacy of the Verde Luz-orange G-acid fuchsin (VOF) stain for differentiating hard and soft connective tissues and compared results with other histochemical staining techniques. Eighty tissue sections comprising developing tooth (30), ossifying fibroma (30) and miscellaneous pathologies (20) expected to contain varying types of calcified tissues were stained with H & E, VOF, and Masson's trichrome (MT). In developing tooth, VOF demonstrated better differentiation of hard tissues, while it was comparable to MT for ossifying fibroma and miscellaneous pathologies. The intensity of staining was greater with VOF than with the other stains studied. VOF stains hard tissue components distinctly and gives good contrast with the surrounding connective tissue. VOF is comparable to MT, but has added advantages including single step staining, rapid and easy procedures, and it distinguishes the maturity of the tissues. PMID:24830362

  9. Membrane filtration-fluorescent antibody staining procedure for detecting and quantifying Renibacterium salmoninarum in coelomic fluid of Chinook salmon Oncorhynchus tshawytscha)

    USGS Publications Warehouse

    Elliott, D.G.; Barila, T.Y.

    1988-01-01

    e developed a rapid method for detecting and quantifying the pathogen Renibacterium salmoninarum in coelomic fluid of spring chinook salmon (Oncorhynchus tshawytscha) by concentrating the bacteria on 0.2-μm polycarbonate filters and staining them with specific fluorescein-labeled antibody. Centrifugation of samples and resuspension of the sedimented material in phosphate-buffered saline containing Triton X-100 increased the ease of filtration. Background fluorescence was reduced by counterstaining filters with Eriochrome black T. Postfiltration staining, rinsing, and counterstaining were done in the syringe-mounted filter holders, reducing handling of the filters and possible loss of bacteria. The number of bacteria detected by the filtration – fluorescent antibody technique in a broth culture of R. salmoninarum ranged from 6.7 × 107to7.6 × 107/mL and was slightly higher than that determined by plate count (9.6 × 106/mL). Increasing the sample dilution or decreasing the number of microscope fields examined generally increased the variability of filter counts of R. salmoninarum. Using the filtration – fluorescent antibody technique, we detected the bacterium in the coelomic fluid of 85% of spawning female spring chinook salmon sampled from a hatchery population. Membrane Filtration – Fluorescent Antibody Staining Procedure for Detecting and Quantifying Renibacterium salmoninarum in Coelomic Fluid of Chinook Salmon (oncorhynchus tshawytscha) (PDF Download Available). 

  10. A blue fluorescent labeling technique utilizing micro- and nanoparticles for tracking in LIVE/DEAD® stained pathogenic biofilms of Staphylococcus aureus and Burkholderia cepacia

    PubMed Central

    Klinger-Strobel, Mareike; Ernst, Julia; Lautenschläger, Christian; Pletz, Mathias W; Fischer, Dagmar; Makarewicz, Oliwia

    2016-01-01

    Strategies that target and treat biofilms are widely applied to bacterial cultures using popular live/dead staining techniques with mostly red or green fluorescent markers (eg, with SYTO® 9, propidium iodide, fluorescein). Therefore, visualizing drugs or micro- and nanoparticulate delivery systems to analyze their distribution and effects in biofilms requires a third fluorescent dye that does not interfere with the properties of the live/dead markers. The present study establishes and evaluates a model for tracking polymeric particles in fluorescently stained biological material. To this end, poly(d,l-lactide-co-glycolide) (PLGA)-based micro- and nanoparticles were used as well-established model systems, which, because of their favorable safety profiles, are expected to play important future roles with regard to drug delivery via inhalation. PLGA was covalently and stably labeled with 7-amino-4-methyl-3-coumarinylacetic acid (AMCA), after which blue fluorescent poly(ethylene glycol)-block-PLGA (PEG-PLGA) particles were prepared using a mixture of fluorescent AMCA-PLGA and PEG-PLGA. Because chitosan is known to reduce negative surface charge, blue fluorescent PEG-PLGA-particles with chitosan were also prepared. These micro- and nanoparticles were physicochemically characterized and could be clearly distinguished from live/dead stained bacteria in biofilms using confocal laser scanning microscopy. PMID:26917959

  11. Ultrastructural staining with sodium metaperiodate and sodium borohydride.

    PubMed

    Lobo, Maria V T; Alonso, F Javier M; Arenas, Maria I; Caso, Enrique; Fraile, Benito; del Río, Rafael Martín

    2002-01-01

    This article describes new ultrastructural staining methods for osmicated tissues based on the incubation of sections with sodium metaperiodate and sodium borohydride solutions before uranyl/lead staining. Sections incubated with sodium metaperiodate and sodium borohydride, treated with Triton X-100, and stained with ethanolic uranyl acetate/lead citrate showed a good contrast for the nucleolus and the interchromatin region, whereas the chromatin masses were bleached. Chromatin bleaching depended on the incubation with these oxidizing (metaperiodate) and reducing (borohydride) agents. Other factors that influenced the staining of the chromatin masses were the en bloc staining with uranyl acetate, the incubation of sections with Triton X-100, and the staining with aqueous or ethanolic uranyl acetate. The combination of these factors on sections treated with metaperiodate/borohydride provided a different appearance to the chromatin, from bleached to highly contrasted. Most cytoplasmic organelles showed a similar appearance with these procedures than with conventional uranyl/lead staining. However, when sections were incubated with metaperiodate/borohydride and Triton X-100 before uranyl/lead staining, the collagen fibers, and the glycocalix and zymogen granules of pancreatic acinar cells, appeared bleached. The possible combination of these methods with the immunolocalization of the amino acid taurine was also analyzed. (J Histochem Cytochem 50:11-19, 2002) PMID:11748290

  12. OBSERVATION OF SALMONELLA TYPHIMURIUM FIMBRIAE BY NEGATIVE STAIN

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Washed and unwashed overnight cultures of Salmonella typhimurium were examined for the expression of fimbriae using negative stain. In the course of the evaluation, it was noted that the distribution of bacteria on formvar coated grids was dependent on the negative stain utilized for visualization....

  13. 7 CFR 3201.87 - Wood and concrete stains.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 15 2013-01-01 2013-01-01 false Wood and concrete stains. 3201.87 Section 3201.87... Designated Items § 3201.87 Wood and concrete stains. (a) Definition. Products that are designed to be applied as a finish for concrete and wood surfaces and that contain dyes or pigments to change the...

  14. 7 CFR 3201.87 - Wood and concrete stains.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 15 2014-01-01 2014-01-01 false Wood and concrete stains. 3201.87 Section 3201.87... Designated Items § 3201.87 Wood and concrete stains. (a) Definition. Products that are designed to be applied as a finish for concrete and wood surfaces and that contain dyes or pigments to change the...

  15. Dynamic staining of bacteria at a single-cell level

    NASA Astrophysics Data System (ADS)

    Nuñez, Vicente; Upadhyayula, Srigokul; Lin, Adam; Chau, Kenny; Vullev, Valentine I.

    2011-05-01

    Bacterial infectious diseases remain one of the major health hazards nation- and worldwide. The expedience of detection and identification of bacterial pathogens determines how early the diagnosis is, and hence, what the treatment and the outcome of the illness would be. As we have previously reported, the dynamics of fluorescence staining provides venues for the development of expedient assays for detection and identification of bacterial species[1]. We measured the kinetics of bacterial staining with cyanine and thioflavin dyes and investigated their photophysical properties. We demonstrated that the pseudo first-order kinetic constants of the fluorescence staining processes have species specificity without contrition dependence. Combining the dynamics of staining with real-time fluorescence microscopy we characterized the fluorescence staining process at the single-cell level with improved sensitivity and contrast.

  16. Quenching of fluorescein-conjugated lipids by antibodies. Quantitative recognition and binding of lipid-bound haptens in biomembrane models, formation of two-dimensional protein domains and molecular dynamics simulations.

    PubMed Central

    Ahlers, M; Grainger, D W; Herron, J N; Lim, K; Ringsdorf, H; Salesse, C

    1992-01-01

    Three model biomembrane systems, monolayers, micelles, and vesicles, have been used to study the influence of chemical and physical variables of hapten presentation at membrane interfaces on antibody binding. Hapten recognition and binding were monitored for the anti-fluorescein monoclonal antibody 4-4-20 generated against the hapten, fluorescein, in these membrane models as a function of fluorescein-conjugated lipid architecture. Specific recognition and binding in this system are conveniently monitored by quenching of fluorescein emission upon penetration of fluorescein into the antibody's active site. Lipid structure was shown to play a large role in affecting antibody quenching. Interestingly, the observed degrees of quenching were nearly independent of the lipid membrane model studied, but directly correlated with the chemical structure of the lipids. In all cases, the antibody recognized and quenched most efficiently a lipid based on dioctadecylamine where fluorescein is attached to the headgroup via a long, flexible hydrophilic spacer. Dipalmitoyl phosphatidylethanolamine containing a fluorescein headgroup demonstrated only partial binding/quenching. Egg phosphatidylethanolamine with a fluorescein headgroup showed no susceptibility to antibody recognition, binding, or quenching. Formation of two-dimensional protein domains upon antibody binding to the fluorescein-lipids in monolayers is also presented. Chemical and physical requirements for these antibody-hapten complexes at membrane surfaces have been discussed in terms of molecular dynamics simulations based on recent crystallographic models for this antibody-hapten complex (Herron et al., 1989. Proteins Struct. Funct. Genet. 5:271-280). Images FIGURE 7 FIGURE 8 PMID:1420916

  17. Novel Process for Laser Stain Removal from Archaeological Oil Paintings

    NASA Astrophysics Data System (ADS)

    El-Nadi, Lotfia; El-Feky, Osama; Abdellatif, Galila; Darwish, Sawsan

    2013-03-01

    Some samples of oil paintings (5 × 5 cm) were prepared on wooden panel with four types of fungi commonly encountered on oil paintings were selected for this study. Each of the fungi is associated with different colored stains. Fungus Alternaria tenuis is associated by a dense black stain, Chetomium globosum by a brownish gray stain, Aspergillus flavus by a yellowish stain, and Fusaruim oxysporum by a pinkish stain. Fungi growing on oil paintings affect the surface characteristics by forming a variety of colored patches typically composed of many complex chemical substances that are produced during metabolic processes. These colored stains may be encrusted in spores, present in mycelium or secreted to a substance such as oil paintings surfaces. While the fungal stains can sometimes be extracted with appropriate solvents, there are some stains that resist solvent extraction entirely. Developing new solvent system that might attack the paint structure, and is time consuming and requires a great deal of trial and error. Mechanical stain removal is also problematic in that it often produces abrasion of the surface, markedly deteriorating the artwork, and is extra ordinarily fine and tedious. For these reasons, we decided to examine an alternative physical technique as a new approach to deal with stain removal. Since the stains are due to the existence of fungi, we thought it a good idea to remove them by singlet oxygen. We applied the photo dynamic process through which the fungi stains were covered with organic dye derivatives in solution under controlled illumination in the lab. The samples were then irradiated by low power Laser light from a He-Ne laser, the dye will be photodecomposed and produce singlet oxygen. We report in this work the results obtained as a function of: - The concentration and types of the organic dye in solution, - The presence of certain amounts of liquids added to the solution, - The scanning speed of the laser beam on the sample surface, - The irradiation time. For each case fresh samples were used and photographed before and after the treatment. The results obtained will be speculated and discussed. This procedure was applied to the cleaning of archaeological oil paintings for the first time to our knowledge. The method could well be considered as a new field of combined science and technology applied to laser stain removal and represents a significant addition to the techniques available to art conservation.

  18. Automated detection of cells from immunohistochemically-stained tissues: application to Ki-67 nuclei staining

    NASA Astrophysics Data System (ADS)

    Cinar Akakin, Hatice; Kong, Hui; Elkins, Camille; Hemminger, Jessica; Miller, Barrie; Ming, Jin; Plocharczyk, Elizabeth; Roth, Rachel; Weinberg, Mitchell; Ziegler, Rebecca; Lozanski, Gerard; Gurcan, Metin N.

    2012-03-01

    An automated cell nuclei detection algorithm is described to be used for the quantification of immunohistochemicallystained tissues. Detection and segmentation of positively stained cells and their separation from the background and negatively-stained cells is crucial for fast, accurate, consistent and objective analysis of pathology images. One of the major challenges is the identification, hence accurate counting of individual cells, when these cells form clusters. To identify individual cell nuclei within clusters, we propose a new cell nuclei detection method based on the well-known watershed segmentation, which can lead to under- or over-segmentation for this problem. Our algorithm handles oversegmentation by combining H-minima transformed watershed algorithm with a novel region merging technique. To handle under-segmentation problem, we develop a Laplacian-of-Gaussian (LoG) filtering based blob detection algorithm, which estimates the range of the scales from the image adaptively. An SVM classifier was trained in order to separate non-touching single cells and touching cell clusters with five features representing connected region properties such as eccentricity, area, perimeter, convex area and perimeter-to-area ratio. Classified touching cell clusters are segmented with the H-minima based watershed algorithm. The resulting over-segmented regions are improved with the merging algorithm. The remaining under-segmented cell clusters are convolved with LoG filters to detect the cells within them. Cell-by-cell nucleus detection performance is evaluated by comparing computer detections with cell locations manually marked by eight pathology residents. The sensitivity is 89% when the cells are marked as positive at least by one resident and it increases to 99% when the evaluated cells are marked by all eight residents. In comparison, the average reader sensitivity varies between 70% +/- 18% and 95% +/- 11%.

  19. Silver and Cyanine Staining of Oligonucleotides in Polyacrylamide Gel

    PubMed Central

    Li, Wei

    2015-01-01

    To explore why some oligonucleotides in denaturing polyacrylamide gel could not be silver-stained, 134 different oligonucleotides were analyzed using denaturing polyacrylamide gel electrophoresis stained with silver and asymmetric cyanine. As a result, we found that the sensitivity of oligos (dA), (dC), (dG) and (dT) to silver staining could be ranged as (dA) > (dG) > (dC) > (dT) from high to low. It was unexpected that oligo (dT) was hard to be silver-stained. Moreover, the silver staining of an oligonucleotide containing base T could be partially or completely inhibited by base T. The inhibition of silver staining by base T was a competitive inhibition which could be affected by the amounts of the argyrophil nucleobase and base T, the cis-distance between the argyrophil nucleobase and base T, and the gel concentration. The changes of the intensity of an oligonucleotide band caused by the changes of DNA base composition were diverse and interesting. The intensity of some oligonucleotide bands would significantly change when the changes of DNA base composition accumulated to a certain extent (usually ≥ 4 nt). The sensitivity of cyanine staining of ≤ 11-nt long oligonucleotides could be enhanced about 250-fold by fixing the gels with methanol fixing solution. PMID:26650843

  20. Western Blot of Stained Proteins from Dried Polyacrylamide Gels

    NASA Technical Reports Server (NTRS)

    Gruber, Claudia; Stan-Lotter, Helga

    1996-01-01

    Western blotting of proteins is customarily performed following their separation on polyacrylamide gels, either prior to staining (1) or, as recently reported, following staining (2). We describe here Western blotting with stained gels, which had been dried and some of which had been stored for years. This procedure permits immunological analysis of proteins, to which antisera may have become available only later, or where the application of newly developed sensitive detection methods is desired. Once rehydration of the gels is achieved, proteins can be-transferred to blotting membranes by any appropriate protocol. Proteins stained with Coomassie Blue have to be detected with a non-chromogenic method, such as the film-based enhanced chemiluminescence (ECL)2) procedure (3). Silver stained proteins, which transfer in the colorless form, may be visualized by any detection method, although, because of the usually very low amounts of proteins, detection by ECL is preferable. Blotting of stained proteins from rehydrated gels is as rapid and as quantitative as from freshly prepared gels, in contrast to blotting from wet stained gels, which requires extensive washing and results in low transfer efficiency (2). Together with a photographic record of the gel pattern, unambiguous identification of immunoreactive proteins from complex mixtures is possible. Some further applications of this work are discussed.

  1. Microscopic analysis of MTT stained boar sperm cells

    PubMed Central

    van den Berg, B.M.

    2015-01-01

    The ability of sperm cells to develop colored formazan by reduction of MTT was used earlier to develop a spectrophotometric assay to determine the viability of sperm cells for several mammalian species. It was the objective of the present study to visualize microscopically the location of the formazan in boar sperm cells. The MTT staining process of boar sperm cells can be divided into a series of morphological events. Incubation of the sperm cells in the presence of MTT resulted after a few min in a diffuse staining of the midpiece of the sperm cells. Upon further incubation the staining of the midpiece became more intense, and gradually the formation of packed formazan granules became more visible. At the same time, a small formazan stained granule appeared medially on the sperm head, which increased in size during further incubation. After incubation for about 1 h the midpiece granules were intensely stained and more clearly distinct as granules, while aggregation of sperm cells occurred. Around 90% of the sperm cells showed these staining events. At the end of the staining the formazan granules have disappeared from both the sperm cells and medium, whereas formazan crystals appeared as thin crystal threads, that became heavily aggregated in the incubation medium. It was concluded that formazan is taken up by lipid droplets in the cytoplasm. Further, the use of the MTT assay to test for sperm viability should be regarded as a qualitative assay, whereas its practical use at artificial insemination (AI) Stations is limited. PMID:26623368

  2. Staining with methylthioninium chloride for the diagnosis of fungal keratitis

    PubMed Central

    LAN, LAN; WANG, FENG-YUN; ZENG, GUANGWEI

    2013-01-01

    The aim of this study was to investigate the potential efficacy of staining with methylthioninium chloride (MC) for the diagnosis of fungal keratitis. A total of 70 cases of fungal keratitis were included in the study from January 2009 to December 2010. The corneal scraping specimens of the patients were collected and stained with MC or a 10% potassium hydroxide (KOH)-based smear prior to microscopic examination. The staining results were confirmed with fungal culture and strain identification, which are recognized as ‘gold standards’ for the diagnosis of fungal keratitis. Among the 70 cases of fungal keratitis, 58 cases were positive and the positive rate was 82.86%. MC staining showed a positive rate of 62.86%, with higher levels of sensitivity (70.69%) and specificity (34.61%) compared with staining with the 10% KOH-based smear. The KOH-based smear showed a positive rate of 44.29% (P<0.05), a level of sensitivity of 44.83% (P<0.05) and a specificity of 17.95% (P<0.05). In addition, the MC staining showed false positive and negative rates of 25.00 and 29.31%, respectively, an accuracy index of 5.30% and positive and negative predictive values of 93.18 and 34.61%, respectively. The results indicate that MC staining is a fast and effective method for the early diagnosis of fungal keratitis. PMID:24223649

  3. Soft X-ray excited optical luminescence (XEOL) studies of fluorescein isothiocyanate (FITC) and FITC-labeled proteins1

    NASA Astrophysics Data System (ADS)

    Kim, Pil-Sook Grace; Petersen, N. O.; Sham, T. K.; Hu, Y. F.

    2004-07-01

    The electronic and optical properties of fluorescein-isothiocyanate (FITC) and FITC labeled proteins, Concanavalin A lectin and goat anti-rabbit immunoglobulin G have been investigated by X-ray excited optical luminescence (XEOL) using tunable soft X-rays from a synchrotron light source, together with X-ray absorption find structures (XAFS) spectroscopy at the C, N, and O K-edges. It is found that the carbon-localized excitation at the C K-edge is coupled to the luminescence chromophore effectively and excitations at the nitrogen and oxygen K-edge are not. The implication is that XEOL is excitation channel specific and that it can be used as a powerful tool in studying the optical properties of biomolecules.

  4. Blood–brain barrier transport studies, aggregation, and molecular dynamics simulation of multiwalled carbon nanotube functionalized with fluorescein isothiocyanate

    PubMed Central

    Shityakov, Sergey; Salvador, Ellaine; Pastorin, Giorgia; Förster, Carola

    2015-01-01

    In this study, the ability of a multiwalled carbon nanotube functionalized with fluorescein isothiocyanate (MWCNT–FITC) was assessed as a prospective central nervous system-targeting drug delivery system to permeate the blood–brain barrier. The results indicated that the MWCNT–FITC conjugate is able to penetrate microvascular cerebral endothelial monolayers; its concentrations in the Transwell® system were fully equilibrated after 48 hours. Cell viability test, together with phase-contrast and fluorescence microscopies, did not detect any signs of MWCNT–FITC toxicity on the cerebral endothelial cells. These microscopic techniques also revealed presumably the intracellular localization of fluorescent MWCNT–FITCs apart from their massive nonfluorescent accumulation on the cellular surface due to nanotube lipophilic properties. In addition, the 1,000 ps molecular dynamics simulation in vacuo discovered the phenomenon of carbon nanotube aggregation driven by van der Waals forces via MWCNT–FITC rapid dissociation as an intermediate phase. PMID:25784800

  5. Cellular carbohydrate components in human, rabbit and rat lacrimal gland. Studies using fluorescein and peroxidase labelled lectins.

    PubMed

    Ahmed, A; Grierson, I

    1989-01-01

    Orbital lacrimal glands from adult male and female rabbits, rats and humans were examined for the presence of intracellular receptors of four lectins: concanavalin-A agglutinin, lutus tetragonolobus agglutinin, ricinus comunis-60 agglutinin and wheat-germ agglutinin using fluorescein-conjugated lectin and peroxidase labelling methods for fluorescence and electron microscopy, respectively. Lectins were used as specific probes to detect carbohydrate moiety of the lacrimal gland. The pattern of labelling with the lectins suggests that N-acetyl-glucosamine, N-acetyl-D-galactosamine, D-galactose, D-mannose, sialic acid and L-fucose are contained in the lacrimal gland of the three species. The significance of these findings is discussed. PMID:2920911

  6. Integrating photoacoustic ophthalmoscopy with scanning laser ophthalmoscopy, optical coherence tomography, and fluorescein angiography for a multimodal retinal imaging platform

    NASA Astrophysics Data System (ADS)

    Song, Wei; Wei, Qing; Liu, Tan; Kuai, David; Burke, Janice M.; Jiao, Shuliang; Zhang, Hao F.

    2012-06-01

    Photoacoustic ophthalmoscopy (PAOM) is a newly developed retinal imaging technology that holds promise for both fundamental investigation and clinical diagnosis of several blinding diseases. Hence, integrating PAOM with other existing ophthalmic imaging modalities is important to identify and verify the strengths of PAOM compared with the established technologies and to provide the foundation for more comprehensive multimodal imaging. To this end, we developed a retinal imaging platform integrating PAOM with scanning laser ophthalmoscopy (SLO), spectral-domain optical coherence tomography (SD-OCT), and fluorescein angiography (FA). In the system, all the imaging modalities shared the same optical scanning and delivery mechanisms, which enabled registered retinal imaging from all the modalities. High-resolution PAOM, SD-OCT, SLO, and FA images were acquired in both albino and pigmented rat eyes. The reported in vivo results demonstrate the capability of the integrated system to provide comprehensive anatomic imaging based on multiple optical contrasts.

  7. Evaluation of Common Angling-Induced Sources of Epithelial Damage for Popular Freshwater Sport Fish using Fluorescein

    SciTech Connect

    Colotelo, Alison HA; Cooke, Steven J.

    2011-05-01

    Angling is a popular recreational activity across the globe and a large proportion of fish captured by anglers are released due to voluntary or mandatory catch-and-release practices. The handling associated with hook removal and return of the fish to their environment can cause physical damage to the epidermal layer of the fish which may affect the condition and survival of released fish. This study investigated possible sources of epithelial damage associated with several different handling methods (i.e. landing net types, interactions with different boat floor surfaces, tournament procedures) commonly used in recreational angling for two popular freshwater sport fish species, largemouth bass (Micropterus salmoides) and northern pike (Esox lucius). Epithelial damage was examined using fluorescein, a non-toxic dye, which has been shown to detect latent epithelial damage. Northern pike exhibited extensive epithelial damage after exposure to several of the induced treatments (i.e., interaction with a carpeted surface, knotted nylon net, and line rolling) but relatively little epithelial damage when exposed to others (i.e., knotless rubber nets, smooth boat surfaces, or lip gripping devices). Largemouth bass did not show significant epithelial damage for any of the treatments, with the exception of fish caught in a semi-professional live release tournament. The detection of latent injuries using fluorescein can be an important management tool as it provides visual examples of potential damage that can be caused by different handling methods. Such visualizations can be used to encourage fish friendly angler behaviour and enhance the survival and welfare of released fish. It can also be used to test new products that are intended to or claim to reduce injury to fish that are to be released. Future research should evaluate the relationship between different levels of epithelial damage and mortality across a range of environmental conditions.

  8. A new technique for Gram staining paraffin-embedded tissue.

    PubMed

    Engbaek, K; Johansen, K S; Jensen, M E

    1979-02-01

    Five techniques for Gram staining bacteria in paraffin sections were compared on serial sections of pulmonary tissues from eight bacteriological necropsies. Brown and Hopp's method was the most satisfactory for distinguishing Gram-positive and Gram-negative bacteria. However, this method cannot be recommended as the preparations were frequently overstained, and the Gram-negative bacteria were stained indistinctly. A modification of Brown and Hopps' method was developed which stains larger numbers of Gram-negative bacteria and differentiates well between different cell types and connective tissue, and there is no risk of overstaining. PMID:86548

  9. A new technique for Gram staining paraffin-embedded tissue.

    PubMed Central

    Engbaek, K; Johansen, K S; Jensen, M E

    1979-01-01

    Five techniques for Gram staining bacteria in paraffin sections were compared on serial sections of pulmonary tissues from eight bacteriological necropsies. Brown and Hopp's method was the most satisfactory for distinguishing Gram-positive and Gram-negative bacteria. However, this method cannot be recommended as the preparations were frequently overstained, and the Gram-negative bacteria were stained indistinctly. A modification of Brown and Hopps' method was developed which stains larger numbers of Gram-negative bacteria and differentiates well between different cell types and connective tissue, and there is no risk of overstaining. PMID:86548

  10. Improving acid-fast fluorescent staining for the detection of mycobacteria using a new nucleic acid staining approach.

    PubMed

    Ryan, Gavin J; Shapiro, Howard M; Lenaerts, Anne J

    2014-09-01

    Acid fast staining of sputum smears by microscopy remains the prevalent method for detecting Mycobacterium tuberculosis. The sensitivity of microscopy using acid fast stains requires 10(4) bacilli per ml of sputum. Although fluorescent acid fast stains, such as Auramine-O, show improved sensitivity, almost half of culture-positive TB cases are currently estimated to remain smear-negative. These current diagnosis problems provide impetus for improving staining procedures. We evaluated a novel fluorescent acid-fast staining approach using the nucleic acid-binding dye SYBR(®) Gold on mycobacterial in vitro cultures. The SYBR(®) Gold stain detected 99% of MTB in both actively replicating aerobic and non-replicating hypoxic cultures. Transmission light microscopy with Ziehl-Neelsen fuchsin, and fluorescence microscopy with Auramine-O or Auramine-rhodamine detected only 54%-86% of MTB bacilli. SYBR(®) Gold fluoresces more intensely than Auramine-O, and is highly resistant to fading. The signal to noise ratio is exceptionally high due to a >1000-fold enhanced fluorescence after binding to DNA/RNA, thereby reducing most background fluorescence. Although cost and stability of the dye may perhaps limit its clinical use at this time, these results warrant further research into more nucleic acid dye variants. In the meantime, SYBR(®) Gold staining shows great promise for use in numerous research applications. PMID:25130623

  11. Widespread Microbial Adaptation to l-Glutamate-N,N-diacetate (L-GLDA) Following Its Market Introduction in a Consumer Cleaning Product.

    PubMed

    Itrich, Nina R; McDonough, Kathleen M; van Ginkel, Cornelis G; Bisinger, Ed C; LePage, Jim N; Schaefer, Edward C; Menzies, Jennifer Z; Casteel, Kenneth D; Federle, Thomas W

    2015-11-17

    l-Glutamate-N,N-diacetate (L-GLDA) was recently introduced in the United States (U.S.) market as a phosphate replacement in automatic dishwashing detergents (ADW). Prior to introduction, L-GLDA exhibited poor biodegradation in OECD 301B Ready Biodegradation Tests inoculated with sludge from U.S. wastewater treatment plants (WWTPs). However, OECD 303A Activated Sludge WWTP Simulation studies showed that with a lag period to allow for growth (40-50 days) and a solids retention time (SRT) that allows establishment of L-GLDA degraders (>15 days), significant biodegradation (>80% dissolved organic carbon removal) would occur. Corresponding to the ADW market launch, a study was undertaken to monitor changes in the ready biodegradability of L-GLDA using activated sludge samples from various U.S. WWTPs. Initially all sludge inocula showed limited biodegradation ability, but as market introduction progressed, both the rate and extent of degradation increased significantly. Within 22 months, L-GLDA was ready biodegradable using inocula from 12 WWTPs. In an OECD 303A study repeated 18 months post launch, significant and sustained carbon removal (>94%) was observed after a 29-day acclimation period. This study systematically documented field adaptation of a new consumer product chemical across a large geographic region and confirmed the ability of laboratory simulation studies to predict field adaptation. PMID:26465169

  12. Thermodynamic and Spectroscopic Studies of Trivalent f-element Complexation with Ethylenediamine-N,N'-di(acetylglycine)-N,N'-diacetic Acid.

    PubMed

    Heathman, Colt R; Grimes, Travis S; Zalupski, Peter R

    2016-03-21

    The coordination behavior and thermodynamic features of complexation of trivalent lanthanides and americium by ethylenediamine-N,N'-di(acetylglycine)-N,N'-diacetic acid (EDDAG-DA) (bisamide-substituted-EDTA) were investigated by potentiometric and spectroscopic techniques. Acid dissociation constants (Ka) and complexation constants (β) of lanthanides (except Pm) were determined by potentiometric analysis. Absorption spectroscopy was used to determine stability constants for the binding of trivalent americium and neodymium by EDDAG-DA under similar conditions. The potentiometry revealed 5 discernible protonation constants and 3 distinct metal-ligand complexes (identified as ML(-), MHL, and MH2L(+)). Time-resolved fluorescence studies of Eu-(EDDAG-DA) solutions (at varying pH) identified a constant inner-sphere hydration number of 3, suggesting that glycine functionalities contained in the amide pendant arms are not involved in metal complexation and are protonated under more acidic conditions. The thermodynamic studies identified that f-element coordination by EDDAG-DA is similar to that observed for ethylenediamine-N,N,N',N'-tetraacetic acid (EDTA). However, coordination via two amidic oxygens of EDDAG-DA lowers its trivalent f-element complex stability by roughly 3 orders of magnitude relative to EDTA. PMID:26930023

  13. 6. Vick Farm, interior perspective of stained glass window, added ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    6. Vick Farm, interior perspective of stained glass window, added as part of deck addition on west side. - Vick Farm, North side Idlewild Road, 0.2 mile northwest of Idlewild & Maplewood Drive, Burlington, Boone County, KY

  14. [The morphological features of a 20 mcl blood stain].

    PubMed

    Pigolkin, Iu I; Leonova, E N; Leonov, S V; Nagornov, M N

    2015-01-01

    The objective of this work was to study blood stains of small volume (20 mcl). It was shown that the fall of a blood droplet from the height of 5 to 200 cm at the angle of 90 degrees on a smooth non-absorbing surface (glass) leaves round stains. Their size increases with increasing height of the fall. The character of the stain edges also depends on the height of the fall. The edges are even when the drops fall from the height of less than 20 cm but become wave-shaped with blunt projections when the height of the fall is increased to 30-90 cm. The fall from the height of 100 to 200 cm gives rise to the stains with the scalloped edges having the rectangular or nearly rectangular protrusions. From one to three additional Plateau drops can be observed near the main one. PMID:26036072

  15. Oblique view of memorial stained glass window in east loge ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Oblique view of memorial stained glass window in east loge corridor - National Home for Disabled Volunteer Soldiers, Northwestern Branch, Ward Memorial Hall, 5000 West National Avenue, Milwaukee, Milwaukee County, WI

  16. Hydroxyethyl lactamide, a dye solvent useful in vital staining.

    PubMed

    Risso-Dominguez, C J

    1976-03-01

    In a search for new vital stains to reveal the microanatomy of nudibranch mollusks, the slow or very low solubility of many dyes in sea water posed a serious problem. Preliminary dissolution in tap water proved impractical. Hydroxyethyl lactamide, an odorless liquid and dye solvent was found ideal since it permits immediate attainment of saturated solutions of dyes in sea water. Since hydroxyethyl lactamide passed the severe "eolid nudibranch test" and has been found nonirritating for the very sensitive rhinophorial structures, and furthermore since it has been used by the pharmaceutical industry as a vehicle in antibiotic preparations, it appears to be an ideal universal dye solvent for general use in vital staining. It has been used extensively in unpublished research by the writer on vital staining of nudibranchs. It has a low order of physiological activity and can be regarded an essentially inert when used in vital staining. PMID:59418

  17. Steinway piano and stained glass clerestory window in lounge area, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Steinway piano and stained glass clerestory window in lounge area, upper deck. Hot water radiators can be seen at base of wall. These run throughout the houseboat. - Houseboat LA DUCHESSE, The Antique Boat Museum, Clayton, Jefferson County, NY

  18. 18. INTERIOR DETAIL VIEW OF STAINED GLASS WINDOW LOCATED AT ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    18. INTERIOR DETAIL VIEW OF STAINED GLASS WINDOW LOCATED AT SOUTH SIDE OF ALTAR, NOTE INSCRIPTION DEDICATED IN THE MEMORY OF FATHER DAMIEN - St. Francis Catholic Church, Moloka'i Island, Kalaupapa, Kalawao County, HI

  19. A cold staining method for acid-fast bacilli

    PubMed Central

    Vasanthakumari, R.; Jagannath, K.; Rajasekaran, S.

    1986-01-01

    The Ziehl-Neelsen method is probably the best known and most frequently used procedure for staining tubercle bacilli. The method requires controlled heating for its success. However, in developing countries, such as India, where most laboratories rely mainly on spirit lamps as a source of heat, the Ziehl-Neelsen method often cannot be carried out because rectified spirit is difficult to obtain. The study describes a cold staining technique that uses the same staining solutions as the conventional Ziehl-Neelsen method. For direct smears, the correlation of results of the cold staining procedure with those of the Ziehl-Neelsen method was 97% and for concentrated smears was 99%. The method described is suitable for use in basically equipped laboratories. PMID:2433067

  20. INTERIOR VIEW OF ENTRY. SHOWING THE STAINED CONCRETE FLOOR AND ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    INTERIOR VIEW OF ENTRY. SHOWING THE STAINED CONCRETE FLOOR AND WINDOW WITH DIAMOND PATTERN MUNTINS. VIEW FACING NORTHWEST. - Hickam Field, Officers' Housing Type F, 602 Beard Avenue, Honolulu, Honolulu County, HI

  1. VIEW OF THREE NORTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF THREE NORTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ARE LOCATED ADJACENT TO THE ALTAR. - U.S. Naval Base, Pearl Harbor, Chapel, Corner of Oakley & Nimitz Street, Pearl City, Honolulu County, HI

  2. INTERIOR VIEW OF THE LANAI. SHOWING THE ORIGINAL STAINED CONCRETE ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    INTERIOR VIEW OF THE LANAI. SHOWING THE ORIGINAL STAINED CONCRETE FLOOR WITH INCISED LINES, AND HINGED DOOR TO GARAGE WITH VERTICAL BOARD PANELING (BACKGROUND). VIEW FACING NORTHWEST. - Hickam Field, Officers' Housing Type J, 701 Beard Street, Honolulu, Honolulu County, HI

  3. VIEW OF THREE NORTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF THREE NORTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ARE LOCATED JUST BELOW THE CHOIR LOFT. - U.S. Naval Base, Pearl Harbor, Chapel, Corner of Oakley & Nimitz Street, Pearl City, Honolulu County, HI

  4. 18. INTERIOR OF KITCHEN NO. 1 SHOWING STAINED CABINETRY ON ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    18. INTERIOR OF KITCHEN NO. 1 SHOWING STAINED CABINETRY ON OPPOSITE WALL FROM PAINTED CABINETS. VIEW TO NORTHEAST. - Bishop Creek Hydroelectric System, Plant 6, Cashbaugh-Kilpatrick House, Bishop Creek, Bishop, Inyo County, CA

  5. VIEW OF THREE SOUTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    VIEW OF THREE SOUTH FACING STAINED GLASS WINDOWS. THESE WINDOWS ARE LOCATED ADJACENT TO THE ALTER. - U.S. Naval Base, Pearl Harbor, Chapel, Corner of Oakley & Nimitz Street, Pearl City, Honolulu County, HI

  6. 4. September 1969 DETAIL OF STAINED GLASS WINDOWS IN EAST ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    4. September 1969 DETAIL OF STAINED GLASS WINDOWS IN EAST WALL, INTERIOR VIEW FROM BALCONY - Mount Zion United Methodist Church, 1334 Twenty-ninth Street Northwest, Washington, District of Columbia, DC

  7. Immunohistochemical CD3 staining detects additional patients with celiac disease

    PubMed Central

    Mubarak, Amani; Wolters, Victorien M; Houwen, Roderick HJ; ten Kate, Fiebo JW

    2015-01-01

    AIM: To investigate whether performing immunohistochemical CD3 staining, in order to improve the detection of intra-epithelial lymphocytosis, has an additional value in the histological diagnosis of celiac disease. METHODS: Biopsies obtained from 159 children were stained by hematoxylin and eosin (HE) and evaluated using the Marsh classification. CD3 staining was subsequently evaluated separately and independently. RESULTS: Differences in evaluation between the routine HE sections and CD3 staining were present in 20 (12.6%) cases. In 10 (6.3%) patients the diagnosis of celiac disease (Marsh II and III) changed on examination of CD3 staining: in 9 cases, celiac disease had initially been missed on the HE sections, while 1 patient had been over-diagnosed on the routine sections. In all patients, the final diagnosis based on CD3 staining, was concordant with serological results, which was not found previously. In the other 10 (12.3%) patients, the detection of sole intra-epithelial lymphocytosis (Marsh I) improved. Nine patients were found to have Marsh I on CD3 sections, which had been missed on routine sections. Interestingly, the only patient with negative serology had Giardiasis. Finally, in 1 patient with negative serology, in whom Marsh I was suspected on HE sections, this diagnosis was withdrawn after evaluation of the CD3 sections. CONCLUSION: Staining for CD3 has an additional value in the histological detection of celiac disease lesions, and CD3 staining should be performed when there is a discrepancy between serology and the diagnosis made on HE sections. PMID:26140002

  8. Interior detail view, surviving stained glass panel in an east ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Interior detail view, surviving stained glass panel in an east aisle window. Most of the stained glass has been removed from the building and relocated to other area churches. (Similar to HABS No. PA-6694-25). - Acts of the Apostles Church in Jesus Christ, 1400-28 North Twenty-eighth Street, northwest corner of North Twenty-eighth & Master Streets, Philadelphia, Philadelphia County, PA

  9. Spreading Depression Increases Immunohistochemical Staining of Glial Fibrillary Acidic Protein

    PubMed Central

    Kraig, Richard P.; Dong, Liming; Thisted, Ronald; Jaeger, Christine B.

    2009-01-01

    Reactive astrocytosis is a process by which astrocytes respond to brain injury by showing an increase in glial fibrillary acidic protein (GFAP) staining that is associated with hypertrophy and/or hyperplasia of these cells. Because spreading depression (SD) is a perturbation uncomplicated by neuronal necrosis and is seen in both in vivo and in vitro neural structures, we sought to determine whether SD was a sufficient stimulus to induce enhanced GFAP staining. SD was elicited in anesthetized rats by application of KCI to parietal cortex for 3 hr; equimolar NaCl was applied to contralateral cortex. SD was confirmed by monitoring DC potentials in frontal neocortices. Animals were allowed to recover for 48 hr, and their brains were processed for semiquantitative and computer-based analyses of GFAP staining intensity. Experimental GFAP staining was referenced to contralateral control levels. Neocortical SD (1337 SDs) was associated with a significant (p < 10?4), 43% increase in GFAP staining intensity, which remained statistically greater than normal for more than 2 weeks. If SD was inhibited by combined hyperoxia and hypercarbia, only a nonsignificant (p > 0.20), 7% increase in GFAP staining was seen. Thus, SD may be a useful physiologic process with which to begin to explore the cellular mechanisms that induce the transformation of normal astrocytes into reactive species. PMID:1906091

  10. Freeze-fracture of biological specimens prior to conductive staining.

    PubMed

    Iida, N

    1984-03-01

    Liver, kidney, spleen and other organs of the rat were fixed with glutaraldehyde, substituted with absolute ethanol or dimethyl sulfoxide (DMSO), freeze-fractured in liquid nitrogen, stained by the rapid tannin-osmium thiocarbohydrazide-osmium (TaOTO) method (staining with each agent for 10 min), critical-point-dried with liquid carbon dioxide, and observed with the scanning electron microscope. The absolute ethanol or DMSO freeze-fracture method provided flat fracture surfaces (without regard to cell boundaries) of the samples and allowed a good visualization of their inner structures. The fracture surfaces were suitably stained by the rapid TaOTO method, and could be scanned with no charging. Neither maked damage nor undesired dislocation of tissue elements was noted on the freeze-fractured and TaOTO-stained surfaces. This procedure, freeze-fracture prior to conductive staining, has an advantage of eliminating the bulk charging effects that tend to occur in specimens fractured after staining. When substituted with 75% DMSO aqueous solution, the samples spontaneously fractured without any need for razor blades. Fracture planes in this spontaneous fracture sometimes ran along the cell boundaries and allowed a clear visualization in the SEM of the enfaced surfaces of closely associated cells such as hepatocytes. PMID:6204620

  11. Post-staining electroblotting for efficient and reliable peptide blotting.

    PubMed

    Lee, Der-Yen; Chang, Geen-Dong

    2015-01-01

    Post-staining electroblotting has been previously described to transfer Coomassie blue-stained proteins from polyacrylamide gel onto polyvinylidene difluoride (PVDF) membranes. Actually, stained peptides can also be efficiently and reliably transferred. Because of selective staining procedures for peptides and increased retention of stained peptides on the membrane, even peptides with molecular masses less than 2 kDa such as bacitracin and granuliberin R are transferred with satisfactory results. For comparison, post-staining electroblotting is about 16-fold more sensitive than the conventional electroblotting for visualization of insulin on the membrane. Therefore, the peptide blots become practicable and more accessible to further applications, e.g., blot overlay detection or immunoblotting analysis. In addition, the efficiency of peptide transfer is favorable for N-terminal sequence analysis. With this method, peptide blotting can be normalized for further analysis such as blot overlay assay, immunoblotting, and N-terminal sequencing for identification of peptide in crude or partially purified samples. PMID:26044003

  12. One-tube triple staining method for flow cytometric analysis of DNA ploidy and phenotypic heterogeneity of human solid tumors using single laser excitation.

    PubMed

    Corver, W E; Bonsing, B A; Abeln, E C; Vlak-Theil, P M; Cornelisse, C J; Fleuren, G J

    1996-12-01

    We have developed a "one-tube" triple staining procedure that allows the identification of intratumor phenotypic subpopulations by FCM. Solid tumors were dissociated by a combined mechanical/ enzymatic method. Ovarian ascites tumor cell aggregates were enzymatically dissociated using trypsin. An antikeratin 8/18 MAb was used to label the epithelial fraction of these tumor samples. A second MAb directed against the leukocyte common antigen (LCA) was applied to identify nonneoplastic DNA-diploid cells. Other MAbs used as a second marker were directed against a tumor-associate surface, a cytoplasmic, or a nuclear antigen. Cells were stained using subclass-specific fluorescein-isothiocyanate (FITC) or R-phycoerythrin (PE)-conjugated antibodies. DNA was stained with propidium iodide (PI). Triply stained samples were measured on a standard bench-top flow cytometer (FACScan). Keratin 8/18-positive cells, LCA-positive cells, and DNA could be simultaneously detected in dissociated breast carcinomas, mixed Müllerian tumors, and ovarian ascites specimen for refining DNA index (DI) calculations and S phase fraction (SPF) determination. Coefficients of variation (CV) of the G0G1 peak of the DNA histograms obtained ranged from 2.55% to 4.64% and from 2.71% to 4.71% for the DNA-diploid and -aneuploid fractions, respectively. In DNA-diploid tumors, antigen expression (HER-2/Neu, proliferating cell nuclear antigen) could be analyzed without interference of fluorescence signals from nonneoplastic cells. Neoplastic tumor subpopulations were clearly identified based on both DNA-ploidy status and heterogeneity of antigen expression. The present method offers new possibilities for multiparameter DNA FCM on clinical samples and enables the identification of intratumor neoplastic subpopulations based on antigen expression and DNA-ploidy status. PMID:8946143

  13. Application of a Vital Fluorescent Staining Method for Simultaneous, Near-Real-Time Concentration Monitoring of Two Bacterial Strains in an Atlantic Coastal Plain Aquifer in Oyster, Virginia

    PubMed Central

    Fuller, Mark E.; Mailloux, Brian J.; Streger, Sheryl H.; Hall, James A.; Zhang, Pengfei; Kovacik, William P.; Vainberg, Simon; Johnson, William P.; Onstott, Tullis C.; DeFlaun, Mary F.

    2004-01-01

    Two differentially labeled bacterial strains were monitored in near-real time during two field-scale bacterial transport experiments in a shallow aquifer in July 2000 and July 2001. Comamonas sp. strain DA001 and Acidovorax sp. strain OY-107 were grown and labeled with the vital fluorescent stain TAMRA/SE (5 [and -6]-carboxytetramethylrhodamine, succinimidyl ester) or CFDA/SE (5 [and -6]-carboxyfluorescein diacetate, succinimidyl ester). Fluorescently labeled cells and a conservative bromide tracer were introduced into a suboxic superficial aquifer, followed by groundwater collection from down-gradient multilevel samplers. Cells were enumerated in the field by microplate spectrofluorometry, with confirmatory analyses for selected samples done in the laboratory by epifluorescence microscopy, flow cytometry, and ferrographic capture. There was general agreement in the results from all of the vital-stain-based enumeration methods, with differences ranging from <10% up to 40% for the analysis of identical samples between different tracking methods. Field analysis by microplate spectrofluorometry was robust and efficient, allowing thousands of samples to be analyzed in quadruplicate for both of the injected strains. The near-real-time data acquisition allowed adjustments to the predetermined sampling schedule to be made. The microplate spectrofluorometry data sets for the July 2000 and July 2001 experiments allowed the transport of the injected cells to be related to the site hydrogeology and injection conditions and enabled the assessment of differences in the transport of the two strains. This near-real-time method should prove effective for a number of microbial ecology applications. PMID:15006793

  14. Reliability of a rapid hematology stain for sputum cytology*

    PubMed Central

    Gonçalves, Jéssica; Pizzichini, Emilio; Pizzichini, Marcia Margaret Menezes; Steidle, Leila John Marques; Rocha, Cristiane Cinara; Ferreira, Samira Cardoso; Zimmermann, Célia Tânia

    2014-01-01

    Objective: To determine the reliability of a rapid hematology stain for the cytological analysis of induced sputum samples. Methods: This was a cross-sectional study comparing the standard technique (May-Grünwald-Giemsa stain) with a rapid hematology stain (Diff-Quik). Of the 50 subjects included in the study, 21 had asthma, 19 had COPD, and 10 were healthy (controls). From the induced sputum samples collected, we prepared four slides: two were stained with May-Grünwald-Giemsa, and two were stained with Diff-Quik. The slides were read independently by two trained researchers blinded to the identification of the slides. The reliability for cell counting using the two techniques was evaluated by determining the intraclass correlation coefficients (ICCs) for intraobserver and interobserver agreement. Agreement in the identification of neutrophilic and eosinophilic sputum between the observers and between the stains was evaluated with kappa statistics. Results: In our comparison of the two staining techniques, the ICCs indicated almost perfect interobserver agreement for neutrophil, eosinophil, and macrophage counts (ICC: 0.98-1.00), as well as substantial agreement for lymphocyte counts (ICC: 0.76-0.83). Intraobserver agreement was almost perfect for neutrophil, eosinophil, and macrophage counts (ICC: 0.96-0.99), whereas it was moderate to substantial for lymphocyte counts (ICC = 0.65 and 0.75 for the two observers, respectively). Interobserver agreement for the identification of eosinophilic and neutrophilic sputum using the two techniques ranged from substantial to almost perfect (kappa range: 0.91-1.00). Conclusions: The use of Diff-Quik can be considered a reliable alternative for the processing of sputum samples. PMID:25029648

  15. Cell type related differences in staining with pentameric thiophene derivatives.

    PubMed

    Cieślar-Pobuda, Artur; Bäck, Marcus; Magnusson, Karin; Jain, Mayur V; Rafat, Mehrdad; Ghavami, Saeid; Nilsson, K Peter R; Łos, Marek J

    2014-07-01

    Fluorescent compounds capable of staining cells selectively without affecting their viability are gaining importance in biology and medicine. Recently, a new family of optical dyes, denoted luminescent conjugated oligothiophenes (LCOs), has emerged as an interesting class of highly emissive molecules for studying various biological phenomena. Properly functionalized LCOs have been utilized for selective identification of disease-associated protein aggregates and for selective detection of distinct cells. Herein, we present data on differential staining of various cell types, including cancer cells. The differential staining observed with newly developed pentameric LCOs is attributed to distinct side chain functionalities along the thiophene backbone. Employing flow cytometry and fluorescence microscopy we examined a library of LCOs for stainability of a variety of cell lines. Among tested dyes we found promising candidates that showed strong or moderate capability to stain cells to different extent, depending on target cells. Hence, LCOs with diverse imidazole motifs along the thiophene backbone were identified as an interesting class of agents for staining of cancer cells, whereas LCOs with other amino acid side chains along the backbone showed a complete lack of staining for the cells included in the study. Furthermore, for p-HTMI,a LCO functionalized with methylated imidazole moieties, the staining was dependent on the p53 status of the cells, indicating that the molecular target for the dye is a cellular component regulated by p53. We foresee that functionalized LCOs will serve as a new class of optical ligands for fluorescent classification of cells and expand the toolbox of reagents for fluorescent live imaging of different cells. PMID:24500794

  16. Fluorescent multiple staining and CASA system to assess boar sperm viability and membranes integrity in short and long-term extenders

    PubMed Central

    Lange-Consiglio, A.; Meucci, A.; Cremonesi, F.

    2013-01-01

    The aim of this study was to assess the effect on boar spermatozoa quality of in vitro storage in short and long-term extenders by fluorescent multiple staining (FMS) and computer assisted semen analyzer (CASA). Fresh ejaculates from three healthy, sexually mature boars were diluted with equal volumes of six short-term or three long-term commercial extenders and stored at 19°C for 6 days (short-term) or 12 days (long-term). The integrity of spermatozoa membranes was analyzed by FMS using propidium iodide, 5,5’,6,6’-tetrachloro-1,1’,3,3’ tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) and fluorescein isothiocyanate-conjugated peanut agglutinin (PNA). The results obtained from this staining were compared with spermatozoa motility assessed by CASA. Our study showed that the number of viable spermatozoa with non-reacted acrosomes and intact mitochondria was positively correlated with the rate of motile spermatozoa (r2>0.9) irrespective of the extender used. In all extenders the number of motile spermatozoa significantly decreased as preservation period increased (P<0.05). FMS test is a potent indicator of sperm motility because it analyses mitochondrial integrity independently from observable alterations in motility. The best performing extenders were BTS for short-term storage and TRI-x-Cell for long-term storage. PMID:26623308

  17. Histochemical staining of Arabidopsis thaliana secondary cell wall elements.

    PubMed

    Pradhan Mitra, Prajakta; Loqué, Dominique

    2014-01-01

    Arabidopsis thaliana is a model organism commonly used to understand and manipulate various cellular processes in plants, and it has been used extensively in the study of secondary cell wall formation. Secondary cell wall deposition occurs after the primary cell wall is laid down, a process carried out exclusively by specialized cells such as those forming vessel and fiber tissues. Most secondary cell walls are composed of cellulose (40-50%), hemicellulose (25-30%), and lignin (20-30%). Several mutations affecting secondary cell wall biosynthesis have been isolated, and the corresponding mutants may or may not exhibit obvious biochemical composition changes or visual phenotypes since these mutations could be masked by compensatory responses. Staining procedures have historically been used to show differences on a cellular basis. These methods are exclusively visual means of analysis; nevertheless their role in rapid and critical analysis is of great importance. Congo red and calcofluor white are stains used to detect polysaccharides, whereas Mäule and phloroglucinol are commonly used to determine differences in lignin, and toluidine blue O is used to differentially stain polysaccharides and lignin. The seemingly simple techniques of sectioning, staining, and imaging can be a challenge for beginners. Starting with sample preparation using the A. thaliana model, this study details the protocols of a variety of staining methodologies that can be easily implemented for observation of cell and tissue organization in secondary cell walls of plants. PMID:24894795

  18. Histochemical Staining of Arabidopsis thaliana Secondary Cell Wall Elements

    PubMed Central

    Pradhan Mitra, Prajakta; Loqué, Dominique

    2014-01-01

    Arabidopsis thaliana is a model organism commonly used to understand and manipulate various cellular processes in plants, and it has been used extensively in the study of secondary cell wall formation. Secondary cell wall deposition occurs after the primary cell wall is laid down, a process carried out exclusively by specialized cells such as those forming vessel and fiber tissues. Most secondary cell walls are composed of cellulose (40–50%), hemicellulose (25–30%), and lignin (20–30%). Several mutations affecting secondary cell wall biosynthesis have been isolated, and the corresponding mutants may or may not exhibit obvious biochemical composition changes or visual phenotypes since these mutations could be masked by compensatory responses. Staining procedures have historically been used to show differences on a cellular basis. These methods are exclusively visual means of analysis; nevertheless their role in rapid and critical analysis is of great importance. Congo red and calcofluor white are stains used to detect polysaccharides, whereas Mäule and phloroglucinol are commonly used to determine differences in lignin, and toluidine blue O is used to differentially stain polysaccharides and lignin. The seemingly simple techniques of sectioning, staining, and imaging can be a challenge for beginners. Starting with sample preparation using the A. thaliana model, this study details the protocols of a variety of staining methodologies that can be easily implemented for observation of cell and tissue organization in secondary cell walls of plants. PMID:24894795

  19. Methods And Compositions For Chromosome-Specific Staining

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    2003-08-19

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods are provided to disable the hybridization capacity of shared, high copy repetitive sequences and/or remove such sequences to provide for useful contrast. Still further methods are provided to produce chromosome-specific staining reagents which are made specific to the targeted chromosomal material, which can be one or more whole chromosomes, one or more regions on one or more chromosomes, subsets of chromosomes and/or the entire genome. Probes and test kits are provided for use in tumor cytogenetics, in the detection of disease related loci, in analysis of structural abnormalities, such as translocations, and for biological dosimetry. Further, methods and prenatal test kits are provided to stain targeted chromosomal material of fetal cells, including fetal cells obtained from maternal blood. Still further, the invention provides for automated means to detect and analyse chromosomal abnormalities.

  20. Methods of biological dosimetry employing chromosome-specific staining

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    2000-01-01

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods are provided to disable the hybridization capacity of shared, high copy repetitive sequences and/or remove such sequences to provide for useful contrast. Still further methods are provided to produce chromosome-specific staining reagents which are made specific to the targeted chromosomal material, which can be one or more whole chromosomes, one or more regions on one or more chromosomes, subsets of chromosomes and/or the entire genome. Probes and test kits are provided for use in tumor cytogenetics, in the detection of disease related loci, in analysis of structural abnormalities, such as translocations, and for biological dosimetry. Further, methods and prenatal test kits are provided to stain targeted chromosomal material of fetal cells, including fetal cells obtained from maternal blood. Still further, the invention provides for automated means to detect and analyse chromosomal abnormalities.

  1. CD3 immunohistochemical staining in diagnosis of lymphocytic colitis.

    PubMed

    Fiehn, Anne-Marie Kanstrup; Engel, Ulla; Holck, Susanne; Munck, Lars Kristian; Engel, Peter Johan Heiberg

    2016-02-01

    Microscopic colitis (MC) is a common cause of chronic watery diarrhea. Traditionally, MC encompasses the 2 subgroups lymphocytic colitis (LC) and collagenous colitis, but recently, an additional subgroup, MC incomplete, has been introduced. Distinguishing between the subgroups relies exclusively on histopathologic evaluation. In the present study, 4 pathologists evaluated 156 archived biopsies originally diagnosed as LC or LC incomplete (LCi). Each pathologist assigned a diagnosis of LC, LCi, or nonspecific inflammation to all cases at 2 independent assessments. At the first assessment, hematoxylin and eosin (HE) stainings were available. At the second assessment, a supplementary CD3 immunohistochemical staining was also available. The aim was to evaluate whether a supplementary CD3 would increase the diagnostic agreement among pathologists, and whether a CD3 stain would change the diagnosis based on HE staining only. After the complete assessment, the cases were divided into 3 groups, that is, full agreement, partial agreement, and disagreement. The CD3 staining increased the number of cases with full agreement from 60 to 78. One hundred thirty-one cases with agreement or partial diagnostic agreement based on HE + CD3 were compared with the HE diagnoses. In 44 (34%) of 131 cases, CD3 changed the diagnosis. Cases assigned to the LCi category based on HE were often changed by a supplementary CD3. Conclusively, it is recommended to use a CD3 before giving the histopathologic diagnosis of LCi. PMID:26772395

  2. Silver stain for proteins on a cellulose acetate membrane.

    PubMed

    Fujita, T; Toda, T; Ohashi, M

    1984-06-01

    A rapid and sensitive silver staining method to detect proteins on a cellulose acetate membrane has been established. This method is achieved by modification of the silver-based color staining for detection of proteins in polyacrylamide gels [D. W. Sammons, L. D. Adams, and E. E. Nishizawa, Electrophoresis 2, 135-141 (1981)] and applied to our new type of two-dimensional electrophoresis for analysis of proteins on a cellulose acetate sheet [T. Toda, T. Fujita, and M. Ohashi, Anal. Biochem. 119, 167-176 (1982)]. Maximal sensitivity of silver stain for proteins on a cellulose acetate membrane can be obtained by an optimal balance between deposition of silver on the protein and on the background. Certain kinds of proteins are colored red, orange, or grayish-blue. The silver stain is 20-80 times more sensitive than Coomassie blue and some spots are visualized reproducibly by silver only. Densitometric evaluation of standard proteins stained with silver and Coomassie blue is also demonstrated. The method takes only 50 min to perform and is sensitive, simple, and reproducible. PMID:6206749

  3. Detection of reactive oxygen species (ROS) by the oxidant-sensing probe 2',7'-dichlorodihydrofluorescein diacetate in the cyanobacterium Anabaena variabilis PCC 7937

    SciTech Connect

    Rastogi, Rajesh P.; Laboratory of Photobiology and Molecular Microbiology, Centre of Advanced Study in Botany, Banaras Hindu University, Varanasi 221005 ; Singh, Shailendra P.; Haeder, Donat-P.; Sinha, Rajeshwar P.

    2010-07-02

    The generation of reactive oxygen species (ROS) under simulated solar radiation (UV-B: 0.30 Wm{sup -2}, UV-A: 25.70 Wm{sup -2} and PAR: 118.06 Wm{sup -2}) was studied in the cyanobacterium Anabaena variabilis PCC 7937 using the oxidant-sensing fluorescent probe 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). DCFH-DA is a nonpolar dye, converted into the polar derivative DCFH by cellular esterases that are nonfluorescent but switched to highly fluorescent DCF when oxidized by intracellular ROS and other peroxides. The images obtained from the fluorescence microscope after 12 h of irradiation showed green fluorescence from cells covered with 295, 320 or 395 nm cut-off filters, indicating the generation of ROS in all treatments. However, the green/red fluorescence ratio obtained from fluorescence microscopic analysis showed the highest generation of ROS after UV-B radiation in comparison to PAR or UV-A radiation. Production of ROS was also measured by a spectrofluorophotometer and results obtained supported the results of fluorescence microscopy. Low levels of ROS were detected at the start (0 h) of the experiment showing that they are generated even during normal metabolism. This study also showed that UV-B radiation causes the fragmentation of the cyanobacterial filaments which could be due to the observed oxidative stress. This is the first report for the detection of intracellular ROS in a cyanobacterium by fluorescence microscopy using DCFH-DA and thereby suggesting the applicability of this method in the study of in vivo generation of ROS.

  4. Detection of reactive oxygen species (ROS) by the oxidant-sensing probe 2',7'-dichlorodihydrofluorescein diacetate in the cyanobacterium Anabaena variabilis PCC 7937.

    PubMed

    Rastogi, Rajesh P; Singh, Shailendra P; Häder, Donat-P; Sinha, Rajeshwar P

    2010-07-01

    The generation of reactive oxygen species (ROS) under simulated solar radiation (UV-B: 0.30Wm(-2), UV-A: 25.70Wm(-2) and PAR: 118.06Wm(-2)) was studied in the cyanobacterium Anabaena variabilis PCC 7937 using the oxidant-sensing fluorescent probe 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA). DCFH-DA is a nonpolar dye, converted into the polar derivative DCFH by cellular esterases that are nonfluorescent but switched to highly fluorescent DCF when oxidized by intracellular ROS and other peroxides. The images obtained from the fluorescence microscope after 12h of irradiation showed green fluorescence from cells covered with 295, 320 or 395nm cut-off filters, indicating the generation of ROS in all treatments. However, the green/red fluorescence ratio obtained from fluorescence microscopic analysis showed the highest generation of ROS after UV-B radiation in comparison to PAR or UV-A radiation. Production of ROS was also measured by a spectrofluorophotometer and results obtained supported the results of fluorescence microscopy. Low levels of ROS were detected at the start (0h) of the experiment showing that they are generated even during normal metabolism. This study also showed that UV-B radiation causes the fragmentation of the cyanobacterial filaments which could be due to the observed oxidative stress. This is the first report for the detection of intracellular ROS in a cyanobacterium by fluorescence microscopy using DCFH-DA and thereby suggesting the applicability of this method in the study of in vivo generation of ROS. PMID:20570649

  5. Fate of pGFP-bearing Escherichia coli O157:H7 in ground beef at 2 and 10 degrees C and effects of lactate, diacetate, and citrate.

    PubMed

    Ajjarapu, S; Shelef, L A

    1999-12-01

    Although beef has been implicated in the largest outbreaks of Escherichia coli O157:H7 infection in the United States, studies on the fate of this pathogen have been limited. Problems in such studies are associated with detection of the pathogen at levels considerably lower than the levels of the competing microorganisms. In the present study, a green fluorescent protein-expressing E. coli O157:H7 strain was used, and the stable marker allowed us to monitor the behavior of the pathogen in ground beef stored aerobically from freshness to spoilage at 2 and 10 degrees C. In addition, the effects of sodium salts of lactate (SL) (0.9 and 1.8%), diacetate (SDA) (0.1 and 0.2%), and buffered citrate (SC) (1 and 2%) and combinations of SL and SDA were evaluated. SC had negligible antimicrobial activity, and SL delayed microbial growth, while SDA and SL plus SDA were most inhibitory to the total-aerobe population in the meat. At 2 degrees C, the initial numbers of E. coli O157:H7 (3 and 5 log(10) CFU/g) decreased by approximately 1 log(10) CFU/g when spoilage was manifest (>7 log(10) CFU of total aerobes/g), irrespective of the treatment. There was no decline in the numbers of the pathogen during storage at 10 degrees C. Our results showed that the pathogen was resistant to the salts tested and confirmed that refrigerated meat contaminated with the pathogen remains hazardous. PMID:10583994

  6. Spectroscopic studies on H2O2 damaging BSA induced by 1,2-dihydroxy-9, 10-anthraquinone-3-aminomethyl-N, N-diacetate-Ferrous(III)

    NASA Astrophysics Data System (ADS)

    Zou, Mingming; Li, Ying; Wang, Jun; Gao, Jingqun; Wang, Qi; Wang, Baoxin; Fan, Ping

    2013-08-01

    The interaction between 1,2-dihydroxy-9, 10-anthraquinone-3-aminomethyl-N, N-diacetate-Ferrous(III) (Alizarin-DA-Fe(III)) and bovine serum albumin (BSA) was studied by using UV-vis and fluorescence spectra. And then, the H2O2 damage of BSA induced by Alizarin-DA-Fe(III) was examined. The results show that due to the interaction the fluorescence of BSA solution can be obviously quenched by Alizarin-DA-Fe(III) and that the quenching process belongs to the static quenching. In addition, in the presence of Alizarin-DA-Fe(III) the BSA molecules were markedly damaged by H2O2. Meanwhile, the effects of the standing time, Alizarin-DA-Fe(III) concentration and H2O2 concentration on the damage of BSA molecules were also researched. The experimental results demonstrate that the damage degree increase with the increase of standing time, Alizarin-DA-Fe(III) concentration and H2O2 concentration. Finally, the generation of reactive oxygen species (ROS) from H2O2 induced by Alizarin-DA-Fe(III) as Fenton-like reagent was estimated by some quenchers. Because the Iminodiacetic-Ferrous(III) (IDA-Fe(III)) and Nitrilotriacetic-Ferrous(III) (NTA-Fe(III)) can be thought of as the active part of Alizarin-DA-Fe(III), they were used to compare the catalytic activity with Alizarin-DA-Fe(III). Owing to the special plane structure, the experiment results showed that the Alizarin-DA-Fe(III) exhibited higher damage ability than IDA-Fe(III) and NTA-Fe(III). Perhaps, the Alizarin-DA-Fe(III) may be used as a new antitumor compound to induce peroxides in body to kill cancer cells.

  7. Potassium lactate combined with sodium diacetate can inhibit growth of Listeria monocytogenes in vacuum-packed cold-smoked salmon and has no adverse sensory effects.

    PubMed

    Vogel, Birte Fonnesbech; Ng, Yoke Yin; Hyldig, Grethe; Mohr, Mona; Gram, Lone

    2006-09-01

    Growth of Listeria monocytogenes in ready-to-eat fish products such as cold-smoked salmon is an important food safety issue. The objective of this study was to evaluate the antilisterial activity of potassium lactate (PL) in combination with sodium acetate (SA) or sodium diacetate (SDA) in cold-smoked salmon and to determine whether these compounds could be incorporated easily into the formulations and technology currently used by processors. A commercial brine injector was used to inject salmon filets with either saturated saline brine or saturated saline brine supplemented with combinations of PL and SA (PURASAL Opti. Form PA 4) or PL and SDA (PURASAL Opti. Form PD 4). In the brine-injected cold-smoked salmon, 2.1% (water phase) PL and 0.12% (water phase) SDA delayed the growth of L. monocytogenes for up to 42 days of vacuum-packaged storage at 10 degrees C. Storage at 25 degrees C for 6 h resulted in only a 1-log CFU/g increase in L. monocytogenes. Treatments with lower concentrations of PL and SDA or similar concentrations of PL and SA resulted in an extended lag phase and slower growth of L. monocytogenes. It was not possible to incorporate more than 2% (water phase) PL while ensuring a minimum of 3% (water phase) NaCl in the finished product because PL decreased the solubility of NaCl. Sensory analyses revealed that the preservatives did not negatively affect flavor or odor. The combination of PL and SDA is therefore a viable technology for preventing L. monocytogenes growth on cold-smoked salmon. PMID:16995515

  8. Chromosome-specific staining to detect genetic rearrangements

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel; Tkachuk, Douglas; Westbrook, Carol

    2013-04-09

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyzes. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  9. Stain length passive dosimeter for monitoring of carbon monoxide

    SciTech Connect

    Gonzalez, L.A.; Sefton, M.V.

    1983-07-01

    A new passive dosimeter for the personal monitoring of CO exposure in the workplace has been developed. This new type of sampling device does not require follow-up analysis of the collected sample to determine the exposure. Rather, the time-weighted average concentration is determined directly from the length of a coloured stain which is produced instantaneously during the exposure period in a specially prepared indicator tube. The stain length is a function of both contaminant concentration and exposure time. The effects of CO concentration in the range 0.5-2.0 x TLV on the response of the dosimeter were evaluated in a dynamic exposure chamber. Data were fitted to the appropriate model equation with a correlation coefficient of 0.968. By eliminating the need for follow-up analysis, this stain-length dosimeter significantly reduces the cost of monitoring.

  10. Peroxisomes: 40 years of histochemical staining, personal reminiscences.

    PubMed

    Dariush Fahimi, H

    2009-04-01

    The historical circumstances that led to the discovery of the 3,3'-diamino-benzidine (DAB) method for staining of peroxisomes 40 years ago are reviewed. In the course of studies on the uptake and absorption of horse radish peroxidase in mammalian liver, in sections incubated for detection of peroxidase activity in DAB, it was noted that peroxisomes also stained positively for peroxidase activity. Subsequently, it was revealed that the peroxidatic activity of catalase, which is abundantly present in peroxisomes, is responsible for that staining. This notion was confirmed in quantitative biochemical studies with crystalline beef liver catalase and in tracer studies using catalase as an ultrastructural tracer. The application of the DAB method led to the discovery of peroxisomes as a ubiquitous eukaryotic cell organelle, attracting great interest in their investigation in biomedical research. PMID:19219449

  11. Skull Base Meningiomas and Cranial Nerves Contrast Using Sodium Fluorescein: A New Application of an Old Tool

    PubMed Central

    da Silva, Carlos Eduardo; da Silva, Vinicius Duval; da Silva, Jefferson Luis Braga

    2014-01-01

    Objective The identification of cranial nerves is one of the most challenging goals in the dissection of skull base meningiomas. The authors present an application of sodium fluorescein (SF) in skull base meningiomas with the purpose of improving the identification of cranial nerves. Design A prospective study within-subjects design. Setting Hospital Ernesto Dornelles, Porto Alegre, Brazil. Participants Patients with skull base meningiomas. Main Outcomes Measures Cranial nerve identification. Results The group of nine meningiomas was composed of one cavernous sinus, three petroclival, one tuberculum sellae, two sphenoid wing, one olfactory groove, and one temporal floor meningioma. The SF enhancement in all tumors was strong, and the contrast with cranial nerves clearly evident. There were one definite olfactory nerve deficit, one transient abducens deficit, and one definite hemiparesis. All lesions were resected (Simpson grades 1 and 2). The analysis of the difference of the delta SF wavelength between the meningiomas and cranial nerve contrast was performed by the Wilcoxon signed rank test and showed p = 0.011. Conclusions The contrast between the enhanced meningiomas and cranial nerves was evident and assisted in the visualization and microsurgical dissection of these structures. The anatomical preservation of these structures was improved using the contrast.

  12. Fluorescein: A Photo-CIDNP Sensitizer Enabling Hypersensitive NMR Data Collection in Liquids at Low Micromolar Concentration.

    PubMed

    Okuno, Yusuke; Cavagnero, Silvia

    2016-02-01

    Photochemically induced dynamic nuclear polarization (photo-CIDNP) is a powerful approach for sensitivity enhancement in NMR spectroscopy. In liquids, intermolecular photo-CIDNP depends on the transient bimolecular reaction between photoexcited dye and sample of interest. Hence the extent of polarization is sample-concentration dependent. This study introduces fluorescein (FL) as a photo-CIDNP dye whose performance is exquisitely tailored to data collection at extremely low sample concentrations. The photo-CIDNP resonance intensities of tryptophan in the presence of either FL or FMN (i.e., the routinely employed flavin mononucleotide photosensitizer) in the liquid state show that FL yields superior sensitivity and enables rapid data collection down to an unprecedented 1 μM concentration. This result was achieved on a conventional spectrometer operating at 14.1 T and equipped with a room-temperature probe (i.e., noncryogenic). Kinetic simulations show that the excellent behavior of FL arises from its long excited-state triplet lifetime and superior photostability relative to conventional photo-CIDNP sensitizers. PMID:26744790

  13. Liquid chromatographic determination of endogenous phytohormones in vegetable samples based on chemical derivatization with 6-oxy(acetylpiperazine) fluorescein.

    PubMed

    Chen, Hao; Zhang, Zi-Xing; Zhang, Gui-Min; Guo, Xiao-Feng; Zhang, Hua-Shan; Wang, Hong

    2010-04-28

    In phytohormone analysis, mass spectrometry (MS)-based methods are primary and powerful tools. However, complex sample preparation and high cost are problems for their application. As a complement for MS-based methods, a new fluorescent labeling reagent for carboxylic acids, 6-oxy(acetylpiperazine) fluorescein (APF), has been used for the determination of endogenous phytohormones, including indolebutyric acid, 1-naphthaleneacetic acid and 2,4-dichlorophenoxyacetic acid. The derivatization yield was maximized by optimizing derivatization conditions in detail, and the derivatives of three phytohormones could be separated completely in 15 min on a C18 column with fluorescence detection at lambda(ex)/lambda(em) = 467/512 nm. The derivatization limits could reach 0.1 microM, and the detection limits (signal-to-noise ratio = 3) were 4.43-14.2 nM. The proposed method has been applied to the determination of the exogenous phytohormones in the crude extracts of vegetable samples without extra purification and enrichment with recoveries of 94.2-102.4%. PMID:20356302

  14. Sieve-element differentiation and fluoresceine translocation in wound-phloem of pea roots after complete severance of the stele.

    PubMed

    Schulz, A

    1987-03-01

    Experimental interruption of the root stele of Pisum sativum L. induces in the cortex tissue the development of wound-sieve tubes which bridge the wound and reconnect the vascular stumps. Outside the stele, sieve plates arise from primary pit fields. This origin is confirmed by the distribution of future sieve pores over the original parenchyma cell wall and by remnants of the pitfield cavity in developing sieve plates. Differentiation of wound-sieve elements is similar to that of bundle-sieve elements and includes the chromatolytic disintegration of nuclei as well as the development of typical sieve pores arising from pit-field plasmodesmata. The completion of first woundsieve tubes (indicated by a continuous chain of anilin-blue-positive sieve plates by-passing the wound) was observed 55-62 h after wounding. However, effective translocation, visualized with fluoresceine as a phloem-mobile marker, was not found until 10 h (on average) later. It is suggested that this time delay corresponds to the maturing of the last link within a chain of wound-sieve-tube members. Presumably, enucleate sieve elements with widened pores are a prerequisite for effective phloem translocation. PMID:24232957

  15. Fluorescent labeling of cranberry proanthocyanidins with 5-([4,6-dichlorotriazin-2-yl]amino)fluorescein (DTAF).

    PubMed

    Feliciano, Rodrigo P; Heintz, Joseph A; Krueger, Christian G; Vestling, Martha M; Reed, Jess D

    2015-01-01

    A novel methodology was developed to elucidate proanthocyanidins (PAC) interaction with extra-intestinal pathogenic Escherichia coli (ExPEC). PAC inhibit ExPEC invasion of epithelial cells and, therefore, may prevent transient gut colonization, conferring protection against subsequent extra-intestinal infections, such as urinary tract infections. Until now PAC have not been chemically labeled with fluorophores. In this work, cranberry PAC were labeled with 5-([4,6-dichlorotriazin-2-yl]amino) fluorescein (DTAF), detected by high-performance liquid chromatography with diode-array detection and characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). We report single and double fluorescent-labeled PAC with one or two chlorine atoms displaced from DTAF in alkaline pH via nucleophilic substitution. Fluorescent labeling was confirmed by fragmentation experiments using MALDI-TOF/TOF MS. Fluorescent labeled PAC were able to promote ExPEC agglutination when observed with fluorescence microscopy. DTAF tagged PAC may be used to trace the fate of PAC after they agglutinate ExPEC and follow PAC-ExPEC complexes in cell culture assays. PMID:25053065

  16. Quantitative observation of focused-ultrasound-induced vascular leakage and deformation via fluorescein angiography and optical coherence tomography.

    PubMed

    Tsai, Meng-Tsan; Lee, Cheng-Kuang; Lin, Kung-Min; Lin, Yu-Xiang; Lin, Tzu-Han; Chang, Ting-Chia; Lee, Jiann-Der; Liu, Hao-Li

    2013-10-01

    Focused ultrasound (FUS) is a recently discovered noninvasive technique for local and temporal enhancement of vascular permeability, which facilitates drug delivery from the vessels into the surrounding tissue. However, exposure to FUS at a high intensity may cause permanent damage. To investigate the effects of the FUS treatment on blood vessels, we propose to use fluorescein angiography (FA) and optical coherence tomography (OCT) for real-time observation of the diffusion of fluorescence dye from blood vessels and to evaluate the morphological changes of the vessels in vivo. With time-resolved FA imaging, the relationship between the exposed power and the improved permeability of the vessels can be assessed according to the enhancement of the fluorescent intensity due to the dye leakage. Furthermore, the variation of the time-resolved fluorescent intensities can be used to identify the occurrence of dye leakage. In contrast, OCT can be implemented for the reconstruction of tissue microstructures. To quantitatively evaluate the morphological changes of the vessels after the FUS exposure with OCT, a new algorithm was proposed to estimate the vessel area based on the comparison of backscattering properties resulting from the tissue and vascular structures. Results showed that the vessel area increased as the exposed power increased, and the area became significantly larger at a higher FUS exposure power of 10 W. In conclusion, integrated FA and OCT observation can be potentially effective for monitoring the outcome and investigating the effects of FUS treatment. PMID:23812607

  17. The prediction of human skin responses by using the combined in vitro fluorescein leakage/Alamar Blue (resazurin) assay.

    PubMed

    Clothier, Richard; Starzec, Gemma; Pradel, Lionel; Baxter, Victoria; Jones, Melanie; Cox, Helen; Noble, Linda

    2002-01-01

    A range of cosmetics formulations with human patch-test data were supplied in a coded form, for the examination of the use of a combined in vitro permeability barrier assay and cell viability assay to generate, and then test, a prediction model for assessing potential human skin patch-test results. The target cells employed were of the Madin Darby canine kidney cell line, which establish tight junctions and adherens junctions able to restrict the permeability of sodium fluorescein across the barrier of the confluent cell layer. The prediction model for interpretation of the in vitro assay results included initial effects and the recovery profile over 72 hours. A set of the hand-wash, surfactant-based formulations were tested to generate the prediction model, and then six others were evaluated. The model system was then also evaluated with powder laundry detergents and hand moisturisers: their effects were predicted by the in vitro test system. The model was under-predictive for two of the ten hand-wash products. It was over-predictive for the moisturisers, (two out of six) and eight out of ten laundry powders. However, the in vivo human patch test data were variable, and 19 of the 26 predictions were correct or within 0.5 on the 0-4.0 scale used for the in vivo scores, i.e. within the same variable range reported for the repeat-test hand-wash in vivo data. PMID:12405878

  18. Evaluation of cell viability and T2 relaxivity of fluorescein conjugated SPION-PAMAM third generation nanodendrimers for bioimaging.

    PubMed

    Khosroshahi, Mohammad E; Rezvani, Hamideh Alanagh; Keshvari, Hamid; Bonakdar, Shahin; Tajabadi, Maryam

    2016-05-01

    This study has investigated the possibility of using fluorescent dendronized magnetic nanoparticles (FDMNPs) for potential applications in drug delivery and imaging. FDMNPs were first synthesized, characterized and then the effect of Polyamidoamine (PAMAM) dendrimer functionalization and fluorescein isothiocyanate (FITC) conjugation on biocompatibility of superparamagnetic iron oxide nanoparticles (SPIONs) was evaluated. The nanostructures' cytotoxicity tests were performed at different concentrations from 10 to 500μg/mL using MCF-7 and L929 cell lines. IC50 in MTT assay were 139.22 and 201.88μg/mL for DMNP incubated L929 and MCF-7 cell lines respectively, whereas the cell viability for FDMNPs did not decrease to 50%. The results showed that FITC conjugation diminishes the toxicity of dendronized magnetic nanoparticles (DMNPs) mainly due to the reduction of surface charge. DMNP appears to be cytotoxic at the concentration levels being used for both cell lines. On the contrary, FDMNPs showed more biocompatibility and cell viability of MCF-7 and L929 cell lines at all concentrations. The fluorescence microscopy of FDMNPs incubated with MCF-7 cells showed a successful localization of cells indicating their ability for applications such as a magnetic fluorescent probe in cell studies and imaging purposes. T2 relaxivity measurements demonstrated the applicability of the synthesized nanostructures as the contrast agents in tissue differential assessment by altering their relaxation times. In our case, the r2 relaxivity of FDMNPs was measured as 103.67mM(-1)S(-1). PMID:26952457

  19. Fluorescein: a Photo-CIDNP Sensitizer Enabling Hyper-Sensitive NMR Data Collection in Liquids at Low Micromolar Concentration

    PubMed Central

    Okuno, Yusuke; Cavagnero, Silvia

    2016-01-01

    Photochemically induced dynamic nuclear polarization (photo-CIDNP) is a powerful approach for sensitivity enhancement in NMR spectroscopy. In liquids, inter-molecular photo-CIDNP depends on the transient bimolecular reaction between photoexcited dye and sample of interest. Hence the extent of polarization is sample-concentration dependent. This study introduces fluorescein (FL) as a photo-CIDNP dye whose performance is exquisitely tailored to data collection at extremely low sample concentrations. The photo-CIDNP resonance intensities of tryptophan in the presence of either FL or FMN (i.e., the routinely employed flavin mononucleotide photosensitizer) in the liquid state show that FL yields superior sensitivity and enables rapid data collection down to an unprecedented 1 micromolar concentration. This result was achieved on a conventional spectrometer operating at 14.1 Tesla, and equipped with a room-temperature probe (i.e., non-cryogenic). Kinetic simulations show that the excellent behavior of FL arises from its long excited-state triplet lifetime and superior photostability relative to conventional photo-CIDNP sensitizers. PMID:26744790

  20. Stain Specific Standardization of Whole-Slide Histopathological Images.

    PubMed

    Ehteshami Bejnordi, Babak; Litjens, Geert; Timofeeva, Nadya; Otte-Holler, Irene; Homeyer, Andre; Karssemeijer, Nico; van der Laak, Jeroen Awm

    2016-02-01

    Variations in the color and intensity of hematoxylin and eosin (H&E) stained histological slides can potentially hamper the effectiveness of quantitative image analysis. This paper presents a fully automated algorithm for standardization of whole-slide histopathological images to reduce the effect of these variations. The proposed algorithm, called whole-slide image color standardizer (WSICS), utilizes color and spatial information to classify the image pixels into different stain components. The chromatic and density distributions for each of the stain components in the hue-saturation-density color model are aligned to match the corresponding distributions from a template whole-slide image (WSI). The performance of the WSICS algorithm was evaluated on two datasets. The first originated from 125 H&E stained WSIs of lymph nodes, sampled from 3 patients, and stained in 5 different laboratories on different days of the week. The second comprised 30 H&E stained WSIs of rat liver sections. The result of qualitative and quantitative evaluations using the first dataset demonstrate that the WSICS algorithm outperforms competing methods in terms of achieving color constancy. The WSICS algorithm consistently yields the smallest standard deviation and coefficient of variation of the normalized median intensity measure. Using the second dataset, we evaluated the impact of our algorithm on the performance of an already published necrosis quantification system. The performance of this system was significantly improved by utilizing the WSICS algorithm. The results of the empirical evaluations collectively demonstrate the potential contribution of the proposed standardization algorithm to improved diagnostic accuracy and consistency in computer-aided diagnosis for histopathology data. PMID:26353368

  1. Toward Digital Staining using Imaging Mass Spectrometry and Random Forests

    PubMed Central

    Hanselmann, Michael; Köthe, Ullrich; Kirchner, Marc; Renard, Bernhard Y.; Amstalden, Erika R.; Glunde, Kristine; Heeren, Ron M. A.; Hamprecht, Fred A.

    2009-01-01

    We show on Imaging Mass Spectrometry (IMS) data that the Random Forest classifier can be used for automated tissue classification and that it results in predictions with high sensitivities and positive predictive values, even when inter-sample variability is present in the data. We further demonstrate how Markov Random Fields and vector-valued median filtering can be applied to reduce noise effects to further improve the classification results in a post-hoc smoothing step. Our study gives clear evidence that digital staining by means of IMS constitutes a promising complement to chemical staining techniques. PMID:19469555

  2. Port-wine stain--a surgical and psychological problem.

    PubMed

    Malm, M; Carlberg, M

    1988-06-01

    Patients with large and small port-wine stains have been studied using an intensive interview technique. Those with large defects were shown to have reality problems in common. In addition, their experiences were found to be tempered by individual life events and personality dynamics. Those with small port-wine stains who sought treatment showed no reality problems but experienced great individual psychological problems, which in this study are proved to be the same type. The implications of these findings for prophylaxis against mental illness as well as factors to take into consideration when treating these patients are discussed. PMID:3389703

  3. Modeling of alkane emissions from a wood stain

    SciTech Connect

    Chang, J.C.S.; Guo, Z.

    1993-01-01

    The article discusses full-scale residential house tests to evaluate the effects of organic emissions from a wood finishing product--wood stain--on indoor air quality (IAQ). The test house concentrations of three alkane species, nonane, decane, and undecane, were measured as a function of time after the application of the wood stain. It was found that the test house concentrations can be simulated by an integrated IAQ model which takes into consideration source, sink, and ventilation effects. The alkane emissions were controlled by an evaporation-like process.

  4. Evaluation of a fluorescent lectin-based staining technique for some acidophilic mining bacteria

    SciTech Connect

    Fife, D.J.; Bruhn, D.F.; Miller, K.S.; Stoner, D.L.

    2000-05-01

    A fluorescence-labeled wheat germ agglutinin staining technique was modified and found to be effective for staining gram-positive, acidophilic mining bacteria. Bacteria identified by others as being gram positive through 16S rRNA sequence analyses, yet clustering near the divergence of that group, stained weakly. Gram-negative bacteria did not stain. Background staining of environmental samples was negligible, and pyrite and soil particles in the samples did not interfere with the staining procedure.

  5. Lipophilic Dye Staining of Cryptococcus neoformans Extracellular Vesicles and Capsule?

    PubMed Central

    Nicola, Andr Moraes; Frases, Susana; Casadevall, Arturo

    2009-01-01

    Cryptococcus neoformans is an encapsulated yeast that causes systemic mycosis in immunosuppressed individuals. Recent studies have determined that this fungus produces vesicles that are released to the extracellular environment both in vivo and in vitro. These vesicles contain assorted cargo that includes several molecules associated with virulence and implicated in host-pathogen interactions, such as capsular polysaccharides, laccase, urease, and other proteins. To date, visualization of extracellular vesicles has relied on transmission electron microscopy, a time-consuming technique. In this work we report the use of fluorescent membrane tracers to stain lipophilic structures in cryptococcal culture supernatants and capsules. Two dialkylcarbocyanine probes with different spectral characteristics were used to visualize purified vesicles by fluorescence microscopy and flow cytometry. Dual staining of vesicles with dialkylcarbocyanine and RNA-selective nucleic acid dyes suggested that a fraction of the vesicle population carried RNA. Use of these dyes to stain whole cells, however, was hampered by their possible direct binding to capsular polysaccharide. A fluorescent phospholipid was used as additional membrane tracer to stain whole cells, revealing punctate structures on the edge of the capsule which are consistent with vesicular trafficking. Lipophilic dyes provide new tools for the study of fungal extracellular vesicles and their content. The finding of hydrophobic regions in the capsule of C. neoformans adds to the growing evidence for a structurally complex structure composed of polysaccharide and nonpolysaccharide components. PMID:19465562

  6. 5. Downstream elevation, view to southeast. Dark stains on side ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    5. Downstream elevation, view to southeast. Dark stains on side of main girder are from deck drain scuppers, marking deck level within the girders. Compare this view and CA-126-7 to CA-126-19 for indication of severity of siltation of Salt River channel has silted. - Salt River Bridge, Spanning Salt River at Dillon Road, Ferndale, Humboldt County, CA

  7. Image analysis of dye stained patterns in soils

    NASA Astrophysics Data System (ADS)

    Bogner, Christina; Trancón y Widemann, Baltasar; Lange, Holger

    2013-04-01

    Quality of surface water and groundwater is directly affected by flow processes in the unsaturated zone. In general, it is difficult to measure or model water flow. Indeed, parametrization of hydrological models is problematic and often no unique solution exists. To visualise flow patterns in soils directly dye tracer studies can be done. These experiments provide images of stained soil profiles and their evaluation demands knowledge in hydrology as well as in image analysis and statistics. First, these photographs are converted to binary images classifying the pixels in dye stained and non-stained ones. Then, some feature extraction is necessary to discern relevant hydrological information. In our study we propose to use several index functions to extract different (ideally complementary) features. We associate each image row with a feature vector (i.e. a certain number of image function values) and use these features to cluster the image rows to identify similar image areas. Because images of stained profiles might have different reasonable clusterings, we calculate multiple consensus clusterings. An expert can explore these different solutions and base his/her interpretation of predominant flow mechanisms on quantitative (objective) criteria. The complete workflow from reading-in binary images to final clusterings has been implemented in the free R system, a language and environment for statistical computing. The calculation of image indices is part of our own package Indigo, manipulation of binary images, clustering and visualization of results are done using either build-in facilities in R, additional R packages or the LATEX system.

  8. ENHANCED DAPI STAINING FOR CRYPTOSPORIDIUM IN WATER SAMPLES

    EPA Science Inventory

    The U.S. Environmental Protection Agency's Method 1623 is used to detect and quantify the presence of {ital Cryptosporidium} spp. oocysts in water. The protocol consists of concentrating a sample, staining this concentrate with a fluorescent antibody, and examining the sample mi...

  9. Analysis of Oxiclean: An Interesting Comparison of Percarbonate Stain Removers

    ERIC Educational Resources Information Center

    Bracken, Jeffrey D.; Tietz, David

    2005-01-01

    The study focuses on percarbonate-based stain removers since the percarbonate can be heated to produce additional sodium carbonate. An experiment that provides general chemistry students an opportunity to apply their knowledge of basic stoichiometry to solve a relevant, real-world problem is described.

  10. ANEUPLOIDY TEST DEVELOPMENT: KINETOCHORE STAINING IN MAMMALIAN SYSTEMS

    EPA Science Inventory

    The purpose of the project was to determine the feasibility of using human-derived antibodies against the chromosomal kinetochore region coupled with immunofluorescence staining as a method for evaluating the induction of aneuploidy in mammalian cells in vitro and in vivo. The te...

  11. Analytical and microbiological characterization of paper samples exhibiting foxing stains.

    PubMed

    Nunes, Margarida; Relvas, Cátia; Figueira, Francisca; Campelo, Joana; Candeias, António; Caldeira, Ana T; Ferreira, Teresa

    2015-02-01

    This work comprises the use of a multi-analytical approach combined with microbiological studies to characterize six paper samples, containing foxing stains, from the 20th century, regarding their cellulose matrix, fillers, and sizing materials, and to evaluate possible paper degradation that might have occurred during the foxing stains. Photography under different illuminations and optical microscopy were used for morphological characterization of the paper samples and foxing stains. Scanning electron microscopy coupled energy dispersive spectroscopy (SEM-EDS) was of particular importance for defining the presence of fiber disorder and disruption on the surface of some of the stains, and localized accumulations of mineral-like particles on the surface of others. SEM-EDS, attenuated total reflection Fourier transform infrared spectroscopy (ATR-FT-IR), and energy dispersive X-ray fluorescence (EDXRF) were used for the identification of mineral fillers, whereas sizing agents were analyzed using ATR-FT-IR. EDXRF results showed that no differences, within the standard deviation, were found in iron and copper contents between the foxed and unfoxed areas. Fungi belonging to the genus Penicillium spp. were found in all the paper samples. Unfoxed areas presented lower contamination than the foxed areas. PMID:25787782

  12. MODELING OF ALKANE EMISSIONS FROM A WOOD STAIN

    EPA Science Inventory

    The article discusses full-scale residential house tests to evaluate the effects of organic emissions from a wood finishing product--wood stain--on indoor air quality (IAQ). The test house concentrations of three alkane species, nonane, decane, and undecane, were measured as a fu...

  13. Visible fluorescent detection of proteins in polyacrylamide gels without staining.

    PubMed

    Ladner, Carol L; Yang, Jing; Turner, Raymond J; Edwards, Robert A

    2004-03-01

    2,2,2-Trichloroethanol (TCE) incorporated into polyacrylamide gels before polymerization provides fluorescent visible detection of proteins in less than 5min of total processing time. The tryptophans in proteins undergo an ultraviolet light-induced reaction with trihalocompounds to produce fluorescence in the visible range so that the protein bands can be visualized on a 300-nm transilluminator. In a previous study trichloroacetic acid or chloroform was used to stain polyacrylamide gel electrophoresis (PAGE) gels for protein visualization. This study shows that placing TCE in the gel before electrophoresis can eliminate the staining step. The gel is removed from the electrophoresis apparatus and placed on a transilluminator and then the protein bands develop their fluorescence in less than 5min. In addition to being rapid this visualization method provides detection of 0.2microg of typical globular proteins, which for some proteins is slightly more sensitive than the standard Coomassie brilliant blue (CBB) method. Integral membrane proteins, which do not stain well with CBB, are visualized well with the TCE in-gel method. After TCE in-gel visualization the same gel can then be CBB stained, allowing for complementary detection of proteins. In addition, visualization with TCE in the gel is compatible with two-dimensional PAGE, native PAGE, Western blotting, and autoradiography. PMID:14769330

  14. Efficacy of chewing gum in preventing extrinsic tooth staining.

    PubMed

    Yankell, S L; Emling, R C

    1997-01-01

    The purpose of this six-week clinical study was to determine the efficacy of sugar-free chewing gum versus no chewing on preventing Peridex (0.12% chlorhexidine)-associated stain. One-hundred and fifty healthy adult subjects, categorized by tea or coffee intake and smoking, were randomly assigned to a chewing or no chewing gum group. All subjects were given Peridex and an ADA-approved toothbrush and fluoride toothpaste to use twice a day. Gum was chewed for 20 minutes five times each day, after toothbrushing and Peridex rinse in the morning and evening, and after each meal. At baseline, all subjects received a professional cleaning to remove all supragingival deposits and extrinsic strain. At three and six weeks, safety and stain intensity and area were monitored on the anterior teeth and posterior Ramfjord teeth using the Lobene stain scoring method. Seventy-two subjects in each group completed the study. Attrition was unrelated to product use. No untoward reactions were reported or observed at any time in the study. At the six-week evaluations, the chewing gum group exhibited significantly lower (p < 0.05-0.001) total stain scores on both anterior and posterior areas evaluated compared to the no chewing group scores. In addition to the stain evaluations, a randomly selected subset of 60 subjects was evaluated for gingivitis at baseline prior to cleaning, and at three and six weeks, on the buccal and lingual surfaces of the Ramfjord teeth. Both the chewing gum and no chewing gum subset subjects had a significant decrease in gingivitis scores from baseline to three weeks (p < 0.001) and from baseline to six weeks (p < 0.05-0.001). There were no significant statistical differences between the two groups at anytime during the study on gingivitis levels. Chewing gum, after product use, did not reduce the efficacy of chlorhexidine on gingivitis scores. PMID:9586534

  15. Multiple function fluorescein probe performs metal chelation, disaggregation, and modulation of aggregated Aβ and Aβ-Cu complex.

    PubMed

    Muthuraj, B; Layek, Sourav; Balaji, S N; Trivedi, Vishal; Iyer, Parameswar Krishnan

    2015-11-18

    An exceptional probe comprising indole-3-carboxaldehyde fluorescein hydrazone (FI) performs multiple tasks, namely, disaggregating amyloid β (Aβ) aggregates in different biomarker environments such as cerebrospinal fluid (CSF), Aβ1-40 fibrils, β-amyloid lysozyme aggregates (LA), and U87 MG human astrocyte cells. Additionally, the probe FI binds with Cu(2+) ions selectively, disrupts the Aβ aggregates that vary from few nanometers to micrometers, and prevents their reaggregation, thereby performing disaggregation and modulation of amyloid-β in the presence as well as absence of Cu(2+) ion. The excellent selectivity of probe FI for Cu(2+) was effectively utilized to modulate the assembly of metal-induced Aβ aggregates by metal chelation with the "turn-on" fluorescence via spirolactam ring opening of FI as well as the metal-free Aβ fibrils by noncovalent interactions. These results confirm that FI has exceptional ability to perform multifaceted tasks such as metal chelation in intracellular conditions using Aβ lysozyme aggregates in cellular environments by the disruption of β-sheet rich Aβ fibrils into disaggregated forms. Subsequently, it was confirmed that FI had the ability to cross the blood-brain barrier and it also modulated the metal induced Aβ fibrils in cellular environments by "turn-on" fluorescence, which are the most vital properties of a probe or a therapeutic agent. Furthermore, the morphology changes were examined by atomic force microscopy (AFM), polarizable optical microscopy (POM), fluorescence microscopy, and dynamic light scattering (DLS) studies. These results provide very valuable clues on the Aβ (CSF Aβ fibrils, Aβ1-40 fibrils, β-amyloid lysozyme aggregates) disaggregation behavior via in vitro studies, which constitute the first insights into intracellular disaggregation of Aβ by "turn-on" method thereby influencing amyloidogenesis. PMID:26332658

  16. Fluoresceination of FepA during Colicin B Killing: Effects of Temperature, Toxin and TonB

    PubMed Central

    Smallwood, Chuck R.; Marco, Amparo Gala; Xiao, Qiaobin; Trinh, Vy; Newton, Salete M. C.; Klebba, Phillip E.

    2009-01-01

    We studied the reactivity of 35 genetically engineered Cys sulfhydryl groups at different locations in Escherichia coli FepA. Modification of surface loop residues by fluorescein maleimide (FM) was strongly temperature-dependent in vivo, whereas reactivity at other sites was much less affected. Control reactions with bovine serum albumin showed that the temperature dependence of loop residue reactivity was unusually high, indicating that conformational changes in multiple loops (L2, L3, L4, L5, L7, L8, L10) transform the receptor to a more accessible form at 37 °C. At 0 °C colicin B binding impaired or blocked labeling at 8 of 10 surface loop sites, presumably by steric hindrance. Overall, colicin B adsorption decreased the reactivity of more than half of the 35 sites, in both the N - and C- domains of FepA. However, colicin B penetration into the cell at 37 °C did not augment the chemical modification of any residues in FepA. The FM modification patterns were similarly unaffected by the tonB locus. FepA was expressed at lower levels in a tonB host strain, but when we accounted for this decrease its FM-labeling was comparable whether TonB was present or absent. Thus we did not detect TonB-dependent structural changes in FepA, either alone or when it interacted with colicin B at 37 °C. The only changes in chemical modification were reductions from steric hindrance when the bacteriocin bound to the receptor protein. The absence of increases in the reactivity of N-domain residues argues against the idea (Devanathan and Postle, Mol. Microbiol. 65: 441–453, 2007) that the colicin B polypeptide traverses the FepA channel. PMID:19432807

  17. High and Low Molecular Weight Fluorescein Isothiocyanate (FITC)-Dextrans to Assess Blood-Brain Barrier Disruption: Technical Considerations.

    PubMed

    Hoffmann, Angelika; Bredno, Jörg; Wendland, Michael; Derugin, Nikita; Ohara, Peter; Wintermark, Max

    2011-03-01

    This note is to report how histological preparation techniques influence the extravasation pattern of the different molecular sizes of fluorescein isothiocyanate (FITC)-dextrans, typically used as markers for blood-brain barrier leakage. By using appropriate preparation methods, false negative results can be minimized. Wistar rats underwent a 2-h middle cerebral artery occlusion and magnetic resonance imaging. After the last imaging scan, Evans blue and FITC-dextrans of 4, 40, and 70 kDa molecular weight were injected. Different histological preparation methods were used. Sites of blood-brain barrier leakage were analyzed by fluorescence microscopy. Extravasation of Evans blue and high molecular FITC-dextrans (40 and 70 kDa) in the infarcted region could be detected with all preparation methods used. If exposed directly to saline, the signal intensity of these FITC-dextrans decreased. Extravasation of the 4-kDa low molecular weight FITC-dextran could only be detected using freshly frozen tissue sections. Preparations involving paraformaldehyde and sucrose resulted in the 4-kDa FITC-dextran dissolving in these reactants and being washed out, giving the false negative result of no extravasation. FITC-dextrans represent a valuable tool to characterize altered blood-brain barrier permeability in animal models. Diffusion and washout of low molecular weight FITC-dextran can be avoided by direct immobilization through immediate freezing of the tissue. This pitfall needs to be known to avoid the false impression that there was no extravasation of low molecular weight FITC-dextrans. PMID:21423333

  18. Monoclonal antibody-targeted fluorescein-5-isothiocyanate-labeled biomimetic nanoapatites: a promising fluorescent probe for imaging applications.

    PubMed

    Oltolina, Francesca; Gregoletto, Luca; Colangelo, Donato; Gómez-Morales, Jaime; Delgado-López, José Manuel; Prat, Maria

    2015-02-10

    Multifunctional biomimetic nanoparticles (NPs) are acquiring increasing interest as carriers in medicine and basic research since they can efficiently combine labels for subsequent tracking, moieties for specific cell targeting, and bioactive molecules, e.g., drugs. In particular, because of their easy synthesis, low cost, good biocompatibility, high resorbability, easy surface functionalization, and pH-dependent solubility, nanocrystalline apatites are promising candidates as nanocarriers. This work describes the synthesis and characterization of bioinspired apatite nanoparticles to be used as fluorescent nanocarriers targeted against the Met/hepatocyte growth factor receptor, which is considered a tumor associated cell surface marker of many cancers. To this aim the nanoparticles have been labeled with Fluorescein-5-isothiocyanate (FITC) by simple isothermal adsorption, in the absence of organic, possibly toxic, molecules, and then functionalized with a monoclonal antibody (mAb) directed against such a receptor. Direct labeling of the nanoparticles allowed tracking the moieties with spatiotemporal resolution and thus following their interaction with cells, expressing or not the targeted receptor, as well as their fate in vitro. Cytofluorometry and confocal microscopy experiments showed that the functionalized nanocarriers, which emitted a strong fluorescent signal, were rapidly and specifically internalized in cells expressing the receptor. Indeed, we found that, once inside the cells expressing the receptor, mAb-functionalized FITC nanoparticles partially dissociated in their two components, with some mAbs being recycled to the cell surface and the FITC-labeled nanoparticles remaining in the cytosol. This work thus shows that FITC-labeled nanoapatites are very promising probes for targeted cell imaging applications. PMID:25602940

  19. Development of Cell Staining Technique for X-Ray Microscopy

    SciTech Connect

    Tseng, P. Y.; Shih, Y. T.; Liu, C. J.; Hsu, T.; Chien, C. C.; Leng, W. H.; Liang, K. S.; Yin, G. C.; Chen, F. R.; Je, J. H.; Margaritondo, G.; Hwu, Y.

    2007-01-19

    We report a technique for detection of sub-cellular organelles and proteins with hard x-ray microscopy. Several metals were used for enhancing contrast for x-ray microscopy. Osmium tetroxide provides an excellent stain for lipid and can delineate cell membrane. Uranyl acetate has high affinity for nucleotide and can stain nucleus. Immunolocalization of specific proteins and sub-cellular organelles was achieved by 3'3 diaminobenzidine (DAB) with nickel enhancement and nanogold-conjugated secondary antibody with silver enhancement. The x-rays emitted from synchrotron source was monochromatized by double crystal monochromator, the photon energy was fixed at 8 keV to optimize the focusing efficiency of the zone plates. The estimated resolution is about 60 nm. When compared with visible light and conventional confocal microscopy, the X-ray microscopy provides a superior resolution to both conventional optical microscopes.

  20. Selection of ovine oocytes by brilliant cresyl blue staining.

    PubMed

    Wang, Liqin; Lin, Jiapeng; Huang, Juncheng; Wang, Jing; Zhao, Yuncheng; Chen, Tong

    2012-01-01

    Sheep oocytes derived from the ovaries collected from the slaughterhouse are often used for research on in vitro embryo production, animal cloning, transgenesis, embryonic stem cells, and other embryo biotechnology aspects. Improving the in vitro culture efficiency of oocytes can provide more materials for similar studies. Generally, determination of oocyte quality is mostly based on the layers of cumulus cells and cytoplasm or cytoplasm uniformity and colors. This requires considerable experience to better identify oocyte quality because of the intense subjectivity involved (Gordon (2003), Madison et al. (1992) and De Loos et al. (1992)). BCB staining is a function of glucose-6-phosphate dehydrogenase (G6PD) activity, an enzyme synthesized in developing oocytes, which decreases in activity with maturation. Therefore, unstained oocytes (BCB-) are high in G6PD activity, while the less mature oocytes stains are deep blue (BCB+) due to insuffcient G6PD activity to decolorize the BCB dye. PMID:22675245

  1. Selection of Ovine Oocytes by Brilliant Cresyl Blue Staining

    PubMed Central

    Wang, Liqin; Lin, Jiapeng; Huang, Juncheng; Wang, Jing; Zhao, Yuncheng; Chen, Tong

    2012-01-01

    Sheep oocytes derived from the ovaries collected from the slaughterhouse are often used for research on in vitro embryo production, animal cloning, transgenesis, embryonic stem cells, and other embryo biotechnology aspects. Improving the in vitro culture efficiency of oocytes can provide more materials for similar studies. Generally, determination of oocyte quality is mostly based on the layers of cumulus cells and cytoplasm or cytoplasm uniformity and colors. This requires considerable experience to better identify oocyte quality because of the intense subjectivity involved (Gordon (2003), Madison et al. (1992) and De Loos et al. (1992)). BCB staining is a function of glucose-6-phosphate dehydrogenase (G6PD) activity, an enzyme synthesized in developing oocytes, which decreases in activity with maturation. Therefore, unstained oocytes (BCB−) are high in G6PD activity, while the less mature oocytes stains are deep blue (BCB+) due to insuffcient G6PD activity to decolorize the BCB dye. PMID:22675245

  2. Imaging port wine stains by fiber optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Zhao, Shiyong; Gu, Ying; Xue, Ping; Guo, Jin; Shen, Tingmei; Wang, Tianshi; Huang, Naiyan; Zhang, Li; Qiu, Haixia; Yu, Xin; Wei, Xunbin

    2010-05-01

    We develop a fiber optical coherence tomography (OCT) system in the clinical utility of imaging port wine stains (PWS). We use our OCT system on 41 patients with PWS to document the difference between PWS skin and contralateral normal skin. The system, which operates at 4 frames/s with axial and transverse resolutions of 10 and 9 μm, respectively, in the skin tissue, can clearly distinguish the dilated dermal blood vessels from normal tissue. We present OCT images of patients with PWS and normal human skin. We obtain the structural parameters, including epidermal thickness and diameter and depth of dilated blood vessels. We demonstrate that OCT may be a useful tool for the noninvasive imaging of PWS. It may help determine the photosensitizer dose and laser parameters in photodynamic therapy for treating port wine stains.

  3. Identification of active fluorescence stained bacteria by Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Krause, Mario; Beyer, Beatrice; Pietsch, Christian; Radt, Benno; Harz, Michaela; Rösch, Petra; Popp, Jürgen

    2008-04-01

    Microorganisms can be found everywhere e.g. in food both as useful ingredients or harmful contaminations causing food spoilage. Therefore, a fast and easy to handle analysis method is needed to detect bacteria in different kinds of samples like meat, juice or air to decide if the sample is contaminated by harmful microorganisms. Conventional identification methods in microbiology require always cultivation and therefore are time consuming. In this contribution we present an analysis approach to identify fluorescence stained bacteria on strain level by means of Raman spectroscopy. The stained bacteria are highlighted and can be localized easier against a complex sample environment e.g. in food. The use of Raman spectroscopy in combination with chemometrical methods allows the identification of single bacteria within minutes.

  4. Cement line staining in undecalcified thin sections of cortical bone

    NASA Technical Reports Server (NTRS)

    Bain, S. D.; Impeduglia, T. M.; Rubin, C. T.

    1990-01-01

    A technique for demonstrating cement lines in thin, undecalcified, transverse sections of cortical bone has been developed. Cortical bone samples are processed and embedded undecalcified in methyl methacrylate plastic. After sectioning at 3-5 microns, cross-sections are transferred to a glass slide and flattened for 10 min. Sections of cortical bone are stained for 20 sec free-floating in a fresh solution of 1% toluidine blue dissolved in 0.1% formic acid. The section is dehydrated in t-butyl alcohol, cleared in xylene, and mounted with Eukitt's medium. Reversal lines appear as thin, scalloped, dark blue lines against a light blue matrix, whereas bone formation arrest lines are thicker with a smooth contour. With this technique cellular detail, osteoid differentiation, and fluorochrome labels are retained. Results demonstrate the applicability of a one-step staining method for cement lines which will facilitate the assessment of bone remodeling activity in thin sections of undecalcified cortical bone.

  5. Multicolored Stain-free Histopathology with Coherent Raman Imaging

    PubMed Central

    Freudiger, Christian W.; Pfannl, Rolf; Orringer, Daniel A.; Saar, Brian G.; Ji, Minbiao; Zeng, Qing; Ottoboni, Linda; Ying, Wei; Waeber, Christian; Sims, John. R.; De Jager, Philip L.; Sagher, Oren; Philbert, Martin A.; Xu, Xiaoyin; Kesari, Santosh; Xie, X. Sunney; Young, Geoffrey S.

    2013-01-01

    Conventional histopathology with hematoxylin & eosin (H&E) has been the gold standard for histopathological diagnosis of a wide range of diseases. However, it is not performed in vivo and requires thin tissue sections obtained after tissue biopsy, which carries risk, particularly in the central nervous system. Here we describe the development of an alternative, multicolored way to visualize tissue in real time through the use of coherent Raman imaging (CRI), without the use of dyes. CRI relies on intrinsic chemical contrast based on vibrational properties of molecules and intrinsic optical sectioning by nonlinear excitation. We demonstrate that multi-color images originating from CH2 and CH3 vibrations of lipids and protein, as well as two-photon absorption of hemoglobin, can be obtained with subcellular resolution from fresh tissue. These stain-free histopathological images show resolutions similar to those obtained by conventional techniques, but do not require tissue fixation, sectioning or staining of the tissue analyzed. PMID:22906986

  6. Lectins stain cells differentially in the coral, Montipora capitata

    USGS Publications Warehouse

    Work, Thierry M.; Farah, Yael

    2014-01-01

    A limitation in our understanding of coral disease pathology and cellular pathogenesis is a lack of reagents to characterize coral cells. We evaluated the utility of plant lectins to stain tissues of a dominant coral, Montipora capitata, from Hawaii. Of 22 lectins evaluated, nine of these stained structures in the upper or basal body wall of corals. Specific structures revealed by lectins that were not considered distinct or evident on routine hematoxylin and eosin sections of coral tissues included apical and basal granules in gastrodermis and epidermis, cnidoglandular tract and actinopharynx cell surface membranes, capsules of mature holotrichous isorhizas, and perivitelline and periseminal cells. Plant lectins could prove useful to further our understanding of coral physiology, anatomy, cell biology, and disease pathogenesis.

  7. 'Catalysts' for polyacrylamide gel polymerization and detection of proteins by silver staining.

    PubMed

    Hochstrasser, D F; Merril, C R

    1988-01-01

    The crosslinker diacrylyl-piperazine produces polyacrylamide gels which display improved electrophoretic separation of proteins and better physical strength. It also produces gels with improved detection of proteins by ammoniacal silver staining by reducing the background. This reduced background provided us with an opportunity to investigate residual background staining caused by the catalytic reagents utilized in the polymerization of acrylamide gels. The commonly used catalyst system, tetramethyl-ethylenediamine and ammonium persulfate was shown to be responsible for the yellow staining background found after a prolonged development time with silver staining. An alternate catalyst system has been designed to decrease further the formation of this background staining. Dimethyl-piperazine or tetramethylethylenediamine, potassium or ammonium persulfate, and sodium thiosulfate are shown to provide for gels which have excellent mechanical and staining characteristics. These catalytic systems produce little background staining despite prolonged development time with the ammoniacal silver stain, and they reduce background staining with the dichromate silver stain. PMID:2484987

  8. Method and apparatus for staining immobilized nucleic acids

    DOEpatents

    Ramsey, J. Michael; Foote, Robert S.; Jacobson, Stephen C.

    2000-01-01

    A method for staining immobilized nucleic acids includes the steps of affixing DNA probes to a solid substrate, moving target DNA material into proximity with the DNA probes, whereby the target DNA hybridized with specific ones of the DNA probes, and moving a fluorescent dye into proximity with the hybridized target DNA, whereby the fluorescent dye binds to the hybridized DNA to enable subsequent detection of fluorescence.

  9. Genetic Variants Associated with Port-Wine Stains

    PubMed Central

    Wooderchak-Donahue, Whitney; Tan, Oon T.; Margraf, Rebecca; Stevenson, David A.; Grimmer, J. Fredrik; Bayrak-Toydemir, Pinar

    2015-01-01

    Background Port-wine stains (PWS) are capillary malformations, typically located in the dermis of the head and neck, affecting 0.3% of the population. Current theories suggest that port-wine stains are caused by somatic mutations that disrupt vascular development. Objectives Understanding PWS genetic determinants could provide insight into new treatments. Methods Our study used a custom next generation sequencing (NGS) panel and digital polymerase chain reaction to investigate genetic variants in 12 individuals with isolated port-wine stains. Importantly, affected and healthy skin tissue from the same individual were compared. A subtractive correction method was developed to eliminate background noise from NGS data. This allowed the detection of a very low level of mosaicism. Results A novel somatic variant GNAQ, c.547C>G, p.Arg183Gly was found in one case with 4% allele frequency. The previously reported GNAQ c.548G>A, p.Arg183Gln was confirmed in 9 of 12 cases with an allele frequency ranging from 1.73 to 7.42%. Digital polymerase chain reaction confirmed novel variants detected by next generation sequencing. Two novel somatic variants were also found in RASA1, although neither was predicted to be deleterious. Conclusions This is the second largest study on isolated, non-syndromic PWS. Our data suggest that GNAQ is the main genetic determinant in this condition. Moreover, isolated port-wine stains are distinct from capillary malformations seen in RASA1 disorders, which will be helpful in clinical evaluation. PMID:26192947

  10. Coffee Stains from Drops with Receding Contact Lines

    NASA Astrophysics Data System (ADS)

    Freed-Brown, Julian

    2015-03-01

    We present a framework for calculating the surface density profile of a coffee stain deposited by a drying drop with a receding contact line. For standard coffee stains, the fluid pins to the substrate, forces flow towards the exterior of the drop and deposits a thin, concentrated ring of particles. Unlike a pinned drop, a receding drop pushes fluid towards its interior and continuously deposits mass across its substrate as it evaporates. This gives rise to a new class of mountain-like morphologies that are not seen in the standard coffee ring effect but are reminiscent of recent experimental results. For a thin, circular drop with uniform evaporation, we calculate the surface density profile analytically and find that it diverges towards the center of the drop as η ~r - 1 / 2 , where r is the distance from the center. We estimate how this divergence is softened due to solute interactions at the final stage of drying. Our framework can easily be extended numerically or analytically to investigate novel stain morphologies left by drying drops of different shapes and evaporation profiles. This work is part of a thesis project advised by Tom Witten. It was supported in part by the National Science Foundation's MRSEC Program under Award Number DMR 0820054.

  11. Evaluation of immunohistochemical staining for glucagon in human pancreatic tissue

    PubMed Central

    Gurlo, Tatyana; Butle, Peter C.; Butler, Alexandra E.

    2016-01-01

    Immunohistochemistry (IHC) and immunofluorescence (IF) staining techniques are important diagnostic tools of anatomic pathology in the clinical setting and widely used analytical tools in research laboratories. In diabetes research, they are routinely used for the assessment of beta- and alpha-cell mass, for assessment of endocrine cell distribution within the pancreas, for evaluation of islet composition and islet morphology. Here, we present the evaluation of IHC techniques for the detection of alpha-cells in human pancreatic tissue. We compared the Horse Radish Peroxidase (HRP)-based method utilizing DAB Peroxidase Substrate to the Alkaline Phosphatase (AP)-based method utilizing Vector Red substrate. We conclude that HRP–DAB staining is a robust and reliable method for detection of alpha-cells using either rabbit polyclonal or mouse monoclonal anti-glucagon antibodies. However, AP-Vector Red staining should be used with caution, because it is affected by the dehydration with ethanol and toluene preceding the mounting of slides with Permount mounting medium. When AP-Vector Red is a preferable method for alpha-cell labeling, slides should be mounted using aqueous mounting medium or, alternatively, they could be air-dried before permanent mounting PMID:27182095

  12. Standardizing Immunohistochemistry: A New Reference Control for Detecting Staining Problems.

    PubMed

    Sompuram, Seshi R; Vani, Kodela; Tracey, Brian; Kamstock, Debra A; Bogen, Steven A

    2015-09-01

    A new standardized immunohistochemistry (IHC) control for breast cancer testing comprises formalin-fixed human epidermal growth factor receptor 2, estrogen receptor, or progesterone receptor peptide antigens covalently attached to 8-µm glass beads. The antigen-coated beads are suspended in a liquid matrix that hardens upon pipetting onto a glass microscope slide. The antigen-coated beads remain in place through deparaffinization, antigen retrieval, and immunostaining. The intensity of the beads' stain provides feedback regarding the efficacy of both antigen retrieval and immunostaining. As a first report, we tested the sensitivity and specificity of the new IHC controls ("IHControls"). To evaluate sensitivity, various staining problems were simulated. IHControls detected primary and secondary reagent degradation similarly to tissue controls. This first group of IHControls behaved similarly to tissue controls expressing high concentrations of the antigen. The IHControls were also able to detect aberrations in antigen retrieval, as simulated by sub-optimal times or temperatures. Specificity testing revealed that each antigen-coated bead was specific for its cognate IHC test antibody. The data support the conclusion that, like tissue controls, IHControls are capable of verifying the analytic components of an immunohistochemical stain. Unlike tissue controls, IHControls are prepared in large bulk lots, fostering day-to-day reproducibility that can be standardized across laboratories. PMID:25940339

  13. QZ1 and QZ2: Rapid, Reversible Quinoline-Derivatized Fluoresceins for Sensing Biological Zn(II)

    PubMed Central

    Nolan, Elizabeth M.; Jaworski, Jacek; Okamoto, Ken-Ichi; Hayashi, Yasunori; Sheng, Morgan

    2005-01-01

    QZ1, 2-[2-chloro-6-hydroxy-3-oxo-5-(quinolin-8-ylaminomethyl)-3H-xanthen-9-yl]benzoic acid, and QZ2, 2-[6-hydroxy-3-oxo-4,5-bis-(quinolin-8-ylaminomethyl)-3H-xanthen-9-yl]benzoic acid, two fluorescein-based dyes derivatized with 8-aminoquinoline, have been prepared and their photophysical, thermodynamic, and zinc-binding kinetic properties determined. Because of their low background fluorescence and highly emissive Zn(II) complexes, QZ1 and QZ2 have a large dynamic range, with 42- and 150-fold fluorescence enhancements upon Zn(II) coordination, respectively. These dyes have micromolar Kd values for Zn(II) and are selective for Zn(II) over biologically relevant concentrations of the alkali and alkaline earth metals. The Zn(II) complexes also fluoresce brightly in the presence of excess Mn(II), Fe(II), Co(II), Cd(II), and Hg(II), offering improved specificity for Zn(II) over di(2-picolyl)amine-based Zn(II) sensors. Stopped-flow kinetic investigations indicate that QZ1 and QZ2 bind Zn(II) with kon values of (3-4) × 106 M-1 s-1, compared to (6-8) × 105 M-1 s-1 for select ZP (Zinpyr) dyes, at 4.3 °C. Dissociation of Zn(II) from QZ1 and QZ2 occurs with koff values of 150 and 160 s-1, over 5 orders of magnitude larger than those for ZP probes, achieving reversibility on the biological (millisecond) time scale. Laser scanning confocal and two-photon microscopy studies reveal that QZ2 is cell-permeable and Zn(II)-responsive in vivo. Because of its weaker affinity for Zn(II), QZ2 responds to higher concentrations of intracellular Zn(II) than members of the ZP family, illustrating that binding affinity is an important parameter for Zn(II) detection in vivo. PMID:16316228

  14. Cardiac gap junctions and gap junction-associated vesicles: ultrastructural comparison of in situ negative staining with conventional positive staining.

    PubMed

    Chen, L; Goings, G E; Upshaw-Earley, J; Page, E

    1989-03-01

    By comparing in situ negative staining of mammalian heart muscle using La(NO3)3 with conventional positive staining by uranium and lead salts, we showed that 1) the membrane area of rat cardiac gap junctions (GJs) at the intercalated disks is threefold to fourfold greater than previously thought; 2) connexon arrays of cardiac GJ are subdivided into microdomains by connexon-free aisles; 3) profiles of GJ-associated vesicles (GJAVs) of plasmalemmal origin (which are present extracellularly and sharply localized at three extracellular sites) are paired to form GJs with each other and with myocyte plasmalemma; 4) some GJAVs contain arrays of assembled connexons; and 5) myocytes contain intracytoplasmic complexes lying within cylindrical or cigar-shaped membranes and consisting of GJs and multiple vesicles apparently dissociating from these GJs. PMID:2465097

  15. En bloc staining with hydroquinone treatment for block face imaging.

    PubMed

    Togo, Akinobu; Ohta, Keisuke; Higashi, Ryuhei; Nakamura, Kei-Ichiro

    2014-11-01

    IntroductionBecause recent three-dimensional (3D) ultrastructural reconstruction techniques such as serial block face scanning electron microscopy (SBFSEM), obtain their images directly from the flat surface of specimens via material contrast[1], specimens should be strongly stained with heavy metals prior to resin embedding in order to obtain higher material contrast using backscattered electrons (BSEs). To enhance membrane contrast for block face imaging (BFI), we usually stain specimens using the method published by Deerinck[2], and the images obtained show TEM-like contrast.However, recently, our research subjects have required reconstruction of a much larger volume, increasing the total image acquisition time. To reduce the total acquisition time, both high sensitivity detectors and a new specimen preparation method that provides much higher contrast are required. Takahashi et al.[3] have reported that hydroquinone (HQ) treatment during traditional electro-conductive staining increases specimen conductivity and drastically reduces the charge problem for SEM observation. They concluded that HQ treatment might increase the efficiency of secondary electron (SE) generation. Because BFI can be performed using SE as well as BSE, we examined whether addition of HQ treatment to en bloc staining protocols increased the contrast for BFI using SE. Materials & methodsMouse liver tissue was used. Mice were deeply anesthetized by diethyl ether and sodium pentobarbital, and tissues were fixed by transcardial perfusion of 2% paraformaldehyde and 2.5% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.4) through the left ventricle, followed by heparin-containing saline. After perfusion, liver tissues were removed and cut into small cubes approximately 1 mm(3) in the fixative, and were further fixed in the same fixative for 2 h at 4°C. Subsequently, en blocstaining was performed as follows: the specimens were treated using a reduced-OTO staining method (1.5% potassium ferrocyanide-2% OsO4, 1% thiocarbohydrazide, and then 2% OsO4). Subsequently, specimens were treated with 1% HQ solution. Some specimens were exempted from this step and used as controls. Specimens were further stained with 4% uranyl acetate and Walton's lead aspartate solution.After staining, specimens were dehydrated using an ethanol series and embedded in epoxy resin (EPON812, TAAB). Surface of specimens block were cut with a diamond knife, and the newly created flat surfaces of the specimens were coated with evaporated carbon (50 Å) and observed using a SEM (Quanta 3D FEG, FEI).ResultsThe HQ-treated specimens generated a larger amount of SEs than control specimens when subjected to irradiation with the same beam, although BSE numbers were not evidently increased by the treatment. The present results suggest that HQ treatment increases SE generation efficiency, but does not enhance the recruitment of heavy metals into specimens. HQ treatment increased the contrast-to-noise ratio of BFI for images obtained using SEs, and may reduce the total image acquisition time of recently developed 3D reconstruction methods based on SEM. PMID:25359840

  16. A flexible mouse-on-mouse immunohistochemical staining technique adaptable to biotin-free reagents, immunofluorescence, and multiple antibody staining.

    PubMed

    Goodpaster, Tracy; Randolph-Habecker, Julie

    2014-03-01

    Immunohistochemistry on mouse tissue utilizing mouse monoclonal antibodies presents a challenge. Secondary antibodies directed against the mouse monoclonal primary antibody of interest will also detect endogenous mouse immunoglobulin in the tissue. This can lead to significant spurious staining. Therefore, a "mouse-on-mouse" staining strategy is needed to yield credible data. This paper presents a method that is easy to use and highly flexible to accommodate both an avidin-biotin detection system as well as a biotin-free polymer detection system. The mouse primary antibody is first combined with an Fab fragment of an anti-mouse antibody in a tube and allowed sufficient time to form an antibody complex. Any non-complexed secondary antibody is bound up with mouse serum. The mixture is then applied to the tissue. The flexibility of this method is confirmed with the use of different anti-mouse antibodies followed by a variety of detection reagents. These techniques can be used for immunohistochemistry (IHC), immunofluorescence (IF), as well as staining with multiple primary antibodies. This method has also been adapted to other models, such as using human antibodies on human tissue and using multiple rabbit antibodies in dual immunofluorescence. PMID:24152994

  17. [A duplicate staining method for permanent specimen of Trichinella spiralis encapsulated larvae].

    PubMed

    Li, Dan; Yang, Ding; Pi, Ben-Wei; Niu, Li-Na; Zhang, Ying; Wang, Guo-Ying

    2012-04-30

    With single staining method, Trichinella spiralis encapsulated larvae specimens were fixed with formaldehyde alcohol acetic acid fixative solution, and stained with alcohol borax-carmine staining solution (4% borax solution 100 ml, carmine 1 g, and 70% alcohol 100 ml). With duplicate staining, the encapsulated larvae specimens were fixed with formaldehyde alcohol acetic acid fixative solution, and double stained with alcohol borax carmine staining solution and fast green staining solution (fast green 0.1 g, 95% alcohol 100 ml). The results showed that with single staining, it was not clear-cut between the cyst and muscle cells although the larva was differentiable, while with duplicate staining, the larva, cyst and muscle cells were distinguished more clearly. PMID:22908823

  18. Characterization of the indigenous microorganisms in Exter Formation sandstone rock cores obtained during deep drilling and evaluation of contamination by drill mud using fluorescein.

    NASA Astrophysics Data System (ADS)

    Pellizzari, Linda; Neumann, Dominik; Würdemann, Hilke

    2013-04-01

    Microorganisms are very effective catalysts and have an important function in mineral and elemental distribution within geological formations. CO2 injection may influence the microbial activities by affecting the composition of the rock-fluid system. Reactions like mineral dissolution and precipitation, related to biological processes may influence aquifer injectivity or permeability of faults. In subsurface reservoirs, a baseline characterization of pristine rock cores is required to monitor changes in the indigenous microbial communities and to study interactions with geotechnical installations. However, drilling procedures and technical fluids, particularly drill mud, are sources of core contamination. To measure the penetration of drill mud into the cores the tracer fluorescein was tested under laboratory as well as under field conditions. The actual penetration depths seem to be related to differences in geology, such as structural heterogeneities or microfractures. The application of fluorescein was successfully applied during a deep drilling campaign at the CO2 storage pilot site in Ketzin, Germany, in August 2011. During inner coring, crowns of 17.5 mm were removed from the outside. Fluorescein analysis showed that after an inner coring 45% (five samples out of eleven) were not influenced by drill mud. The results highlight that the use of tracers is indispensable to ensuring the quality of core samples for microbiological and biogeochemical analysis. Core samples of the Exter Formation (sandstone above the caprock, 400-440 m depth) were retrieved in order to investigate the indigenous microbial community and to investigate the interaction between CO2, fluid formation, rock substrate and microorganisms in long term experiments with geochemical and molecularbiological techniques. The microbial baseline characterization for rock cores of Exter Formation before CO2 exposure revealed a similar bacterial community composition in all samples. First results of sequence analyses indicated the presence of bacteria related to Acinetobacter sp., Comamonadaceae and Sphingobium sp. which can be found in various deep subsurface or soil habitats (e.g. Ochrobactrum sp.).

  19. Restoration of Fluorosis Stained Teeth: A Case Study.

    PubMed

    Slaska, Barbara; Liebman, Arnold I; Kukleris, Diana

    2015-07-01

    Dental fluorosis manifests by too much ingestion of fluoride resulting in disturbances in enamel mineralization. The result is intrinsic discolorations in the maxillary and mandibular teeth with a poor esthetic appearance. In challenging cases, an esthetic result may be achieved only by a combination of techniques. This case report demonstrates a combination of modalities used to treat a patient presenting with atypical staining as a result of high-level exposure to ingested fluoride present in the drinking water as a child. Conservative treatment consisted of a combination of in-office bleaching to reduce the discoloration and porcelain veneers to create an esthetic result. PMID:26140966

  20. Machine vision system for automated detection of stained pistachio nuts

    NASA Astrophysics Data System (ADS)

    Pearson, Tom C.

    1995-01-01

    A machine vision system was developed to separate stained pistachio nuts, which comprise of about 5% of the California crop, from unstained nuts. The system may be used to reduce labor involved with manual grading or to remove aflatoxin contaminated product from low grade process streams. The system was tested on two different pistachio process streams: the bi- chromatic color sorter reject stream and the small nut shelling stock stream. The system had a minimum overall error rate of 14% for the bi-chromatic sorter reject stream and 15% for the small shelling stock stream.

  1. Neurovascular Stains in Two Girls with Neurofibromatosis 1.

    PubMed

    Crall, Cary S; Rork, Jillian; Maguiness, Sheliagh; Schmidt, Birgitta; Gellis, Stephen

    2016-03-01

    We report two girls, ages 9 and 17 years, with a clinical diagnosis of neurofibromatosis 1 (NF-1) who presented with ill-defined, blanchable, erythematous patches on the dorsal feet and ankles. We hypothesized that these patches were a rare cutaneous finding in NF-1 that exist on a clinical and histopathologic spectrum with previously described NF-1 vascular lesions including blue-red macules and skin ulceration due to NF-1 vasculopathy. We suggest that these lesions be called neurovascular stains (NVSs) to unify their clinical and histopathologic features. PMID:26861968

  2. 10. Photocopy of an engraving of a stained glass window ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    10. Photocopy of an engraving of a stained glass window design by Johann Friedrich Overbeck (1789-1869) on which two of the chancel windows in the Church of the Holy Cross are thought to have been based. This copy is of a photocopy obtained from the Treasury of Notre Dame de Paris, Paris, France, by the late Mrs. Walter C. White of Stateburg, South Carolina. Mrs. White's photocopy is in the possession of Mrs. Richard K. Anderson of the Borough House at Stateburg. - Church of the Holy Cross, State Route 261, Stateburg, Sumter County, SC

  3. DNA Stains as Surrogate Nucleobases in Fluorogenic Hybridization Probes.

    PubMed

    Hövelmann, Felix; Seitz, Oliver

    2016-04-19

    The increasing importance assigned to RNA dynamics in cells and tissues calls for probe molecules that enable fluorescence microscopy imaging in live cells. To achieve this goal, fluorescence dyes are conjugated with oligonucleotides so as to provide strong emission upon hybridization with the target molecule. The impressive 10(3)-fold fluorescence intensification observed when DNA stains such as thiazole orange (TO) interact with double-stranded DNA is intriguing and prompted the exploration of oligonucleotide conjugates. However, nonspecific interactions of DNA stains with polynucleotides tend to increase background, which would affect the contrast achievable in live-cell imaging. This Account describes the development of DNA-stain-labeled hybridization probes that provide high signal-to-background. We focus on our contributions in context with related advances from other laboratories. The emphasis will be on the requirements of RNA imaging in live cells. To reduce background, intercalator dyes such as TO were appended to peptide nucleic acid (PNA), which is less avidly recognized by DNA stains than DNA/RNA. Constraining the TO dye as a nucleobase surrogate in "forced intercalation (FIT) probes" improved the target specificity, presumably by helping to prevent unspecific interactions. The enforcement of TO intercalation between predetermined base pairs upon formation of the probe-target duplex provided for high brightness and enabled match/mismatch selectivity beyond stringency of hybridization. We show examples that highlight the use of PNA FIT probes in the imaging of mRNA, miRNA, and lncRNA in living cells. The "FIT approach" was recently extended to DNA probes. Signal brightness can become limiting when low-abundance targets ought to be visualized over cellular autofluorescence. We discuss strategies that further the brightness of signaling by FIT probes. Multilabeling with identical dyes does not solve the brightness issue. To avoid self-quenching, we combined two different yet spectrally overlapping fluorescent base surrogates. A hybridization-sensitive dye serves as a light collector that transfers energy to a brightly emissive acceptor dye. To improve the brilliance of single-dye probes, the "TO-nucleotide" was accompanied by an adjacent locked nucleic acid (LNA) unit. The LNA-constrained FIT probes are responsive and bright, enabling the tracking of mRNA transport in living tissue. We also show that the color repertoire of FIT probes is not restricted to the green-emissive TO but can be expanded to cyan and red. A new base surrogate (4,4-linked bisquinoline) provided up to 195-fold enhancement of the fluorescence. PMID:26963493

  4. High resolution structures of the 4-4-20 Fab-fluorescein complex in two solvent systems: effects of solvent on structure and antigen-binding affinity.

    PubMed Central

    Herron, J N; Terry, A H; Johnston, S; He, X M; Guddat, L W; Voss, E W; Edmundson, A B

    1994-01-01

    Three-dimensional structures were determined for three crystal forms of the antigen binding fragment (Fab) of anti-fluorescein antibody 4-4-20 in complex with fluorescein. These included 1) a triclinic (P1) form crystallized in 47% (v/v) 2-methyl-2,4-pentanediol (MPD); 2) a triclinic (P1) form crystallized in 16% (w/v) poly(ethylene glycol), molecular weight 3350 (PEG); and 3) a monoclinic (P21) form crystallized in 16% PEG. Solvent molecules were added to the three models and the structures were refined to their diffraction limits (1.75-A, 1.78-A, and 2.49-A resolution for the MPD, triclinic PEG, and monoclinic PEG forms, respectively). Comparisons of these structures were interesting because 4-4-20 exhibited a lower antigen-binding affinity in 47% MPD (Ka = 1.3 x 10(8) M-1) than in either 16% PEG (Ka = 2.9 x 10(9) M-1) or phosphate-buffered saline (Ka = 1.8 x 10(10) M-1). Even though the solution behavior of the antibody was significantly different in MPD and PEG, the crystal structures were remarkably similar. In all three structures, the fluorescein-combining site was an aromatic slot formed by tyrosines L32, H96, and H97 and tryptophans L96 and H33. In addition, several active site constituents formed an electrostatic network with the ligand. These included a salt link between arginine L34 and one of fluorescein's enolate oxygen atoms, a hydrogen bond between histidine L27d and the second enolic group, a hydrogen bond between tyrosine L32 and the phenylcarboxylate group, and two medium range (approximately 5 A) electrostatic interactions with lysine L50 and arginine H52. The only major difference between the triclinic MPD and PEG structures was the degree of hydration of the antigen-combining site. Three water molecules participated in the above electrostatic network in the MPD structure, while eight were involved in the PEG structure. Based on this observation, we believe that 4-4-20 exhibits a lower affinity in MPD due to the depletion of the hydration shell of the antigen-combining site. Images FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 6 FIGURE 7 PMID:7696460

  5. Ring stains in the presence of electromagnetohydrodynamic interactions.

    PubMed

    Das, Siddhartha; Mitra, Sushanta K; Chakraborty, Suman

    2012-11-01

    In a recent paper [Das et al., Phys. Rev. E 85, 046311 (2012)], we delineated the role of electrokinetic transport in modifying the classical "coffee stain" effect. In this study, we extend this calculation to incorporate the consequences of a generalized electromagnetohydrodynamic transport in the coffee stain phenomenon. The magnetohydrodynamic (MHD) effect enhances the velocities at the beginning of the drop life, whereas the electrokinetic effect increases the "disordering" effect in particle deposition at the end of the drop, triggered by a velocity divergence. For a suitable combination of the strength of the MHD and electrokinetic transport, however, this disordering effect is substantially enhanced, and, most nonintuitively, such velocity divergence and the disordering effect may occur at a time that is much earlier than the end of the drop life, or may occur even instantaneously after the start of the drop evaporation. This work will provide useful insight in the understanding of the dynamics of mesoscopic patterns formed as the magnetic nanocrystals deposit in the presence of a combined transport driven by evaporation and magnetic field effects. PMID:23214885

  6. Immunocytochemical staining on ThinPrep processed smears.

    PubMed

    Leung, S W; Bédard, Y C

    1996-03-01

    This study reports our first-year experience in the immunocytochemical staining of specimens prepared by the ThinPrep processor. The avidin-biotin-complex technique was applied to smears of 70 cytological specimens, including fine-needle aspiration samples from various sites (56 specimens), body fluids (13 specimens) and urine (one specimen). The specimens were collected in CytoLyt solution to lyse red blood cells and then resuspended in the processing solution. The smears were made by the processor and fixed in 95% alcohol. The panel of antibodies used included alpha-fetoprotein, carcinoembryonic antigen, cytokeratin, chromogranin A, HMB-45, beta-human chorionic gonadotropin (beta-HCG), Kappa, Lambda, Ki-1, leucocyte common antigen, L26, neuron-specific enolase, 013, prostatic acid phosphatase, prostatic-specific antigen, S100, thyroglobulin, UCHL-1, and vimentin. The various antigens could be easily demonstrated in the ThinPrep smears, except for some lymphoma markers. Nonspecific background staining caused by blood and proteinaceous debris was virtually absent. In general, the monolayer preparation made the interpretation easier than conventional preparations. The only problem with this technique was in the classification of lymphomas where immunotyping was inconclusive. Our preliminary results indicate that ThinPrep smears are suitable for immunocytochemical studies, except for the classification of lymphomas. PMID:8685232

  7. Analysis of surface stains on modern gold coins

    NASA Astrophysics Data System (ADS)

    Corregidor, V.; Alves, L. C.; Cruz, J.

    2013-07-01

    It is a mandatory practice in the European Mint Houses to provide a certificate of guarantee of their products specially when issuing commemorative gold or silver coins. This practise should assure satisfaction and trust both for the mint house and for the demanding numismatic collector. For these reasons the Mint Houses follow a strict quality control in all the production steps in order to ensure a no-defect, fully supervised output. In spite of all the undertaken precautions, different surface stains with diverse origin on gold coins recently minted in Europe were observed. Those were compositionally studied by means of IBA techniques at the end-stage nuclear microprobe installed at IST/ITN. From this study it was possible to identify several possible sources for these stains. The presence of defects at the surface of these commemorative coins address the need of improving the quality control system and the results here presented point out where these improvements should occur, in order to reduce/eliminate them and give the customer a product that with time probably will be revalued.

  8. Sperm DNA fragmentation is related to sperm morphological staining patterns.

    PubMed

    Sá, Rosália; Cunha, Mariana; Rocha, Eduardo; Barros, Alberto; Sousa, Mário

    2015-10-01

    In this prospective comparative study, sperm DNA fragmentation (sDNAfrag) was compared at each step of a sequential semen preparation, with semen parameters according to their degree of severity. At each step (fractions) of the sequential procedure, sDNAfrag was determined: fresh (Raw), after gradient centrifugation, washing, and swim-up (SU) for 70 infertile men enrolled in intracytoplasmic sperm injection cycles. sDNAfrag significantly (P = 0.04; P < 0.0001) decreased throughout the steps of semen preparation, with centrifugation and washing not increasing it. A negative correlation to sperm motility was observed in Raw and SU fractions, and a higher sDNAfrag was observed in samples with lower semen quality. Our results confirm that the steps of the sequential procedure do not compromise sperm DNA integrity and progressively decreased sDNAfrag regardless of the sperm abnormality and that semen parameters with lower quality present higher sDNAfrag. Four distinct patterns were observed, of which the entire sperm head staining was the pattern most expressed in all studied fractions. Additionally, the sperm head gene-rich region staining pattern was reduced by the procedure. This suggests that pattern quantification might be a useful adjunct when performing sDNAfrag testing for male infertility. PMID:26278809

  9. Coproduction of detergent compatible bacterial enzymes and stain removal evaluation.

    PubMed

    Niyonzima, Francois N; More, Sunil S

    2015-10-01

    Most of the detergents that are presently produced contain the detergent compatible enzymes to improve and accelerate the washing performance by removing tough stains. The process is environment friendly as the use of enzymes in the detergent formulation reduces the utilization of toxic detergent constituents. The current trend is to use the detergent compatible enzymes that are active at low and ambient temperature in order to save energy and maintain fabric quality. As the detergent compatible bacterial enzymes are used together in the detergent formulation, it is important to co-produce the detergent enzymes in a single fermentation medium as the enzyme stability is assured, and production cost gets reduced enormously. The review reports on the production, purification, characterization and application of detergent compatible amylases, lipases, and proteases are available. However, there is no specific review or minireview on the concomitant production of detergent compatible amylases, lipases, and proteases. In this minireview, the coproduction of detergent compatible enzymes by bacterial species, enzyme stability towards detergents and detergent components, and stain release analysis were discussed. PMID:26011283

  10. The Golgi Stain: invention, diffusion and impact on neurosciences.

    PubMed

    Pannese, E

    1999-08-01

    The black reaction, invented in 1873 by Camillo Golgi (1843-1926, was the first technique to reveal neurons in their entirety, i.e. with all their processes. This important development passed unnoticed at first and only received wide international attention after a long delay. The Golgi stain was widely employed for almost thirty years and was directly responsible for major advances in our knowledge of the microscopic anatomy of the nervous system, as well as in other fields of study. In the hands of other researchers, the black reaction provided vital evidence that helped to establish the neuron theory. The Golgi stain was almost forgotten in the period between the two World Wars, but the introduction of the electron microscope to neurocytological resarch revived its use around the middle of the twentieth century. Today, the black reaction is still used extensively not only in combination with electron microscopy, but also as an autonomous technique in studies on the evolution, ontogeny, and organization of the nervous system. PMID:11624294

  11. Color stability of ceramic brackets immersed in potentially staining solutions

    PubMed Central

    Guignone, Bruna Coser; Silva, Ludimila Karsbergen; Soares, Rodrigo Villamarim; Akaki, Emilio; Goiato, Marcelo Coelho; Pithon, Matheus Melo; Oliveira, Dauro Douglas

    2015-01-01

    OBJECTIVE: To assess the color stability of five types of ceramic brackets after immersion in potentially staining solutions. METHODS: Ninety brackets were divided into 5 groups (n = 18) according to brackets commercial brands and the solutions in which they were immersed (coffee, red wine, coke and artificial saliva). The brackets assessed were Transcend (3M/Unitek, Monrovia, CA, USA), Radiance (American Orthodontics, Sheboygan, WI, USA), Mystique (GAC International Inc., Bohemia, NY, USA) and Luxi II (Rocky Mountain Orthodontics, Denver, CO, USA). Chromatic changes were analyzed with the aid of a reflectance spectrophotometer and by visual inspection at five specific time intervals. Assessment periods were as received from the manufacturer (T0), 24 hours (T1), 72 hours (T2), as well as 7 days (T3) and 14 days (T4) of immersion in the aforementioned solutions. Results were submitted to statistical analysis with ANOVA and Bonferroni correction, as well as to a multivariate profile analysis for independent and paired samples with significance level set at 5%. RESULTS: The duration of the immersion period influenced color alteration of all tested brackets, even though these changes could not always be visually observed. Different behaviors were observed for each immersion solution; however, brackets immersed in one solution progressed similarly despite minor variations. CONCLUSIONS: Staining became more intense over time and all brackets underwent color alterations when immersed in the aforementioned solutions. PMID:26352842

  12. Rapid, sensitive, and simple method for quantification of both neurotoxic and neurotrophic effects of NMDA on cultured cerebellar granule cells.

    PubMed

    Didier, M; Heaulme, M; Soubrié, P; Bockaert, J; Pin, J P

    1990-09-01

    A simple and sensitive method adapted from the staining of living cells with fluorescein diacetate was developed to rapidly estimate the number of living cells remaining in a culture dish 24 hr after a few min of NMDA treatment of cerebellar neurons. This method consists of the measurement, after cell lysis, of the total amount of fluorescein produced from fluorescein diacetate by the living granule cells present in each culture dish. We show that this method can also be used to quantify the survival effect of chronic exposure of granule cells to either K+ or NMDA. In both cases, the fluorescence measured was found to be proportional to the number of fluorescein-labelled cells counted under a fluorescence microscope, indicating that the present method can be used to quantify both toxic and trophic effects of NMDA on cerebellar granule cells. This study confirms that these two NMDA effects occur at the same NMDA concentration, and both are inhibited by MK 801 in the same concentration range. We showed, moreover, that granule neurons developed in the presence of NMDA are much less sensitive to NMDA toxicity than neurons developed in K(+)-enriched medium. PMID:1979352

  13. No Evidence for Interference of Hematoxylin and Eosin (HE) Staining in DNA Testing: Utility of DNA Extraction from HE-Stained Archival Tissue Sections

    PubMed Central

    Morikawa, Teppei; Shima, Kaori; Kuchiba, Aya; Yamauchi, Mai; Tanaka, Noriko; Imamura, Yu; Liao, Xiaoyun; Qian, Zhi Rong; Brahmandam, Mohan; Longtine, Janina A.; Lindeman, Neal I.; Fuchs, Charles S.; Ogino, Shuji

    2012-01-01

    Although histochemical staining has been believed to inhibit DNA amplification reaction, no previous study has systematically evaluated the influence of histochemical staining on downstream molecular assays. To evaluate an influence of hematoxylin and eosin (HE) staining on DNA testing, we isolated DNA from 10 unstained, 10 hematoxylin-stained, 10 eosin-stained or 10 HE-stained tissue sections (ie, 4 groups), from each of 5 colon cancers. Among those 4 groups, we did not observe any significant or appreciable difference in DNA fragmentation by agarose gel electrophoresis; in DNA amplification by real-time PCR; in microsatellite PCR fragment analyses; or in PCR-Pyrosequencing. As a proof-of-principle study, we successfully performed microsatellite instability analysis and sequencing of KRAS and BRAF on over 1300 colorectal cancers using DNA extracted from HE stained tissue sections. Our data provide no evidence for interfering effect of HE staining on DNA testing, suggesting that DNA from HE-stained sections can be effectively used for routine DNA testing. PMID:22706867

  14. Sensitive, quantitative, and fast modifications for Coomassie Blue staining of polyacrylamide gels.

    PubMed

    Westermeier, Reiner

    2006-09-01

    In spite of the high sensitivity of silver staining and the wide dynamic range of various fluorescent detection methods, Coomassie Brilliant Blue staining is still the most widely used protein detection technique for proteins separated by polyacrylamide gel electrophoresis. There are several reasons: Low price, Visible with the eye, Desk top scanners can be employed for image acquisition, Better for quantitative analysis than silver staining, Possible modifications for fast or highly sensitive staining, Mass spectrometry compatible. PMID:17031800

  15. Substituent effect study on experimental ¹³C NMR chemical shifts of (3-(substituted phenyl)-cis-4,5-dihydroisoxazole-4,5-diyl)bis(methylene)diacetate derivatives.

    PubMed

    Kara, Yesim S

    2015-12-01

    Eleven novel (3-(substituted phenyl)-cis-4,5-dihydroisoxazole-4,5-diyl)bis(methylene) diacetate derivatives were synthesized in the present study. These dihydroisoxazole derivatives were characterized by IR, (1)H NMR, (13)C NMR and elemental analyses. Their (13)C NMR spectra were measured in Deuterochloroform (CDCl3). The correlation analysis for the substituent-induced chemical shift (SCS) with Hammett substituent constant (σ), inductive substituent constant (σI), different of resonance substituent constants (σR, σR(o)) and Swain-Lupton substituent parameters (F, R) were performed using SSP (single substituent parameter), and DSP (dual substituent parameter) methods, as well as single and multiple regression analysis. From the result of regression analysis, the effect of substituent on the (13)C NMR chemical shifts was explained. PMID:26172459

  16. 19 CFR 10.52 - Painted, colored or stained glass windows for religious institutions.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Painted, colored or stained glass windows for.... General Provisions Works of Art § 10.52 Painted, colored or stained glass windows for religious institutions. When painted, colored, or stained glass windows or parts thereof, are claimed free of duty...

  17. 19 CFR 10.52 - Painted, colored or stained glass windows for religious institutions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 19 Customs Duties 1 2012-04-01 2012-04-01 false Painted, colored or stained glass windows for.... General Provisions Works of Art § 10.52 Painted, colored or stained glass windows for religious institutions. When painted, colored, or stained glass windows or parts thereof, are claimed free of duty...

  18. 19 CFR 10.52 - Painted, colored or stained glass windows for religious institutions.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 19 Customs Duties 1 2011-04-01 2011-04-01 false Painted, colored or stained glass windows for.... General Provisions Works of Art § 10.52 Painted, colored or stained glass windows for religious institutions. When painted, colored, or stained glass windows or parts thereof, are claimed free of duty...

  19. 19 CFR 10.52 - Painted, colored or stained glass windows for religious institutions.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 19 Customs Duties 1 2014-04-01 2014-04-01 false Painted, colored or stained glass windows for.... General Provisions Works of Art § 10.52 Painted, colored or stained glass windows for religious institutions. When painted, colored, or stained glass windows or parts thereof, are claimed free of duty...

  20. 19 CFR 10.52 - Painted, colored or stained glass windows for religious institutions.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 19 Customs Duties 1 2013-04-01 2013-04-01 false Painted, colored or stained glass windows for.... General Provisions Works of Art § 10.52 Painted, colored or stained glass windows for religious institutions. When painted, colored, or stained glass windows or parts thereof, are claimed free of duty...

  1. A Comparison of Heat versus Methanol Fixation for Gram Staining Bacteria

    ERIC Educational Resources Information Center

    Minnerath, Jeanne M.; Roland, Jenna M.; Rossi, Lucas C.; Weishalla, Steven R.; Wolf, Melissa M.

    2009-01-01

    Gram staining bacteria is a fundamental technique introduced in general biology and microbiology laboratory courses. Two common problems students encounter when Gram staining bacteria are (1) having a difficult time locating bacterial cells on the microscope slide and (2) over-decolorizing bacterial cells during the staining procedure such that…

  2. Apparatus for fixing, staining, and rinsing of tissue cultures for fluorescent-antibody testing.

    PubMed

    Poole, G M

    1972-08-01

    A staining tray and tray-housing container have been developed to facilitate fluorescent-antibody staining of tissue cultures on cover slips, which allows fixing, staining, and rinsing with a minimum of handling. Breakage and loss of cells were negligible. PMID:16349928

  3. OIL RED AS A HISTOCHEMICAL STAIN FOR NATURAL FIBERS AND PLANT CUTICLE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Oil Red was evaluated as a histochemical stain for the cuticle of plants and for components in cotton and flax fibers. A positive reaction for arachidyl stearate and as well as differential staining of plants after sequential extraction of fatty acids and alcohols confirmed that Oil Red stained wa...

  4. Ink and Vinegar, a Simple Staining Technique for Arbuscular-Mycorrhizal Fungi

    PubMed Central

    Vierheilig, Horst; Coughlan, Andrew P.; Wyss, Urs; Piché, Yves

    1998-01-01

    We developed a reliable, inexpensive, and simple method for staining arbuscular-mycorrhizal fungal colonizations in root tissues. Apart from applications in research, this nontoxic, high-quality staining method also could be of great utility in teaching exercises. After adequate clearing with KOH, an ink-vinegar solution successfully stained all fungal structures, rendering them clearly visible. PMID:9835596

  5. Facilitating normal physiology in the presence of meconium stained liquor.

    PubMed

    Hudson, Julika

    2015-06-01

    There is sufficient evidence to support the practice of optimal cord clamping in normal labour and birth. In this paper, the physiology of meconium stained liquor (MSL), meconium aspiration syndrome and the practice of optimal cord clamping in babies born through MSL, is discussed. Guidelines suggest not stimulating babies born through MSL, at birth, to avoid aspiration. However, the obvious stimulation resulting from early clamping and cutting the cord, leaves a baby with no choice but to inhale, but this appears to be overlooked in practice. Midwives in their role as supporters of normal physiology are in a position to question this routine intervention in the absence of any evidence to support it. PMID:26320331

  6. Method for Whole Mount Antibody Staining in Chick

    PubMed Central

    Psychoyos, Delphine; Finnell, Richard

    2009-01-01

    The chick embryo is a valuable tool in the study of early embryonic development. Its transparency, accessibility and ease of manipulation, make it an ideal tool for studying antibody expression in developing brain, neural tube and somite. This video demonstrates the different steps in whole-mount antibody staining using HRP conjugated secondary antibodies; First, the embryo is dissected from the egg and fixed in paraformaldehyde. Second, endogenous peroxidase is inactivated; The embryo is then exposed to primary antibody. After several washes, the embryo is incubated with secondary antibody conjugated to HRP. Peroxidase activity is revealed using reaction with diaminobenzidine substrate. Finally, the embryo is fixed and processed for photography and sectioning. The advantage of this method over the use of fluorescent antibodies is that embryos can be processed for wax sectioning, thus enabling the study of antigen sites in cross section. This method was originally introduced by Jane Dodd and Tom Jessell 1. PMID:19488030

  7. Photonic Crystal Hydrogel Enhanced Plasmonic Staining for Multiplexed Protein Analysis.

    PubMed

    Mu, Zhongde; Zhao, Xiangwei; Huang, Yin; Lu, Meng; Gu, Zhongze

    2015-12-01

    Plasmonic nanoparticles are commonly used as optical transducers in sensing applications. The optical signals resulting from the interaction of analytes and plamsonic nanoparticles are influenced by surrounding physical structures where the nanoparticles are located. This paper proposes inverse opal photonic crystal hydrogel as 3D structure to improve Raman signals from plasmonic staining. By hybridization of the plasmonic nanoparticles and photonic crystal, surface-enhanced Raman spectroscopy (SERS) analysis of multiplexed protein is realized. It benefits the Raman analysis by providing high-density "hot spots" in 3D and extra enhancement of local electromagnetic field at the band edge of PhC with periodic refractive index distribution. The strong interaction of light and the hybrid 3D nanostructure offers new insights into plasmonic nanoparticle applications and biosensor design. PMID:26436833

  8. Vegetable cells in Papanicolaou-stained cervical smears.

    PubMed

    Rivasi, Francesco; Tosi, Giovanni; Ruozi, Barbara; Curatola, Carlo

    2006-01-01

    Vegetable cells are unusual findings in Papanicolaou-stained cervical smears; these structures could be wrongly mistaken for abnormal human cells, worm eggs, or spores by a cytologist encountering the possibility of meeting those elements in cytological analysis. Five cervicovaginal smears showing similar vegetable cells have been detected over a 3-yr period (2002-2004) in the course of a population screening program for cancer of the uterine cervix in Modena (Italy) involving 32,500 women. According to the clinical histories of the patients, the vaginal pharmaceutical drugs or appliances used were of different types: vaginal lavages, pessaries, and vaginal creams. Following a careful investigation, the only substance that can lead to vegetal elements has been identified as polysaccharide galactomannan, which is one of the excipient present in the drugs used. The authors have identified the origin of these contaminants and the means of pollution, using cytological and pharmaceutical investigation. PMID:16355382

  9. Early morphea mimicking acquired port-wine stain.

    PubMed

    Pickert, Amanda J; Carpentieri, David; Price, Harper; Hansen, Ronald C

    2014-01-01

    We report the case of a 2.5-year-old girl with linear morphea initially diagnosed as an acquired port-wine stain (PWS). She underwent three treatments to the right face using the pulsed dye laser (PDL) before sclerotic changes were observed and the correct diagnosis was confirmed with histopathology. Treatment using the PDL reduced the skin erythema but did not prevent subsequent sclerosis. The sclerosis became most prominent superior to the patient's right ear in an area not treated using the laser. A review of the English-language medical literature identified no cases of morphea triggered using a PDL, but there were several reports of early morphea misdiagnosed as an acquired PWS. Briefly, we review those cases, as well as morphea subtypes, and comment on how the pathophysiology of morphea may lend itself to an early underrecognized inflammatory presentation, delaying diagnosis. PMID:23627630

  10. Effect of droplet shape on ring stains from dried liquid

    NASA Astrophysics Data System (ADS)

    Santiago, Melvin; Brown, Katherine; Mathur, Harsh

    A landmark experimental paper on coffee stains by Deegan et al included a simple theoretical analysis of circular droplets. The analysis was based on a model informally called the Maxwell House equations. It describes the evolving height profile of the droplet, the evaporation of the solvent and the outflow of solute to the rim of the droplet. Since typical droplets are not circles, here we extend the analysis to more general shapes. We find that for thin droplets the height profile may be determined by solving Poisson's equation in a domain corresponding to the footprint of the droplet. Evaporation is treated in a simple approximation via an electrostatic analogy and is dominated by the sharp edges of the droplet. Assuming zero vorticity allows us to analyze the solvent flow in droplets of arbitrary shape. We compare circular droplets to other shapes including long linear droplets, ring shaped droplets and droplets with an elliptical footprint

  11. [DNA quantification in nuclei of cultivated mushroom with DAPI staining].

    PubMed

    Pancheva, E V; Volkova, V N; Kamzolkina, O V

    2004-01-01

    Agaricus bisporus (Lange) Imbach is actively cultivated amphithallic basidiomycete, in which various strains are primary homothallic, heterothallic or secondary homothallic. Countings of relative nuclear DNA content by means of DAPI stain and its comparison in different strains can help to understand the mushroom's life cycle features. The authors for the first time observed change of nuclear phases in basidia of A. bisporus strains with different types of life cycle and revealed that DNA content in diploid nuclei is about 1.3 times higher than in haploid ones. The method is highly sensitive and can be used for quantitative measurings of nuclear DNA even in objects with nuclei of about 1 mkm in diameter. PMID:15346797

  12. Sizing of single fluorescently stained DNA fragments by scanning microscopy

    PubMed Central

    Laib, Stephan; Rankl, Michael; Ruckstuhl, Thomas; Seeger, Stefan

    2003-01-01

    We describe an approach to determine DNA fragment sizes based on the fluorescence detection of single adsorbed fragments on specifically coated glass cover slips. The brightness of single fragments stained with the DNA bisintercalation dye TOTO-1 is determined by scanning the surface with a confocal microscope. The brightness of adsorbed fragments is found to be proportional to the fragment length. The method needs only minute amount of DNA, beyond inexpensive and easily available surface coatings, like poly-l-lysine, 3-aminoproyltriethoxysilane and polyornithine, are utilizable. We performed DNA-sizing of fragment lengths between 2 and 14 kb. Further, we resolved the size distribution before and after an enzymatic restriction digest. At this a separation of buffers or enzymes was unnecessary. DNA sizes were determined within an uncertainty of 7–14%. The proposed method is straightforward and can be applied to standardized microtiter plates. PMID:14602931

  13. Fat tissue staining and photodynamic/photothermal effects

    NASA Astrophysics Data System (ADS)

    Tuchin, Valery V.; Altshuler, Gregory B.; Yanina, Irina Yu.; Kochubey, Vyacheslav I.; Simonenko, Georgy V.

    2010-02-01

    Cellulite is considered as a disease of the subcutaneous fat layer that appears mostly in women and consists of changes in fat cell accumulation together with disturbed lymphatic drainage, affecting the external appearance of the skin. The photodynamic and selective photothermal treatments may provide reduction the volume of regional or sitespecific accumulations of subcutaneous adipose tissue on the cellular level. We hypothesize that light irradiation of stained fat tissue at selected temperature leads to fat cell lypolytic activity (the enhancement of lipolysis of cell triglycerides due to expression of lipase activity and cell release of free fat acids (FFAs) due to temporal cell membrane porosity), and cell killing due to apoptosis caused by the induced fat cell stress and/or limited cell necrosis.

  14. An indoxyl compound 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, suppresses activation of Fyn kinase in mast cells and IgE-mediated allergic responses in mice

    SciTech Connect

    Lee, Jun Ho; Kim, Tae Hyung; Kim, Hyuk Soon; Kim, A-Ram; Kim, Do-Kyun; Nam, Seung Taek; Kim, Hyun Woo; Park, Young Hwan; Her, Erk; Park, Yeong Min; Kim, Hyung Sik; Kim, Young Mi; Choi, Wahn Soo

    2015-06-15

    Mast cells, constituents of virtually all organs and tissues, are critical cells in IgE-mediated allergic responses. The aim of this study was to investigate the effect and mechanism of an indoxyl chromogenic compound, 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, on IgE-mediated mast cell activation and allergic responses in mice. CAC-0982 reversibly suppressed antigen-stimulated degranulation in murine mast cells (IC{sub 50}, ~ 3.8 μM) and human mast cells (IC{sub 50}, ~ 3.0 μM). CAC-0982 also inhibited the expression and secretion of IL-4 and TNF-α in mast cells. Furthermore, CAC-0982 suppressed the mast cell-mediated allergic responses in mice in a dose-dependent manner (ED{sub 50} 27.9 mg/kg). As for the mechanism, CAC-0982 largely suppressed the phosphorylation of Syk and its downstream signaling molecules, including LAT, Akt, Erk1/2, p38, and JNK. Notably, the tyrosine kinase assay of antigen-stimulated mast cells showed that CAC-0982 inhibited Fyn kinase, one of the upstream tyrosine kinases for Syk activation in mast cells. Taken together, these results suggest that CAC-0982 may be used as a new treatment for regulating IgE-mediated allergic diseases through the inhibition of the Fyn/Syk pathway in mast cells. - Highlights: • The anti-allergic effect of 5-bromo-4-chloro-3-indolyl 1,3-diacetate, CAC-0982, was measured. • CAC-0982 reversibly suppressed the activation of mast cells by IgE and antigen. • CAC-0982 inhibited passive cutaneous anaphylaxis in mice. • CAC-0982 suppresses mast cells through inhibition of Fyn activation in mast cells.

  15. A study to evaluate the efficacy of xylene-free hematoxylin and eosin staining procedure as compared to the conventional hematoxylin and eosin staining: An experimental study

    PubMed Central

    Ankle, Madhuri R; Joshi, Priya S

    2011-01-01

    Context: Use of diluted dish washing solution (DWS) has been experimented successfully as a substitute for xylene to deparaffinize tissue sections during hematoxylin and eosin (H and E) staining. Aims: (1) Test the hypothesis that xylene- and methanol-free sections (XMF) deparaffinized with diluted DWS are better than or at par with conventional H and E sections. (2) To compare the efficacy of xylene-free sections with the conventional H and E sections. Settings and Design: Single blinded experimental study. Materials and Methods: Sixty paraffin blocks were considered. One section was stained with conventional H and E method (Group A) and the other with XMF H and E (Group B). Slides were scored for parameters; nuclear staining, cytoplasmic staining (adequate = score1, inadequate = score0), uniformity, clarity, crispness (present = score1, absent = score0). Score >/= 2 was inadequate for diagnosis and 3-5 was adequate for diagnosis. Statistical analysis used: Z test. Results: Adequate nuclear staining, 96.66% sections in group A and 98.33% in Group B (Z = 0.59, P>0.05); adequate cytoplasmic staining, 93.33% in group A and 83.33% in Group B (Z = 1.97, P<0.05); uniform staining, 70% in group A, 50% in group B (Z = 1.94, P<0.05), clarity present in 85% of group A, 88.33% of group B sections (Z = 0.27, P>0.05), crisp staining in 76.66% in group A and 83.33% in Group B (Z = 1.98, P<0.05), 88.33% Group A sections stained adequately for diagnosis as compared with 90% in Group B (Z = 0.17, P>0.05). Conclusion: Xylene- and methanol-free H and E staining is a better alternative to the conventional H and E staining procedure. PMID:22529574

  16. Surface discoloration of composite resins: Effects of staining and bleaching

    PubMed Central

    Poggio, Claudio; Beltrami, Riccardo; Scribante, Andrea; Colombo, Marco; Chiesa, Marco

    2012-01-01

    Background: The purpose of this in vitro study was to evaluate surface discoloration of three microhybrid composite resins (Esthet•X HD, Clearfil AP-X, Gradia Direct) and five nanohybrid composite resins (Ceram•X, GC Kalore, G-aenial, Grandio, GrandioSO), after staining and bleaching procedures. Materials and Methods: The composite resins were polymerized with a curing light (Celalux II, Voco, Cuxhaven, Germany) into 160 silicon molds (6,4 mm in diameter and 2 mm in thickness) to obtain identical specimens. Twenty samples for each composite resin were prepared. The specimens were polished using an automated polishing machine with the sequence of 600-, 800-, 1000-grit abrasive paper under water irrigation. The specimens were immersed in tea and distilled water: the specimens were dipped for 20 min, once a day (every 24 h), for 14 days into the drinks. The specimens were then bleached with carbamide peroxide at 17% (Perfect Bleach-Voco). The color of specimens was measured with a spectrophotometer according to the CIE L*a*b* system after light-polymerization of composite resin specimens, after 7 days, after 14 days, and after bleaching. The color difference h index (DEab*) between each measurement was calculated. Statistical analysis was made using analysis of variance (ANOVA). Results: All specimens showed a significant increase in staining with a similar trend and no significant differences between microhybrid and nanohybrid composite resins. After whitening procedures, materials tested showed both significant and unsignificant differences of the h index. Conclusions: Microhybrid and nanohybrid composite resins had similar in vitro surface discoloration in tea. After bleaching, discoloration was removed from some composite resins tested. PMID:23559921

  17. Multi-stained whole slide image alignment in digital pathology

    NASA Astrophysics Data System (ADS)

    Déniz, Oscar; Toomey, David; Conway, Catherine; Bueno, Gloria

    2015-03-01

    In Digital Pathology, one of the most simple and yet most useful feature is the ability to view serial sections of tissue simultaneously on a computer monitor. This enables the pathologist to evaluate the histology and expression of multiple markers for a patient in a single review. However, the rate limiting step in this process is the time taken for the pathologist to open each individual image, align the sections within the viewer, with a maximum of four slides at a time, and then manually move around the section. In addition, due to tissue processing and pre-analytical steps, sections with different stains have non-linear variations between the two acquisitions, that is, they will stretch and change shape from section to section. To date, no solution has come close to a workable solution to automatically align the serial sections into one composite image. This research work address this problem to obtain an automated serial section alignment tool enabling the pathologists to simply scroll through the various sections in a single viewer. To this aim a multi-resolution intensity-based registration method using mutual information as a similarity metric, an optimizer based on an evolutionary process and a bilinear transformation has been used. To characterize the performance of the algorithm 40 cases x 5 different serial sections stained with hematoxiline-eosine (HE), estrogen receptor (ER), progesterone receptor (PR), Ki67 and human epidermal growth factor receptor 2 (Her2), have been considered. The qualitative results obtained are promising, with average computation time of 26.4s for up to 14660x5799 images running interpreted code.

  18. A simple system for staining protein and nucleic acid electrophoresis gels.

    PubMed

    Raymer, Dorian M; Smith, Douglas E

    2007-03-01

    Researchers in molecular biology spend a significant amount of time tending to the staining and destaining of electrophoresis gels. Here we describe a simple system, costing approximately $100 and taking approximately 1 h to assemble, that automates standard nucleic acid and protein gel staining protocols. Staining is done in a tray or, with DNA gels, in the electrophoresis chamber itself following automatic detection of the voltage drop. Miniature pumps controlled by a microcontroller chip exchange the necessary solutions at programmed time intervals. We demonstrate efficient and highly reproducible ethidium bromide and methylene blue staining of DNA in agarose gels and Coomassie blue and silver staining of proteins in polyacrylamide gels. PMID:17265540

  19. Applications of immunocolloids in light microscopy. IV. Use of photochemical silver staining in a simple and efficient double-staining technique.

    PubMed

    Manigley, C; Roth, J

    1985-12-01

    We report the development of a new light-microscopic double-staining technique using colloidal gold as sole marker. The contrasting color to the red of colloidal gold is achieved by the application of photochemical silver reaction. The silver reaction, which is principally performed at the end of the first staining sequence, converts the red color of a gold-labeled reagent into black. This contrasts clearly with the red coloration that results from the second incubation sequence without silver reaction. For antigen double staining, the same protein A-gold complex can be used to provide the black and the red color, thus rendering the technique very economical. Alternatively, combination of protein A-gold immunolocalization and lectin-gold staining is possible, as is combined lectin-gold staining. PMID:2415576

  20. Identification and isolation of slow-dividing cells in human glioblastoma using carboxy fluorescein succinimidyl ester (CFSE).

    PubMed

    Deleyrolle, Loic P; Rohaus, Mark R; Fortin, Jeff M; Reynolds, Brent A; Azari, Hassan

    2012-01-01

    Tumor heterogeneity represents a fundamental feature supporting tumor robustness and presents a central obstacle to the development of therapeutic strategies(1). To overcome the issue of tumor heterogeneity, it is essential to develop assays and tools enabling phenotypic, (epi)genetic and functional identification and characterization of tumor subpopulations that drive specific disease pathologies and represent clinically relevant targets. It is now well established that tumors exhibit distinct sub-fractions of cells with different frequencies of cell division, and that the functional criteria of being slow cycling is positively associated with tumor formation ability in several cancers including those of the brain, breast, skin and pancreas as well as leukemia(2-8). The fluorescent dye carboxyfluorescein succinimidyl ester (CFSE) has been used for tracking the division frequency of cells in vitro and in vivo in blood-borne tumors and solid tumors such as glioblastoma(2,7,8). The cell-permeant non-fluorescent pro-drug of CFSE is converted by intracellular esterases into a fluorescent compound, which is retained within cells by covalently binding to proteins through reaction of its succinimidyl moiety with intracellular amine groups to form stable amide bonds(9). The fluorescent dye is equally distributed between daughter cells upon divisions, leading to the halving of the fluorescence intensity with every cell division. This enables tracking of cell cycle frequency up to eight to ten rounds of division(10). CFSE retention capacity was used with brain tumor cells to identify and isolate a slow cycling subpopulation (top 5% dye-retaining cells) demonstrated to be enriched in cancer stem cell activity(2). This protocol describes the technique of staining cells with CFSE and the isolation of individual populations within a culture of human glioblastoma (GBM)-derived cells possessing differing division rates using flow cytometry(2). The technique has served to identify and isolate a brain tumor slow-cycling population of cells by virtue of their ability to retain the CFSE labeling. PMID:22565048