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Parathyroid hormone-related protein in normal and neoplastic canine tissues: immunohistochemical localization and biochemical extraction.

PubMed

Gröne, A; Werkmeister, J R; Steinmeyer, C L; Capen, C C; Rosol, T J

1994-05-01

Two polyclonal antibodies, directed against N-terminal amino acids (1-36) or the midregion (amino acids 34-53) of parathyroid hormone-related protein (PTHrP), were used to localize PTHrP in a variety of normal and neoplastic canine tissues. Parathyroid hormone (PTH) immunoreactivity was demonstrated using anti-bovine PTH (amino acids 14-34). The following tissues (among others) stained strongly positive for PTHrP: all layers of epidermal keratinocytes, with the most intense staining of the basal layer; hair follicle keratinocytes; myoepithelial cells of dermal apocrine glands, mammary glands, and apocrine glands of the anal sac; anal sac epithelium; mammary duct epithelium; and thyroid C cells. Adenocarcinomas of the anal sac stained moderately positive (5/22 dogs), weakly positive (11/22 dogs), or did not stain (6/22 dogs). Most parathyroid gland adenomas stained moderately (2/6 dogs) or weakly positive (3/6 dogs) for PTHrP. Squamous cell carcinomas (6/6 dogs) stained strongly positive. Lymphomas stained weakly positive (2/10 dogs) or did not stain (8/10 dogs). There was no consistent relationship between the staining intensity of the tumors and serum calcium concentrations of the dogs. The anti-PTH antibodies stained only parathyroid chief cells strongly positive. Concentrations of PTHrP were measured by radioimmunoassay in protein extracts from an adenocarcinoma derived from the apocrine glands of the anal sac, pancreas, kidney, liver, heart, thyroid, adrenal, and parathyroid glands. PTHrP concentrations varied from undetectable up to 150 pg/mg in normal tissues as compared with 2,000 pg/mg in apocrine adenocarcinoma of the anal sac.(ABSTRACT TRUNCATED AT 250 WORDS)
Rapid demonstration of Legionella pneumophila in unembedded tissue. An adaptation of the Giménez stain.

PubMed

Greer, P W; Chandler, F W; Hicklin, M D

1980-06-01

The Giménez stain, originally developed for demonstrating rickettsiae, readily stained the Legionnaires' disease bacterium (Legionella pneumophila) in frozen tissue sections and smears of fresh or formalin-fixed lung tissue from patients who had confirmed Legionnaires' disease. With the Giménez procedure, the bacterium stained bright red against a blue-green background. The tissue Gram procedures also stained L. pneumophila in frozen sections and smears, but the staining reaction was weak, and these stains were neither as sensitive nor a consistent as the Giménez procedure.
Steroid hormone receptors ERalpha and PR characterised by immunohistochemistry in the mare adrenal gland.

PubMed

Alm, Ylva Hedberg; Sukjumlong, Sayamon; Kindahl, Hans; Dalin, Anne-Marie

2009-07-22

Sex steroid hormone receptors have been identified in the adrenal gland of rat, sheep and rhesus monkey, indicating a direct effect of sex steroids on adrenal gland function. In the present study, immunohistochemistry using two different mouse monoclonal antibodies was employed to determine the presence of oestrogen receptor alpha (ERalpha) and progesterone receptor (PR) in the mare adrenal gland. Adrenal glands from intact (n = 5) and ovariectomised (OVX) (n = 5) mares, as well as uterine tissue (n = 9), were collected after euthanasia. Three of the OVX mares were treated with a single intramuscular injection of oestradiol benzoate (2.5 mg) 18-22 hours prior to euthanasia and tissue collection (OVX+Oe). Uterine tissue was used as a positive control and showed positive staining for both ERalpha and PR. ERalpha staining was detected in the adrenal zona glomerulosa, fasciculata and reticularis of all mare groups. Ovariectomy increased cortical ERalpha staining intensity. In OVX mares and one intact mare, positive ERalpha staining was also detected in adrenal medullary cells. PR staining of weak intensity was present in a low proportion of cells in the zona fasciculata and reticularis of all mare groups. Weak PR staining was also found in a high proportion of adrenal medullary cells. In contrast to staining in the adrenal cortex, which was always located within the cell nuclei, medullary staining for both ERalpha and PR was observed only in the cell cytoplasm. The present results show the presence of ERalpha in the adrenal cortex, indicating oestradiol may have a direct effect on mare adrenal function. However, further studies are needed to confirm the presence of PR as staining in the present study was only weak and/or minor. Also, any possible effect of oestradiol treatment on the levels of steroid receptors cannot be determined by the present study, as treatment time was of a too short duration.
Heparanase expression in periapical granulomas and radicular cysts.

PubMed

Elad, S; Sherman, Y; Palmon, A; Vlodavsky, I; Or, R

2013-01-01

Heparanase is an endo-β-D-glucuronidase enzyme which degrades heparan sulfate glycosaminoglycan side chains of proteoglycans in the extracellular matrix and in basement membranes. The aim of this study was to evaluate the expression of heparanase in periapical granulomas (PGs) and radicular cysts (RCs). Immunohistochemistry was used to assess heparanase expression in PGs and RCs. Parameters including stain intensity, location and cell type were used to characterize heparanase expression in the periapical lesions. Ordered categories (from weak to strong) were used to compare the level of heparanase staining in the PG and RC groups. Both epithelial cells and inflammatory cells were positive for heparanase. The relative staining of the epithelial cells was strong, whereas the relative staining of the inflammatory cells was weak. Significant differences in immunohistochemical staining of epithelial cells were observed between RCs and PGs (p = 0.002). The relative expression of heparanase in epithelial cells in RCs was strong. In PGs, lesions with few or no epithelial cells, heparanase was predominantly expressed weakly by inflammatory cells. PGs and RCs have the same infectious origin. Therefore, the different cellular sources of heparanase in these periapical lesions may imply that this enzyme has specific pathogenetic functions in RCs and PGs.
Fascin and EMMPRIN expression in primary mucinous tumors of ovary: a tissue microarray study.

PubMed

Alici, Omer; Kefeli, Mehmet; Yildiz, Levent; Baris, Sancar; Karagoz, Filiz; Kandemir, Bedri

2014-12-01

The aim of this study was to compare the expressions of fascin and EMMPRIN in primary malignant, borderline and benign mucinous ovarian tumors, and to investigate the relationship of these markers with tumor progression and their applicability to differential diagnosis. An immunohistochemical study was performed for fascin and EMMPRIN using the tissue microarray technique. Eighty-one cases were included in the study; there were 37 benign, 25 borderline and 19 malignant primary mucinous ovarian tumors. For each case, a total staining score was determined, consisting of scores for extent of staining and intensity of staining. The cases were allocated to negative, weakly positive and strongly positive staining categories, according to the total staining score. Both of the markers were significantly negative in benign tumors as compared with borderline and malignant tumors. There was no significant difference between borderline and malignant groups for both markers. Sixty-eight percent of malignant tumors were stained positive by fascin, while this rate was 40% for borderline mucinous tumors. All malignant tumors were strongly stained positive for EMMPRIN, while this rate was 92% for borderline mucinous tumors. The rest of the cases stained weakly positive. No significant difference in staining score was found between fascin and EMMPRIN expression. In ovarian primary mucinous tumors, fascin and EMMPRIN may play an important role in tumor progression from benign tumor to carcinoma. In that context, EMMPRIN and fascin expression may have potential application in the differential diagnosis of some diagnostically problematic mucinous ovarian tumors. However, the differential diagnostic applicability of EMMPRIN appears to be more limited than that of fascin due to its wide spectrum of staining in mucinous ovarian tumors. Copyright © 2014 Elsevier GmbH. All rights reserved.
Circular Mixture Modeling of Color Distribution for Blind Stain Separation in Pathology Images.

PubMed

Li, Xingyu; Plataniotis, Konstantinos N

2017-01-01

In digital pathology, to address color variation and histological component colocalization in pathology images, stain decomposition is usually performed preceding spectral normalization and tissue component segmentation. This paper examines the problem of stain decomposition, which is a naturally nonnegative matrix factorization (NMF) problem in algebra, and introduces a systematical and analytical solution consisting of a circular color analysis module and an NMF-based computation module. Unlike the paradigm of existing stain decomposition algorithms where stain proportions are computed from estimated stain spectra using a matrix inverse operation directly, the introduced solution estimates stain spectra and stain depths via probabilistic reasoning individually. Since the proposed method pays extra attentions to achromatic pixels in color analysis and stain co-occurrence in pixel clustering, it achieves consistent and reliable stain decomposition with minimum decomposition residue. Particularly, aware of the periodic and angular nature of hue, we propose the use of a circular von Mises mixture model to analyze the hue distribution, and provide a complete color-based pixel soft-clustering solution to address color mixing introduced by stain overlap. This innovation combined with saturation-weighted computation makes our study effective for weak stains and broad-spectrum stains. Extensive experimentation on multiple public pathology datasets suggests that our approach outperforms state-of-the-art blind stain separation methods in terms of decomposition effectiveness.
The usefulness of changing focus during examination using Gram staining as initial diagnostic clue for infective tuberculosis.

PubMed

Atsukawa, Yoshiko; Kawakami, Sayoko; Asahara, Miwa; Ishigaki, Shinobu; Tanaka, Takashi; Ono, Yasuo; Nishiya, Hajime; Fujisaki, Ryuichi; Koga, Ichiro; Ota, Yasuo; Miyazawa, Yukihisa

2011-08-01

Gram staining is a useful technique for detecting bacteria but is highly questionable in detecting Mycobacterium tuberculosis. Its detection generally requires special staining, such as Ziehl-Neelsen staining. We experienced three cases in which tuberculosis was first suggested by Gram staining of sputum or pus, confirmed by Ziehl-Neelsen staining, and diagnosed by polymerase chain reaction or culture. To find colorless tubercle bacilli in clinical samples with various organisms, varying the focus to slightly longer and shorter during study of the slides is indispensable. We present criteria for detecting infective pulmonary tuberculosis in Gram staining. First, in the ordinary focus, weakly stained, thin, gram-positive bacilli are found; second, with a slightly longer focus distance, the thin, cord-like, conspicuous gram-positive bacilli can be observed; and third, with a shorter focus distance, the gram-positive bacilli have changed into the brightened, colorless, or ghost ones. Four laboratory technologists each evaluated 20 Gram-stained samples after being lectured on the criteria, with no prior information about the sample. They accurately evaluated the presence of the bacilli in Gram-stained preparations in more than 90% of samples containing 3+ bacilli on Ziehl-Neelsen staining. Gram staining is available as an easy and rapid initial clue to recognize highly infective tuberculosis.
The minimal carcinoma triple stain is superior to commercially available multiplex immunohistochemical stains: breast triple stain and LC/DC breast cocktail.

PubMed

Ginter, Paula S; Varma, Sonal; Liu, Yi-Fang; Shin, Sandra J

2015-12-01

The Minimal Carcinoma (MC) Triple Stain is a tri-chromogen multiplex immunostain (CK7, p63, and E-cadherin) helpful in classifying morphologically ambiguous and/or small carcinomas as either ductal or lobular and/or in situ or invasive. We compared the utility of this stain with two commercially available duplex/multiplex immunostains: Breast Triple Stain (BTS) (Clarient, Aliso Viejo, CA; CK5, p63, and CK8/18) and LC/DC Breast Cocktail (LCDC) (Biocare, Concord, CA; E-cadherin and p120). Ninety-seven mammary carcinomas stained with the MC Triple Stain, BTS, and LCDC were compared. The MC Triple Stain, LCDC, and BTS were diagnostic in 90 (93%) of 97, 82 (85%) of 97, and 85 (88%) of 97 of cases, respectively. All stains showed decreased diagnostic utility due to variability in tissue integrity, quality of the staining, and/or ease of interpretation. In cases where all immunostains were interpretable, the MC Triple Stain yielded the most information. When technically sufficient, all three immunostains demonstrated relative strengths and weaknesses in their ability to provide diagnostic information with the highest consistency and ease of use. Many cases stained with LCDC were technically insufficient due to a suboptimal staining protocol provided by the company. Overall, the MC Triple Stain outperformed BTS and LCDC by more consistently providing more diagnostic information. The MC Triple Stain is a viable alternative to other multiplex immunostains in evaluating small foci of carcinoma, particularly when both the histologic type and extent of disease (in situ vs invasive) require clarification. Copyright© by the American Society for Clinical Pathology.
Young and intense: FoxP2 immunoreactivity in Area X varies with age, song stereotypy, and singing in male zebra finches

PubMed Central

Thompson, Christopher K.; Schwabe, Fabian; Schoof, Alexander; Mendoza, Ezequiel; Gampe, Jutta; Rochefort, Christelle; Scharff, Constance

2013-01-01

FOXP2 is a transcription factor functionally relevant for learned vocalizations in humans and songbirds. In songbirds, FoxP2 mRNA expression in the medium spiny neurons of the basal ganglia song nucleus Area X is developmentally regulated and varies with singing conditions in different social contexts. How individual neurons in Area X change FoxP2 expression across development and in social contexts is not known, however. Here we address this critical gap in our understanding of FoxP2 as a link between neuronal networks and behavior. We used a statistically unbiased analysis of FoxP2-immunoreactivity (FoxP2-IR) on a neuron-by-neuron basis and found a bimodal distribution of FoxP2-IR neurons in Area X: weakly-stained and intensely-stained. The density of intensely-stained FoxP2-IR neurons was 10 times higher in juveniles than in adults, exponentially decreased with age, and was negatively correlated with adult song stability. Three-week old neurons labeled with BrdU were more than five times as likely to be intensely-stained than weakly-stained. The density of FoxP2-IR putative migratory neurons with fusiform-shaped nuclei substantially decreased as birds aged. The density of intensely-stained FoxP2-IR neurons was not affected by singing whereas the density of weakly-stained FoxP2-IR neurons was. Together, these data indicate that young Area X medium spiny neurons express FoxP2 at high levels and decrease expression as they become integrated into existing neural circuits. Once integrated, levels of FoxP2 expression correlate with singing behavior. Together, these findings raise the possibility that FoxP2 levels may orchestrate song learning and song stereotypy in adults by a common mechanism. PMID:23450800
Young and intense: FoxP2 immunoreactivity in Area X varies with age, song stereotypy, and singing in male zebra finches.

PubMed

Thompson, Christopher K; Schwabe, Fabian; Schoof, Alexander; Mendoza, Ezequiel; Gampe, Jutta; Rochefort, Christelle; Scharff, Constance

2013-01-01

FOXP2 is a transcription factor functionally relevant for learned vocalizations in humans and songbirds. In songbirds, FoxP2 mRNA expression in the medium spiny neurons of the basal ganglia song nucleus Area X is developmentally regulated and varies with singing conditions in different social contexts. How individual neurons in Area X change FoxP2 expression across development and in social contexts is not known, however. Here we address this critical gap in our understanding of FoxP2 as a link between neuronal networks and behavior. We used a statistically unbiased analysis of FoxP2-immunoreactivity (FoxP2-IR) on a neuron-by-neuron basis and found a bimodal distribution of FoxP2-IR neurons in Area X: weakly-stained and intensely-stained. The density of intensely-stained FoxP2-IR neurons was 10 times higher in juveniles than in adults, exponentially decreased with age, and was negatively correlated with adult song stability. Three-week old neurons labeled with BrdU were more than five times as likely to be intensely-stained than weakly-stained. The density of FoxP2-IR putative migratory neurons with fusiform-shaped nuclei substantially decreased as birds aged. The density of intensely-stained FoxP2-IR neurons was not affected by singing whereas the density of weakly-stained FoxP2-IR neurons was. Together, these data indicate that young Area X medium spiny neurons express FoxP2 at high levels and decrease expression as they become integrated into existing neural circuits. Once integrated, levels of FoxP2 expression correlate with singing behavior. Together, these findings raise the possibility that FoxP2 levels may orchestrate song learning and song stereotypy in adults by a common mechanism.
Use of Oil Red O stain in the cytologic diagnosis of canine liposarcoma.

PubMed

Masserdotti, Carlo; Bonfanti, Ugo; De Lorenzi, Davide; Ottolini, Nicola

2006-03-01

Oil Red O, a stain commonly used to demonstrate lipid in frozen tissue, also may be used to stain air-dried cytologic specimens. The purpose of this study was to prospectively evaluate the value of Oil Red O in identifying lipid to aid in the differentiation of liposarcomas from other types of sarcoma. Twelve tumor specimens from dogs were evaluated. The tumors were included in the study if initial cytologic evaluation indicated a sarcoma, and if histologic confirmation was available. Oil Red O was applied to all cytologic specimens. Tumor specimens were diagnosed histologically as liposarcoma (3 well-differentiated, 1 pleomorphic), hemangiopericytoma (n = 3), fibrosarcoma (n = 3), malignant fibrous histiocytoma (n = 1), and undifferentiated sarcoma (n = 1). Cytologic specimens from all liposarcomas showed strong positive staining of cytoplasmic vacuoles for lipid. Specimens from other sarcomas stained negative for Oil Red O, with the exception of weak, irregular positive staining in 1 hemangiopericytoma. Our results suggest that Oil Red O staining may be an easy, inexpensive, and useful diagnostic tool for the differentiation of liposarcoma from other mesenchymal neoplasms.
Glutathione S-transferase (placental) as a marker of transformation in the human cervix uteri: an immunohistochemical study.

PubMed Central

Randall, B. J.; Angus, B.; Akiba, R.; Hall, A.; Cattan, A. R.; Proctor, S. J.; Jones, R. A.; Horne, C. H.

1990-01-01

Using an indirect immunohistochemical technique on paraffin sections, employing a polyclonal antibody to the acidic (placental) form of glutathione-S-transferase (GST), we have evaluated cytoplasmic and nuclear staining in a series of 67 cervical biopsies including normal non neoplastic tissue, immature squamous metaplasia, all grades of cervical intraepithelial neoplasia (CIN) and invasive carcinomas of keratinising and non-keratinising types. No differences in cytoplasmic staining between the varied lesions studied were seen. However, there were marked differences in nuclear staining. While normal non-neoplastic stratified squamous epithelium showed weak staining of the lower one-third of the epithelium only, in immature squamous metaplasia and in all grades of CIN there was intense nuclear staining in all layers of the epithelium. Invasive carcinomas showed generally less intense nuclear staining than CIN lesions. Endocervical cell nuclei also showed intense nuclear staining. These findings indicate that GST is of limited use as a marker of transformation in the human cervix uteri. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:2223578
Loss of histochemical identity in mast cells lacking carboxypeptidase A.

PubMed

Feyerabend, Thorsten B; Hausser, Heinz; Tietz, Annette; Blum, Carmen; Hellman, Lars; Straus, Anita H; Takahashi, Hélio K; Morgan, Ellen S; Dvorak, Ann M; Fehling, Hans Jörg; Rodewald, Hans-Reimer

2005-07-01

Mast cell carboxypeptidase A (Mc-cpa) is a highly conserved secretory granule protease. The onset of expression in mast cell progenitors and lineage specificity suggest an important role for Mc-cpa in mast cells. To address the function of Mc-cpa, we generated Mc-cpa-null mice. Mc-cpa-/- mast cells lacked carboxypeptidase activity, revealing that Mc-cpa is a nonredundant enzyme. While Mc-cpa-/- peritoneal mast cells were ultrastructurally normal and synthesized normal amounts of heparin, they displayed striking histochemical and biochemical hallmarks of immature mast cells. Wild-type peritoneal mast cells had a mature phenotype characterized by differential histochemical staining with proteoglycan-reactive dyes (cells do not stain with alcian blue but stain with safranin and with berberine) and a high side scatter to forward scatter ratio by flow cytometry and were detergent resistant. In contrast, Mc-cpa-/- peritoneal mast cells, like immature bone marrow-derived cultured mast cells, stained with alcian blue normally or weakly and either did not stain with safranin and berberine or stained weakly, had a low side scatter to forward scatter ratio, and were detergent sensitive. This phenotype was partially ameliorated with age. Thus, histochemistry and flow cytometry, commonly used to measure mast cell maturation, deviated from morphology in Mc-cpa-/- mice. The Mc-cpa-/- mast cell phenotype was not associated with defects in degranulation in vitro or passive cutaneous anaphylaxis in vivo. Collectively, Mc-cpa plays a crucial role for the generation of phenotypically mature mast cells.
Loss of Histochemical Identity in Mast Cells Lacking Carboxypeptidase A

PubMed Central

Feyerabend, Thorsten B.; Hausser, Heinz; Tietz, Annette; Blum, Carmen; Hellman, Lars; Straus, Anita H.; Takahashi, Hélio K.; Morgan, Ellen S.; Dvorak, Ann M.; Fehling, Hans Jörg; Rodewald, Hans-Reimer

2005-01-01

Mast cell carboxypeptidase A (Mc-cpa) is a highly conserved secretory granule protease. The onset of expression in mast cell progenitors and lineage specificity suggest an important role for Mc-cpa in mast cells. To address the function of Mc-cpa, we generated Mc-cpa-null mice. Mc-cpa−/− mast cells lacked carboxypeptidase activity, revealing that Mc-cpa is a nonredundant enzyme. While Mc-cpa−/− peritoneal mast cells were ultrastructurally normal and synthesized normal amounts of heparin, they displayed striking histochemical and biochemical hallmarks of immature mast cells. Wild-type peritoneal mast cells had a mature phenotype characterized by differential histochemical staining with proteoglycan-reactive dyes (cells do not stain with alcian blue but stain with safranin and with berberine) and a high side scatter to forward scatter ratio by flow cytometry and were detergent resistant. In contrast, Mc-cpa−/− peritoneal mast cells, like immature bone marrow-derived cultured mast cells, stained with alcian blue normally or weakly and either did not stain with safranin and berberine or stained weakly, had a low side scatter to forward scatter ratio, and were detergent sensitive. This phenotype was partially ameliorated with age. Thus, histochemistry and flow cytometry, commonly used to measure mast cell maturation, deviated from morphology in Mc-cpa−/− mice. The Mc-cpa−/− mast cell phenotype was not associated with defects in degranulation in vitro or passive cutaneous anaphylaxis in vivo. Collectively, Mc-cpa plays a crucial role for the generation of phenotypically mature mast cells. PMID:15988029
Lack of prognostic significance of C-erbB-2 expression in low- and high- grade astrocytomas.

PubMed

Muallaoglu, Saik; Besen, Ali Ayberk; Ata, Alper; Mertsoylu, Huseyin; Arican, Ali; Kayaselcuk, Fazilet; Ozyilkan, Ozgur

2014-01-01

Astrocytic tumors, the most common primary glial tumors of the central nervous system, are classified from low to high grade according to the degree of anaplasia and presence of necrosis. Despite advances in therapeutic management of high grade astrocytic tumors, prognosis remains poor. In the present study, the frequency and prognostic significance of c-erb-B2 in astrocytic tumors was investigated. Records of 72 patients with low- and high-grade astrocytic tumors were evaluated. The expression of C-erbB-2 was determined immunohistochemically and intensity was recorded as 0 to 3+. Tumors with weak staining (1+) or no staining (0) were considered Her-2 negative, while tumors with moderate (2+) and strong (3+) staining were considered Her-2 positive. Of the 72 patients, 41 (56.9%) had glioblastoma (GBM), 10 (13.9%) had diffuse astrocytoma, 15 (20.8%) had anaplastic astrocytoma, 6 (8.3%) had pilocytic astrocytoma. C-erbB-2 overexpression was detected in the tumor specimens of 17 patients (23.6%). Six (8.3%) tumors, all GBMs, exhibited strong staining, 2 (2.7%) specimens, both GBMs, exhibited moderate staining, and 9 specimens, 5 of them GBMs (12.5%), exhibited weak staining. No staining was observed in diffuse astrocytoma and pilocytic astrocytoma specimens. Median overall survival of patients with C-erbB-2 negative and C-erbB-2 positive tumors were 30 months (95%CI: 22.5-37.4 months) and 16.9 months (95%CI: 4.3-29.5 months), respectively (p=0.244). Although there was no difference in survival, C-erbB-2 overexpression was observed only in the GBM subtype.
Improved method for combination of immunocytochemistry and Nissl staining.

PubMed

Kádár, Andrea; Wittmann, Gábor; Liposits, Zsolt; Fekete, Csaba

2009-10-30

Nissl staining is a widely used method to study morphology and pathology of neural tissue. After standard immunocytochemistry, the Nissl staining labels only the nucleus of neurons and the characteristic staining of the neuronal perikarya is absent or very weak. We hypothesized that the RNA degradation during the immunocytochemical treatment results in the loss of cytoplasmic staining with Nissl-dyes. To test this hypothesis, we used RNAse-free conditions for all steps of immunostaining. To further prevent the RNA-degradation by RNAse contaminations, the RNAse inhibitor heparin was added to all antibody-containing solutions. The efficiency of Nissl staining after standard and RNAse-free double-labeling immunocytochemistry was compared using antibodies against c-Fos and neuropeptide Y (NPY) on tissues of rats refed after 3 days of fasting. After standard immunocytochemistry, the Nissl-staining labeled the nuclei of neurons and only very faintly the cytoplasm of these cells. The RNAse-free treatment did not alter the distribution of immunoreaction signal, but preserved the staining of neuronal perikarya by the Nissl-dyes. In conclusion, the RNAse-free conditions during immunocytochemistry allow the labeling of neuronal perikarya by Nissl-dyes. The described method facilitates the mapping of immunocytochemical signals and makes possible the light microscopic examination of the innervation of neurons identified by their nuclear protein content.
Improved method for combination of immunocytochemistry and Nissl staining

PubMed Central

Kádár, Andrea; Wittmann, Gábor; Liposits, Zsolt; Fekete, Csaba

2009-01-01

Nissl-staining is a widely used method to study morphology and pathology of neural tissue. After standard immunocytochemistry, the Nissl-staining labels only the nucleus of neurons and the characteristic staining of the neuronal perikarya is absent or very weak. We hypothesized that the RNA degradation during the immunocytochemical treatment results in the loss of cytoplasmic staining with Nissl-dyes. To test this hypothesis, we used RNAse-free conditions for all steps of immunostaining. To further prevent the RNA-degradation by RNAse contaminations, the RNAse inhibitor heparin was added to all antibody-containing solutions. The efficiency of Nissl-staining after standard and RNAse-free double-labeling immunocytochemistry was compared using antibodies against c-Fos and neuropeptide Y (NPY) on tissues of rats refed after three days of fasting. After standard immunocytochemistry, the Nissl-staining labeled the nuclei of neurons and only very faintly the cytoplasm of these cells. The RNAse-free treatment did not alter the distribution of immunoreaction signal, but preserved the staining of neuronal perikarya by the Nissl-dyes. In conclusion, the RNAse-free conditions during immunocytochemistry, allows the labeling of neuronal perikarya by Nissl-dyes. The described method facilitates the mapping of immunocytochemical signals and makes possible the light microscopic examination of the innervation of neurons identified by their nuclear protein content. PMID:19615409
Localization of cytochrome P4501A in liver and extrahepatic organs of the pilot whale, Globiocephala melaena

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

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

1994-12-31

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

PubMed

Noda, Hiroshi; Yokota, Makoto; Tatsumi, Shinji; Sugiyama, Shizuyuki

2002-03-01

Using the highly sensitive immunohistochemical staining method EnVision+, which employs a dextran polymer reagent for the secondary antibody, the detection of the ABH antigens was attempted in the oral squamous epithelium. This new technique uses monoclonal antibody as a primary antibody and it takes about three hours for staining. The time is much shorter than conventional absorption-elution testing or absorption-inhibition testing for the determination of ABO blood grouping. Secretor saliva samples were stained at strong intensity by the antibody, which corresponded to its blood group and anti-H. On the one hand, nonsecretor saliva samples were stained at strong intensity only by the antibody that corresponded to its blood group, and at weak intensity only by anti-H. Since human oral squamous epithelium antigens were stained specifically by this method, we can examine the ABO blood group of saliva samples and perform cytodiagnosis at the same time. Our research suggested that the EnVision+ Method is a useful technique for ABO blood grouping of saliva in forensic cases.
Address substrates as promising targets for laser histochemical surgery as a nontraditional line in medicine

NASA Astrophysics Data System (ADS)

Piruzyan, L. A.; Mikhailovskiy, Ye. M.; Piruzyan, A. L.

1999-12-01

The priority concept of the laser histochemical surgery as a potentially novel line in medicine is presented. The histochemical stains, selectively coloring some targets (address substrates), that are cells or their biochemical ingredients, sensitize them to the laser irradiation. Such sensitization to laser irradiation by staining turns the colored targets into targets for the laser beam. The action of the irradiation onto its specific targets beats out of the cell its ingredients which participate in a pathology process. In particular, the beating of a stained ferment out of the general stage of biochemical processes characteristic for the pathology interrupts their currence. The laser beam, when beating out its stained targets without any damage of the unstained tissues, acts like a scalpel that cuts off affected tissues not brushing healthy ones. A scheme for testing stains as sensitizers of the `address substrates' to the laser irradiation is presented. As the criterion of the stain sensitization the fact was chosen of absence or weakness of pathomorphologic and biochemical signs of the disease in an experimental model of the pathology irradiated with laser after a stain use, while the pathology signs are present in a control sample. The basis is done for study of the histochemical stains as potential means for the laser histochemical surgery of disseminated sclerosis, mucopolysaccharidosis, hypercholesterolemia, myocardial infarction, cardiosclerosis, caries and parodontosis.

Immunohistochemical analysis of the gingiva with periodontitis of type I plasminogen deficiency compared to gingiva with gingivitis and periodontitis and healthy gingiva.

PubMed

Kurtulus Waschulewski, Idil; Gökbuget, Aslan Y; Christiansen, Nina M; Ziegler, Maike; Schuster, Volker; Wahl, Gerhard; Götz, Werner

2016-12-01

Type I plasminogen deficiency (Plgdef) is an uncommon chronic inflammation of mucous membranes. Gingival enlargements usually proceed with progressive periodontal destruction and tooth-loss. Plasmin(ogen)-independent enzymatic mechanisms for fibrin clearance have already been discussed in the literature. Our primary objective was to verify, immunohistochemically, the occurrence of different enzymatic factors involved in tissue breakdown of inflamed compared to healthy gingiva. Secondly, we tried to find out, if these patients have a similar microbiological profile to the patients with known gingivitis and periodontitis. Immunohistochemical analysis of enzymes elastase, plasminogen (plg), cathepsin G, matrix-metalloproteinase (MMP)-3 and MMP-7 and of glycoprotein fibrinogen were performed with gingival tissues from 3 healthy controls, 8 patients with Plgdef and 3 patients with gingivitis and periodontitis. Furthermore, plaque from 5 patients with plasminogen deficiency were also obtained to determine the microbiological profile. Significantly high numbers of elastase positive leukocytes were detected in all samples. Staining for MMP-3 and MMP-7 was seen in samples with gingivitis and periodontitis with a stronger staining in samples with periodontitis by Plgdef. Fibrinogen was detectable in all samples. Staining for plg was stronger in samples with periodontitis than in other samples. Staining for cathepsin G was weak in gingivitis and periodontitis. Subgingival microbial flora showed elevated colony forming units of Prevotella intermedia/nigrescens, Fusobacterium spp., Eikenella corrodens, Porphyromonas gingivalis and viridans streptococci. Strong staining of elastase, MMP-3 and MMP-7 and weak staining of plg in Plgdef samples supports the plasmin(ogen) - independent fibrin clearance. Similar subgingival microbiological flora was observed in periodontitis with Plgdef as in other periodontal diseases. Further investigations should determine the exact pathomechanism and focus on effective treatment methods of this entity. Copyright © 2016. Published by Elsevier Ltd.
Ulex Europaeus lectin and anti-CD31 staining in squamous cell carcinoma of the uterine cervix: potential prognostic markers.

PubMed

Davidson, B; Goldberg, I; Gotlieb, W H; Lerner-Geva, L; Ben-Baruch, G; Kopolovic, J

1998-07-01

Seventy-five squamous cell carcinomas of the uterine cervix and 10 controls were stained for Ulex Europaeus lectin 1 (UEA-1) and anti-CD31, and the results were analyzed with respect to patient age, clinical stage, tumor grade, and survival during a follow-up period of 1 to 13 years. The patients' mean age at the time of diagnosis was 47.8 years (range, 27 to 83). Seventeen patients died of disease, 2 had disease recurrence, and 51 patients remained free of disease; 5 patients were lost to follow-up. Twenty-eight cases (37.3%) showed focal membranous staining for UEA-1 and 9 cases (12%) showed a diffuse pattern; 38 cases (50.7%) were UEA-1 negative. Poor survival was related to diffuse membranous UEA-1 immunoreactivity (p = 0.02), age (p = 0.014), grade (p = 0.02), and stage (p = 0.0002). CD31-positive neoplastic cells displayed a cytoplasmic pattern. Fifteen cases (20%) had diffuse staining and another 15 (20%) stained focally; 45 cases (60%) were CD31-negative. The adjacent nonneoplastic epithelium and all 10 controls were uniformly negative for CD31. Variable staining of the endocervical epithelium and weak or negative staining of ectocervical epithelium for UEA-1 were observed. However, the epithelium in all controls was negative for UEA-1. Poor survival was related to both focal and diffuse staining for CD31 (p = 0.01 and p = 0.03, respectively). Staining by both UEA-1 and anti-CD31 retained its correlation with survival after exclusion of stage la tumors.
[Significance of CEA in gastric and colorectal cancer].

PubMed

Uehara, K; Miyamoto, Y; Izuo, M; Shiozaki, H; Aiba, S; Matsumoto, H

1985-04-01

The determination of serum CEA (Sandwich method) and CEA staining (PAP method) of excised specimens were performed in patients with gastric or colorectal cancer, and the biological characteristics of each cancer and the factors to increase serum CEA were studied with the following results: As colonic cancer has strong CEA productivity, serum CEA can be useful for the detection of cancer, and especially effective for the postoperative observation. Gastric cancer has weak CEA productivity, and serum CEA is not so useful in the detection of cancer and the judgement of resectability. The CEA positive rate of tissue with CEA staining was 80% in gastric cancer, 100% in colonic cancer, and were nearly equal to the CEA positive rate of serum in the group of terminal stage. In the mode of CEA staining of cancerous cells, IV type was observed most frequently in gastric cancer, and I type in colonic cancer. Among the resected cases showing more than 7ng/ml serum CEA, differentiated type, lymph node metastasis (+), the degree of tissue staining with CEA staining, the mode of cell staining O or I type in gastric cancer and I type in colonic cancer were observed in common.
GATA3: a promising marker for metastatic breast carcinoma in serous effusion specimens.

PubMed

Shield, Paul W; Papadimos, David J; Walsh, Michael D

2014-04-01

The usefulness of GATA3 (GATA-binding protein 3 to DNA sequence [A/T]GATA[A/G]) as a marker for metastatic breast carcinoma in serous effusion specimens was investigated. Cell block sections from 74 serous effusion specimens (32 ascitic, 2 pericardial, and 40 pleural fluids) were stained with an anti-GATA3 murine monoclonal antibody. The specimens included 62 confirmed metastatic carcinomas from the breast (30 specimens), female genital tract (13 specimens), gastrointestinal tract (7 specimens), lung adenocarcinoma (9 specimens), pancreas (1 specimen), kidney (1 specimen), and bladder (1 specimen). The breast carcinoma cases included 15 ductal carcinomas and 8 lobular carcinomas; the histology subtype was not available for 7 specimens. Twelve cases containing florid reactive mesothelial cells were also stained. The breast carcinoma cases were also stained for mammaglobin and gross cystic disease fluid protein of 15 kilodaltons (GCDFP-15) to compare their sensitivity with GATA3. Positive nuclear staining for GATA3 was found to be present in 90% of metastatic breast carcinoma specimens (27 of 30 specimens). All nonbreast metastatic carcinomas tested were negative with the exception of the single case of metastatic urothelial carcinoma. No staining was observed in any of the benign reactive cases or in benign mesothelial cells present in the malignant cell block preparations. Two cases demonstrated weak positivity of benign lymphoid cells. Staining results were unambiguous because all positive cases demonstrated intense nuclear staining in > 50% of tumor cells. Mammaglobin (57% staining; 17 of 30 cases) and GCDFP-15 (33% staining; 10 of 30 cases) were found to be less sensitive markers of breast carcinoma. If used in a panel, mammaglobin and GCFP-15 staining would have identified only 1 additional case compared with those stained with GATA3. GATA3 may be a useful addition to immunostaining panels for serous effusion specimens when metastatic breast carcinoma is a consideration. © 2014 American Cancer Society.
Histochemical analysis of glycoconjugates in the skin of a catfish (arius tenuispinis, day).

PubMed

Al-Banaw, A; Kenngott, R; Al-Hassan, J M; Mehana, N; Sinowatz, F

2010-02-01

A histochemical study using conventional carbohydrate histochemistry (periodic-acid staining including diastase controls, alcian blue staining at pH 1 and 2.5) as well as using a battery of 14 fluorescein isothiocyanate (FITC)-labelled lectins to identify glycoconjugates present in 10 different areas of the skin of a catfish (Arius tenuispinis) was carried out. The lectins used were: mannose-binding lectins (Con A, LCA and PSA), galactose-binding lectins (PNA, RCA), N-acetylgalactosamine-binding lectins (DBA, SBA, SJA and GSL I), N-acetylglucosamine-binding lectins (WGA and WGAs), fucose-binding lectins (UEA) and lectins which bind to complex carbohydrate configurations (PHA E, PHA L). Conventional glycoconjugate staining (PAS staining, alcian blue at pH 1 and 2.5) showed that the mucous goblet cells contain a considerable amount of glycoconjugates in all locations of the skin, whereas the other unicellular gland type, the club cells, lacked these glycoconjugates. The glycoproteins found in goblet cells are neutral and therefore stain magenta when subjected to PAS staining. Alcian blue staining indicating acid glycoproteins was distinctly positive at pH 1, but gave only a comparable staining at pH 2.5. The mucus of the goblet cells therefore also contains acid glycoproteins rich in sulphate groups. Using FITC-labelled lectins, the carbohydrate composition of the glycoproteins of goblet cells could be more fully characterized. A distinct staining of the mucus of goblet cells was found with the mannose-binding lectins LCA and PSA; the galactosamine-binding lectins DBA, SBA and GLS I; the glucosamine-binding lectin WGA; and PHA E which stains glycoproteins with complex carbohydrate configurations. No reaction occurred with the fucose-binding lectin UEA and the sialic acid-specific lectin SNA. In addition, the galactose-binding lectins PNA and RCA showed only a weak or completely negative staining of the mucus in the goblet cells. The specificity of the lectin staining could be proved by inhibiting binding of the lectins by competitive inhibition with the corresponding sugars. From these data, we can conclude that the mucus produced by the epidermal goblet cells of A. tenuispinis is rich in mannose, N-acetylgalactosamine and N-acetylglucosamine residues.
Immunohistochemical analysis of an ectopic endometriosis in the uterine round ligament.

PubMed

Terada, Shinichi; Miyata, Yachiyo; Nakazawa, Hiroaki; Higashimori, Takamitsu; Arai, Takanari; Kikuchi, Yuji; Nozaki, Motohiro

2006-09-09

A rare case of the inguinal endometriosis was reported with immunohistochemical analysis. A 28-year-old woman had a thumb-sized tumor in the right groin for two years with a gradual increase in size and pain. An operation revealed an elastic hard tumor with an unclear margin and adhesion to the uterine round ligament. The histology showed irregular proliferation of the endometrial glands and stroma. The glandular epithelium stained weakly positive against CD125 antibody and the stromal matrix stained strongly positive against CD10 antibody. The nucleus in both the epithelial and stromal cells stained strongly positive against progesterone and estrogen receptor antibodies, and the cytoplasm in both types of cells stained moderately positive against COX-2 (cyclooxygenase-2) antibody. In conclusion, the combination of estrogen or progesterone receptor antibody for the nucleus and CD10 or COX-2 antibody for the cytoplasm could enhance the accuracy of diagnosis for ectopic endometriosis.
Lethal cardiac amyloidosis: Modification of the Congo Red technique on a forensic case.

PubMed

Rancati, A; Andreola, S; Bailo, P; Boracchi, M; Fociani, P; Gentile, G; Zoja, R

2018-05-26

Congo Red staining is usually used in diagnosing amyloidosis, a pathology characterized by the storage of abnormal proteins in several human organs. When assessed on samples fixated in formalin and embended in paraffin, this staining can undergo several artefacts, causing diagnostic and interpretative difficulties due to its weak stainability and a consequent reduced visibility of the amyloid. These complications, in time, requested several variations of this staining technique, especially in clinical practice, while in the forensic field no protocols has ever been adapted to cadaveric samples, a material that is already characteristically burdened by a peculiar stainability. In our work, studying a sudden death caused by cardiac amyloidosis and diagnosed only with post-mortem exams, we present a modified Congo Red staining used with the purpose to demonstrate amyloid in cadaveric material after the unsuccessfully use of all standard protocols. Copyright © 2018. Published by Elsevier B.V.
Synovial chondromatosis of the temporomandibular joint: Immunohistochemical examinations regarding the role of insulin-like growth factors and their binding proteins in the etiology of this disease.

PubMed

Wilms, Christian T; Heim, Nils; Teschke, Marcus; Reich, Rudolf R; Götz, Werner

2017-02-01

Synovial chondromatosis (SC) is a benign disease of the joints without a known cause. It sometimes affects the temporomandibular joint (TMJ) and is accompanied by pain, swelling, malocclusion, and crepitation. It has been divided into three stages by Milgram and is supposed to originate from the synovia and cartilage of a joint (Milgram, 1977b). The aim of this study was to examine an involvement of the insulin-like growth factors (IGF-I/-II) and their binding proteins (IGFBP-1 to -6) in the etiology of this disease. Therefore 23 specimen of SC from 16 patients were immunohistochemically stained and microscopically examined. Staining was assessed semiquantitatively: negative (-), weakly positive ((+)), moderately positive (+), strongly positive (++) and very strongly positive (+++). It could be seen that especially the chondro- and fibrocytes and the synovia showed positive staining for almost all IGFs and IGFBPs. The underlying tissue, consisting of connective tissue or chondroid matrix, was stained as well but more weakly so. We conclude that the IGF/IGFBP system seems to contribute to the pathogenesis of SC, especially IGF-I and -II, and their effects enhancing binding protein 5. Copyright © 2016 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.
Methylene blue-aided cholangioscopy in patients with biliary strictures: feasibility and outcome analysis.

PubMed

Hoffman, A; Kiesslich, R; Bittinger, F; Galle, P R; Neurath, M F

2008-07-01

Chromoendoscopy using methylene blue is employed in the gastrointestinal tract to delineate neoplastic lesions. We tested the value of chromoendoscopy during choledochoscopy for characterization of local inflammation, neoplasias, and other alterations in patients with biliary strictures. Patients with suspected biliary lesions were scheduled for endoscopic retrograde cholangiography with subsequent cholangioscopy. After initial inspection of the bile duct, 15 ml methylene blue (0.1 %) was administered via the working channel of the cholangioscope. Newly appearing circumscribed or unstained lesions were judged according to their macroscopic type and staining features. Methylene-blue-aided diagnosis was compared with either clinical follow-up of the patients or, in some cases, with the results of targeted biopsies. A total of 55 patients [biliary stenosis/cholestasis of unknown origin (n = 24), stenosis after orthotopic liver transplantation (n = 11), primary sclerosing cholangitis (n = 20)] were included. Methylene blue unmasked subtle mucosal changes and permitted macroscopic characterization of circumscribed lesions. Characteristic surface staining patterns were seen in chronic inflammation, dysplasia, and ischemic-type biliary lesions. Nondysplastic mucosa appeared homogeneously stained, whereas scarred strictures showed a weak uptake of methylene blue. In this prospective feasibility study, methylene-blue-aided cholangioscopy was used for the first time to define different staining patterns of the bile duct. The differences in staining patterns identified normal, dysplastic, and inflamed mucosa of the bile duct, as was proved by follow-up or, in some cases, histology. Whereas homogeneous staining predicted the presence of normal mucosa, absence of staining of circumscribed lesions, or diffused staining of such lesions, represented neoplastic changes or inflammation.
CDX2 immunostaining in primary and metastatic germ cell tumours of the testis.

PubMed

Oz Atalay, Fatma; Aytac Vuruskan, Berna; Vuruskan, Hakan

2016-12-01

Objective To evaluate the immunohistochemical staining pattern of caudal type homeobox 2 (CDX2) protein in germ cell tumours (GCTs) of the testis. Methods This study reassessed archival tissue samples collected from patients diagnosed with primary and metastatic testicular GCTs for CDX2 immunoreactivity using standard immunohistochemical techniques. Positive nuclear immunostaining was evaluated with regard to both the staining intensity and the extent of the staining. Results Tissue sections from primary and metastatic testicular GCTs ( n = 104), germ cell neoplasia in situ (GCNis) ( n = 5) and benign testicles ( n = 15) were analysed. The GCNis and benign testicular tissues showed no immunoreactivity for CDX2. Strong and diffuse staining of CDX2 was demonstrated only in the mature colonic epithelium of teratomas in both primary and metastatic GCTs. CDX2 positivity in other tumours (one pure yolk sac tumour, one yolk sac component of a mixed GCT and one pure seminoma) was infrequent, and was only weak and focal. Conclusions CDX2 immunostaining should be interpreted based on both the staining intensity and the extent of staining so as not to cause misdiagnosis. Teratomas with colonic-type epithelium should be considered in the differential diagnosis if a metastatic tumour with an unknown primary shows prominent CDX2 immunostaining.
GATA3 Expression in Normal Skin and in Benign and Malignant Epidermal and Cutaneous Adnexal Neoplasms

PubMed Central

de Peralta-Venturina, Mariza N.; Balzer, Bonnie L.; Frishberg, David P.

2015-01-01

Abstract: Initial investigations reported GATA3 to be a sensitive and relatively specific marker for mammary and urothelial carcinomas. Recently, GATA3 expression has been described in several other epithelial tumors. However, there has been only limited investigation of GATA3 expression in cutaneous epithelial tumors. The objective of this study was to examine the immunohistochemical expression of GATA3 in a wide variety of cutaneous epithelial neoplasms. GATA3 expression was evaluated in 99 benign and 63 malignant cutaneous epithelial tumors. GATA3 was consistently and usually strongly expressed in clear cell acanthoma, trichofolliculoma, trichoepithelioma, trichilemmoma, sebaceous adenoma, sebaceoma, apocrine hidrocystoma, apocrine tubular papillary adenoma, hidradenoma papilliferum, and syringocystadenoma papilliferum. Hidradenomas exhibited variable positive staining. Most poromas, syringomas, chondroid syringomas, cylindromas, and spiradenomas were negative or only focally and weakly positive. Focal staining was present in all pilomatrixomas. Thirteen of 14 basal cell carcinomas, 21 of 24 squamous carcinomas, and all 6 sebaceous carcinomas exhibited positive staining. The 1 apocrine carcinoma, both mucinous carcinomas, and 2 of 3 microcystic adnexal carcinomas also exhibited positive staining, whereas the 1 eccrine porocarcinoma and the 1 adenoid cystic carcinoma were negative. One of 11 Merkel cell carcinomas exhibited focal weak staining. Our findings demonstrate that GATA3 is expressed in a wide variety of benign and malignant cutaneous epithelial neoplasms. In addition to carcinomas of breast and urothelial origin and other more recently described GATA3-positive tumors, the differential diagnosis of a metastatic tumor of unknown primary origin that expresses GATA3 should also include a carcinoma of cutaneous epithelial origin. PMID:26595821
GATA3 Expression in Normal Skin and in Benign and Malignant Epidermal and Cutaneous Adnexal Neoplasms.

PubMed

Mertens, Richard B; de Peralta-Venturina, Mariza N; Balzer, Bonnie L; Frishberg, David P

2015-12-01

Initial investigations reported GATA3 to be a sensitive and relatively specific marker for mammary and urothelial carcinomas. Recently, GATA3 expression has been described in several other epithelial tumors. However, there has been only limited investigation of GATA3 expression in cutaneous epithelial tumors. The objective of this study was to examine the immunohistochemical expression of GATA3 in a wide variety of cutaneous epithelial neoplasms. GATA3 expression was evaluated in 99 benign and 63 malignant cutaneous epithelial tumors. GATA3 was consistently and usually strongly expressed in clear cell acanthoma, trichofolliculoma, trichoepithelioma, trichilemmoma, sebaceous adenoma, sebaceoma, apocrine hidrocystoma, apocrine tubular papillary adenoma, hidradenoma papilliferum, and syringocystadenoma papilliferum. Hidradenomas exhibited variable positive staining. Most poromas, syringomas, chondroid syringomas, cylindromas, and spiradenomas were negative or only focally and weakly positive. Focal staining was present in all pilomatrixomas. Thirteen of 14 basal cell carcinomas, 21 of 24 squamous carcinomas, and all 6 sebaceous carcinomas exhibited positive staining. The 1 apocrine carcinoma, both mucinous carcinomas, and 2 of 3 microcystic adnexal carcinomas also exhibited positive staining, whereas the 1 eccrine porocarcinoma and the 1 adenoid cystic carcinoma were negative. One of 11 Merkel cell carcinomas exhibited focal weak staining. Our findings demonstrate that GATA3 is expressed in a wide variety of benign and malignant cutaneous epithelial neoplasms. In addition to carcinomas of breast and urothelial origin and other more recently described GATA3-positive tumors, the differential diagnosis of a metastatic tumor of unknown primary origin that expresses GATA3 should also include a carcinoma of cutaneous epithelial origin.
Evaluation of 5 different labeled polymer immunohistochemical detection systems.

PubMed

Skaland, Ivar; Nordhus, Marit; Gudlaugsson, Einar; Klos, Jan; Kjellevold, Kjell H; Janssen, Emiel A M; Baak, Jan P A

2010-01-01

Immunohistochemical staining is important for diagnosis and therapeutic decision making but the results may vary when different detection systems are used. To analyze this, 5 different labeled polymer immunohistochemical detection systems, REAL EnVision, EnVision Flex, EnVision Flex+ (Dako, Glostrup, Denmark), NovoLink (Novocastra Laboratories Ltd, Newcastle Upon Tyne, UK) and UltraVision ONE (Thermo Fisher Scientific, Fremont, CA) were tested using 12 different, widely used mouse and rabbit primary antibodies, detecting nuclear, cytoplasmic, and membrane antigens. Serial sections of multitissue blocks containing 4% formaldehyde fixed paraffin embedded material were selected for their weak, moderate, and strong staining for each antibody. Specificity and sensitivity were evaluated by subjective scoring and digital image analysis. At optimal primary antibody dilution, digital image analysis showed that EnVision Flex+ was the most sensitive system (P < 0.005), with means of 8.3, 13.4, 20.2, and 41.8 gray scale values stronger staining than REAL EnVision, EnVision Flex, NovoLink, and UltraVision ONE, respectively. NovoLink was the second most sensitive system for mouse antibodies, but showed low sensitivity for rabbit antibodies. Due to low sensitivity, 2 cases with UltraVision ONE and 1 case with NovoLink stained false negatively. None of the detection systems showed any distinct false positivity, but UltraVision ONE and NovoLink consistently showed weak background staining both in negative controls and at optimal primary antibody dilution. We conclude that there are significant differences in sensitivity, specificity, costs, and total assay time in the immunohistochemical detection systems currently in use.
Sensitivity of Helicobacter pylori detection by Giemsa staining is poor in comparison with immunohistochemistry and fluorescent in situ hybridization and strongly depends on inflammatory activity.

PubMed

Kocsmár, Éva; Szirtes, Ildikó; Kramer, Zsófia; Szijártó, Attila; Bene, László; Buzás, György Miklós; Kenessey, István; Bronsert, Peter; Csanadi, Agnes; Lutz, Lisa; Werner, Martin; Wellner, Ulrich Friedrich; Kiss, András; Schaff, Zsuzsa; Lotz, Gábor

2017-08-01

Conventional stainings (including H&E and special stains like Giemsa) are the most widely applied histopathologic detection methods of Helicobacter pylori (HP). We aimed to compare the diagnostic performance of Giemsa staining with immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) on a monocentric cohort of 2896 gastric biopsies and relate results to histologic alterations in order to find such histopathologic subgroups in which these methods underperform. All cases were categorized regarding presence or absence of chronic gastritis, inflammatory activity, and mucosal structural alterations. Giemsa revealed 687 cases (23.7%), IHC 795 cases (27.5%), and FISH 788 cases (27.2%) as being HP positive. Giemsa showed significantly lower overall sensitivity (83.3%) compared to IHC (98.8%) and FISH (98.0%). Moreover, the sensitivity of Giemsa dramatically dropped to 33.6% in the nonactive cases. We found that sensitivity of Giemsa strongly depends on HP density and, accordingly, on the presence of activity. Structural alterations (intestinal metaplasia, atrophy, etc.) had only no or weak effect on sensitivity of the three stainings. Both IHC and FISH proved to be equally reliable HP detecting techniques whose diagnostic performance is minimally influenced by mucosal inflammatory and structural alterations contrary to conventional stainings. We highly recommend immunohistochemistry for clinically susceptible, nonactive chronic gastritis cases, if the conventional stain-based HP detection is negative. Moreover, we recommend to use IHC more widely as basic HP stain. Helicobacter pylori FISH technique is primarily recommended to determine bacterial clarithromycin resistance. Furthermore, it is another accurate diagnostic tool for HP. © 2017 John Wiley & Sons Ltd.
Ber-H2 (CD30) immunohistochemical staining in malignant melanoma.

PubMed

Polski, J M; Janney, C G

1999-09-01

Malignant melanoma can be included in the differential diagnosis of Hodgkin's disease, anaplastic large cell lymphoma, or embryonal carcinoma These malignancies express CD30, a marker of diagnostic value. A retrospective immunohistochemical study was undertaken to determine the frequency of immunoreactivity of Ber-H2 (anti-CD30 monoclonal antibody) in malignant melanoma Archival paraffin-embedded tissue from 24 primary and metastatic lesions was used. No Ber-H2 labeling was observed in the majority of the studied cases. Variable weak cytoplasmic staining was present in only one case. The findings are compared with the previous reports claiming frequent CD30 expression in malignant melanoma. We discuss issues pertaining to the interpretation of the Ber-H2 IHC staining in formalin-fixed, paraffin-embedded tissue.
Miki (Mitotic Kinetics Regulator) Immunoexpression in Normal Liver, Cirrhotic Areas and Hepatocellular Carcinomas: a Preliminary Study with Clinical Relevance.

PubMed

Fernández-Vega, Iván; Santos-Juanes, Jorge; Camacho-Urkaray, Emma; Lorente-Gea, Laura; García, Beatriz; Gutiérrez-Corres, Francisco Borja; Quirós, Luis M; Guerra-Merino, Isabel; Aguirre, José Javier

2018-02-12

Hepatocellular carcinoma (HCC) is the most common type of primary malignant tumor in the liver. One of the main features of cancer survival is the generalized loss of growth control exhibited by cancer cells, and Miki is a protein related to the immunoglobulin superfamily that plays an important role in mitosis. We aim to study protein expression levels of Miki in non-tumoral liver and 20 HCCs recruited from a Pathology Department. Clinical information was also obtained. A tissue microarray was performed, and immunohistochemical techniques applied to study protein expression levels of Miki. In normal liver, Miki was weakly expressed, showing nuclear staining in the hepatocytes. Cirrhotic areas and HCCs showed a variety of staining patterns. Most HCC samples showed positive expression, with three different staining patterns being discernible: nuclear, cytoplasmic and mixed. Statistical analysis showed a significant association between grade of differentiation, Ki-67 proliferative index, survival rates and staining patterns. This study has revealed the positive expression of Miki in normal liver, cirrhotic areas and HCCs. Three different staining patterns of Miki expression with clinical relevance were noted in HCCs.
Morin Stain Detects Aluminum-Containing Macrophages in Macrophagic Myofasciitis and Vaccination Granuloma With High Sensitivity and Specificity.

PubMed

Chkheidze, Rati; Burns, Dennis K; White, Charles L; Castro, Diana; Fuller, Julie; Cai, Chunyu

2017-04-01

Macrophagic myofasciitis (MMF) is an inflammatory condition associated with the intramuscular (i.m.) injection of aluminum adjuvant-containing vaccines. It is clinically characterized by myalgia, weakness, and chronic fatigue and histologically by aggregates of cohesive macrophages with abundant basophilic, periodic acid-Schiff (PAS)-positive, diastase-resistant granules that percolate through the peri- and endomysium without eliciting substantial myofiber damage. The definitive diagnosis of MMF requires demonstration of aluminum within these macrophages. We evaluated the Morin stain, a simple, 2-step histochemical stain for aluminum, as a confirmatory diagnostic tool for MMF. Among 2270 muscle biopsies processed at UTSW between 2010 and 2015, a total of 12 MMF cases and 1 subcutaneous vaccination granuloma case were identified (11 pediatric, 2 adults). With the Morin stain, all 13 cases showed strong granular reactivity within the cytoplasm of macrophages but not in myofibers or connective tissue. Three cases of inflammatory myopathy with abundant macrophages (IMAM), 8 cases of granulomatous inflammation and 23 other deltoid muscle biopsies used as controls were all negative. Morin stain could be used in both formalin-fixed paraffin-embedded and cryostat sections. Thus, Morin stain detects aluminum with high sensitivity and specificity in human muscle and soft tissue and may improve the diagnostic yield of MMF and vaccination granuloma. © 2017 American Association of Neuropathologists, Inc.
Atypical nuclear localization of VIP receptors in glioma cell lines and patients

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

Barbarin, Alice; Séité, Paule; Godet, Julie

Highlights: • The VIP receptor VPAC1 contains a putative NLS signal. • VPAC1 is predominantly nuclear in GBM cell lines but not VPAC2. • Non-nuclear VPAC1/2 protein expression is correlated with glioma grade. • Nuclear VPAC1 is observed in 50% of stage IV glioma (GBM). - Abstract: An increasing number of G protein-coupled receptors, like receptors for vasoactive intestinal peptide (VIP), are found in cell nucleus. As VIP receptors are involved in the regulation of glioma cell proliferation and migration, we investigated the expression and the nuclear localization of the VIP receptors VPAC1 and VPAC2 in this cancer. First, bymore » applying Western blot and immunofluorescence detection in three human glioblastoma (GBM) cell lines, we observed a strong nuclear staining for the VPAC1 receptor and a weak nuclear VPAC2 receptor staining. Second, immunohistochemical staining of VPAC1 and VPAC2 on tissue microarrays (TMA) showed that the two receptors were expressed in normal brain and glioma tissues. Expression in the non-nuclear compartment of the two receptors significantly increased with the grade of the tumors. Analysis of nuclear staining revealed a significant increase of VPAC1 staining with glioma grade, with up to 50% of GBM displaying strong VPAC1 nuclear staining, whereas nuclear VPAC2 staining remained marginal. The increase in VPAC receptor expression with glioma grades and the enhanced nuclear localization of the VPAC1 receptors in GBM might be of importance for glioma progression.« less
Low vacuum scanning electron microscopy for paraffin sections utilizing the differential stainability of cells and tissues with platinum blue.

PubMed

Inaga, Sumire; Hirashima, Sayuri; Tanaka, Keiichi; Katsumoto, Tetsuo; Kameie, Toshio; Nakane, Hironobu; Naguro, Tomonori

2009-07-01

The present study introduces a novel method for the direct observation of histological paraffin sections by low vacuum scanning electron microscopy (LVSEM) with platinum blue (Pt-blue) treatment. Pt-blue was applied not only as a backscattered electron (BSE) signal enhancer but also as a histologically specific stain. In this method, paraffin sections of the rat tongue prepared for conventional light microscopy (LM) were stained on glass slides with a Pt-blue staining solution (pH 9) and observed in a LVSEM using BSE detector. Under LVSEM, overviews of whole sections as well as three-dimensional detailed observations of individual cells and tissues could be easily made at magnifications from x40 to x10,000. Each kind of cell and tissue observed in the section could be clearly distinguished due to the different yields of BSE signals, which depended on the surface structures and different affinities to Pt-blue. Thus, we roughly classified cellular and tissue components into three groups according to the staining intensity of Pt-blue observed by LM and LVSEM: 1) a strongly stained (deep blue by LM and brightest by LVSEM) group which included epithelial tissue, endothelium and mast cells; 2) a moderately stained (light blue and bright) group which included muscular tissue and nervous tissue; 3) an unstained or weakly stained (colorless and dark) group which included elastic fibers and collagen fibers. We expect that this method will prove useful for the three-dimensional direct observation of histological paraffin sections of various tissues by LVSEM with higher resolutions than LM.
Improvement of malaria diagnostic system based on acridine orange staining.

PubMed

Kimura, Masatsugu; Teramoto, Isao; Chan, Chim W; Idris, Zulkarnain Md; Kongere, James; Kagaya, Wataru; Kawamoto, Fumihiko; Asada, Ryoko; Isozumi, Rie; Kaneko, Akira

2018-02-07

Rapid diagnosis of malaria using acridine orange (AO) staining and a light microscope with a halogen lamp and interference filter was deployed in some malaria-endemic countries. However, it has not been widely adopted because: (1) the lamp was weak as an excitation light and the set-up did not work well under unstable power supply; and, (2) the staining of samples was frequently inconsistent. The halogen lamp was replaced by a low-cost, blue light-emitting diode (LED) lamp. Using a reformulated AO solution, the staining protocol was revised to make use of a concentration gradient instead of uniform staining. To evaluate this new AO diagnostic system, a pilot field study was conducted in the Lake Victoria basin in Kenya. Without staining failure, malaria infection status of about 100 samples was determined on-site per one microscopist per day, using the improved AO diagnostic system. The improved AO diagnosis had both higher overall sensitivity (46.1 vs 38.9%: p = 0.08) and specificity (99.0 vs 96.3%) than the Giemsa method (N = 1018), using PCR diagnosis as the standard. Consistent AO staining of thin blood films and rapid evaluation of malaria parasitaemia with the revised protocol produced superior results relative to the Giemsa method. This AO diagnostic system can be set up easily at low cost using an ordinary light microscope. It may supplement rapid diagnostic tests currently used in clinical settings in malaria-endemic countries, and may be considered as an inexpensive tool for case surveillance in malaria-eliminating countries.

Rasch analysis of three dry eye questionnaires and correlates with objective clinical tests.

PubMed

McAlinden, Colm; Gao, Rongrong; Wang, Qinmei; Zhu, Senmiao; Yang, Jing; Yu, Ayong; Bron, Anthony J; Huang, Jinhai

2017-04-01

To assess the psychometric properties of Chinese versions of the Ocular Comfort Index (OCI), Ocular Surface Disease Index (OSDI) and McMonnies questionnaires. Further, to assess the correlation between questionnaire scores and objective dry eye disease (DED) clinical tests. Translated versions of the OCI, OSDI and McMonnies questionnaires were completed in a random order by 238 participants with DED. Objective clinical tests included visual acuity (VA), fluorescein tear film break-up time (TBUT), corneal fluorescein staining, Schirmer I testing and meibomian gland grading. Rasch analysis was used to assess questionnaire psychometrics and spearman rank for correlations. For the OCI, the person separation was 2.31, item infit and outfit statistics ranged from 0.74-1.14 and 0.75-1.32, respectively, and targeting 1.54 logits. For the OSDI, person separation was 0.94. None of the three subscales provided valid measurements based on Rasch analysis. For the McMonnies questionnaire, person separation was 1.17, item infit and outfit statistics ranged from 0.7 to 1.21 and 0.51-3.49, respectively. There were weak correlations between questionnaire scores and clinical tests. There were weak correlations between OSDI scores and VA, fluorescein TBUT, Schirmer I testing and corneal fluorescein staining. There were weak correlations between McMonnies scores and VA, fluorescein TBUT, Schirmer I testing, and corneal fluorescein staining and meibomian gland grading. The OCI questionnaire was the only questionnaire that provided valid measurement on the basis of Rasch analysis, although slight multidimensionality was found. There were weak correlations between OCI scores and fluorescein TBUT, Schirmer I testing, and corneal fluorescein staining. Due to this paradoxical disconnect between symptoms and signs and the repeatability of tests, the use of both subjective and objective markers in the clinical management of patients or as endpoints in clinical trials would appear prudent. Copyright © 2017 Elsevier Inc. All rights reserved.
Distinct T cell interactions with HLA class II tetramers characterize a spectrum of TCR affinities in the human antigen-specific T cell response.

PubMed

Reichstetter, S; Ettinger, R A; Liu, A W; Gebe, J A; Nepom, G T; Kwok, W W

2000-12-15

The polyclonal nature of T cells expanding in an ongoing immune response results in a range of disparate affinities and activation potential. Recently developed human class II tetramers provide a means to analyze this diversity by direct characterization of the trimolecular TCR-peptide-MHC interaction in live cells. Two HSV-2 VP16(369-379)-specific, DQA1*0102/DQB1*0602 (DQ0602)-restricted T cell clones were compared by means of T cell proliferation assay and HLA-DQ0602 tetramer staining. These two clones were obtained from the same subject, but show different TCR gene usage. Clone 48 was 10-fold more sensitive to VP16(369-379) peptide stimulation than clone 5 as assayed by proliferation assays, correlating with differences in MHC tetramer binding. Clone 48 gave positive staining with the DQ0602/VP16(369-379) tetramer at either 23 or 37 degrees C. Weak staining was also observed at 4 degrees C. Clone 5 showed weaker staining compared with clone 48 at 37 degrees C, and no staining was observed at 23 degrees C or on ice. Receptor internalization was not required for positive staining. Competitive binding indicates that the cell surface TCR of clone 48 has higher affinity for the DQ0602/VP16(369-379) complex than clone 5. The higher binding affinity of clone 48 for the peptide-MHC complex also correlates with a slower dissociation rate compared with clone 5.
Classification of phytoplankton cells as live or dead using the vital stains fluorescein diacetate and 5-chloromethylfluorescein diacetate.

PubMed

MacIntyre, Hugh L; Cullen, John J

2016-08-01

Regulations for ballast water treatment specify limits on the concentrations of living cells in discharge water. The vital stains fluorescein diacetate (FDA) and 5-chloromethylfluorescein diacetate (CMFDA) in combination have been recommended for use in verification of ballast water treatment technology. We tested the effectiveness of FDA and CMFDA, singly and in combination, in discriminating between living and heat-killed populations of 24 species of phytoplankton from seven divisions, verifying with quantitative growth assays that uniformly live and dead populations were compared. The diagnostic signal, per-cell fluorescence intensity, was measured by flow cytometry and alternate discriminatory thresholds were defined statistically from the frequency distributions of the dead or living cells. Species were clustered by staining patterns: for four species, the staining of live versus dead cells was distinct, and live-dead classification was essentially error free. But overlap between the frequency distributions of living and heat-killed cells in the other taxa led to unavoidable errors, well in excess of 20% in many. In 4 very weakly staining taxa, the mean fluorescence intensity in the heat-killed cells was higher than that of the living cells, which is inconsistent with the assumptions of the method. Applying the criteria of ≤5% false negative plus ≤5% false positive errors, and no significant loss of cells due to staining, FDA and FDA+CMFDA gave acceptably accurate results for only 8-10 of 24 species (i.e., 33%-42%). CMFDA was the least effective stain and its addition to FDA did not improve the performance of FDA alone. © 2016 The Authors. Journal of Phycology published by Wiley Periodicals, Inc. on behalf of Phycological Society of America.
Cytochemical evaluation of the Guard procedure a regressive staining method for demonstrating chromosomal basic proteins. I. Effects of fixation, blocking reactions, selective extractions, and polyacid "differentiation".

PubMed

Cowden, R R; Rasch, E M; Curtis, S K

1976-08-12

Appropriately fixed preparations stained by a modification of the Guard (1959) reaction for "sex chromatin" display selective staining of interphase chromatin and mitotic or meiotic chromosomes. This is a regressive staining method which seems to depend on the selective displacement of an acidic dye from less basic structures, and retention of the dye at more basic sites. The results obtained with the reaction can be controlled by the length of time that the preparations are "differentiated" in solutions containing phosphomolybdic and phosphotungstic acids (polyacids). After three- or four-hour exposures to polyacid solutions, all chromatin is stained. However, with longer differentiation, "condensed" chromatin can be stained preferentially. Of a number of fixatives investigated, only 10% formalin, ethanol-acetic acid (3:1), and Bouin's solution proved useful. Others resulted in diminished specificity or a total loss of selectivity. The most intense staining was obtained after formalin fixation. Less intense dyebinding was observed after fixation in 3:1 - probably due to extraction of some histone fractions-and the least amount of dye was bound in Bouin's-fixed chromatin - probably due to blockage of arginine residues by picric acid. The reaction was not affected by enzymatic removal of nucleic acids or the extraction of lipids. It was diminished by treatment with trypsin or weak acetylation, and it was completely prevented by strong acetylation, deamination, or extraction of basic proteins with HCl. The results presented suggest that the modified Guard (1959) procedure selectively demonstrates basic nucleoproteins. Further, by the use of regressive differentiation in polyacid solutions, the retention of dye in more condensed chromatin can be favored.
Strong FGFR3 staining is a marker for FGFR3 fusions in diffuse gliomas

PubMed Central

Annala, Matti; Lehtinen, Birgitta; Kesseli, Juha; Haapasalo, Joonas; Ruusuvuori, Pekka; Yli-Harja, Olli; Visakorpi, Tapio; Haapasalo, Hannu; Nykter, Matti; Zhang, Wei

2017-01-01

Abstract Background Inhibitors of fibroblast growth factor receptors (FGFRs) have recently arisen as a promising treatment option for patients with FGFR alterations. Gene fusions involving FGFR3 and transforming acidic coiled-coil protein 3 (TACC3) have been detected in diffuse gliomas and other malignancies, and fusion-positive cases have responded well to FGFR inhibition. As high FGFR3 expression has been detected in fusion-positive tumors, we sought to determine the clinical significance of FGFR3 protein expression level as well as its potential for indicating FGFR3 fusions. Methods We performed FGFR3 immunohistochemistry on tissue microarrays containing 676 grades II–IV astrocytomas and 116 grades II–III oligodendroglial tumor specimens. Fifty-one cases were further analyzed using targeted sequencing. Results Moderate to strong FGFR3 staining was detected in gliomas of all grades, was more common in females, and was associated with poor survival in diffuse astrocytomas. Targeted sequencing identified FGFR3-TACC3 fusions and an FGFR3-CAMK2A fusion in 10 of 15 strongly stained cases, whereas no fusions were found in 36 negatively to moderately stained cases. Fusion-positive cases were predominantly female and negative for IDH and EGFR/PDGFRA/MET alterations. These and moderately stained cases show lower MIB-1 proliferation index than negatively to weakly stained cases. Furthermore, stronger FGFR3 expression was commonly observed in malignant tissue regions of lower cellularity in fusion-negative cases. Importantly, subregional negative FGFR3 staining was also observed in a few fusion-positive cases. Conclusions Strong FGFR3 protein expression is indicative of FGFR3 fusions and may serve as a clinically applicable predictive marker for treatment regimens based on FGFR inhibitors. PMID:28379477
Autoantibodies in melanoma-associated retinopathy target TRPM1 cation channels of retinal ON bipolar cells.

PubMed

Dhingra, Anuradha; Fina, Marie E; Neinstein, Adam; Ramsey, David J; Xu, Ying; Fishman, Gerald A; Alexander, Kenneth R; Qian, Haohua; Peachey, Neal S; Gregg, Ronald G; Vardi, Noga

2011-03-16

Melanoma-associated retinopathy (MAR) is characterized by night blindness, photopsias, and a selective reduction of the electroretinogram b-wave. In certain cases, the serum contains autoantibodies that react with ON bipolar cells, but the target of these autoantibodies has not been identified. Here we show that the primary target of autoantibodies produced in MAR patients with reduced b-wave is the TRPM1 cation channel, the newly identified transduction channel in ON bipolar cells. Sera from two well characterized MAR patients, but not from a control subject, stained human embryonic kidney cells transfected with the TRPM1 gene, and Western blots probed with these MAR sera showed the expected band size (∼180 kDa). Staining of mouse and primate retina with MAR sera revealed immunoreactivity in all types of ON bipolar cells. Similar to staining for TRPM1, staining with the MAR sera was strong in dendritic tips and somas and was weak or absent in axon terminals. This staining colocalized with GFP in Grm6-GFP transgenic mice, where GFP is expressed in all and only ON bipolar cells, and also colocalized with Gα(o), a marker for all types of ON bipolar cells. The staining in ON bipolar cells was confirmed to be specific to TRPM1 because MAR serum did not stain these cells in a Trpm1(-/-) mouse. Evidence suggests that the recognized epitope is likely intracellular, and the sera can be internalized by retinal cells. We conclude that the vision of at least some patients with MAR is compromised due to autoantibody-mediated inactivation of the TRPM1 channel.
Short-term in vivo evaluation of novel vital dyes for intraocular surgery.

PubMed

Haritoglou, Christos; Tadayoni, Ramin; May, Christian A; Gass, Carolin A; Freyer, Wolfgang; Priglinger, Siegfried G; Kampik, Anselm

2006-01-01

To evaluate the staining characteristics and safety of potential new dyes for intraocular surgery in porcine eyes. Four dyes in different solutions (light green SF yellowish [LGSF]: 2%; copper(II) phthalocyanine-tetrasulfonic acid [E68]: 2% and 0.5%; bromophenol blue [BPB]: 2%, 1%, and 0.2%; and Chicago blue [CB]: 2% and 0.5%) were included in this investigation. All dyes were dissolved and diluted using balanced salt solution (BSS plus; Alcon Laboratories, Inc., Fort Worth, TX). After triamcinolone-assisted vitrectomy on 10 porcine eyes in vivo, the dyes were first injected into the air-filled vitreous cavity. After 1 minute, the dye was removed by irrigation with BSS, and the staining effect was graded by two examiners. After vitrectomy, the same dyes and concentrations were injected in the air-filled anterior chamber to stain the lens capsule of the same eye. After surgery, the eyes were enucleated and underwent fixation for light and electron microscopy. The animals were killed by injection of pentobarbital (50 mg/kg). For controls, each BSS plus alone and indocyanine green 0.5% were applied in one eye. On the retinal surface, bright staining of the retinal surface was seen after application of BPB 2% and 1%. The staining effect was less pronounced but still very good using E68 2%, and CB 2% and weak using BPB 0.2%, E68 0.5% and CB 0.5% as well as indocyanine green 0.5%. No staining of the retinal surface but of the vitreous was seen after application of LGSF 2%. The lens capsule stained very well with E68 2%, CB 2% and 0.5%, and BPB 2%, 1%, and 0.2% but not with LGSF. No histologic abnormalities were seen after the application in any eye after dye injection. No dye-related complications occurred during surgery. In this study, we identified three dyes with satisfying staining characteristics in both anterior and posterior segments. Because BPB stained the retinal surface and lens capsule at a low concentration (0.2%) with no signs of toxicity, this dye seems to be the most promising candidate for application in humans.
Real-Time Binding Kinetic Analyses of the Interaction of the Dietary Stain Orange II with Dentin Matrix.

PubMed

Alraies, Amr; Cole, David K; S Rees, Jeremy; Glasse, Carl; Young, Nigel; Waddington, Rachel J; Sloan, Alastair J

2018-06-09

Dietary stains can be adsorbed into the dentin of teeth. Using Orange II as a model dietary stain, this study investigated the strength of its interaction with the mineral and protein components of dentin matrix and how hydrogen peroxide (H 2 O 2 ) treatment influences this interaction. Dentin slices were prepared from human teeth and were either deproteinized (5.6% sodium hypochlorite, 12 days), demineralised (0.5 M EDTA, 3 days) or left as intact control samples. Samples were stained with Orange II for 1-168 h, during which staining intensity was quantified by image analysis. Similarly, uptake of stain by deproteinized / demineralized samples treated with 10 or 30% H 2 O 2 was investigated. Using surface plasmon resonance technology, real-time binding kinetics were determined assessing the interaction of orange II with the dentin matrix protein constituents, collagen type I, biglycan, decorin, dentin sialoprotein and osteopontin. Deproteinization of dentin matrix reduced the uptake of the orange II compared to the intact control. Conversely, demineralization of dentin samples increased the uptake of the dye. Treatment of samples for 48 h with H 2 O 2 reduced subsequent uptake of the orange II. Real-time kinetic analysis indicated moderate strength of binding for Orange II with collagen type I, weak binding with decorin and biglycan and negligible binding with dentine sialoprotein and osteopontin. These results indicate a predominant role for collagen type I, which accounts for 90% of the organic protein matrix of teeth, for attracting dietary stains. Binding analyses indicate that the interaction is highly dissociable, and further binding is reduced following H 2 O 2 treatment. This study provides new information regarding adsorption of dietary stains into tooth dentin, suggesting that they are attracted and moderately bound to the collagen type I matrix. This study also contributes valuable information for discussion for considering the effect of H 2 O 2 on bleaching teeth and its influence on subsequent uptake of dietary stains following whitening treatments. Copyright © 2018. Published by Elsevier Ltd.
Amplification and overexpression of cyclin D1 in breast cancer detected by immunohistochemical staining.

PubMed

Gillett, C; Fantl, V; Smith, R; Fisher, C; Bartek, J; Dickson, C; Barnes, D; Peters, G

1994-04-01

Immunohistochemical staining with a monoclonal antibody against human cyclin D1 can be used to identify breast cancers that have an amplification of the q13 region of chromosome 11. In general, the intensity of staining is directly proportional to the degree of DNA amplification. In two unusual tumors, in which the CCND1 locus is highly amplified but staining is relatively weak, it appears that the DNA has undergone rearrangement and that the amplified/rearranged CCND1 allele may have reduced transcriptional activity. More significantly, the immunohistochemical technique identifies additional tumors in which the cyclin D1 gene is overexpressed with only marginal or undetectable increases in copy number, implying that other mechanisms can lead to deregulated expression. These results suggest that the frequency of overexpression is much higher than previously concluded from DNA-based analyses and that more than one-third of human breast cancers may contain excessive levels of cyclin D1. The technique we describe should facilitate the detection of this abnormality in a clinical setting and clarify its prognostic significance.
Automated image analysis for quantification of reactive oxygen species in plant leaves.

PubMed

Sekulska-Nalewajko, Joanna; Gocławski, Jarosław; Chojak-Koźniewska, Joanna; Kuźniak, Elżbieta

2016-10-15

The paper presents an image processing method for the quantitative assessment of ROS accumulation areas in leaves stained with DAB or NBT for H 2 O 2 and O 2 - detection, respectively. Three types of images determined by the combination of staining method and background color are considered. The method is based on the principle of supervised machine learning with manually labeled image patterns used for training. The method's algorithm is developed as a JavaScript macro in the public domain Fiji (ImageJ) environment. It allows to select the stained regions of ROS-mediated histochemical reactions, subsequently fractionated according to the weak, medium and intense staining intensity and thus ROS accumulation. It also evaluates total leaf blade area. The precision of ROS accumulation area detection is validated by the Dice Similarity Coefficient in the case of manual patterns. The proposed framework reduces the computation complexity, once prepared, requires less image processing expertise than the competitive methods and represents a routine quantitative imaging assay for a general histochemical image classification. Copyright © 2016 Elsevier Inc. All rights reserved.
Nuclear β-Catenin Expression is Frequent in Sinonasal Hemangiopericytoma and Its Mimics.

PubMed

Jo, Vickie Y; Fletcher, Christopher D M

2017-06-01

Sinonasal hemangiopericytoma (HPC) is a tumor showing pericytic myoid differentiation and which arises in the nasal cavity and paranasal sinuses. CTNNB1 mutations appear to be a consistent aberration in sinonasal HPC, and nuclear expression of β-catenin has been reported. Our aim was to evaluate the frequency of β-catenin expression in sinonasal HPC and its histologic mimics in the upper aerodigestive tract. Cases were retrieved from the surgical pathology and consultation files. Immunohistochemical staining for β-catenin was performed on 50 soft tissue tumors arising in the sinonasal tract or oral cavity, and nuclear staining was recorded semiquantitatively by extent and intensity. Nuclear reactivity for β-catenin was present in 19/20 cases of sinonasal HPC; 17 showed moderate-to-strong multifocal or diffuse staining, and 2 had moderate focal nuclear reactivity. All solitary fibrous tumors (SFT) (10/10) showed focal-to-multifocal nuclear staining, varying from weak to strong in intensity. Most cases of synovial sarcoma (9/10) showed nuclear β-catenin expression in the spindle cell component, ranging from focal-weak to strong-multifocal. No cases of myopericytoma (0/10) showed any nuclear β-catenin expression. β-catenin expression is prevalent in sinonasal HPC, but is also frequent in SFT and synovial sarcoma. Our findings indicate that β-catenin is not a useful diagnostic tool in the evaluation of spindle cell tumors with a prominent hemangiopericytoma-like vasculature in the sinonasal tract and oral cavity, and that definitive diagnosis relies on the use of a broader immunohistochemical panel.
Functional anatomy of the lacrimal gland in African black ostrich Struthio camelus domesticus in the embryonic and postnatal period.

PubMed

Klećkowska-Nawrot, Joanna; Goździewska-Harłajczuk, Karolina; Nowaczyk, Renata; Krasucki, Krzysztof

2015-03-25

The aim of the present study was morphological and histochemical analysis of the lacrimalgland (LG) in African black ostrich Struthio camelus domesticus in the embryonic and postnatalperiod. Studies were conducted on 50 ostriches aged between the 28th day of incubation until7 months old. Tissue sections were stained with haematoxylin and eosin, Azan trichrome,periodic acid-Schiff, Alcian blue pH 2.5, aldehyde fuchsin and Hale's dialysed iron. The LGin ostrich was classified as a tubulo-acinar type. The primordia of the lobes were determinedin the LG structure on the 28th day of incubation, whilst the weakly visible lobes with aciniand tubules were observed on the 40th day of incubation. Morphometric studies of the LGshowed steady growth, characterised by an increase in both length and width. Histometricmeasurements of lobe size showed little difference between the first, second and third agegroups, whilst in the fourth age group a marked increase in size of lobes was observed.The study showed that, apart from morphological changes, during the growth of the LGthe character of acid mucopolysaccharides changed. Sulphated acid mucopolysaccharideswere indicated, particularly with aldehyde fuchsin (AF) staining in the fourth age group.The Hale's dialysed iron (HDI) staining showed a low concentration of carboxylated acidmucopolysaccharides in the first and second age groups and a higher concentration in thethird and fourth age groups. Periodic acid-Schiff staining (PAS)-positive cells were observedin each age group, but only a small number of cells with a weakly PAS-positive reaction weredemonstrated in the first age group.
Primary cell culture and morphological characterization of canine dermal papilla cells and dermal fibroblasts.

PubMed

Bratka-Robia, Christine B; Mitteregger, Gerda; Aichinger, Amanda; Egerbacher, Monika; Helmreich, Magdalena; Bamberg, Elmar

2002-02-01

Skin biopsies were taken from female dogs, the primary hair follicles isolated and the dermal papilla dissected. After incubation in supplemented Amniomax complete C100 medium in 24-well culture plates, the dermal papilla cells (DPC) grew to confluence within 3 weeks. Thereafter, they were subcultivated every 7 days. Dermal fibroblast (DFB) cultures were established by explant culture of interfollicular dermis in serum-free medium, where they reached confluence in 10 days. They were subcultivated every 5 days. For immunohistochemistry, cells were grown on cover slips for 24 h, fixed and stained with antibodies against collagen IV and laminin. DPC showed an aggregative growth pattern and formation of pseudopapillae. Intensive staining for collagen IV and laminin could be observed until the sixth passage. DFB grew as branching, parallel lines and showed only weak staining for collagen IV and laminin.
[Expression and distribution of xenoantigen alpha-Gal in intervertebral disk of Chinese banna minipig inbred line].

PubMed

Shou, Jian-guo; Mi, Jian-hong; Ying, Da-jun

2002-09-01

To investigate the expression and distribution of xenoantigen in intervertebral disk of Chinese banna minipig inbred line, and to study the availability of xenograft transplantation of intervertebral disk. Samples of intervertebral disk were collected from six Banna pigs of 8 to 11-month-old. The fixation, embedment and slice were performed. alpha-Gal specific binding lection (BSI-B4) were used as affinity reagents and affinity-immunohistochemistry assays (SABC methods and DAB stain) were conducted to detect the expression and distribution of xenoantigen (alpha-Gal). alpha-Gal was found in chondrocyte cell and chondrocyte-like cell in intervertebral disk which have the positive yellow-stained particulate aggradation. There was no stain in the matrix, elastic fiber and collagen fiber. The distribution of xenoantigen is locally in the tissue of intervertebral disk and its expression is weak. This suggests that the intervertebral disk of Banna pig may be alternative donor for xenotransplantation.
Characteristics of weak base-induced vacuoles formed around individual acidic organelles.

PubMed

Hiruma, Hiromi; Kawakami, Tadashi

2011-01-01

We have previously found that the weak base 4-aminopyridine induces Brownian motion of acidic organelles around which vacuoles are formed, causing organelle traffic disorder in neurons. Our present study investigated the characteristics of vacuoles induced by weak bases (NH(4)Cl, aminopyridines, and chloroquine) using mouse cells. Individual vacuoles included acidic organelles identified by fluorescent protein expression. Mitochondria and actin filaments were extruded outside the vacuoles, composing the vacuole rim. Staining with amine-reactive fluorescence showed no protein/amino acid content in vacuoles. Thus, serous vacuolar contents are probably partitioned by viscous cytosol, other organelles, and cytoskeletons, but not membrane. The weak base (chloroquine) was immunochemically detected in intravacuolar organelles, but not in vacuoles. Early vacuolization was reversible, but long-term vacuolization caused cell death. The vacuolization and cell death were blocked by the vacuolar H(+)-ATPase inhibitor and Cl--free medium. Staining with LysoTracker or LysoSensor indicated that intravacuolar organelles were strongly acidic and vacuoles were slightly acidic. This suggests that vacuolization is caused by accumulation of weak base and H(+) in acidic organelles, driven by vacuolar H(+)-ATPase associated with Cl(-) entering, and probably by subsequent extrusion of H(+) and water from organelles to the surrounding cytoplasm.
The cytochemical and ultrastructural characteristics of phagocytes in the Pacific oyster Crassostrea gigas.

PubMed

Jiang, Shuai; Jia, Zhihao; Xin, Lusheng; Sun, Ying; Zhang, Ran; Wang, Weilin; Wang, Lingling; Song, Linsheng

2016-08-01

Phagocytes have been proved to play vital roles in the innate immune response. However, the cellular characteristics of phagocytes in invertebrates, especially in molluscs, remain largely unknown. In the present study, fluorescence activated cell sorting (FACS) was employed to sort the phagocytes from the non-phagocytic haemocytes of the Pacific oyster Crassostrea gigas. The cytochemical staining analysis revealed that phagocytes were positive staining for α-naphthyl acetate esterase and myeloperoxidase, while negative staining for toluidine blue and periodic acid-Schiff. The non-phagocytic haemocytes exhibited positive staining for periodic acid-Schiff, weak positive staining for toluidine blue, but negative staining for α-naphthyl acetate esterase and myeloperoxidase. In addition, phagocytes exhibited ultrastructural cellular features similar to those of macrophages, with large cell diameter, rough cell membrane and extended pseudopodia revealed by the scanning electron microscopy, while the non-phagocytic haemocytes exhibited small cell diameter, smooth cell surface and round spherical shape. Transmission electron microscopy further demonstrated that phagocytes were abundant of cytoplasmic bodies and mitochondria, while non-phagocytic haemocytes were characterized as the comparatively large cell nucleus with contorted and condensed heterochromatin adherent to the nuclear envelope. Moreover, compared with non-phagocytic haemocytes, phagocytes exhibited significantly higher levels of intracellular cytokines, including tumor necrosis factor, interferon-like protein and interleukin-17, and significantly higher abundance of lysosome and reactive oxygen species, which were of great importance to the activation of immune response and pathogen clearance. Taken together, these findings revealed the different cytochemical and ultrastructural features between phagocytes and non-phagocytic haemocytes in C. gigas, which would provide an important clue to investigate the mechanism of phagocytosis underlying the innate immune response. Copyright © 2016 Elsevier Ltd. All rights reserved.
Developmental expression of the glutathione S-transferase Yo subunit in the rat testis and epididymis using light microscope immunocytochemistry.

PubMed

Papp, S; Robaire, B; Hermo, L

1994-11-01

Glutathione S-transferases (GSTs) are a family of isozymes that catalyze the conjugation of glutathione with various toxic electrophilic compounds. GSTs are composed of several classes based on the degree of sequence homology of their subunits. The Yo subunit, a member of the mu class, is expressed at high levels in the testis and epididymis. The purpose of this study was to immunolocalize the GST-Yo in these tissues during development. The testes and epididymides of rats aged 7, 15, 21, 28, 39, 42, 45, 49, and 56 days were fixed in Bouin's fixative, and immunostained for light microscopic analysis. In the testis the cytoplasm of all germ cells was unreactive until day 39. At that time, step 18 spermatids appeared moderately reactive, while the few observed step 19 spermatids were intensely reactive as were their residual bodies. The presence of residual bodies indicates that spermiation takes place as early as day 39; however, the number of step 19 spermatids is low at this age. A progressive increase in the size of the tubule and number of elongating spermatids was seen between days 42 and 49. In addition, by day 49, a weak staining was observed in steps 12-15, moderate in steps 16-17, and intense in steps 18-19 spermatids. In terms of the intensity of staining, cell types stained, size of the tubules, and number of elongating spermatids, no difference was noted between day 49, 56, and adult animals. Thus Yo protein expression in germ cells reached maturity by day 49. The epithelial cells of the rete testis were intensely reactive at day 7 and remained so throughout development. In contrast, while the epithelial cells of the efferent ducts at day 7 were intensely reactive, they were weakly reactive by day 39 and remained so at later ages. Along the entire epididymis, the columnar epithelial cells showed a moderate apical/supranuclear reaction from day 7 to 28. By day 39 principal cells of the initial segment became weakly reactive, while those in the caput and corpus were moderately stained, a situation seen at later ages including adults. Only by day 49 did principal cells of the proximal cauda become moderately stained as seen in adult animals. Thus the expression of the Yo protein in the principal cells of the proximal cauda may be regulated by different factors than those of the caput and corpus epididymidis. Alternatively, the expression of the Yo subunit in principal cells of the proximal cauda may develop later since this region would be the last to receive luminally derived testicular products. In the initial segment, the decrease in staining of principal cells at day 39 may be due to an inhibiting factor emanating from the testis. Spermatozoa appeared in the lumen of each epididymal region well after the expression of Yo had reached its adult staining pattern indicating that they are not a factor. Overall these results suggest that the expression of GST-Yo in the various cells of the testis and epididymis are controlled by different factors during postnatal development.
Development of a stain shade guide to aid the measurement of extrinsic dental stain.

PubMed

Gadhia, K; Shah, R; Swaminathan, D; Wetton, S; Moran, J

2006-05-01

Accurate and reproducible assessment of extrinsic staining is pivotal to determining efficacy of some tooth whitening oral hygiene products. The aim of this study was: (1) to produce a stain shade guide to aid the in vitro and in vivo stain assessment (2) to assess intra- and inter-examiner reproducibility of stain assessment using the stain shade guide. Using chlorhexidine and tea, perspex and acrylic teeth specimens were stained. The amount of staining on the perspex was measured with a spectrophotometer and the values obtained were assigned to the stained acrylic teeth, which were made into a stain guide. Using clinical photographs and a group of 10 volunteers, stain area and intensity were assessed using the stain guide and the recognized Lobene stain index by two examiners. The degree of intra- and inter-examiner reproducibility for these measurements were assessed using Cohen's kappa statistics. For both the clinical examination and use of photographs, intra-examiner reproducibility for stain intensity was improved when using the stain guide compared with the Lobene Index. Similarly, when assessing inter-examiner reproducibility, stain intensity kappa values were greater using the stain guide (kappa = 0.82) compared with the Lobene Index (kappa = 0.57). The findings of this study would suggest that the use of the stain guide could be of importance in the assessment of extrinsic dental stain.
Characterization of the Mycobacterium tuberculosis phagosome and evidence that phagosomal maturation is inhibited

PubMed Central

1995-01-01

We have used the cryosection immunogold technique to study the composition of the Mycobacterium tuberculosis phagosome. We have used quantitative immunogold staining to determine the distribution of several known markers of the endosomal-lysosomal pathway in human monocytes after ingestion of either M. tuberculosis, Legionella pneumophila, or polystyrene beads. Compared with the other phagocytic particles studied, the M. tuberculosis phagosome exhibits delayed clearance of major histocompatibility complex (MHC) class I molecules, relatively intense staining for MHC class II molecules and the endosomal marker transferrin receptor, and relatively weak staining for the lysosomal membrane glycoproteins, CD63, LAMP-1, and LAMP-2 and the lysosomal acid protease, cathepsin D. In contrast to M. tuberculosis, the L. pneumophila phagosome rapidly clears MHC class I molecules and excludes all endosomal-lysosomal markers studied. In contrast to both live M. tuberculosis and L. pneumophila phagosomes, phagosomes containing either polystyrene beads or heat-killed M. tuberculosis stain intensely for lysosomal membrane glycoproteins and cathepsin D. These findings suggest that (a) M. tuberculosis retards the maturation of its phagosome along the endosomal-lysosomal pathway and resides in a compartment with endosomal, as opposed to lysosomal, characteristics; and (b) the intraphagosomal pathway, i.e., the pathway followed by several intracellular parasites that inhibit phagosome-lysosome fusion, is heterogeneous. PMID:7807006
PIM-1 kinase expression in adipocytic neoplasms: diagnostic and biological implications

PubMed Central

Nga, Min En; Swe, Nu Nu Ma; Chen, Kang Ting; Shen, Liang; Lilly, Michael B; Chan, Siew Pang; Salto-Tellez, Manuel; Das, Kakoli

2010-01-01

The differential diagnosis of soft tissue tumours poses a considerable challenge for pathologists, especially adipocytic tumours, as these may show considerable overlap in clinical presentation and morphological features with many other mesenchymal neoplasms. Hence, a specific and reliable marker that identifies adipocytic differentiation is much sought. We investigated the immunohistochemical expression of PIM-1 kinase in 35 samples of soft tissue tumours using tissue microarray technology and 49 full sections of adipocytic (n = 26) and non-adipocytic tumours (n = 23). Benign and malignant adipocytic tumours showed strong expression of PIM-1 while the non-adipocytic tumours were either negative or showed only weak staining for the protein. In myxoid liposarcomas, PIM-1 showed a distinct, unique vacuolar staining pattern, clearly outlining fine cytoplasmic lipid vacuoles. By contrast, non-adipocytic myxoid tumours (myxoma, chordoma and myxoid chondrosarcoma) did not show this vacuolar pattern of PIM-1 staining, although vacuolated cells were present on H&E. This differential expression was confirmed at a gene expression level in selected cases. Our results indicate that the expression of PIM-1 in adipose tissue may be a useful marker of adipocytic differentiation, in particular if the staining is both of high intensity and present in a unique, vacuolar pattern. PMID:19878356

The CD200-tolerance signaling molecule associated with pregnancy success is present in patients with early-stage breast cancer but does not favor nodal metastasis.

PubMed

Clark, David A; Dhesy-Thind, Sukhbinder; Ellis, Peter; Ramsay, Jennifer

2014-11-01

The CD200-tolerance signaling molecule prevents pregnancy failure and is also expressed by a wide variety of malignant tumors. The effect of CD200 mRNA expression on progression of human tumors has been variable. A cross-sectional study was performed to examine the correlation between CD200 protein expression in the primary tumors from postoperative Stage I-IIIA human breast cancer and the likelihood of regional lymph node metastasis. Fifty-eight percentage of patients had strong CD200(+) tumor staining (71% of Stage I and 53% Stage II-IIIA). Strong staining was associated with large T2-3 primary tumors compared to T1 tumors (64 versus 50%) and T2-3 N(+) versus T1 N(-) tumors (70 versus 63%), but this was not statistically significant. Nodal metastases were not more frequent in patients with strong CD200(+) staining (57% compared to 58% for weak/negative staining cases), and the metastatic tumor cells in regional lymph nodes were often CD200(-) when the primary tumor was CD200(+). CD200 expression by early-stage human breast cancer cells in primary tumors did not correlate with increased regional lymph node metastasis. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Fluorometric method of quantitative cell mutagenesis

DOEpatents

Dolbeare, Frank A.

1982-01-01

A method for assaying a cell culture for mutagenesis is described. A cell culture is stained first with a histochemical stain, and then a fluorescent stain. Normal cells in the culture are stained by both the histochemical and fluorescent stains, while abnormal cells are stained only by the fluorescent stain. The two stains are chosen so that the histochemical stain absorbs the wavelengths that the fluorescent stain emits. After the counterstained culture is subjected to exciting light, the fluorescence from the abnormal cells is detected.
Fluorometric method of quantitative cell mutagenesis

DOEpatents

Dolbeare, F.A.

1980-12-12

A method for assaying a cell culture for mutagenesis is described. A cell culture is stained first with a histochemical stain, and then a fluorescent stain. Normal cells in the culture are stained by both the histochemical and fluorescent stains, while abnormal cells are stained only by the fluorescent stain. The two stains are chosen so that the histochemical stain absorbs the wavelengths that the fluorescent stain emits. After the counterstained culture is subjected to exciting light, the fluorescence from the abnormal cells is detected.
Methods for validating the presence of and characterizing proteins deposited onto an array

DOEpatents

Schabacker, Daniel S.

2010-09-21

A method of determining if proteins have been transferred from liquid-phase protein fractions to an array comprising staining the array with a total protein stain and imaging the array, optionally comparing the staining with a standard curve generated by staining known amounts of a known protein on the same or a similar array; a method of characterizing proteins transferred from liquid-phase protein fractions to an array including staining the array with a post-translational modification-specific (PTM-specific) stain and imaging the array and, optionally, after staining the array with a PTM-specific stain and imaging the array, washing the array, re-staining the array with a total protein stain, imaging the array, and comparing the imaging with the PTM-specific stain with the imaging with the total protein stain; stained arrays; and images of stained arrays.
Modified Field's staining--a rapid stain for Trichomonas vaginalis.

PubMed

Afzan, M Yusuf; Sivanandam, S; Kumar, G Suresh

2010-10-01

Trichomonas vaginalis, a flagellate protozoan parasite commonly found in the human genitourinary tract, is transmitted primarily by sexual intercourse. Diagnosis is usually by in vitro culture method and staining with Giemsa stain. There are laboratories that use Gram stain as well. We compared the use of modified Field's (MF), Giemsa, and Gram stains on 2 axenic and xenic isolates of T. vaginalis, respectively. Three smears from every sediment of spun cultures of all 4 isolates were stained, respectively, with each of the stains. We showed that MF staining, apart from being a rapid stain (20 s), confers sharper staining contrast, which differentiates the nucleus and the cytoplasm of the organism when compared to Giemsa and Gram staining especially on parasites from spiked urine samples. The alternative staining procedure offers in a diagnostic setting a rapid stain that can easily visualize the parasite with sharp contrasting characteristics between organelles especially the nucleus and cytoplasm. Vacuoles are more clearly visible in parasites stained with MF than when stained with Giemsa. Copyright © 2010 Elsevier Inc. All rights reserved.
Glucose transporter-1 (GLUT-1) immunoreactivity in benign, premalignant and malignant lesions of the gallbladder.

PubMed

Legan, Mateja; Tevžič, Spela; Tolar, Ana; Luzar, Boštjan; Marolt, Vera Ferlan

2011-03-01

GLUT-1 is a transmembrane glucose transport protein that allows the facilitated transport of glucose into cells, normally expressed in tissues which depend mainly on glucose metabolism. Enhanced expression of GLUT-1 can also be found in a large spectrum of carcinomas. This study aimed to investigate GLUT-1 expression in gallbladder tissue: from normal tissue samples, hyperplasias, low-grade and high-grade dysplasias to gallbladder carcinomas. In all, 115 archived samples of gallbladder tissue from 68 patients, presented after cholecystectomy, were immunohistochemically stained for GLUT-1. According to the intensity of GLUT-1 immunoreactivity, samples were divided into negative (stained 0-10% of cells stained), positive with weak to moderate (10-50%) and positive with strong (>50%) GLUT-1 expression. The GLUT-1 immunoreactivity of the samples showed a characteristic increase from premalignant lesions to carcinomas. Normal gallbladder tissue samples did not express GLUT-1 (100%). Weak expression was shown only focally in hyperplasias, but to a greater extent with low-grade dysplasias (20%), high-grade dysplasias (40%) and carcinomas (51.8%). Normal gallbladder tissue is GLUT-1 negative. GLUT-1 expression in carcinoma tissue is significantly higher than in dysplastic lesions. Strong GLUT-1 expression indicates 100% specificity for detecting gallbladder carcinomas. Therefore, GLUT-1 is a candidate as a diagnostic as well as a tissue prognostic marker in gallbladder carcinoma patients.
The lower expression of gonadotropin-releasing hormone receptor associated with poor prognosis in gastric cancer

PubMed Central

Lu, Mingzhu; Zhu, Jing; Ling, Yang; Shi, Wenping; Zhang, Changsong; Wu, Haorong

2015-01-01

Aims: Expression of gonadotropin-releasing hormone receptor (GnRHR) has been demonstrated in a number of malignancies. The aim is to investigate the expression of GnRHR and prognosis in gastric cancer. Methods and materials: GnRHR mRNA was examined in tumor and non-tumor tissues from 48 gastric cancer patients by Real-time PCR. The GnRHR protein expression was performed by immunohistochemical analysis. Results: The expression of GnRHR mRNA was higher (mean ± SD, -10.06 ± 1.28) in gastric tumor tissues than matched non-tumor tissues (mean ± SD, -12.43 ± 1.33). GnRHR mRNA expression was associated with lymph node metastasis, distant metastasis, and TNM stage. We found the decreased expression of GnRHR mRNA were significantly correlated with poor overall survival (P = 0.003). Immunocytochemical staining of GnRHR in tumor tissues showed mainly weak staining (43.48%, 10/23) and moderate staining (21.74%, 5/23) in high GnRHR mRNA patients, and mainly negative staining in low GnRHR mRNA patients. And the staining of GnRHR was not detection in tumor tissues for more than half of gastric patients (52.08%, 25/48). These results implied that the loss of GnRHR protein could be a main event in gastric cancer. Conclusion: The GnRHR expression is very low in gastric cancer, and the loss of GnRHR expression could be a poor prognostic factor, which implied that GnRHR could play an important role in the development of gastric cancer. PMID:26550267
Expression of GLUT-1 glucose transporter in borderline and malignant epithelial tumors of the ovary.

PubMed

Cantuaria, G; Magalhaes, A; Penalver, M; Angioli, R; Braunschweiger, P; Gomez-Marin, O; Kanhoush, R; Gomez-Fernandez, C; Nadji, M

2000-10-01

Cancer cells have increased rates of glucose metabolism when compared to normal cells. One of the mechanisms proposed for the accelerated glucose use in malignant cells is the overexpression of glucose transporters. In this study we evaluated the expression of the GLUT-1 glucose transporter in borderline and malignant epithelial neoplasms of the ovary. Histologic sections of tumor tissues from 21 borderline and 82 malignant epithelial neoplasms of the ovary were stained for GLUT-1 using polyclonal GLUT-1 antibody (Dako, Carpinteria, CA) and the labeled streptavidin biotin procedure. DAB was used as chromagen and tissues were counterstained with hematoxylin. Normal ovarian surface epithelial cells were either negative or weakly positive. Of the 82 carcinomas, 81 (98.8%) were positive for GLUT-1. The staining intensity was significantly associated with the grade of tumor (P = 0.001). Of the 21 borderline neoplasms, 20 (95.2%) were positive for GLUT-1. Carcinomas had a significantly stronger stain than borderline tumors (P = 0.0001). The intensity of the stain was also stronger in serous carcinomas compared to other subtypes (P = 0. 0001). Positive cells demonstrated a cytoplasmic membrane staining that was more intense in tumor cells farther away from blood supply. Overexpression of the GLUT-1 transporter is associated with the histology and grade of the tumors. Our findings show a progressive increase in the expression of the GLUT-1 transporter from the borderline tumor to the high-grade carcinomas. These data suggest that the expression of this transporter may be closely related to the malignant transformation of epithelial ovarian tumors. Copyright 2000 Academic Press.
A useful single-solution polychrome stain for plant material...Brook Cyte-Chrome I.

Treesearch

Stanley L Krugman; Julia F. Littlefield

1968-01-01

Fresh and chemically fixed sectioned plant material can be quickly stained by applying a Brook Cyte Chrome I polychrome stain. Staining time averaged only about 10 minutes. And exact timing of staining and de-staining is not as critical as with most of the commonly used stains. The overall quality is comparable to that of the traditional stains.
Nonclassical MHC-E (Mamu-E) expression in the rhesus monkey placenta

PubMed Central

Dambaeva, Svetlana V.; Bondarenko, Gennadiy I.; Grendell, Richard L.; Kravitz, Rachel H.; Durning, Maureen; Golos, Thaddeus G.

2009-01-01

The aim of this study was to characterize the expression of the rhesus HLA-E ortholog Mamu-E, particularly at the maternal-fetal interface. Mamu-E expression was confirmed by locus-specific RT-PCR in the placenta as well as in peripheral blood mononuclear cells (PBMC) and other organs. We evaluated the utility of antibodies recognizing HLA-E (MEM-E/06 against native HLA-E, MEM-E/02 against denatured HLA-E) to detect Mamu-E by flow cytometry/immunofluorescence, Western blot, and immunohistochemistry (IHC). Western blot analysis of cells and selected transfectants confirmed the recognition of Mamu-E but not Mamu-AG by antibodies MEM-E/06 and HC10 but not MEM-E/02. Immunohistochemical staining of frozen sections of rhesus placenta with the MEM-E/06 antibody demonstrated expression in most populations of rhesus monkey trophoblast cells, including villous cytotrophoblasts (strong positive staining), apical membrane of syncytiotrophoblasts (light to moderate staining) and extravillous cytotrophoblasts (moderate to strong staining, especially endovascular trophoblasts in early pregnancy). Expression was not trophoblast cell-specific, especially at term, when endothelial cells in both the chorionic plate and placental villi showed strong staining for Mamu-E. Staining of rhesus extravillous trophoblast cells suggested the co-expression of Mamu-E and Mamu-AG (the rhesus HLA-G homolog) on these cells. MEM-E/06 was shown also to react with differentiating rhesus placental syncytiotrophoblasts in primary culture, detecting intracellular and weak surface expression of Mamu-E. We conclude that the gestation-dependent co-expression of Mamu-E with Mamu-AG in villous and extravillous trophoblast cells suggests important and perhaps complementary but distinct roles of these two nonclassical MHC class I loci in pregnancy at the maternal-fetal interface. In addition, the MEM-E/06 antibody will be useful for the detection of Mamu-E at the maternal-fetal interface in the rhesus monkey. PMID:17996936
Asporin stably expressed in the surface layer of mandibular condylar cartilage and augmented in the deeper layer with age.

PubMed

Miyamoto, Yutaka; Kanzaki, Hiroyuki; Wada, Satoshi; Tsuruoka, Sari; Itohiya, Kanako; Kumagai, Kenichi; Hamada, Yoshiki; Nakamura, Yoshiki

2017-12-01

Mandibular condylar cartilage (MCC) exhibits dual roles both articular cartilage and growth center. Of many growth factors, TGF-β has been implicated in the growth of articular cartilage including MCC. Recently, Asporin, decoy to TGF-β, was discovered and it blocks TGF-β signaling. Asporin is expressed in a variety of tissues including osteoarthritic articular cartilage, though there was no report of Asporin expression in MCC. In the present study, we investigated the temporal and spatial expression of Asporin in MCC. Gene expression profile of MCC and epiphyseal cartilage in tibia of 5 weeks old ICR mice were firstly compared with microarray analysis using the laser capture microdissected samples. Variance of gene expression was further confirmed by real-time RT-PCR and immunohistochemical staining at 1,3,10, and 20 weeks old. TGF-β and its signaling molecule, phosphorylated Smad-2/3 (p-Smad2/3), were also examined by immunohistochemical staining. Microarray analysis revealed that Asporin was highly expressed in MCC. Real-time RT-PCR analysis confirmed that the fibrous layer of MCC exhibited stable higher Asporin expression at any time points as compared to epiphyseal cartilage. This was also observed in immunohistochemical staining. Deeper layer in MCC augmented Asporin expression with age. Whereas, TGF-β was stably highly observed in the layer. The fibrous layer of MCC exhibited weak staining of p-Smad2/3, though the proliferating layer of MCC was strongly stained as compared to epiphyseal cartilage of tibia at early time point. Consistent with the increase of Asporin expression in the deeper layer of MCC, the intensity of p-Smad-2/3 staining was decreased with age. In conclusion, we discovered that Asporin was stably expressed at the fibrous layer of MCC, which makes it possible to manage both articular cartilage and growth center at the same time.
Differential contribution of Kv4-containing channels to A-type, voltage-gated potassium currents in somatic and visceral dorsal root ganglion neurons.

PubMed

Yunoki, Takakazu; Takimoto, Koichi; Kita, Kaori; Funahashi, Yasuhito; Takahashi, Ryosuke; Matsuyoshi, Hiroko; Naito, Seiji; Yoshimura, Naoki

2014-11-15

Little is known about electrophysiological differences of A-type transient K(+) (KA) currents in nociceptive afferent neurons that innervate somatic and visceral tissues. Staining with isolectin B4 (IB4)-FITC classifies L6-S1 dorsal root ganglion (DRG) neurons into three populations with distinct staining intensities: negative to weak, moderate, and intense fluorescence signals. All IB4 intensely stained cells are negative for a fluorescent dye, Fast Blue (FB), injected into the bladder wall, whereas a fraction of somatic neurons labeled by FB, injected to the external urethral dermis, is intensely stained with IB4. In whole-cell, patch-clamp recordings, phrixotoxin 2 (PaTx2), a voltage-gated K(+) (Kv)4 channel blocker, exhibits voltage-independent inhibition of the KA current in IB4 intensely stained cells but not the one in bladder-innervating cells. The toxin also shows voltage-independent inhibition of heterologously expressed Kv4.1 current, whereas its inhibition of Kv4.2 and Kv4.3 currents is voltage dependent. The swapping of four amino acids at the carboxyl portion of the S3 region between Kv4.1 and Kv4.2 transfers this characteristic. RT-PCRs detected Kv4.1 and the long isoform of Kv4.3 mRNAs without significant Kv4.2 mRNA in L6-S1 DRGs. Kv4.1 and Kv4.3 mRNA levels were higher in laser-captured, IB4-stained neurons than in bladder afferent neurons. These results indicate that PaTx2 acts differently on channels in the Kv4 family and that Kv4.1 and possibly Kv4.3 subunits functionally participate in the formation of KA channels in a subpopulation of somatic C-fiber neurons but not in visceral C-fiber neurons innervating the bladder. Copyright © 2014 the American Physiological Society.
Differential contribution of Kv4-containing channels to A-type, voltage-gated potassium currents in somatic and visceral dorsal root ganglion neurons

PubMed Central

Yunoki, Takakazu; Takimoto, Koichi; Kita, Kaori; Funahashi, Yasuhito; Takahashi, Ryosuke; Matsuyoshi, Hiroko; Naito, Seiji

2014-01-01

Little is known about electrophysiological differences of A-type transient K+ (KA) currents in nociceptive afferent neurons that innervate somatic and visceral tissues. Staining with isolectin B4 (IB4)-FITC classifies L6-S1 dorsal root ganglion (DRG) neurons into three populations with distinct staining intensities: negative to weak, moderate, and intense fluorescence signals. All IB4 intensely stained cells are negative for a fluorescent dye, Fast Blue (FB), injected into the bladder wall, whereas a fraction of somatic neurons labeled by FB, injected to the external urethral dermis, is intensely stained with IB4. In whole-cell, patch-clamp recordings, phrixotoxin 2 (PaTx2), a voltage-gated K+ (Kv)4 channel blocker, exhibits voltage-independent inhibition of the KA current in IB4 intensely stained cells but not the one in bladder-innervating cells. The toxin also shows voltage-independent inhibition of heterologously expressed Kv4.1 current, whereas its inhibition of Kv4.2 and Kv4.3 currents is voltage dependent. The swapping of four amino acids at the carboxyl portion of the S3 region between Kv4.1 and Kv4.2 transfers this characteristic. RT-PCRs detected Kv4.1 and the long isoform of Kv4.3 mRNAs without significant Kv4.2 mRNA in L6-S1 DRGs. Kv4.1 and Kv4.3 mRNA levels were higher in laser-captured, IB4-stained neurons than in bladder afferent neurons. These results indicate that PaTx2 acts differently on channels in the Kv4 family and that Kv4.1 and possibly Kv4.3 subunits functionally participate in the formation of KA channels in a subpopulation of somatic C-fiber neurons but not in visceral C-fiber neurons innervating the bladder. PMID:25143545
Immunohistochemichal Assessment of the CrkII Proto-oncogene Expression in Common Malignant Salivary Gland Tumors and Pleomorphic Adenoma.

PubMed

Askari, Mitra; Darabi, Masoud; Jahanzad, Esa; Mostakhdemian Hosseini, Zahra; Musavi Chavoshi, Marjan; Darabi, Maryam

2015-01-01

Background and aims. Various morphologies are seen in different salivary gland tumorsor within an individual tumor, and the lesions show divers biological behaviors. Experimental results support the hypothesis that increased CrkII proto-oncogene is associated with cytokine-induced tumor initiation and progression by altering cell motility signaling pathway. The aim of this study was to assess the CrkII expression in common malignant salivary gland tumors and pleomorphic ade-noma. Materials and methods. Immunohistochemical analysis of CrkII expression was performed on paraffin blocks of 64 car-cinomas of salivary glands, 10 pleomorphic adenomas, and 10 normal salivary glands. Biopsies were subjected to immu-nostaining with EnVision detection system using monoclonal anti-CrkII. Evaluation of immunoreactivity of CrkII was based on the immunoreaction intensity and percentage of stained tumor cells which were scored semi-quantitatively on a scale with four grades 0 to 3. Kruskal-wallis test and additional Mann-Whitney statistical test were used for analysis of CrkII expression levels. Results. Increased expression of CrkII was seen (P=0.005) in malignant tumors including: mucoepidermoid carcinoma, adenoid cystic carcinoma, and carcinoma ex pleomorphic adenoma, but CrkII expression in acinic cell carcinoma was weak. CrkII expression in pleomorphic adenoma was weak or negative. A weak staining was sparsely seen in normal acinar serous cell. Conclusion. Increased expression of CrkII and its higher intensity of staining in tumors with more aggressive biologic behavior in carcinomas of salivary gland is consistent with a role for this proto-oncogene in salivary gland tumorigenesis and cancer progression.
Crystal violet stain as a selective stain for the assessment of mitotic figures in oral epithelial dysplasia and oral squamous cell carcinoma.

PubMed

Jadhav, Kiran B; Ahmed Mujib, B R; Gupta, Nidhi

2012-01-01

Assessment of mitotic figures (MFs) is routinely practiced as prognostic indicator in oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC), but identification of MFs poses a problem in terms of staining characteristics. To evaluate effectiveness of crystal violet stain for staining of MFs and its comparison with hematoxylin and eosin (H and E) stain. Study sample includes archival tissues embedded in paraffin blocks diagnosed as OED (n = 30) and OSCC (n = 30). The control group comprised of tissue specimen from oral mucosa of healthy volunteers (n = 30). Two serial sections of each tissue specimen were stained separately with H and E stain and 1% crystal violet stain. The stained sections were observed under microscope for identification and counting of MFs. Data obtained was statistically analyzed by using the Man-Whitney U test. A significant increase in number of MFs was observed in OED and OSCC in comparison with normal oral mucosa. There was a highly significant increase in number of MFs in crystal violet stained tissue sections when compared with H and E stain. Metaphase is the most commonly observed phase of mitosis in crystal violet stain when compared with H and E stain for all three groups. Crystal violet stain can be considered as selective stain for mitotic figures.
Brilliant Blue G double staining enhances successful internal limiting membrane peeling with minimal adverse effect by low cellular permeability into live cells.

PubMed

Hisatomi, Toshio; Notomi, Shoji; Tachibana, Takashi; Oishi, Seiichiro; Asato, Ryo; Yamashita, Takehiro; Murakami, Yusuke; Ikeda, Yasuhiro; Enaida, Hiroshi; Sakamoto, Taiji; Ishibashi, Tatsuro

2015-02-01

Brilliant Blue G is used as a surgical adjuvant for retinal surgery. Although BBG double or multiple staining was reported, the effectiveness and safety of repeated staining is still elusive. To further examine the effectiveness and safety, we examined BBG in clinical cases in vivo, primary cell culture in vitro, and surgically resected specimen ex vivo. A retrospective interventional case series with in vitro and ex vivo studies were performed. Vitrectomy was performed in 28 cases of epiretinal membrane with BBG single to multiple staining. The surgically resected membranes were stained by BBG with or without cellular fixation. Primary cell cultures were examined with BBG and live/death cell markers, such as Calcein AM and TUNEL. Single staining provided satisfactory staining in seven cases. Double or multiple staining substantially visualized internal limiting membrane (21 cases), especially the edges of remaining internal limiting membrane (11 cases). Adverse retinal staining was not noted and the final visual acuity showed no difference with multiple staining. The live cells barely stained with BBG, while some dead cells were stained. Brilliant Blue G multiple staining substantially enhanced the visualization of internal limiting membrane. The absence of abnormal staining supports the safety of repeated BBG staining.
HMB-45, S-100, NK1/C3, and MART-1 in metastatic melanoma.

PubMed

Zubovits, Judit; Buzney, Elizabeth; Yu, Lawrence; Duncan, Lyn M

2004-02-01

The diagnosis of melanoma metastatic to lymph node remains a difficult problem given its histological diversity. We examined the staining patterns of S-100, NK1/C3, HMB-45, and MART-1 (DC10) in melanoma metastases to lymph nodes. Immunohistochemical stains were performed on tissue sections of 126 formalin-fixed lymph nodes from 126 patients with an established diagnosis of metastatic melanoma. A total of 98% of cases (123 of 126) stained positive for S-100, 93% (117 of 125) stained positive for NK1/C3, 82% (103 of 126) stained positive for MART-1, and 76% (95 of 125) stained positive for HMB-45. The distribution and intensity of staining varied among these markers. A diffuse staining pattern, defined as >50% of tumor cells stained, was observed in 83% of MART-1-positive cases but in only 56% of S-100-positive cases, 48% of NK1/C3-positive cases, and 34% of HMB-45-positive cases. A maximally intense signal was almost always observed for MART-1 (83% of positive cases) but was rarely observed for NK1/C3 (20%). S-100 and HMB-45 showed maximally intense staining in 50% and 54% of cases, respectively. S-100 and NK1/C3 stained both histiocytes and melanocytes, whereas MART-1 and HMB-45 stained only melanocytes. Seventy-eight cases (63%) stained positive for all 4 markers, 17 cases (14%) stained for all markers except HMB-45, 13 cases (10%) stained for all markers except MART-1, 6 cases (5%) stained only with S-100 and NK1/C3, 4 cases (3%) stained only with S-100 and HMB-45, and 2 cases stained for all markers except S-100. One case each stained for the following: only S-100, only S-100 and HMB-45, and all markers except NK1/C3. One case exhibited absence of staining for any of these markers. We demonstrate that lymph node metastases of melanoma are heterogeneous with regard to tumor marker expression. S-100 and NK1/C3 were the most sensitive stains for detecting metastatic melanoma; however, they both also stain other nontumor cells in lymph nodes. MART-1 did not stain histiocytes and exhibited a more frequently intense and diffuse staining pattern than NK1/C3. HMB-45 was less sensitive and demonstrated less diffuse staining than MART-1.
Differential distribution of annexins-I, -II, -IV, and -VI in synovium.

PubMed Central

Goulding, N J; Dixey, J; Morand, E F; Dodds, R A; Wilkinson, L S; Pitsillides, A A; Edwards, J C

1995-01-01

OBJECTIVES--To examine the distribution of four annexins in non-inflamed rheumatoid arthritic and osteoarthritic synovial tissue. METHODS--Frozen sections were stained with monoclonal antibodies (MAb) specific for annexins-I, -II, -IV, and -VI, and for cell lineage related markers including CD68 and CD14 (macrophages), prolyl hydroxylase (fibroblasts), and CD3 (T cells). RESULTS--Each of the annexins was present in synovial tissues in significant amounts in the three groups studied. Annexin-I was predominantly found within the synovial lining layer and double labelling showed it to be present predominantly in cells of the macrophage lineage. In rheumatoid specimens there was increased staining within the lining layer, perivascularly and on macrophages within the tissue stroma. Annexin-II was present in a distribution similar to that of annexin-I, but with more prominent perivascular staining. Annexins-IV and -VI were seen chiefly in association with areas of lymphocyte infiltration in rheumatoid tissue, whereas annexins-I and -II were absent from these areas. Endothelial cells stained weakly positive for annexins-I and -II, and more strongly for -IV and -VI. CONCLUSIONS--This study demonstrates that annexins (particularly annexin-I, a putative mediator of the anti-inflammatory activities of glucocorticoids) are abundant in rheumatoid and non-rheumatoid synovial tissue, annexins-IV and -VI having a distribution distinct from that of -I and -II. Images PMID:7492225
An epitope localized in c-Src negative regulatory domain is a potential marker in early stage of colonic neoplasms.

PubMed

Sakai, T; Kawakatsu, H; Fujita, M; Yano, J; Owada, M K

1998-02-01

In previous work, we established a new monoclonal antibody that specifically recognizes the active form of c-Src tyrosine kinase (Kawakatsu et al, 1996). To determine whether c-Src is active in colorectal tumorigenesis, we examined the expression of an active form of c-Src in human normal mucosa, hyperplastic polyps, adenomas, and adenocarcinomas. The tissue distribution of the active form of c-Src was studied by immunohistochemistry using this antibody, termed Clone 28. Among 66 cases of adenoma tested, 61 (92%) showed positive staining (adenoma with mild atypia, 3 of 3; adenoma with moderate atypia, 38 of 42; adenoma with severe atypia, 20 of 21). In contrast to the frequent and intense staining in adenomas, adenocarcinoma showed weak staining with less frequency in 4 of 16 (25%) cases. The number of specimens with positive staining in well- and moderately differentiated adenocarcinomas was limited to an early stage. The active form of c-Src mainly localized to the nuclear membrane and the perinuclear region. These results provide the first direct evidence that the activation of c-Src appears to be an early event in colonic carcinogenesis in situ. The findings of the present study thus allow us to propose a molecular mechanism involving c-Src activation in the process of malignant transformation of the human colonic neoplastic cells.
Leptospirosis pulmonary haemorrhage syndrome is associated with linear deposition of immunoglobulin and complement on the alveolar surface.

PubMed

Croda, J; Neto, A N D; Brasil, R A; Pagliari, C; Nicodemo, A C; Duarte, M I S

2010-06-01

Leptospirosis is a zoonotic infection associated with severe diseases such as leptospirosis pulmonary haemorrhage syndrome (LPHS). The cause of pulmonary haemorrhage is unclear. Understanding which mechanisms and processes are involved in LPHS will be important in treatment regimens under development for this life-threatening syndrome. In the present study, we evaluated 30 lung specimens from LPHS patients and seven controls using histology and immunohistochemistry (detection of IgM, IgG, IgA and C3) in order to describe the pathological features associated with this syndrome. Immunoglobulin deposits were detected on the alveolar surface in 18/30 LPHS patients. Three staining patterns were observed for the immunoglobulins and C3 in the lung tissues of LPHS patients: AS, delicate linear staining adjacent to the alveolar surface, which was indicative of a membrane covering the luminal surface of type I and II pneumocyte cells; S, heterogeneous staining which was sporadically distributed along the alveolar septum; and IA, weak, focal intra-alveolar granular staining. Human LPHS is associated with individual and unique histological patterns that differ from those of other causes of pulmonary haemorrhage. In the present study, it was found that the linear deposition of immunoglobulins (IgA, IgG and IgM) and complement on the alveolar surface may play a role in the pathogenesis of pulmonary haemorrhage in human leptospirosis.

Exit Site Infection due to Mycobacterium chelonae in an Elderly Patient on Peritoneal Dialysis.

PubMed

Hibi, Arata; Kasugai, Takahisa; Kamiya, Keisuke; Ito, Chiharu; Kominato, Satoru; Miura, Toshiyuki; Koyama, Katsushi

2018-01-01

Nontuberculous mycobacteria (NTM) are rarely isolated from peritoneal dialysis (PD)-associated catheter infections. However, NTM infection is usually difficult to treat and leads to catheter loss. Prompt diagnosis is essential for appropriate treatment. A 70-year-old Japanese man who had been on PD for 2 years and with a medical history of 2 episodes of exit site infections (ESIs) due to methicillin-resistant Staphylococcus aureus was admitted to the hospital due to suspected ESI recurrence. However, Gram staining of the pus revealed no gram-positive cocci. Instead, weakly stained gram-positive rods were observed after 7 days of incubation, which were also positive for acid-fast staining. Rapidly growing NTM Mycobacterium chelonae was isolated on day 14. Despite administering a combination antibiotic therapy, ESI could not be controlled, and catheter removal surgery was performed on day 21. Although PD was discontinued temporarily, the patient did not require hemodialysis, without any uremic symptoms. The catheter was reinserted on day 48, and PD was reinitiated on day 61. The patient was discharged on day 65. Antibiotic therapy was continued for 3 months after discharge, with no indications of recurrent infections observed. It is important to consider the risk of NTM infections in patients on PD. Acid-fast staining could be a key test for prompt diagnosis and provision of an appropriate treatment.
Immunophenotyping by slide-based cytometry and by flow cytometry are comparable

NASA Astrophysics Data System (ADS)

Gerstner, Andreas O.; Laffers, Wiebke; Mittag, Anja; Daehnert, Ingo; Lenz, Domnik; Bootz, Friedrich; Bocsi, Jozsef; Tarnok, Attila

2005-03-01

Immunophenotyping of peripheral blood leukocytes (PBLs) is performed by flow cytometry (FCM) as the golden standard. Slide based cytometry systems for example laser scanning cytometer (LSC) can give additional information (repeated staining and scanning, morphology). In order to adequately judge on the clinical usefulness of immunophenotyping by LSC it is obligatory to compare it with the long established FCM assays. We performed this study to systematically compare the two methods, FCM and LSC for immunophenotyping and to test the correlation of the results. Leucocytes were stained with directly labeled monoclonal antibodies with whole blood staining method. Aliquots of the same paraformaldehyde fixed specimens were analyzed in a FACScan (BD-Biosciences) using standard protocols and parallel with LSC (CompuCyte) after placing to glass slide, drying and fixation by aceton and 7-AAD staining. Calculating the percentage distribution of PBLs obtained by LSC and by FCM shows very good correlation with regression coefficients close to 1.0 for the major populations (neutrophils, lymphocytes, and monocytes), as well as for the lymphocyte sub-populations (T-helper-, T-cytotoxic-, B-, NK-cells). LSC can be recommended for immunophenotyping of PBLs especially in cases where only very limited sample volumes are available or where additional analysis of the cells" morphology is important. There are limitations in the detection of rare leucocytes or weak antigens where appropriate amplification steps for immunofluorescence should be engaged.
MyD88 and TLR4 Expression in Epithelial Ovarian Cancer

PubMed Central

Block, Matthew S.; Vierkant, Robert A.; Rambau, Peter F.; Winham, Stacey J.; Wagner, Philipp; Traficante, Nadia; Tołoczko, Aleksandra; Tiezzi, Daniel G.; Taran, Florin Andrei; Sinn, Peter; Sieh, Weiva; Sharma, Raghwa; Rothstein, Joseph H.; Cajal, Teresa Ramón y; Paz-Ares, Luis; Oszurek, Oleg; Orsulic, Sandra; Ness, Roberta B.; Nelson, Gregg; Modugno, Francesmary; Menkiszak, Janusz; McGuire, Valerie; McCauley, Bryan M.; Mack, Marie; Lubiński, Jan; Longacre, Teri A.; Li, Zheng; Lester, Jenny; Kennedy, Catherine J.; Kalli, Kimberly R.; Jung, Audrey Y.; Johnatty, Sharon E.; Jimenez-Linan, Mercedes; Jensen, Allan; Intermaggio, Maria P.; Hung, Jillian; Herpel, Esther; Hernandez, Brenda Y.; Hartkopf, Andreas D.; Harnett, Paul R.; Ghatage, Prafull; García-Bueno, José M.; Gao, Bo; Fereday, Sian; Eilber, Ursula; Edwards, Robert P.; de Sousa, Christiani B.; de Andrade, Jurandyr M.; Chudecka-Głaz, Anita; Chenevix-Trench, Georgia; Cazorla, Alicia; Brucker, Sara Y.; Alsop, Jennifer; Whittemore, Alice S.; Steed, Helen; Staebler, Annette; Moysich, Kirsten B.; Menon, Usha; Koziak, Jennifer M.; Kommoss, Stefan; Kjaer, Susanne K.; Kelemen, Linda E.; Karlan, Beth Y.; Huntsman, David G.; Høgdall, Estrid; Gronwald, Jacek; Goodman, Marc T.; Gilks, Blake; García, María José; Fasching, Peter A.; de Fazio, Anna; Deen, Suha; Chang-Claude, Jenny; Candido dos Reis, Francisco J.; Campbell, Ian G.; Brenton, James D.; Bowtell, David D.; Benítez, Javier; Pharoah, Paul D.P.; Köbel, Martin; Ramus, Susan J.; Goode, Ellen L.

2018-01-01

Objective To evaluate myeloid differentiation primary response gene 88 (MyD88) and Toll-like receptor 4 (TLR4) expression in relation to clinical features of epithelial ovarian cancer, histologic subtypes, and overall survival. Patients and Methods We conducted centralized immunohistochemical staining, semi-quantitative scoring, and survival analysis in 5263 patients participating in the Ovarian Tumor Tissue Analysis consortium. Patients were diagnosed between January 1, 1978, and December 31, 2014, including 2865 high-grade serous ovarian carcinomas (HGSOCs), with more than 12,000 person-years of follow-up time. Tissue microarrays were stained for MyD88 and TLR4, and staining intensity was classified using a 2-tiered system for each marker (weak vs strong). Results Expression of MyD88 and TLR4 was similar in all histotypes except clear cell ovarian cancer, which showed reduced expression compared with other histotypes (P<.001 for both). In HGSOC, strong MyD88 expression was modestly associated with shortened overall survival (hazard ratio [HR], 1.13; 95% CI, 1.01–1.26; P=.04) but was also associated with advanced stage (P<.001). The expression of TLR4 was not associated with survival. In low-grade serous ovarian cancer (LGSOC), strong expression of both MyD88 and TLR4 was associated with favorable survival (HR [95% CI], 0.49 [0.29–0.84] and 0.44 [0.21–0.89], respectively; P=.009 and P=.02, respectively). Conclusion Results are consistent with an association between strong MyD88 staining and advanced stage and poorer survival in HGSOC and demonstrate correlation between strong MyD88 and TLR4 staining and improved survival in LGSOC, highlighting the biological differences between the 2 serous histotypes. PMID:29502561
An investigation of the feasibility of applying Raman microscopy for exploring stained glass

NASA Astrophysics Data System (ADS)

Bouchard, Michel; Smith, David C.; Carabatos-Nédelec, Constantin

2007-12-01

Raman microscopy (RM) is widely used in archaeometrical studies of pigments, geomaterials and biomaterials in the Cultural Heritage, but one domain has received relatively less attention: the colouring of stained glass. This feasibility study investigates the advantages and disadvantages of employing RM alone in this field by means of a study of modern commercial glasses, modern commercial pigments, and a few archaeological stained glasses, but especially by an experimental project whereby the authors created stained glass. The different kinds of possible unreacted or reacted material are rigorously established. The distinction between Na, K, Ca glasses was explored, as well as the red colouring of an industrial glass which was proved to be due to the presence of (Zn, Cd)S xSe 1- x. Yellow, green, blue and maroon pigments were studied before and after an initial firing and then after heating on glass. The quality of the Raman spectra varied enormously and was sometimes disappointing. Nevertheless RM successfully identified various coloured products such as bindheimite, crocoite, cobalt aluminate, haematite; relict reactants such as corundum, eskolaite and oxides of Co or Pb; and provided indications of other phases such as maghemite or Co-olivine. One conclusion is that the amount of chemical reaction between the pigments and the glass is small compared to the amount in between the pigments. Comments are made on the potential for dating archaeological glass from the known age of synthesis of the pigments, and of the dangers of this approach. Overall it has been shown that RM can be useful for studying stained glass, especially for remote in situ analytical operations with mobile RM, but one must expect some problems either with fluorescence or weak spectra.
Significant expression of thyroid transcription factor-1 in pulmonary squamous cell carcinoma detected by SPT24 monoclonal antibody and CSA-II system.

PubMed

Kashima, Kenji; Hashimoto, Hisashi; Nishida, Haruto; Arakane, Motoki; Yada, Naomi; Daa, Tsutomu; Yokoyama, Shigeo

2014-01-01

In contrast to the usefulness of thyroid transcription factor-1 (TTF-1) in distinguishing primary adenocarcinoma of the lung from metastatic lesions, TTF-1 expression in pulmonary squamous cell carcinoma is reported to be at low level and not a suitable immunohistochemical marker. We hypothesized that the highly sensitive detection system, CSA-II, can visualize even faint expression of TTF-1 in pulmonary squamous cell carcinoma. In this study, 2 commercially available clones of TTF-1 monoclonal antibody, 8G7G3/1 and SPT24, were used for staining 38 cases of pulmonary squamous cell carcinoma, in combination with the CSA-II and the conventional detection system, EnVision. The combined use of the 8G7G3/1 clone with EnVision and CSA-II showed a positive reaction in only 1 and 4 cases, respectively. The use of SPT24 clone showed positive staining in 5 cases with EnVision and in 20 of 38 cases (52.6%) with the CSA-II. Interestingly, positive staining by the SPT24-CSA-II technique of samples from tissue blocks preserved for <2 years was 73.6% compared with only 31.5% in those preserved for >2 years. In addition, a 6-month preservation of the cut sections resulted in stain fading and decreased positivity (50%), compared with freshly cut sections. We conclude that the use of the SPT24 monoclonal antibody with the CSA-II system can detect even weak expression of TTF-1 in pulmonary squamous cell carcinoma. This staining technique can potentially allow the discrimination of primary squamous cell carcinoma of the lung from metastatic lesions, especially in freshly prepared paraffin sections.
MyD88 and TLR4 Expression in Epithelial Ovarian Cancer.

PubMed

Block, Matthew S; Vierkant, Robert A; Rambau, Peter F; Winham, Stacey J; Wagner, Philipp; Traficante, Nadia; Tołoczko, Aleksandra; Tiezzi, Daniel G; Taran, Florin Andrei; Sinn, Peter; Sieh, Weiva; Sharma, Raghwa; Rothstein, Joseph H; Ramón Y Cajal, Teresa; Paz-Ares, Luis; Oszurek, Oleg; Orsulic, Sandra; Ness, Roberta B; Nelson, Gregg; Modugno, Francesmary; Menkiszak, Janusz; McGuire, Valerie; McCauley, Bryan M; Mack, Marie; Lubiński, Jan; Longacre, Teri A; Li, Zheng; Lester, Jenny; Kennedy, Catherine J; Kalli, Kimberly R; Jung, Audrey Y; Johnatty, Sharon E; Jimenez-Linan, Mercedes; Jensen, Allan; Intermaggio, Maria P; Hung, Jillian; Herpel, Esther; Hernandez, Brenda Y; Hartkopf, Andreas D; Harnett, Paul R; Ghatage, Prafull; García-Bueno, José M; Gao, Bo; Fereday, Sian; Eilber, Ursula; Edwards, Robert P; de Sousa, Christiani B; de Andrade, Jurandyr M; Chudecka-Głaz, Anita; Chenevix-Trench, Georgia; Cazorla, Alicia; Brucker, Sara Y; Alsop, Jennifer; Whittemore, Alice S; Steed, Helen; Staebler, Annette; Moysich, Kirsten B; Menon, Usha; Koziak, Jennifer M; Kommoss, Stefan; Kjaer, Susanne K; Kelemen, Linda E; Karlan, Beth Y; Huntsman, David G; Høgdall, Estrid; Gronwald, Jacek; Goodman, Marc T; Gilks, Blake; García, María José; Fasching, Peter A; de Fazio, Anna; Deen, Suha; Chang-Claude, Jenny; Candido Dos Reis, Francisco J; Campbell, Ian G; Brenton, James D; Bowtell, David D; Benítez, Javier; Pharoah, Paul D P; Köbel, Martin; Ramus, Susan J; Goode, Ellen L

2018-03-01

To evaluate myeloid differentiation primary response gene 88 (MyD88) and Toll-like receptor 4 (TLR4) expression in relation to clinical features of epithelial ovarian cancer, histologic subtypes, and overall survival. We conducted centralized immunohistochemical staining, semi-quantitative scoring, and survival analysis in 5263 patients participating in the Ovarian Tumor Tissue Analysis consortium. Patients were diagnosed between January 1, 1978, and December 31, 2014, including 2865 high-grade serous ovarian carcinomas (HGSOCs), with more than 12,000 person-years of follow-up time. Tissue microarrays were stained for MyD88 and TLR4, and staining intensity was classified using a 2-tiered system for each marker (weak vs strong). Expression of MyD88 and TLR4 was similar in all histotypes except clear cell ovarian cancer, which showed reduced expression compared with other histotypes (P<.001 for both). In HGSOC, strong MyD88 expression was modestly associated with shortened overall survival (hazard ratio [HR], 1.13; 95% CI, 1.01-1.26; P=.04) but was also associated with advanced stage (P<.001). The expression of TLR4 was not associated with survival. In low-grade serous ovarian cancer (LGSOC), strong expression of both MyD88 and TLR4 was associated with favorable survival (HR [95% CI], 0.49 [0.29-0.84] and 0.44 [0.21-0.89], respectively; P=.009 and P=.02, respectively). Results are consistent with an association between strong MyD88 staining and advanced stage and poorer survival in HGSOC and demonstrate correlation between strong MyD88 and TLR4 staining and improved survival in LGSOC, highlighting the biological differences between the 2 serous histotypes. Copyright © 2017 Mayo Foundation for Medical Education and Research. Published by Elsevier Inc. All rights reserved.
Immunohistochemical expression of Notch signaling in the lining epithelium of periapical cysts.

PubMed

Meliou, Eleni; Kerezoudis, Nikolaos; Tosios, Konstantinos; Lafkas, Daniel; Kiaris, Hippokratis

2011-02-01

In this study we evaluated the immunohistochemical expression of the receptors Notch 1 and Notch 2, the ligand Delta 1, and the transcription factors HES 1 and HES 5 in the epithelium of well-defined periapical cysts. Immunohistochemistry was carried out on 55 formalin-fixed and paraffin-embedded, well-defined periapical cysts with minimum inflammation, obtained from the archival tissue database of the Department of Oral Pathology and Surgery. Western blotting was performed to evaluate the specificity of the anti-Notch antibody and the expression of Notch signaling in 5 fresh-frozen periapical cysts. The levels of staining intensity were estimated by the performance of a semiautomated image analysis system. Descriptive statistic of mean values obtained by computerized image analysis method was performed. Immunostaining reaction of all Notch signaling components was observed in the cytoplasm and/or the cytoplasmic membrane in the majority of epithelial cells of periapical cysts. Nuclear staining was observed occasionally in all cases. Notch 2 showed strong staining in 52.83% of the cases, followed by Notch 1 (35.85%), HES 1 and HES 5 moderate staining in 72.73% and 57.69% of the cases, respectively, and Delta 1 weak staining in 58.33% of the cases. No statistical correlation was found between the antibodies and the sex or the age of the study group. Notch is an evolutionarily conserved signaling mechanism that regulates cell fate decisions during development and postnatal life in organisms as diverse as worms, flies, and humans. The present observations indicate that Notch pathway is active downstream in the lining epithelium of periapical cysts, suggesting an involvement of this pathway in periapical cyst growth and expansion. Copyright Â© 2011 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.
Expression of the gastrin-releasing peptide receptor, the prostate stem cell antigen and the prostate-specific membrane antigen in lymph node and bone metastases of prostate cancer.

PubMed

Ananias, Hildo J K; van den Heuvel, Marius C; Helfrich, Wijnand; de Jong, Igle J

2009-07-01

Cell membrane antigens like the gastrin-releasing peptide receptor (GRPR), the prostate stem cell antigen (PSCA), and the prostate-specific membrane antigen (PSMA), expressed in prostate cancer, are attractive targets for new therapeutic and diagnostic applications. Therefore, we investigated in this study whether these antigens are expressed in metastasized prostate cancer. Formalin-fixed, paraffin-embedded specimens of 15 patients with uni- or bilateral lymph node metastases of prostate cancer (totaling 21 cases) and 17 patient-cases of bone metastases were processed for immunohistochemistry with anti-GRPR, anti-PSCA, and anti-PSMA antibodies. A pathologist blinded to clinical and pathological data scored the immunoreactivity for these antibodies on a four-point scale (0 = no staining; 1+ = weak staining; 2+ = moderate staining; 3+ = strong staining) and documented the distribution pattern. GRPR staining in lymph node metastases was seen in 85.7% of cases (18 of 21 cases), PSCA in 95.2% (20/21), and PSMA in 100% (21/21). GRPR in bone metastases was seen in 52.9% of cases (9/17), PSCA in 94.1% (16/17), and PSMA in 100% (17/17). We have shown for the first time that GRPR is expressed in the vast majority of lymph node metastases and in 52.9% of bone metastases of prostate cancer. PSCA and PSMA are both highly expressed in lymph node and bone metastases. Although PSCA and PSMA are mostly expressed in prostate cancer metastases, GRPR offers an interesting alternative target as it can be targeted relatively easy with peptide-based (radio)pharmaceuticals. (c) 2009 Wiley-Liss, Inc.
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
Structure-Preserving Color Normalization and Sparse Stain Separation for Histological Images.

PubMed

Vahadane, Abhishek; Peng, Tingying; Sethi, Amit; Albarqouni, Shadi; Wang, Lichao; Baust, Maximilian; Steiger, Katja; Schlitter, Anna Melissa; Esposito, Irene; Navab, Nassir

2016-08-01

Staining and scanning of tissue samples for microscopic examination is fraught with undesirable color variations arising from differences in raw materials and manufacturing techniques of stain vendors, staining protocols of labs, and color responses of digital scanners. When comparing tissue samples, color normalization and stain separation of the tissue images can be helpful for both pathologists and software. Techniques that are used for natural images fail to utilize structural properties of stained tissue samples and produce undesirable color distortions. The stain concentration cannot be negative. Tissue samples are stained with only a few stains and most tissue regions are characterized by at most one effective stain. We model these physical phenomena that define the tissue structure by first decomposing images in an unsupervised manner into stain density maps that are sparse and non-negative. For a given image, we combine its stain density maps with stain color basis of a pathologist-preferred target image, thus altering only its color while preserving its structure described by the maps. Stain density correlation with ground truth and preference by pathologists were higher for images normalized using our method when compared to other alternatives. We also propose a computationally faster extension of this technique for large whole-slide images that selects an appropriate patch sample instead of using the entire image to compute the stain color basis.
Histological Stains: A Literature Review and Case Study.

PubMed

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

2015-06-25

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.
Short Nissl staining for incubated cryostat sections of the brain.

PubMed

Lindroos, O F

1991-01-01

Nissl stain often binds poorly to cryostat sections which have been incubated in solutions of radiolabeled ligands. Such incubation is used in receptor autoradiography of the brain when using the in vitro method. We have developed a rapid (16 min) modification of Nissl staining for sections that bind stain poorly, e.g., incubated sections. The method stains well sections which cannot be stained with other rapid Nissl staining methods.
The Bacterial Endospore Stain on Schaeffer Fulton using Variation of Methylene Blue Solution

NASA Astrophysics Data System (ADS)

Oktari, A.; Supriatin, Y.; Kamal, M.; Syafrullah, H.

2017-02-01

Endospores staining is the type of staining to recognize the presence spore in bacterial vegetative cells. The bacterial endospores need a staining which can penetrate wall thickness of spore bacteria. A method of endospores staining is Schaeffer Fulton method that used Malachite Green. It is an alkaline substance staining that can staining the spore bacteria. In this research, it have found the alternative staining that can replace Malachite Green solution in spore bacterial stain. The alternative staining used is Methylene Blue solution (0,5 %, 0,7%, and 1% concentration) with pH variation (10, 11, and 12), and varyous heating time (3, 4, and 5 minutes). The all treatments staining have been effect on bacterial spores staining results. The warming time greatly affect the dye to penetrate the walls of bacterial spores, this can be seen in the results with various concentration at pH 10, indicates that the not long warm-up time 3 and 4 minutes, bacterial spores are not stained, while in the longer heating time is 5 minutes bacterial spores stained. This is caused because the longer heating time can make the pores of spore wall is open so that can facilitate the dye to get into the bacterial spores.
Epidermal Growth Factor Receptor Expression As Prognostic Marker in Patients With Anal Carcinoma Treated With Concurrent Chemoradiation Therapy

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

Fraunholz, Ingeborg, E-mail: inge.fraunholz@kgu.de; Rödel, Franz; Kohler, Daniela

Purpose: To investigate the prognostic value of epidermal growth factor receptor (EGFR) expression in pretreatment tumor biopsy specimens of patients with anal cancer treated with concurrent 5-fluorouracil and mitomycin C-based chemoradiation therapy (CRT). Methods and Materials: Immunohistochemical staining for EGFR was performed in pretreatment biopsy specimens of 103 patients with anal carcinoma. EGFR expression was correlated with clinical and histopathologic characteristics and with clinical endpoints, including local failure-free survival (LFFS), colostomy-free survival (CFS), distant metastases-free survival (DMFS), cancer-specific survival (CSS), and overall survival (OS). Results: EGFR staining intensity was absent in 3%, weak in 23%, intermediate in 36% and intensemore » in 38% of the patients. In univariate analysis, the level of EGFR staining was significantly correlated with CSS (absent/weak vs intermediate/intense expression: 5-year CSS, 70% vs 86%, P=.03). As a trend, this was also observed for DMFS (70% vs 86%, P=.06) and LFFS (70% vs 87%, P=.16). In multivariate analysis, N stage, tumor differentiation, and patients’ sex were independent prognostic factors for CSS, whereas EGFR expression only reached borderline significance (hazard ratio 2.75; P=.08). Conclusion: Our results suggest that elevated levels of pretreatment EGFR expression could be correlated with favorable clinical outcome in anal cancer patients treated with CRT. Further studies are warranted to elucidate how EGFR is involved in the response to CRT.« less
Comparative Assessment of Conventional Papanicolaou and Modified Ultrafast Papanicolaou Stains in Fine Needle Aspiration Samples and Body Fluids.

PubMed

Arul, P; Eniya, S; Pushparaj, Magesh; Masilamani, Suresh; Kanmani, P; Lingasamy, C

2018-01-01

Conventional Papanicolaou (Pap) stain has undergone many modifications; of these, ultrafast Pap stain is the most popular as it shortens the turnaround time of reporting. Application of modified ultrafast Pap (MUFP) stain in the evaluation of fine needle aspiration (FNA) samples and body fluids are scanty. To evaluate the utility of MUFP stain in various FNA samples and body fluids and compare the findings with those of conventional Pap stain. In this cross-sectional study, two wet-fixed and two airdried smears from each sample [301 samples (255 FNA samples and 46 body fluids)] were prepared and stained by the conventional Pap and MUFP stains, respectively. Concordant and discordant rate was calculated. Quality index (QI) of MUFP stain was assessed by background, overall staining, cell morphology, and nuclear characteristics. MUFP-stained smears were also categorized into excellent, good, and fair. The concordance rate for MUFP stain was 100%. QI of MUFP stain for breast, thyroid, lymph node, soft tissue, salivary gland, and body fluids was 0.9, 0.93, 0.95, 1, 0.94, and 1, respectively. Excellent quality of stain was noted in 53.2% and good in 24.6% of the cases allowing easy diagnosis. In 22.2% of fair cases, diagnosis was possible with some difficulties. Our study concluded that MUFP stain could be considered as a rapid and reliable diagnostic tool and can be applied on a regular basis in FNA samples and body fluids to offer immediate diagnosis. However, caution should be taken while reporting certain MUFP-stained smears to avoid over/under diagnosis.
Propidium iodide (PI) stains Nissl bodies and may serve as a quick marker for total neuronal cell count.

PubMed

Niu, Junfei; Li, Chunman; Wu, Haihui; Feng, Xianling; Su, Qingning; Li, Shihe; Zhang, Lihong; Yew, David Tai Wai; Cho, Eric Yu Pang; Sha, Ou

2015-03-01

Propidium iodide (PI) reacts with both DNA and RNA and is a commonly used fluorescent reagent for nucleic acid staining. The aim of the study was to compare the cellular staining patterns of PI with that of Nissl staining in rat nervous tissues and to report a modified staining method that selectively labels Nissl bodies in neurons. Cryosections and paraffin sections of different tissues of normal Sprague-Dawley rats, including trigeminal ganglia, dorsal root ganglia, spinal cord, liver, and small intestine, were stained by either PI or the hematoxylin and eosin method. Some sections were treated with RNase or DNase before the above staining, and some were double stained with PI and a Nissl stain. The sections were observed by light, fluorescence or confocal microscopy. Results showed strong PI signals detected as patterns of granules in the neuronal cytoplasm of all nervous tissues, whereas the staining of neuronal nuclei was weaker. In contrast, nuclei of neuroglial cells were strongly stained by PI, while the cytoplasm was not obviously stained. Pretreatment of the neural tissue with RNase abolished the PI signals. Furthermore, the PI positive granules in neuronal cytoplasm co-localized with Nissl bodies stained by the fluorescent Nissl stain. When the tissue was pretreated with DNase, PI only stained the cytoplasmic granules of neurons, but not that of glial cells. Our results show that PI stains Nissl bodies and may serve as an economical and convenient neuron marker for neuronal cell counting when specific neural markers such as antibodies are not readily available. Copyright © 2015. Published by Elsevier GmbH.
Gram staining with an automatic machine.

PubMed

Felek, S; Arslan, A

1999-01-01

This study was undertaken to develop a new Gram-staining machine controlled by a micro-controller and to investigate the quality of slides that were stained in the machine. The machine was designed and produced by the authors. It uses standard 220 V AC. Staining, washing, and drying periods are controlled by a timer built in the micro-controller. A software was made that contains a certain algorithm and time intervals for the staining mode. One-hundred and forty smears were prepared from Escherichia coli, Staphylococcus aureus, Neisseria sp., blood culture, trypticase soy broth, direct pus and sputum smears for comparison studies. Half of the slides in each group were stained with the machine, the other half by hand and then examined by four different microbiologists. Machine-stained slides had a higher clarity and less debris than the hand-stained slides (p < 0.05). In hand-stained slides, some Gram-positive organisms showed poor Gram-positive staining features (p < 0.05). In conclusion, we suggest that Gram staining with the automatic machine increases the staining quality and helps to decrease the work load in a busy diagnostic laboratory.
Papanicolaou stain: Is it economical to switch to rapid, economical, acetic acid, papanicolaou stain?

PubMed

Dighe, Swati B; Ajit, Dulhan; Pathuthara, Saleem; Chinoy, Roshni

2006-01-01

To standardize an inexpensive and rapid Papanicolaou staining technique with limited ethanol usage. Smears from 200 patients were collected (2 per patient) and fixed in methanol. Half were subjected to conventional Papanicolaou and half to stain ing with rapid, economical, acetic acid Papanicolaou (REAP) stain. In REAP, pre-OG6 and post-OG6 and post-EA36 ethanol baths were replaced by 1% acetic acid and Scott's tap water with tap water. Hematoxylin was preheated to 60 degrees C. Final dehydration was with methanol. REAP smears were compared with Papanicolaou smears for optimal cytoplasmic and nuclear staining, stain preservation, cost and turnaround time. With the REAP method, cytoplasmic and nuclear staining was optimal in 181 and 192 cases, respectively. The staining time was considerably reduced, to 3 minutes, and the cost per smear was reduced to one fourth. The staining quality remained good in all the smears for > 2 years. REAP is a rapid, cost-effective alternative to Papanicolaou stain. Though low stain penetration in large cell clusters is a limitation, final interpretation was not compromised.
Newer applications of the histological stain prepared from Pterocarpus santalinus.

PubMed

Sen Gupta, P C; Mukherjee, A K

1981-03-01

A histological stain prepared from the heartwood of Pterocarpus santalinus Linn. has been found to be an excellent nuclear stain for various cells of animal and plant origin. As an elastic tissue stain, the results are comparable to standard elastic tissue stains. The striations of voluntary muscle fibers are well shown. The Nissl granules and fibers of cranial nerves in the pons are visualized. When counterstained with light green, it differentially stains muscle and fibrous tissue. The stain can be used as counterstain with certain histochemical procedures with satisfactory results. The preparation and use of this versatile stain are described.
Uniform staining of Cyclospora oocysts in fecal smears by a modified safranin technique with microwave heating.

PubMed Central

Visvesvara, G S; Moura, H; Kovacs-Nace, E; Wallace, S; Eberhard, M L

1997-01-01

Cyclospora, a coccidian protist, is increasingly being identified as an important, newly emerging parasite that causes diarrhea, flatulence, fatigue, and abdominal pain leading to weight loss in immunocompetent persons with or without a recent travel history as well as in patients with AIDS. Modified Kinyoun's acid-fast stain is the most commonly used stain to identify the oocyst of this parasite in fecal smears. Oocysts of Cyclospora stain variably by the modified acid-fast procedure, resulting in the possible misidentification of this parasite. We examined fecal smears stained by six different procedures that included Giemsa, trichrome, chromotrope, Gram-chromotrope, acid-fast, and safranin stains. We report on safranin-based stain that uniformly stains oocysts of Cyclospora a brilliant reddish orange, provided that the fecal smears are heated in a microwave oven prior to staining. This staining procedure, besides being superior to acid-fast staining, is fast, reliable, and easy to perform in most clinical laboratories. PMID:9041421

Pleural fluid Gram stain

MedlinePlus

Gram stain of pleural fluid ... mixing it with a violet stain (called a Gram stain). A laboratory specialist uses a microscope to ... reveals an abnormal collection of pleural fluid. The Gram stain can help identify the bacteria that might ...
Selection and application of exterior stains for wood

Treesearch

R. Sam Williams; William C. Feist

1999-01-01

Exterior stains for wood protect the wood surface from sunlight and moisture. Because stains are formulated to penetrate the wood surface, they are not prone to crack or peel as can film-forming finishes, such as paints. This publication describes the properties of stains and wood, methods for applying stains, and the expected service life of stains.
Sebaceous gland carcinoma of the ocular adnexa - variability in clinical and histological appearance with analysis of immunohistochemical staining patterns.

PubMed

Schmitz, Eva Janine; Herwig-Carl, Martina C; Holz, Frank G; Loeffler, Karin U

2017-11-01

The purpose of the study was to evaluate the characteristics of sebaceous gland carcinoma (SGC) of the ocular adnexae, which is due to a high variability in clinical, histological and immunohistochemical characteristics often challenging to diagnose. Records of six patients with SGC were reviewed, who underwent surgical excision and who were histologically diagnosed with SGC. For comparison, there were specimens from four patients with basal cell carcinoma (BCC) and four patients with squamous cell carcinoma (SCC). Histological and immunohistochemical analysis included stains for HE, cytokeratins (CKpan, Cam5.2), epithelial membrane antigen (EMA), androgen receptor (AR441), perforin and adipophilin. SGC's were located in the upper (n = 2) or lower (n = 4) eyelid and were associated with various presenting clinical signs including chalazion-like lesions with pyogenic granuloma (n = 1), papillomatous conjunctival tumors (n = 3), a hyperkeratotic exophytic neoplasm (n = 1) and an ulcerating crusted lesion resembling chronic blepharitis (n = 1). The treatment was tumor resection, followed (if necessary) by adjuvant therapy with topical Mitomycin C (n = 2). Histologic characteristics included basophilic pleomorphic cells with vacuolated cytoplasm, prominent nucleoli, mitotic figures and in some cases pagetoid spread (n = 2). CKpan, EMA and Cam5.2 showed strong positive immunoreactivity in all specimens (SGC, BCC, SCC). Perforin immunostaining showed a varying, but overall weak, non-specific cytoplasmatic staining reaction in all lesions. AR441 positivity was noted with variable intensity in almost all lesions and in particular in pagetoid spread in contrast to non-tumor cells. Adipophilin showed an annular staining of lipid granules in immature sebaceous cells in SGC in contrast to a more granular staining pattern in BCC and SCC. SGCs display a variety of clinical signs and may mimic many other lesions. Tumor resection, followed by histological and immunohistochemical analysis, leads to the diagnosis and initiation of the proper treatment regimen. Herein, immunohistochemistry showed an unequivocal profile in SGC and did not allow for an exact differentiation from BCC and SCC by immunohistochemical means only. An extended evaluation of HE stains remains essential. However, immunohistochemistry can make relevant contributions to the diagnosis of SGC, especially in cases of inconclusive histology, by positive staining for adipophilin in immature sebaceous cells or by AR441 labeling in cases of pagetoid spread.
Digital simulation of staining in histopathology multispectral images: enhancement and linear transformation of spectral transmittance.

PubMed

Bautista, Pinky A; Yagi, Yukako

2012-05-01

Hematoxylin and eosin (H&E) stain is currently the most popular for routine histopathology staining. Special and/or immuno-histochemical (IHC) staining is often requested to further corroborate the initial diagnosis on H&E stained tissue sections. Digital simulation of staining (or digital staining) can be a very valuable tool to produce the desired stained images from the H&E stained tissue sections instantaneously. We present an approach to digital staining of histopathology multispectral images by combining the effects of spectral enhancement and spectral transformation. Spectral enhancement is accomplished by shifting the N-band original spectrum of the multispectral pixel with the weighted difference between the pixel's original and estimated spectrum; the spectrum is estimated using M < N principal component (PC) vectors. The pixel's enhanced spectrum is transformed to the spectral configuration associated to its reaction to a specific stain by utilizing an N × N transformation matrix, which is derived through application of least mean squares method to the enhanced and target spectral transmittance samples of the different tissue components found in the image. Results of our experiments on the digital conversion of an H&E stained multispectral image to its Masson's trichrome stained equivalent show the viability of the method.
Comparison of Gram and Kopeloff stains in the diagnosis of bacterial vaginosis in pregnancy.

PubMed

Libman, Michael D; Kramer, Michael; Platt, Robert

2006-03-01

Bacterial vaginosis (BV) is commonly diagnosed by using the Nugent score, a semiquantitative scoring system to evaluate bacterial morphotypes on Gram stain of vaginal secretions. Some authors have suggested using the Kopeloff modification of the Gram stain. Asymptomatic BV in pregnancy has been associated with adverse outcomes. We performed both stains on simultaneously collected vaginal smears from 2652 women at 24-26 weeks of gestation. Gram staining gave significantly higher (more abnormal) Nugent scores than Kopeloff staining. Compared to the Kopeloff stain, the number of specimens graded as indeterminate or consistent with BV by Gram stain increased by 29% (469 versus 364, P<.001). Interrater reliability of the Nugent score (n=413) for Kopeloff staining was significantly better than Gram staining (agreement=74% versus 63%, intraclass correlation coefficient=0.87 versus 0.79, P<.05, 95% confidence intervals 0.85-0.89 and 0.75-0.82, respectively).
Evaluation of Papanicolaou stain for studying micronuclei in buccal cells under field conditions.

PubMed

Ayyad, Sohair B A; Israel, Ebenezer; El-Setouhy, Maged; Nasr, Ghada Radwan; Mohamed, Mostafa K; Loffredo, Christopher A

2006-01-01

To compare Papanicolaou (Pap) and May-Grünwald Giemsa (MGG) stain as 2 techniques for staining for buccal mucosal cells to detect micronuclei (MN) infield studies. Eighty cytologic smears (2 per individual) were taken from the buccal mucosa of 40 cigarette smokers recruited at a rural village in Egypt. Forty smears were stained with Pap stain and 40 with MGG stain. All were assessed for cellularity and scored for MN. Pap stain was faster and easier to process and transport in the field study than was MGG stain. Regarding MGG smears, bacteria and cell debris masked the MN as compared to Pap smears, in which the fixative destroyed the bacteria and made the cell boundaries clearly demarcated. Using Pap stain, MN were seen easily in transparent cytoplasm. Pap stain is the preferred method infield studies for scoring and detecting MN in cells of buccal mucosa.
EFFECTS OF THE GRAM STAIN ON MICROSPHERES FROM THERMAL POLYAMINO ACIDS1

PubMed Central

Fox, Sidney W.; Yuyama, Shuhei

1963-01-01

Fox, Sidney W. (The Florida State University, Tallahassee) and Shuhei Yuyama. Effects of the Gram stain on microspheres from thermal polyamino acids. J. Bacteriol. 85:279–283. 1963.—Microspheres produced from acid proteinoid accept the Gram stain. The stain is negative, but microspheres produced from mixtures containing a sufficient proportion of lysine proteinoid stain positive. Microspheres produced from mixtures containing the appropriate proportions contain individuals which stain positive and others which stain negative. Images PMID:13959050
Nucleic acid stains as indicators of Giardia muris viability following cyst inactivation.

PubMed

Taghi-Kilani, R; Gyürék, L L; Millard, P J; Finch, G R; Belosevic, M

1996-06-01

A reliable viability assay for Giardia is required for the development of disinfection process design criteria and pathogen monitoring by water treatment utilities. Surveys of single-staining nucleic acid dyes (stain dead parasites only), and double-staining vital dye kits from Molecular Probes (stain live and dead parasites) were conducted to assess the viability of untreated, heat-killed, and chemically inactivated Giardia muris cysts. Nucleic acid staining results were compared to those of in vitro excystation and animal infectivity. Nucleic acid stain, designated as SYTO-9, was considered the best among the single-staining dyes for its ability to stain dead cysts brightly and its relatively slow decay rate of visible light emission following DNA binding. SYTO-9 staining was correlated to animal infectivity. A Live/Dead BacLight was found to be the better of 2 double-staining viability kits tested. Logarithmic survival ratios based on SYTO-9 and Live/Dead BacLight were compared to excystation and infectivity results for G. muris cysts exposed to ozone or free chlorine. The results indicate that SYTO-9 and Live/Dead BacLight staining is stable following treatment of cysts with chemical disinfectants.
Utility of Modified Ultrafast Papanicolaou Stain in Cytological Diagnosis

PubMed Central

Arakeri, Surekha Ulhas

2017-01-01

Introduction Need for minimal turnaround time for assessing Fine Needle Aspiration Cytology (FNAC) has encouraged innovations in staining techniques that require lesser staining time with unequivocal cell morphology. The standard protocol for conventional Papanicolaou (PAP) stain requires about 40 minutes. To overcome this, Ultrafast Papanicolaou (UFP) stain was introduced which reduces staining time to 90 seconds and also enhances the quality. However, reagents required for this were not easily available hence, Modified Ultrafast Papanicolaou (MUFP) stain was introduced subsequently. Aim To assess the efficacy of MUFP staining by comparing the quality of MUFP stain with conventional PAP stain. Materials and Methods FNAC procedure was performed by using 10 ml disposable syringe and 22-23 G needle. Total 131 FNAC cases were studied which were lymph node (30), thyroid (38), breast (22), skin and soft tissue (24), salivary gland (11) and visceral organs (6). Two smears were prepared and stained by MUFP and conventional PAP stain. Scores were given on four parameters: background of smears, overall staining pattern, cell morphology and nuclear staining. Quality Index (QI) was calculated from ratio of total score achieved to maximum score possible. Statistical analysis using chi square test was applied to each of the four parameters before obtaining the QI in both stains. Students t-test was applied to evaluate the efficacy of MUFP in comparison with conventional PAP stain. Results The QI of MUFP for thyroid, breast, lymph node, skin and soft tissue, salivary gland and visceral organs was 0.89, 0.85, 0.89, 0.83, 0.92, and 0.78 respectively. Compared to conventional PAP stain QI of MUFP smears was better in all except visceral organ cases and was statistically significant. MUFP showed clear red blood cell background, transparent cytoplasm and crisp nuclear features. Conclusion MUFP is fast, reliable and can be done with locally available reagents with unequivocal morphology which is the need of the hour for a cytopathology set-up. PMID:28511391
Utility of Modified Ultrafast Papanicolaou Stain in Cytological Diagnosis.

PubMed

Sinkar, Prachi; Arakeri, Surekha Ulhas

2017-03-01

Need for minimal turnaround time for assessing Fine Needle Aspiration Cytology (FNAC) has encouraged innovations in staining techniques that require lesser staining time with unequivocal cell morphology. The standard protocol for conventional Papanicolaou (PAP) stain requires about 40 minutes. To overcome this, Ultrafast Papanicolaou (UFP) stain was introduced which reduces staining time to 90 seconds and also enhances the quality. However, reagents required for this were not easily available hence, Modified Ultrafast Papanicolaou (MUFP) stain was introduced subsequently. To assess the efficacy of MUFP staining by comparing the quality of MUFP stain with conventional PAP stain. FNAC procedure was performed by using 10 ml disposable syringe and 22-23 G needle. Total 131 FNAC cases were studied which were lymph node (30), thyroid (38), breast (22), skin and soft tissue (24), salivary gland (11) and visceral organs (6). Two smears were prepared and stained by MUFP and conventional PAP stain. Scores were given on four parameters: background of smears, overall staining pattern, cell morphology and nuclear staining. Quality Index (QI) was calculated from ratio of total score achieved to maximum score possible. Statistical analysis using chi square test was applied to each of the four parameters before obtaining the QI in both stains. Students t-test was applied to evaluate the efficacy of MUFP in comparison with conventional PAP stain. The QI of MUFP for thyroid, breast, lymph node, skin and soft tissue, salivary gland and visceral organs was 0.89, 0.85, 0.89, 0.83, 0.92, and 0.78 respectively. Compared to conventional PAP stain QI of MUFP smears was better in all except visceral organ cases and was statistically significant. MUFP showed clear red blood cell background, transparent cytoplasm and crisp nuclear features. MUFP is fast, reliable and can be done with locally available reagents with unequivocal morphology which is the need of the hour for a cytopathology set-up.
Evaluation of detection methods for Campylobacter infections among under-fives in Mwanza City, Tanzania.

PubMed

Mushi, Martha Fidelis; Paterno, Laurent; Tappe, Dennis; Deogratius, Anna Pendo; Seni, Jeremiah; Moremi, Nyambura; Mirambo, Mariam Mwijuma; Mshana, Stephen Eliatosha

2014-01-01

Campylobacter species are recognized as a major cause of acute gastroenteritis in humans throughout the world. The diagnosis is mainly based on stool culture. This study was done to evaluate the effectiveness of staining methods (Gram stain using 0.3% carbol fuchsin as counter stain and 1% carbol fuchsin direct stain) versus culture as the gold standard. A total of 300 children attending Bugando Medical Centre (BMC) and the Sekou Toure regional hospital with acute watery diarrhea were enrolled. Two sets of slides were prepared stained with 1% carbol fuchsin for 30 seconds first set, and the second set stained with Gram's stain using 0.3% carbol fuchsin as counter stain for five minutes. Concurrently, stool samples were inoculated on Preston Agar selective. Of 300 stool specimens, 14(4.7%) showed positive culture after 48 hours of incubation and 28 (9.3%) shows typical morphology of Campylobacter species by both Gram stain and direct stain. The sensitivity of the Gram stain using 0.3% carbol fuchsin as counter stain and 1% carbol fuchsin simple stain versus culture as gold standard was 64.3%, with a specificity of 93.4%. The positive predictive value and negative predictive value were 32.1% and 98.2% respectively. The detection of Campylobacter by 1% carbol fuchsin is simple, inexpensive, and fast, with both a high sensitivity and specificity. Laboratories in settings with high prevalence of campylobacteriosis and/or limited resources can employ 1% carbol fuchsin direct stain in detecting campylobacter infections.
Comparison of methylene blue/gentian violet stain to Gram's stain for the rapid diagnosis of gonococcal urethritis in men.

PubMed

Taylor, Stephanie N; DiCarlo, Richard P; Martin, David H

2011-11-01

We compared a simple, one-step staining procedure using a mixture of methylene blue and gentian violet to Gram stain for the detection of gonococcal urethritis. The sensitivity and specificity of both Gram stain and methylene blue/gentian violet stain were 97.3% and 99.6%, respectively. There was a 100% correlation between the 2 methods.
Corneal and conjunctival epithelial staining in hydrogel contact lens wearers.

PubMed

Brautaset, Rune L; Nilsson, Maria; Leach, Norman; Miller, William L; Gire, Anisa; Quintero, Sam; Bergmanson, Jan P G

2008-11-01

The purpose of this study was to investigate the prevalence of conjunctival and corneal epithelial staining in soft contact lens wearers and to see if staining could be associated with factors such as type of lens worn, wearing time, care system, age, and sex. The records of 338 adapted hydrogel contact lens wearers were examined retrospectively. Conjunctival staining was found to be present in 32.5% of the subjects and corneal staining was found to be present in 19.5% of subjects. None of the subjects had staining above grade 2 using the Cornea and Contact Lens Research Unit scale. Because of the low prevalence of staining, the low grading of staining found and the large variation in refractive power, lens type worn, wearing modality, and solution used statistical analysis for association between staining and different factors could only be performed for the association between sex and staining and between corneal and conjunctival staining. However, no statistical significant association could be demonstrated. Despite the low prevalence of staining the conjunctiva and cornea should be examined carefully in contact lens wearers and prospective wearers because the conjunctival and corneal epithelium serve as protective barriers for the underlying layers of the cornea and conjunctiva. To allow comparison of data obtained in different studies assessing corneal staining, it is recommended that clinicians develop and adopt a universal standard protocol for this measure.
Double immunohistochemical staining with MUC4/p53 is useful in the distinction of pancreatic adenocarcinoma from chronic pancreatitis: a tissue microarray-based study.

PubMed

Bhardwaj, Atul; Marsh, William L; Nash, Jason W; Barbacioru, Catalin C; Jones, Susie; Frankel, Wendy L

2007-04-01

Immunohistochemical stains have been used for the distinction of pancreatic adenocarcinoma from chronic pancreatitis. To determine if a double stain for MUC/p53 improved specificity and sensitivity for distinction of pancreatic adenocarcinoma from chronic pancreatitis by comparing maspin, mucin 4 (MUC4), p53, Smad4, and the double stain MUC4/p53. Seventy-four pancreatic adenocarcinomas and 19 chronic pancreatitis cases were retrieved from archival files. Tissue cores were arrayed to create a tissue microarray of 2-mm cores. Sections were stained with antibodies against maspin, MUC4, p53, and Smad4. Additionally, a 2-color, double stain for MUC4 and p53 was developed and evaluated. Five percent or greater staining in either of the cores was considered positive. Intensity (0, 1, 2) and extent (%) of tumor cells staining was also determined. The sensitivity for distinction of pancreatic adenocarcinoma from chronic pancreatitis with maspin, MUC4, p53, and Smad4 was 90%, 77%, 60%, and 63%, respectively; the specificity was 67%, 78%, 88%, and 88%, respectively. When MUC4 and p53 were combined in a double stain, and positive staining for either considered a positive result, the sensitivity increased to 96% but specificity was 73%. When immunoreactivity for both antibodies was necessary for a positive result, sensitivity fell to 39% but specificity was 100%. No correlation was found between intensity or extent of staining with any of the individual stains and tumor differentiation. The double immunohistochemical stain for MUC4/p53 can be a useful diagnostic tool in conjunction with the hematoxylin-eosin-stained section for pancreatic adenocarcinoma, particularly when limited tumor is available for multiple stains.
Counterion dye staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and tryptic gel digestion of stained protein for mass spectrometry.

PubMed

Cong, Wei-Tao; Wang, Xu; Hwang, Sun-Young; Jin, Li-Tai; Choi, Jung-Kap

2012-01-01

A fast and matrix-assisted laser desorption/ionization mass spectrometry compatible protein staining method in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis is described. It is based on the counterion dye staining method that employs oppositely charged two dyes, zincon and ethyl violet, to form an ion-pair complex. The protocol, including fixing, staining, and quick washing steps, can be completed in 1-1.5 h, depending upon gel thickness. It has the sensitivity comparable to the colloidal Coomassie Brilliant Blue G stain using phosphoric acid as a component of staining solution (4-8 ng). The counterion dye stain does not induce protein modifications that complicate interpretation of peptide mapping data from mass spectrometry. Considering the speed, sensitivity, and compatibility with mass spectrometry, the counterion dye stain may be more practical than any other dye-based protein stains for routine proteomic researches.
Multispectral image enhancement for H&E stained pathological tissue specimens

NASA Astrophysics Data System (ADS)

Bautista, Pinky A.; Abe, Tokiya; Yamaguchi, Masahiro; Ohyama, Nagaaki; Yagi, Yukako

2008-03-01

The presence of a liver disease such as cirrhosis can be determined by examining the proliferation of collagen fiber from a tissue slide stained with special stain such as the Masson's trichrome(MT) stain. Collagen fiber and smooth muscle, which are both stained the same in an H&E stained slide, are stained blue and pink respectively in an MT-stained slide. In this paper we show that with multispectral imaging the difference between collagen fiber and smooth muscle can be visualized even from an H&E stained image. In the method M KL bases are derived using the spectral data of those H&E stained tissue components which can be easily differentiated from each other, i.e. nucleus, cytoplasm, red blood cells, etc. and based on the spectral residual error of fiber weighting factors are determined to enhance spectral features at certain wavelengths. Results of our experiment demonstrate the capability of multispectral imaging and its advantage compared to the conventional RGB imaging systems to delineate tissue structures with subtle colorimetric difference.
Facial nerve hemangiomas: vascular tumors or malformations?

PubMed

Benoit, Margo McKenna; North, Paula E; McKenna, Michael J; Mihm, Martin C; Johnson, Matthew M; Cunningham, Michael J

2010-01-01

To reclassify facial nerve hemangiomas in the context of presently accepted vascular lesion nomenclature by examining histology and immunohistochemical markers. Cohort analysis of patients diagnosed with a facial nerve hemangioma between 1990 and 2008. Collaborative analysis at a specialty hospital and a major academic hospital. Seven subjects were identified on composite review of office charts, a pathology database spanning both institutions, and an encrypted patient registry. Clinical data were compiled, and hematoxylin-eosin-stained specimens were reviewed. For six patients, archived pathological tissue was available for immunohistochemical evaluation of markers specific for infantile hemangioma (glucose transporter protein isoform 1 [GLUT1] and Lewis Y antigen) and for lymphatic endothelial cells (podoplanin). All patients clinically presented with slowly progressive facial weakness at a mean age of 45 years without prior symptomatology. Hemotoxylin-eosin-stained histopathological slides showed irregularly shaped, dilated lesional vessels with flattened endothelial cells, scant smooth muscle, and no internal elastic lamina. Both podoplanin staining for lymphatic endothelial cells and GLUT1 and LewisY antigen staining for infantile hemangioma endothelial cells were negative in lesional vessels in all specimens for which immunohistochemical analysis was performed. Lesions of the geniculate ganglion historically referred to as "hemangiomas" do not demonstrate clinical, histopathological, or immunohistochemical features consistent with a benign vascular tumor, but instead are consistent with venous malformation. We propose that these lesions be classified as "venous vascular malformations of the facial nerve." This nomenclature should more accurately predict clinical behavior and guide therapeutic interventions.
First report on C-banding, fluorochrome staining and NOR location in holocentric chromosomes of Elasmolomus (Aphanus) sordidus Fabricius, 1787 (Heteroptera, Rhyparochromidae)

PubMed Central

Suman, Vikas; Kaur, Harbhajan

2013-01-01

Abstract In spite of various cytogenetic works on suborder Heteroptera, the chromosome organization, function and its evolution in this group is far from being fully understood. Cytologically, the family Rhyparochromidae constitutes a heterogeneous group differing in chromosome numbers. This family possesses XY sex mechanism in the majority of the species with few exceptions. In the present work, multiple banding techniques viz., C-banding, base-specific fluorochromes (DAPI/CMA3) and silver nitrate staining have been used to cytologically characterize the chromosomes of the seed plant pest Elasmolomus (Aphanus) sordidus Fabricius, 1787 having 2n=12=8A+2m+XY. One pair of the autosomes was large while three others were of almost equal size. At diplotene, C-banding technique revealed, that three autosomal bivalents show terminal constitutive heterochromatic bands while one medium sized bivalent was euchromatic. Microchromosomes (m-chromosomes) were positively heteropycnotic. After DAPI and CMA3 staining, all the autosomal bivalents showed equal fluorescence, except CMA3 positive signals, observed at both telomeric heterochromatic regions of one medium sized autosomal bivalent. Silver nitrate staining further revealed that this chromosome pair carries Nucleolar Organizer Regions (NORs) at the location of CMA3 positive signals. The X chromosome showed a thick C-band, positive to both DAPI /CMA3 while Y, otherwise C-negative, was weakly positive to DAPI and negative to CMA3, m-chromosomes were DAPI bright and CMA3 dull. PMID:24039525
Similar PDK1-AKT-mTOR pathway activation in balloon cells and dysmorphic neurons of type II focal cortical dysplasia with refractory epilepsy.

PubMed

Lin, Yuan-xiang; Lin, Kun; Kang, De-zhi; Liu, Xin-xiu; Wang, Xing-fu; Zheng, Shu-fa; Yu, Liang-hong; Lin, Zhang-ya

2015-05-01

Dysmorphic neurons and balloon cells constitute the neuropathological hallmarks of type II focal cortical dysplasias (FCDs) with refractory epilepsy. The genesis of these cells may be critical to the histological findings in type II FCD. Recent work has shown enhanced activation of the mTOR cascade in both balloon cells and dysmorphic neurons, suggesting a common pathogenesis for these two neuropathological hallmarks. A direct comparative analysis of balloon cells and dysmorphic neurons might identify a molecular link between balloon cells and dysmorphic neurons. Here, we addressed whether PDK1-AKT-mTOR activation differentiates balloon cells from dysmorphic neurons. We used immunohistochemistry with antibodies against phosphorylated (p)-PDK1 (Ser241), p-AKT (Thr308), p-AKT (Ser473), p-mTOR (Ser2448), p-P70S6K (Thr229), and p-p70S6 kinase (Thr389) in balloon cells compared with dysmorphic neurons. Strong or moderate staining for components of the PDK1-AKT-mTOR signaling pathway was observed in both balloon cells and dysmorphic neurons. However, only a few pyramidal neurons displayed weak staining in control group (perilesional neocortex and histologically normal neocortex). Additionally, p-PDK1 (Ser241) and p-AKT (Thr308) staining in balloon cells were stronger than in dysmorphic neurons, whereas p-P70S6K (Thr229) and p-p70S6 kinase (Thr389) staining in balloon cells was weaker than in dysmorphic neurons. In balloon cells, p-AKT (Ser473) and p-mTOR (Ser2448) staining was comparable with the staining in dysmorphic neurons. Our data support the previously suggested pathogenic relationship between balloon cells and dysmorphic neurons concerning activation of the PDK1-AKT-mTOR, which may play important roles in the pathogenesis of type II FCD. Differential expression of some components of the PDK1-AKT-mTOR pathway between balloon cells and dysmorphic neurons may result from cell-specific gene expression. Copyright © 2015 Elsevier B.V. All rights reserved.
Distribution of calmodulin in pea seedlings: immunocytochemical localization in plumules and root apices

NASA Technical Reports Server (NTRS)

Dauwalder, M.; Roux, S. J.; Hardison, L.

1986-01-01

Immunofluorescence techniques have been used to study the distribution of calmodulin in several tissues in young etiolated pea (Pisum sativum L.) seedlings. A fairly uniform staining was seen in the nucleoplasm and background cytoplasm of most cell types. Cell walls and nucleoli were not stained. In addition, patterned staining reactions were seen in many cells. In cells of the plumule, punctate staining of the cytoplasm was common, and in part this stain appeared to be associated with the plastids. A very distinctive staining of amyloplasts was seen in the columella of the root cap. Staining associated with cytoskeletal elements could be shown in division stages. By metaphase, staining of the spindle region was quite evident. In epidermal cells of the stem and along the underside of the leaf there was an intense staining of the vacuolar contents. Guard cells lacked this vacuolar stain. Vacuolar staining was sometimes seen in cells of the stele, but the most distinctive pattern in the stele was associated with young conducting cells of the xylem. These staining patterns are consistent with the idea that the interactions of plastids and the cytoskeletal may be one of the Ca(2+)-mediated steps in the response of plants to environmental stimuli. Nuclear functions may also be controlled, at least in part, by Ca2+.

The Luna stain, an improved selective stain for detection of microsporidian spores in histologic sections

PubMed Central

Peterson, Tracy S.; Spitsbergen, Jan M.; Feist, Stephen W.; Kent, Michael L.

2014-01-01

Microsporidia in histologic sections are most often diagnosed by observing spores in host tissues. Spores are easy to identify if they occur in large aggregates or xenomas when sections are stained with hematoxylin and eosin (H&E). However, individual spores are not frequently detected in host tissues with conventional H&E staining, particularly if spores are scattered within the tissues, areas of inflammation or small spores in nuclei (i.e., Nucleospora salmonis). Hence, a variety of selective stains that enhance visualization of spores are recommended. We discovered that the Luna stain, used to highlight eosinophils, red blood cells and chitin in arthropods and other invertebrates, also stains spores of Pseudoloma neurophilia. We compared this stain to the Gram, Fite’s acid fast, Giemsa, and H&E stains on eight aquatic microsporidian organisms that were readily available in our two laboratories: Loma salmonae, Glugea anomala, Pseudoloma neurophilia, Pleistophora hyphessobryconis, Pleistophora vermiformis, Glugea sp., Steinhausia mytilovum and an unidentified microsporidian from E. sinensis, UK. Based on tinctorial properties and background staining, the Luna stain performed better for detection of 6 of the 8 microsporidia. Gram stain was superior for the two microsporidia from invertebrates, Steinhausia mytilovum and the unidentified microsporidian from E. sinensis. PMID:21848126
Luna stain, an improved selective stain for detection of microsporidian spores in histologic sections.

PubMed

Peterson, Tracy S; Spitsbergen, Jan M; Feist, Stephen W; Kent, Michael L

2011-06-16

Microsporidia in histologic sections are most often diagnosed by observing spores in host tissues. Spores are easy to identify if they occur in large aggregates or xenomas when sections are stained with hematoxylin and eosin (H&E). However, individual spores are not frequently detected in host tissues with conventional H&E staining, particularly if spores are scattered within the tissues, areas of inflammation, or small spores in nuclei (i.e. Nucleospora salmonis). Hence, a variety of selective stains that enhance visualization of spores is recommended. We discovered that the Luna stain, used to highlight eosinophils, red blood cells, and chitin in arthropods and other invertebrates, also stains spores of Pseudoloma neurophilia. We compared this stain to the Gram, Fite's acid fast, Giemsa, and H&E stains on 8 aquatic microsporidian organisms that were readily available in our 2 laboratories: Loma salmonae, Glugea anomala, Pseudoloma neurophilia, Pleistophora hyphessobryconis, Pleistophora vermiformis, Glugea sp., Steinhausia mytilovum, and an unidentified microsporidian from UK mitten crabs Eriocheir sinensis. Based on tinctorial properties and background staining, the Luna stain performed better for detection of 6 of the 8 microsporidia. Gram stain was superior for the 2 microsporidia from invertebrates: S. mytilovum and the unidentified microsporidian from E. sinensis.
Evaluation of detection methods for Campylobacter infections among under-fives in Mwanza City, Tanzania

PubMed Central

Mushi, Martha Fidelis; Paterno, Laurent; Tappe, Dennis; Deogratius, Anna Pendo; Seni, Jeremiah; Moremi, Nyambura; Mirambo, Mariam Mwijuma; Mshana, Stephen Eliatosha

2014-01-01

Introduction Campylobacter species are recognized as a major cause of acute gastroenteritis in humans throughout the world. The diagnosis is mainly based on stool culture. This study was done to evaluate the effectiveness of staining methods (Gram stain using 0.3% carbol fuchsin as counter stain and 1% carbol fuchsin direct stain) versus culture as the gold standard. Methods A total of 300 children attending Bugando Medical Centre (BMC) and the Sekou Toure regional hospital with acute watery diarrhea were enrolled. Two sets of slides were prepared stained with 1% carbol fuchsin for 30 seconds first set, and the second set stained with Gram's stain using 0.3% carbol fuchsin as counter stain for five minutes. Concurrently, stool samples were inoculated on Preston Agar selective. Results Of 300 stool specimens, 14(4.7%) showed positive culture after 48 hours of incubation and 28 (9.3%) shows typical morphology of Campylobacter species by both Gram stain and direct stain. The sensitivity of the Gram stain using 0.3% carbol fuchsin as counter stain and 1% carbol fuchsin simple stain versus culture as gold standard was 64.3%, with a specificity of 93.4%. The positive predictive value and negative predictive value were 32.1% and 98.2% respectively. Conclusion The detection of Campylobacter by 1% carbol fuchsin is simple, inexpensive, and fast, with both a high sensitivity and specificity. Laboratories in settings with high prevalence of campylobacteriosis and/or limited resources can employ 1% carbol fuchsin direct stain in detecting campylobacter infections. PMID:25995788
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 of...
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 of...
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 of...
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 of...
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 of...
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
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.
Discriminative staining methods for the nervous system: luxol fast blue--periodic acid-Schiff--hematoxylin triple stain and subsidiary staining methods.

PubMed

Goto, N

1987-09-01

This paper describes a new series of staining methods which can discriminatively demonstrate every structure of the nervous system, including axons and capillaries, in animal and human materials. Methods described in this paper consist of one primary stain, luxol fast blue-periodic acid Schiff-hematoxylin (LPH) and six different subsidiary staining methods. The LPH triple stain can precisely differentiate the following structures: neurons (Nissl bodies, cytoplasm, nuclear membrane and nucleolus), various kinds of nuclei (glia, ependyma, endothelium, leucocyte, connective tissue, etc.), myelin sheaths, neuronal processes (axons and dendrites), reacted glial cell bodies (protoplasmic astrocytes, foamy cells, etc.), blood vessels (arteries, veins and capillaries), meninges, intervening connective tissue, erythrocytes, lipofuscin granules, amyloid bodies, and others. Subsidiary staining methods are also described briefly. Applications are discussed in the context of staining technology and neuromorphological research.
Comparison of acridine orange and Gram stains for detection of microorganisms in cerebrospinal fluid and other clinical specimens.

PubMed Central

Lauer, B A; Reller, L B; Mirrett, S

1981-01-01

Acridine orange, a fluorochrome strain, is potentially superior to the Gram stain in the direct microscopic examination of clinical specimens because it gives striking differential staining between bacteria and background cells and debris. Its value in clinical laboratories was evaluated by testing 209 cerebrospinal fluids and 288 other body fluids, tissues, and exudates by both techniques. Smears were made in duplicate, fixed with methanol, stained, and examined without knowledge of the result of the companion smear or culture. Overall, acridine orange was slightly more sensitive than the Gram stain (acridine orange, 59.9%; Gram stain, 55.8%) and equally specific in detecting microorganisms. One smear was falsely positive by the Gram stain; none was falsely positive by the acridine orange stain. We conclude that acridine orange staining is a sensitive method for screening clinical specimens and reviewing selected specimens that are purulent, but negative by the Gram stain. Bloody fluids, thick exudates, and other normally difficult-to-read specimens were easily and quickly examined. We recommend, however, that positive smears be reexamined with the Gram stain to confirm the result and determine the Gram reaction of the microorganisms. PMID:6168652
Morphological characteristics of developmental stages of Acanthamoeba and Naegleria species before and after staining by various techniques.

PubMed

Ithoi, Init; Ahmad, Arine-Fadzlun; Mak, J W; Nissapatorn, Veeranoot; Lau, Yee-Ling; Mahmud, Rohela

2011-11-01

Seven stains were studied to determine the best color and contrast for staining the developmental stages of free living pathogenic Acanthamoeba and Naegleria species. The acid-fast bacilli stain (AFB) produced a blue color without contrast; trichrome-eosin and modified Field's showed various color contrasts; Giemsa, iron-hematoxylin, modified AFB and Gram produced only one color which distinguished the nucleus, nucleolus, cytoplasm, food- and water-vacuoles. The motile organs (acanthopodia, pseudopodia, lobopodia and flagella) were also clearly differentiated but produced a similar color as the cytoplasm. These motile organelles were first induced by incubating at 37 degrees C for at least 15 minutes and then fixing with methanol in order to preserve the protruding morphology prior to staining. The trichrome-eosin and iron-hematoxylin stains showed good color contrast for detecting all three stages, the trophozoite, cyst and flagellate; Giemsa and Gram stained the trophozoite and flagellate stages; the modified Field's and modified AFB stains stained only the trophozoite stage. Depending on the purpose, all these stains (except the AFB stain) can be used to identify the developmental stages of Acanthamoeba and Naegleria for clinical, epidemiological or public health use.
Detection of erythrocyte membrane proteins, sialoglycoproteins, and lipids in the same polyacrylamide gel using a double-staining technique.

PubMed Central

Dzandu, J K; Deh, M E; Barratt, D L; Wise, G E

1984-01-01

A silver/Coomassie brilliant blue R-250 staining technique that permits a color-coded differentiation of erythrocyte membrane proteins, sialoglycoproteins, and lipids in a single one-dimensional NaDodSO4/polyacrylamide gel has been described. Gels stained first with silver stain and then with Coomassie blue (CB) showed the characteristic blue staining of all conventional CB-sensitive membrane polypeptides, whereas periodic acid-Schiff reagent-sensitive sialoglycoproteins and lipids stained yellow. Several yellow Ag-stained bands corresponding to major and minor sialoglycoproteins were detected at Mr X 10(-3) of 88, 72, 65, 41, 35, 31, 28, 24, and 20. Neuraminidase treatment of intact erythrocytes caused shifts in the electrophoretic mobilities of several yellow-stained bands without affecting the CB-stained polypeptide pattern. These observations afforded evidence that the yellow-staining bands were sialoglycoproteins and lipids. The double-staining technique was used in a topological analysis of the membrane surface of the erythrocyte using protease digestion and selective solubilization. Trypsin cleaved the yellow bands at Mr 88,000 and 41,000. Membrane-associated cleavage products were noted at Mr 58,000 and 38,000. Pronase treatment of intact cells gave membrane-associated cleavage products at Mr 38,000 (yellow) and two CB-stained bands at Mr 58,000 and 60,000. These results suggested that the double-staining technique may be applicable in compositional and topological analyses of other biological membranes. Images PMID:6200882
Protocol for vital dye staining of corneal endothelial cells.

PubMed

Park, Sunju; Fong, Alan G; Cho, Hyung; Zhang, Cheng; Gritz, David C; Mian, Gibran; Herzlich, Alexandra A; Gore, Patrick; Morganti, Ashley; Chuck, Roy S

2012-12-01

To describe a step-by-step methodology to establish a reproducible staining protocol for the evaluation of human corneal endothelial cells. Four procedures were performed to determine the best protocol. (1) To determine the optimal trypan blue staining method, goat corneas were stained with 4 dilutions of trypan blue (0.4%, 0.2%, 0.1%, and 0.05%) and 1% alizarin red. (2) To determine the optimal alizarin red staining method, goat corneas were stained with 2 dilutions of alizarin red (1% and 0.5%) and 0.2% trypan blue. (3) To ensure that trypan blue truly stains damaged cells, goat corneas were exposed to either 3% hydrogen peroxide or to balanced salt solution, and then stained with 0.2% trypan blue and 0.5% alizarin red. (4) Finally, fresh human corneal buttons were examined; 1 group was stained with 0.2% trypan blue and another group with 0.4% trypan blue. For the 4 procedures performed, the results are as follows: (1) trypan blue staining was not observed in any of the normal corneal samples; (2) 0.5% alizarin red demonstrated sharper cell borders than 1% alizarin red; (3) positive trypan blue staining was observed in the hydrogen peroxide exposed tissue in damaged areas; (4) 0.4% trypan blue showed more distinct positive staining than 0.2% trypan blue. We were able to determine the optimal vital dye staining conditions for human corneal endothelial cells using 0.4% trypan blue and 0.5% alizarin red.
Contact lens deposits, adverse responses, and clinical ocular surface parameters.

PubMed

Zhao, Zhenjun; Naduvilath, Thomas; Flanagan, Judith L; Carnt, Nicole A; Wei, Xiaojia; Diec, Jennie; Evans, Vicki; Willcox, Mark D P

2010-09-01

To correlate clinical responses during contact lens wear with the amount of protein or cholesterol extracted from lenses after wear. Clinical parameters, including adverse response rates and corneal staining, and symptomatology rating during lens wear were collected from a series of clinical tests comprising four different silicone hydrogel lenses with four different multipurpose solutions. To test for correlates, the amount of total protein or cholesterol extracted from lenses after daily wear were compared statistically to clinical parameters. The amount of protein (p = 0.008) or cholesterol (p = 0.01) extracted from lenses was higher for those subjects who showed solution-induced corneal staining. Amount of protein extracted was correlated (p < 0.01) with conjunctival staining (R = -0.23), lens front surface wetting (r = 0.14), and lens fit tightness (R = -0.20). These clinical parameters accounted for 48% of lens protein deposition. The amount of cholesterol extracted from lenses was much more weakly associated with clinical variables. Amount of protein or cholesterol extracted from lenses was not associated with the production of any corneal infiltrative or mechanical adverse event during wear and was only very weakly correlated with insertion comfort of lenses. These results suggest that there may be no physiologically relevant consequence of cholesterol depositing on silicone hydrogel lenses. The amount of protein that deposits onto silicone hydrogel lenses during wear may have more affect on lens performance on-eye. However, the correlations were generally small and may still not indicate any causative relevant physiological response. Further work is required to determine whether there is any direct causative effect to support these correlative findings.
COX-2 Expression Correlates With Survival in Patients With Osteosarcoma Lung Metastases

PubMed Central

Rodriguez, Nidra I.; Hoots, William Keith; Koshkina, Nadezhda V.; Morales-Arias, Jaime A.; Arndt, Carola A.; Inwards, Carrie Y.; Hawkins, Douglas S.; Munsell, Mark F.; Kleinerman, Eugenie S.

2009-01-01

Summary The purpose of this study was to determine whether a correlation exists between tumor cyclooxygenase (COX)-2 expression and disease-specific survival in patients with osteosarcoma lung metastases. Thirty-six patients diagnosed with osteosarcoma lung metastases between the years 1990 and 2001 were included in this retrospective study. The majority of the patients (72%) presented newly -diagnosed osteosarcoma lung metastases whereas the remaining patients (28%) presented recurrent disease. Clinicopathologic parameters were obtained from patients’ clinical records. Tissue samples were obtained at the time of resection of the lung metastases and stained for COX-2 using immunohistochemistry. Samples were graded according to the intensity of COX-2 staining (grade 0: negative, grade 1: very weak, grade 2: weak, grade 3: moderate, and grade 4: strong). COX-2 staining was correlated with disease-specific survival and clinicopathologic parameters using the Jonckheere-Terpstra and the Kruskal-Wallis tests. All patients with grade 3 or 4 COX-2 expression died of osteosarcoma lung metastases. Ten percent of patients with grade 2 COX-2 expression and 29% of patients with grade 1 expression were alive and free of disease at the last follow-up. By contrast, 60% of the patients with grade 0 COX-2 expression were alive and free of disease at the last follow-up. No association between COX-2 expression and clinicopathologic parameters was found. However, COX-2 expression correlated inversely with disease-specific survival in patients with osteosarcoma lung metastases. Our data indicate that COX-2 expression in metastatic osteosarcoma may have prognostic significance. PMID:18797196
Two-color immunostaining of liver fine needle aspiration biopsies with CD34 and carcinoembryonic antigen. Potential utilization in the diagnosis of primary hepatocellular carcinoma vs. metastatic tumor.

PubMed

Yoder, Michael; Zimmerman, Robert L; Bibbo, Marluce

2004-04-01

To examine immunohistochemical staining of cell block material with antibodies against vascular marker CD34 and polyclonal carcinoembryonic antigen (pCEA) for their clinical utility as part of a 2-color staining protocol in fine needle aspiration (FNA) biopsy of liver masses to distinguish metastases from primary hepatocellular carcinoma (HCC). The authors obtained cell block material from 96 liver FNAs and performed simultaneous (i.e., "dual-color") immunohistochemical staining utilizing antibodies against vascular marker CD34 and pCEA. Cases were blinded and evaluated by the authors for staining pattern and intensity. A consensus was obtained, the results were unblinded, and the diagnoses were correlated. After staining, 89 cases had sufficient tissue for evaluation. Of the 19 HCC cases, 16 (84%) showed peripheral staining with CD34, and 13 (68%) showed a canalicular or mixed canalicular-cytoplasmic staining pattern for pCEA. Thirteen cases (68%) showed staining for both antigens. All HCC exhibited immunostaining for at least 1 antibody in an appropriate staining pattern. Of the 67 cases of metastatic malignancy, 5 (7%) showed a predominantly transgressing pattern of CD34 staining, 43 (64%) showed a predominantly cytoplasmic or mixed cytoplasmic-canalicular pattern of pCEA staining, and 2 cases (3%) showed staining for both antigens in a transgressing CD34 pattern and cytoplasmic pCEA pattern. None of the 3 normal liver tissue blocks showed staining with either antigen. Two-color immunohistochemical staining of liver cell block material obtained by FNA with antibodies to CD34 and pCEA can be helpful in differentiating metastatic tumors vs. primary HCC.
Role of Local Cytokines in Increased Gastric Expression of the Secretory Component in Helicobacter pylori Infection

PubMed Central

Ahlstedt, Ingela; Lindholm, Catharina; Lönroth, Hans; Hamlet, Annika; Svennerholm, Ann-Mari; Quiding-Järbrink, Marianne

1999-01-01

Using immunohistochemical staining, we examined the presence of secretory component (SC) on epithelial cells in gastric and duodenal biopsy specimens collected from Helicobacter pylori-infected individuals and healthy controls. Gastric epithelial cells from healthy volunteers expressed low, but detectable, levels of SC. In contrast, significantly higher level of expression of SC (P < 0.001) was observed on epithelial cells in the antra of H. pylori-infected individuals. The antral SC expression correlated with staining for gamma interferon of intraepithelial and lamina propria lymphocytes (rs = 0.76 and 0.69, respectively, P < 0.001) and correlated weakly with production of tumor necrosis factor alpha (rs = 0.43, P < 0.05), but it did not correlate at all with interleukin-4 production. PMID:10456951
[Histopathological and immunohistochemical studies on mucous cysts].

PubMed

Kuroda, N

1989-01-01

The present study investigated the histopathology, histochemistry of mucopolysaccharides, and immunohistochemistry of oral mucous cysts. The materials were obtained from ninety cases that were histopathologically diagnosed as oral mucous cysts at the Department of Oral Pathology, Meikai University School of Dentistry. Mucopolysaccharide staining was done with PAS, alcian blue (AB, pH 2.5) and high iron diamine (HID). Immunohistochemical studies were focused on secretory component (SC), lactoferrin (Lf), alpha-amylase (Am), IgA, lysozyme (Ly), and keratin (Kr). The following results were obtained: 1. Histopathological findings. (1) Retention and/or retention-like type cysts occurred in was twenty-six cases and the extravasation type in sixty-four cases. (2) Cases showing epithelial lining of the cystic wall were only eight in number, and many cystic walls were contained granulation tissue (fifty cases). (3) As for inflammation of the cystic wall, the degree was slight, and infiltrated cells were mainly macrophages (so-called mucinophages) and lymphocytes. (4) Regarding adjoining salivary glands, acinar cells showed atrophic changes, and hypertrophy of mucous acinar cells was evident. Many ducts showed dilatation, and stromal connective tissue showed fibrosis and hyalinization. 2. Histochemical findings on mucopolysaccharides. (1) Mucous materials in cystic cavity, mucous acinar cells, and secretory materials in ductal lumens were intensely stained by PAS and AB. But stainability with AB was less than that with PAS staining. Serous acinar cells and ductal epithelium were negative to PAS and AB staining. (2) Stainability of the above with HID was less than at with PAS or AB. Cystic walls were not stained by HID. Mucous acinar cells reactive with HID were intensely stained, but the number of the positive cells was limited when compared with the numbers of PAS-and AB-positive cells. 3. Immunohistochemical findings. (1) As for mucous materials in the cystic cavity, reactions for Sc, Am, IgA, and Ly were all positive, whereas those for Lf and Kr were negative. (2) Staining of cystic walls was generally weak: the walls were negative for IgA, Ly and Kr showed and borderline or slightly positive for Sc, Lf, and Am. (3) Mucous acinar cells were negative for all markers examined in this study, but serous acinar cells and/or demilunes were markedly positive for Sc, Ly, and Am. (4) In ductal epithelial cells, Ly and Kr were negative, but IgA was borderline or slightly positive. Sc and Am gave intensely positive staining. (5) Secretory materials in ductal lumens, Sc were intensely positive for Lf, and Ly; slightly or moderately positive for IgA; and slightly positive or negative for Am.(ABSTRACT TRUNCATED AT 400 WORDS)

Comparison of selective staining of fungi in paraffin sections by light microscopy, SEM and BEI

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

Berman, E.L.; Laudate, A.; Carter, H.W.

Paraffin-embedded sections from human tissues with fungi or organisms classified with fungi were studied by light microscopy (LM), scanning electron microscopy (SEM), and the backscatter electron imaging (BEI) mode of the SEM. The fungal organisms selected for study were those familiar to the pathologist on the basis of their appearance in paraffin-embedded material stained with the Gomori-Grocott Chromic Acid Methenamine Silver Stain (GMS). The organisms were Actinomyces, Rhizopus, Cryptococcus, Histoplasma capsulatum, and Coccidia imitis. Sections were stained with the GMS Stain and/or the Becker modification of the GMS Stain (BGMS) and examined in the secondary electron imaging mode (SEI) andmore » BEI mode with an annular backscatter electron detector. This silver staining technique accentuated the wall of fungal organisms, in the backscatter mode. Depending on the fungal organism and type of silver stain employed, the GMS seemed the preferable stain. The advantages of SEM over LM were greater depth of focus and potential range of magnifications. BEI may also be used in conjunction with LM stain for microorganisms to establish their presence.« less
Demonstration of lipofuscin and Nissl bodies in crystal violet stained sections using a fluorescence technique or pyronin Y stain.

PubMed

Terr, L I

1986-09-01

This paper presents two simple, reliable methods for identification of lipofuscin and Nissl bodies in the same section. One method shows that lipofuscin stained with crystal violet retains its ability to fluoresce and can be observed under the fluorescence microscope after the stain has faded. Fading is accompanied by a gradual increase in the intensity of the fluorescence and is complete in about 5 min. Exciting illumination from this part of the spectrum also substantially fades staining of other autofluorescing tissue elements, such as lipids. Nonfluorescing structures, such as Nissl bodies, remain stained. By changing from transillumination with tungsten light to epifluorescent illumination and vice versa, both types of structures--Nissl bodies and lipofuscin--can be identified in the same section. The second technique uses pyronin Y for staining Nissl bodies in preparations previously stained with crystal violet. Nissl bodies are stained pink but lipofuscin remains violet. Lipofuscin in these sections also remains autofluorescent after the crystal violet stain has faded under violet or near-UV light.
Comparing The Efficacy of Hematoxylin and Eosin, Periodic Acid Schiff and Fluorescent Periodic Acid Schiff-Acriflavine Techniques for Demonstration of Basement Membrane in Oral Lichen Planus: A Histochemical Study

PubMed Central

Pujar, Ashwini; Pereira, Treville; Tamgadge, Avinash; Bhalerao, Sudhir; Tamgadge, Sandhya

2015-01-01

Background: Basement membrane (BM) is a thick sheet of extracellular matrix molecules, upon which epithelial cells attach. Various immunohistochemical studies in the past have been carried out but these advanced staining techniques are expensive and not feasible in routine laboratories. Although hematoxylin and eosin (H-E) is very popular among pathologists for looking at biopsies, the method has some limitations. This is where special stains come handy. Aims and Objectives: The aim of the present study was to demonstrate and compare the efficacy of H-E, periodic acid Schiff (PAS) and fluorescent periodic acid–acriflavine staining techniques for the basement membrane and to establish a histochemical stain which could be cost effective, less time consuming, and unambiguous for observation of the basement membrane zone. Materials and Methods: A total number of 40 paraffin-embedded tissue sections of known basement membrane containing tissues including 10 – Normal oral mucosa (NOM) and 30 – oral lichen planus (OLP) were considered in the study. Four-micron-thick sections of each block were cut and stained with H-E stain, PAS and fluorescent periodic acid–acriflavine stain. Sections were evaluated by three oral pathologists independently for continuity, contrast and pattern. Results: Though all the three stains showed favorable features at different levels, acriflavine stain was better than the other stains in demonstrating BM continuity, contrast and also the pattern followed by PAS stain. Acriflavine stain was the better in demonstrating a fibrillar pattern of a BM. Acriflavine stains a BM distinctly and is less time consuming and easy to carry out using readily available dyes as compared to other stains. Conclusion: The continuity and contrast along with the homogenous pattern and the afibrillar pattern of the BM was better demonstrated by acriflavine followed by the PAS stain. PMID:26538690
Expression of Fra-1 in human hepatocellular carcinoma and its prognostic significance.

PubMed

Gao, Xiao-Qiang; Ge, Yong-Sheng; Shu, Qing-Hua; Ma, Hua-Xing

2017-06-01

This study aimed to explore the clinical significance and prognostic value of Fra-1 in hepatocellular carcinoma patients after curative resection. Fra-1 expression was investigated using a combination of techniques: immunohistochemistry for 66 samples of hepatocellular carcinoma and quantitative real-time polymerase chain reaction and western blotting assays for 19 matched hepatocellular carcinoma specimens. Fra-1 was present in 38 of 66 (57.6%) tumor tissues, with intense staining in the nuclei. There was also positive staining in 14 of 66 (21.2%) adjacent peritumoral tissues, with weak staining in the cytoplasm. Quantitative real-time polymerase chain reaction and western blotting assays confirmed higher expression of Fra-1 messenger RNA and Fra-1 protein in tumor tissues than adjacent non-tumor tissues for 19 hepatocellular carcinoma samples (p < 0.001). Positive expression of Fra-1 was significantly related to vascular invasion and serum alpha-fetoprotein. Kaplan-Meier survival analysis found that overexpressed Fra-1 was correlated with poor overall survival and disease-free survival. Multivariate analysis identified Fra-1 as an independent prognostic factor. Fra-1 may be involved in the progress of hepatocellular carcinoma and could be a promising molecular candidate in the diagnosis and treatment of hepatocellular carcinoma.
A case of primary mucosa-associated lymphoid tissue lymphoma of the vagina.

PubMed

Yoshinaga, Kousuke; Akahira, Jun-Ichi; Niikura, Hitoshi; Ito, Kiyoshi; Moriya, Takuya; Murakami, Takashi; Kameoka, Jun-Ichi; Ichinohasama, Ryo; Okamura, Kunihiro; Yaegashi, Nobuo

2004-09-01

We report the first case of primary mucosa-associated lymphoid tissue (MALT) lymphoma of the vagina, the diagnosis of which is supported by genetic and immunophenotypic studies. A 65-year-old, para 2 woman presented to our hospital in July 1997 with a history of prolonged vaginal discharge. Although cytologic examination suggested possible malignancy, a biopsy of the vaginal wall was diagnosed as chronic inflammation. In June 2000, she underwent gynecologic examination because of anuria. Excisional biopsy revealed subepithelial infiltration of atypical lymphoid cells that stained for CD20, CD79a, and BCL-2; stained weakly for IgM; and did not stain for CD3, CD5, CD7, CD10, CD56, CD23, and IgD, suggesting marginal zone B-cell lineage. Monoclonality was detected by Southern blot analysis, and this patient was finally diagnosed as having primary MALT lymphoma of the vagina. She received 3 cycles of chemotherapy (THP-COP) and concurrent radiation to the whole pelvis. The patient is alive and well 40 months after treatment. Because the vagina is one of the mucosa-associated tissues, MALT lymphoma, though rare, must be included in the differential diagnosis of the vaginal neoplasms.
'Ease of interpretation' of cytological smears stained with modified ultrafast papanicolaou stain: Interobserver agreement and reproducibility.

PubMed

Uthamalingam, Preithy; Sathish Kumar, Thabasum; Venus, Albina; Sekar, Preethi; Muthusamy, Rajeshwari K; Mehta, Sangita

2018-04-01

Since its inception in 1995, the Ultrafast Papanicoloau (UFPAP) cytological stain has undergone a number of modifications to suit the local availability of reagents and cost in different set ups. However, the reported results have been uniformly encouraging. We designed a study to investigate the inter-observer agreement in 'perceived ease of interpretation' of cytological smears stained with Modified Ultrafast Papanicoloau stain (MUFPAP). After a small pilot study, we prospectively stained air-dried fine needle aspirate smears (FNACs) and Body Fluid smears with the standardized MUFPAP stain. The MUFPAP stained slides were evaluated in tandem with other routine cytological stains as well as independently by two pathologists. Two rater kappa was used to determine the agreement. The study included 93 fluids and 34 FNACs. A vast majority of the cases stained with MUFPAP were rated 'better' than the routine stains in terms of 'overall ease of interpretation' with considerable agreement. The agreement tended to be better for FNACs than fluid specimens. Cases with malignant pathology demonstrated a perfect agreement (kappa = 1) between the raters in terms of 'overall ease of interpretation' (91.7% cases were rated 'very good' by each pathologist) when compared to cases with benign pathology (kappa = 0.52). Nuclear characteristics were appreciated with a better agreement than other parameters. Modified UFPAP stain appears to be quick, reliable, cost-effective alternative in cytology, especially for detecting malignant cells in smears with low cellularity. Its specific advantage is robust nuclear staining against a clear background. © 2018 Wiley Periodicals, Inc.
Crossover clinical investigation of a whitening chewing gum for inhibiting dental stain formation in conjunction with tooth brushing.

PubMed

Milleman, Jeffery L; Milleman, Kimberly R; Kleber, Carl J; Proskin, Howard M; Dodds, Michael; Kelley, Michael; Ramirez, Lilian

2014-01-01

The purpose of this clinical investigation was to evaluate the effectiveness of a marketed whitening chewing gum compared to a no-gum control in preventing the formation of extrinsic stains on the teeth of stain-forming subjects when chewed over a 12-week period of regular unsupervised use in conjunction with daily tooth brushing. This was a single-center, examiner-blind, randomized, 12-week crossover clinical trial. Stain-forming (after smoking or drinking coffee or tea) adults, starting with a stain-free baseline, either chewed the test gum (Orbit White) unsupervised four times per day, 15 minutes/chew, or used no gum along with daily brushing with a commercially available toothbrush and dentifrice for 12 weeks. At the crossover, all procedures were repeated with subjects assigned the opposite treatment. Extrinsic stain was measured at six and 12 weeks by both the Lobene Stain Index (LSI) and the Modified Lobene Stain Index (MLSI) using separate experienced examiners. After 12 weeks, LSI stain scores showed a significant 25% reduction (p = 0.0008) in new stain formation for subjects using the test chewing gum along with tooth brushing versus tooth brushing alone (no-gum control). The corresponding MLSI stain scores demonstrated a 36% reduction (p < 0.0001) in the formation of extrinsic stain on the teeth. The overall findings of this clinical study demonstrated that regular use of Orbit White chewing gum, soon after smoking or drinking coffee or tea, will supplement daily tooth brushing in preventing unsightly stains from forming on the anterior teeth compared to brushing alone.
Centrifuge-operated specimen staining method and apparatus

NASA Technical Reports Server (NTRS)

Feeback, Daniel L. (Inventor); Clarke, Mark S. F. (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.
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.
Is the gram stain useful in the microbiologic diagnosis of VAP? A meta-analysis.

PubMed

O'Horo, John C; Thompson, Deb; Safdar, Nasia

2012-08-01

In a meta-analysis examining respiratory specimen Gram stain for diagnosis of ventilator-associated pneumonia, absence of bacteria on Gram stain had a high negative predictive value, but a positive Gram stain correlated poorly with organisms recovered in culture. Rapid and accurate diagnosis of ventilator-associated pneumonia (VAP) is a major challenge and no generally accepted gold standard exists for VAP diagnosis. We conducted a meta-analysis to examine the role of respiratory specimen Gram stain to diagnose VAP, and the correlation with final culture results. In 21 studies, pooled sensitivity of Gram stain for VAP was 0.79 (95% confidence interval [CI], .77-0.81; P < .0001) and specificity was 0.75 (95% CI, .73-.78; P < .0001). Negative predictive value of Gram stain for a VAP prevalence of 20%-30% was 91%, suggesting that VAP is unlikely with a negative Gram stain but the positive predictive value of Gram stain was only 40%. Pooled kappa was 0.42 for gram-positive organisms and 0.34 for gram-negative organisms, suggesting fair concordance between organisms on Gram stain and recovery by culture. Therefore, a positive Gram stain should not be used to narrow anti-infective therapy until culture results become available.
Detection of the multiphoton signals in stained tissue using nonlinear optical microscopy

NASA Astrophysics Data System (ADS)

Zeng, Yaping; Xu, Jian; Kang, Deyong; Lin, Jiangbo; Chen, Jianxin

2016-10-01

Multiphoton microscopy (MPM) based on two-photon excited fluorescence (TPEF) and second harmonic generation (SHG) imaging, has become a powerful, important tool for tissue imaging at the molecular level. Recently, MPM is also used to image hematoxylin and eosin (H and E)-stained sections in cancer diagnostics. However, several studies have showed that the MPM images of tissue stained with H and E are significantly different from unstained tissue sections. Our aim was to detect of the multiphoton signals in stained tissue by using MPM. In this paper, MPM was used to image histological sections of esophageal invasive carcinoma tissues stained with H, E, H and E and fresh tissue. To detect of the multiphoton signals in stained tissue, the emission spectroscopic of tissue stained with H, E, H and E were obtained. For comparison, the fresh tissues were also investigated. Our results showed that the tissue stained with H, E, H and E could be detected by their TPEF signals. While the tissue stained with H and fresh tissue could be detected by their TPEF and SHG signals. In this work, we detect of the multiphoton signals in stained tissue. These findings will be useful for choosing suitable staining method so to improve the quality of MPM imaging in the future.
A robust, efficient and flexible method for staining myelinated axons in blocks of brain tissue.

PubMed

Wahlsten, Douglas; Colbourne, Frederick; Pleus, Richard

2003-03-15

Previous studies have demonstrated the utility of the gold chloride method for en bloc staining of a bisected brain in mice and rats. The present study explores several variations in the method, assesses its reliability, and extends the limits of its application. We conclude that the method is very efficient, highly robust, sufficiently accurate for most purposes, and adaptable to many morphometric measures. We obtained acceptable staining of commissures in every brain, despite a wide variety of fixation methods. One-half could be stained 24 h after the brain was extracted and the other half could be stained months later. When staining failed because of an exhausted solution, the brain could be stained successfully in fresh solution. Relatively small changes were found in the sizes of commissures several weeks after initial fixation or staining. A half brain stained to reveal the mid-sagittal section could then be sectioned coronally and stained again in either gold chloride for myelin or cresyl violet for Nissl substance. Uncertainty, arising from pixelation of digitized images was far less than errors arising from human judgments about the histological limits of major commissures. Useful data for morphometric analysis were obtained by scanning the surface of a gold chloride stained block of brain with an inexpensive flatbed scanner.
The use of a differential fluorescent staining method to detect bacteriuria.

PubMed

Ciancaglini, Ettore; Fazii, Paolo; Sforza, Giuseppe Riario

2004-01-01

This report describes a differential staining method which distinguishes gram-positive from gram-negative bacteria in fluorescence. Gram-positive bacteria appear yellow and gram-negative bacteria appear green. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein, which together form a red/ green system. In this report we compared the accuracy of the differential fluorescent staining method and the Gram stain in screening for bacteriuria, as detected by conventional cultures. A total of 1487 urine samples were tested. 289 cultures were positive. 237 specimens grew a single organism at 10(5) and 10(4) CFU/ml. 224 smears were detected by the differential fluorescent staining method and 162 were detected by Gram stain. 1198 samples failed to grow organisms at 10(5) and 10(4) CFU/ml. 107 smears were falsely positive by the fluorescent staining procedure and 289 were falsely positive by the Gram stain. On the basis of the culture results, the sensitivity of the differential fluorescent staining method was 94.5% and that of the Gram stain 68.3%. The specificity of the fluorescent staining procedure was 91.6% and that of the Gram stain 75.8%. The positive predictive value and the negative predictive value of the fluorescent staining method were 67.6% and 98.8%, respectively. Those of the Gram stain were 35.9% and 92.3%, respectively. A wide range of microbiological and chemical techniques are available to identify bacteria in urine. This fluorescent staining method represents a simple, rapid, reliable method with low-running costs. The main advantage of this technique is that it enables the microbiologist to exclude the presence of bacteria in the urine within a short time after specimen receipt and to eliminate a large number of specimens for culture with significant cost saving. Another advantage of the method is that it allows to distinguish gram-positive from gram-negative bacteria in positive slides on the same day the sample is obtained. The stained smears were easily interpreted, even when the bacterial counts in the specimen were low.
Paraffin immunoreactivity of CD10, CDw75, and Bcl-6 in follicle center cell lymphoma.

PubMed

Dunphy, C H; Polski, J M; Lance Evans, H; Gardner, L J

2001-05-01

Follicle center cell lymphoma(FCCL) has the following immunophenotype(IP): sIg+, Pan B+, CD10+/-, CD5-, CD23-/+, CD43-, CD11c-, CD25-. In addition, reactivities of a malignant lymphoma with CDw75(LN-1) and bcl-6 are considered indicators of FCCL. Bcl-6 expression is common in Grade 1 FCCL (100%) and rare in other indolent B-cell lymphomas(BCL). In contrast, bcl-2 expression is common in FCCL (80%) and in other BCL subtypes. Since no previous study has correlated paraffin immunoreactivity(PIR) of CD10, CDw75, and bcl-6 in FCCL (Grades 1-3), this is this study's purpose. Twenty-nine FCCL's were identified and reviewed (6, Grade 1; 10, Grade 2; 13, Grade 3) from the Division of Hematopathology, St. Louis University. The diagnoses were based on morphology and immunohistochemistry(IH)(21 cases) +/- the flow cytometric IP(14 cases). The paraffin blocks were stained for CD10 (Novacastra, Vector Laboratories, Burlingame, CA), CDw75 and bcl-6 (DAKO Corporation, Carpinteria, CA). Results showed that, CD10 by paraffin IH(PIH) was positive in 23 [18(strong); 3(moderate); 2(weak)] and negative in 6(3, Grade 2; 3, Grade 3). All CD10-cases were CDw75+; 4, bcl-6+. The two CD10-, bcl-6-cases were Grade 2. CDw75 was positive in 28 cases [16(strong); 11(moderate); 1(weak)] and negative in 1 (Grade 3; CD10+, bcl-2+, bcl-6+). Bcl-6 was positive in 26 [16(strong); 6(moderate); 4(weak)] and negative in 3(Grade 2's). Thus, the sensitivity of CD10, CDw75, and bcl-6 by PIH for FCCL was 79%, 97%, and 90%, respectively. Of the three stains evaluated by PIH in FCCL, CDw75 was the most sensitive, closely followed by bcl-6. CD10 was least sensitive-79%. By combining these 3 stains, the sensitivity was 100%; thus, a combined approach is recommended.
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.
A Versatile Giemsa Protocol for Permanent Nuclear Staining of Fungi

Treesearch

A. Dan Wilson

1992-01-01

A variety of cytological stains and staining procedures including Giemsa-HCL, acetic-or-cein, propionic-carmine, iron haema-toxylin, safranin O, aniline blue or trypan blue, toluidine blue and basic fuchsin or Feulgen stain have been used to investigate the nuclear condition of reproductive and somatic structures of fungi. Fluorescent stains such as acriflavin acridine...
Pericardial fluid Gram stain

MedlinePlus

... stain To use the sharing features on this page, please enable JavaScript. Pericardial fluid Gram stain is a method of staining a sample of fluid taken from the pericardium. This is the sac surrounding ...
Fluorescein-labeled β-Glucosidase as a Bacterial Stain

PubMed Central

Pital, Abe; Janowitz, Sheldon L.; Hudak, Charles E.; Lewis, Evelyn E.

1967-01-01

Fluorescein isothiocyanate-labeled β-glucosidase was used as a simple staining reagent with selected gram-positive and gram-negative organisms. Staining in situ appeared to be dependent on the presence of accessible glycosidic-type linkages in the bacterial cell wall. Extensive wall damage or lysis did not occur when stained cells were suspended in washing and mounting solutions. The apparent specificity of labeled enzyme for wall substance was tested by blocking reactions, staining of isolated cell walls, and failure to stain substances lacking appropriate glycosidic linkages. Severe cell wall lesions were produced after prolonged contact with labeled enzyme, and this phenomenon may also be related to staining specificity. Gram-negative organisms and spores were poorly stained unless protected glycopeptide substrate was previously exposed by treatment of cells with thioglycolic acid or dilute alkaline sodium hypochlorite solution. A potential for staining tissues and cell lines may also exist. Some possible applications of labeled enzymes are briefly discussed. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:4169543
Bulbar conjunctival fluorescein staining in hydrogel contact lens wearers.

PubMed

Lakkis, C; Brennan, N A

1996-07-01

To evaluate the clinical usefulness of bulbar conjunctival staining in assessing hydrogel contact lens patients. Overall staining, as well as staining at five separate sites (limbal, nasal band, temporal band, superior, and inferior) was graded on an analog scale in 48 contact lens wearing subjects and 50 controls. The degree to which subjects experienced sensations of dryness, wateriness, itchiness, grittiness, and comfort was also assessed using analog scales. Some measure of conjunctival staining was noted in 98% of subjects, with contact lens wearers showing statistically significant greater staining than controls. Only 12% of controls showed staining of greater than grade 1 (equivalent), whereas 62% of contact lens wearers were above this level. Regression analysis showed that overall staining was a function of whether contact lenses were worn, the degree of dryness, and the amount of itchiness. Conjunctival fluorescein staining appears to serve some clinical usefulness as a composite indicator for certain factors and symptoms and, in addition, provides information which is unique for each individual.
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.

A novel histological technique for distinguishing between epithelial cells in forensic casework.

PubMed

French, Claire E V; Jensen, Cynthia G; Vintiner, Susan K; Elliot, Douglas A; McGlashan, Susan R

2008-06-10

There are a number of forensic cases in which the identification of the epithelial cell type from which DNA originated would provide important probative evidence. This study aimed to develop a technique using histological staining of fixed cells to distinguish between skin, buccal and vaginal epithelium. First, 11 different stains were screened on formalin-fixed, wax-embedded cells from five women. Samples were analysed qualitatively by examining staining patterns (colour) and morphology (absence or presence of nuclei). Three of the staining methods--Dane's, Csaba's and Ayoub-Shklar--were successful in distinguishing skin epithelial cells from buccal and vaginal. Second, cells were smeared directly onto slides, fixed with one of five fixatives and stained with one of the three stains mentioned above. Methanol fixation, coupled with the Dane's staining method, specific to keratin, was the only technique that distinguished between all three cell types. Skin cells stained magenta, red and orange and lacked nuclei; buccal cells stained predominantly orange-pink with red nuclei; while vaginal cells stained bright orange with orange nuclei and a blue extracellular hue. This staining pattern in vaginal cells was consistent in samples collected from 50 women aged between 18 and 67. Identification of cell type from unlabelled micrographs by 10 trained observers showed a mean success rate of 95%. The results of this study demonstrate that histological staining may provide forensic scientists with a technique for distinguishing between skin, buccal and vaginal epithelial cells and thus would enable more conclusive analyses when investigating sexual assault cases.
Frequency of satellite association of human chromosomes is correlated with amount of Ag-staining of the nucleolus organizer region.

PubMed Central

Miller, D A; Tantravahi, R; Dev, V G; Miller, O J

1977-01-01

Methaphase chromosomes from karyotypically normal adult humans (three males, six females) and one male with a 13p - chromosome were stained by quinacrine and then by the Ag-AS silver staining method to reveal nucleolus organizer regions (NORs). Each person had a characteristic number of Ag-stained chromosomes per cell, always fewer than 10. Determination of the mean Ag-size of each chromosome showed that each of the 10 individuals had a unique distribution of Ag-stain. Within each individual, there was some variation from cell to cell in the number of acrocentric chromosomes that were Ag-stained; this was not random, and the same chromosomes (those that had at most a small amount of Ag-stain) tended to be unstained in every cell. Satellite associations were scored on the same cells. Chromosomes that had no Ag-stain were involved in satellite association less than 20% as often as those that had some Ag-stain. Chromosomes that had a small amount of Ag-stain were involved in association about 50% as often as those that had a large amount of stain. Regression analysis of the 50 (of a total of 100) acrocentric chromosomes which could be individually identified by quinacrine markers showed that the frequency with which a chromosome was involved in satellite association was strongly correlated with the amount of Ag-stained material in the NOR. Images Fig. 1 Fig. 3 PMID:70995
Differences in staining intensities affect reported occurrences and concentrations of Giardia spp. in surface drinking water sources.

PubMed

Alderisio, K A; Villegas, L F; Ware, M W; McDonald, L A; Xiao, L; Villegas, E N

2017-12-01

USEPA Method 1623, or its equivalent, is currently used to monitor for protozoan contamination of surface drinking water sources worldwide. At least three approved staining kits used for detecting Cryptosporidium and Giardia are commercially available. This study focuses on understanding the differences among staining kits used for Method 1623. Merifluor and EasyStain labelling kits were used to monitor Cryptosporidium oocyst and Giardia cyst densities in New York City's raw surface water sources. In the year following a change to the approved staining kits for use with Method 1623, an anomaly was noted in the occurrence of Giardia cysts in New York City's raw surface water. Specifically, Merifluor-stained samples had higher Giardia cyst densities as compared with those stained with EasyStain. Side by side comparison revealed significantly lower fluorescence intensities of Giardia muris as compared with Giardia duodenalis cysts when labelled with EasyStain. This study showed very poor fluorescence intensity signals by EasyStain on G. muris cysts resulting in lower cyst counts, while Merifluor, with its broader Giardia cyst staining specificity, resulted in higher cyst counts, when using Methods 1623. These results suggest that detected Giardia cyst concentrations are dependent on the staining kits used, which can result in a more or less conservative estimation of occurrences and densities of zoonotic Giardia cysts by detecting a broader range of Giardia species/Assemblages. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.
[Myofibroma/myofibromatosis: a clinicopathologic analysis of 9 cases].

PubMed

Fu, Y; Guan, W Y; Wu, H Y; Wu, H Y; Fan, Z W; Ye, Q; Meng, F Q

2018-01-08

Objective: To investigate the clinical and histological features, diagnosis and differential diagnosis of myofibroma/myofibromatosis. Methods: The clinical data and pathology features of nine cases of myofibroma/myofibromatosis were collected from August 2011 to November 2016 in Affiliated Drum Tower Hospital, Nanjing University Medical School and Children's Hospital of Nanjing Medical University. Immunohistochemistry(IHC), PDGFRB molecular analysis and ETV6-NTRK3 gene fusion were performed and relevant literature reviewed. Results: There were 7 males and 2 females, with age ranging from 3 days to 18 years (mean 5 years). The tumors were located in head and neck (eight cases) and trunk (one case). Clinically, the tumors presented as freely movable nodules. Microscopically, they appeared biphasic with alternating light- and dark-staining areas. The light-staining area consisted mainly of plump myoid spindle cells with eosinophilic cytoplasm arranged in nodules, short fascicles, or whorls.The dark-staining area was composed of round or polygonal cells with slightly hyperchromatic nuclei or small spindle cells arranged around a distinct hemangiopericytoma-like vascular pattern. IHC showed the tumor cells in the light-staining area were strongly positive for vimentin and SMA, while cells in dark-staining area were strongly positive for vimentin, and weakly for SMA. Tumor cells were negative for desmin, S-100 protein, h-Caldesmon, CD34 and STAT6. Analysis of PDGFRB mutations was performed in seven cases. Two cases showed 12 exon point mutation c. 1681 c>T(p.R561C), one case showed 14 exon point mutation c. 1998C>G (p.N666K). ETV6-NTRK3 gene fusion was not detected by fluorescence in situ hybridization in four patients under three years old. All cases were followed for 6 to 68 months, with two recurrences. Conclusions: Myofibroma/myofibromatosis is an uncommon benign myofibroblastic tumor of infancy and childhood. The tumor can appear biphasic, and may show PDGFRB point mutation which is of potential diagnostic value.
Evidence for alternative pathways of granulosa cell death in healthy and slightly atretic bovine antral follicles.

PubMed

Van Wezel, I L; Dharmarajan, A M; Lavranos, T C; Rodgers, R J

1999-06-01

Granulosa cell death is an early feature of atresia; however, there are many apparent contradictions in the literature concerning the mode of granulosa cell death. We have therefore examined this process in bovine healthy and atretic antral follicles, using a variety of established techniques. Light and electron microscopic observations indicated the presence of pyknotic or shrunken nuclei in both the membrana granulosa and the antrum. In the membrana granulosa, these nuclei were frequently crescent shaped and uniformly electron dense and were approximately the same size as healthy nuclei, all of which are typical of early apoptosis. However, these nuclei were within the membranes of a healthy granulosa cell, suggesting that phagocytosis by a neighboring granulosa cell is an unusually early event in the apoptotic pathway of granulosa cells. In the membrana granulosa, pyknotic nuclei stained intensely with hematoxylin but weakly with the DNA-intercalating stain propidium iodide. A percentage of these pyknotic nuclei stained by TUNEL (terminal deoxy-UTP nick end-labeling). However, in the antrum, the pyknotic nuclei and larger globules of DNA stained intensely with both hematoxylin and propidium iodide, but were not TUNEL positive. The comet assay of cell death produced a streak tail of randomly nicked DNA, rather than the plume of low mol wt apoptotic DNA. Globules collected from fresh follicular fluid stained intensely with propidium iodide and were shown by PAGE to contain DNA, the majority of which was high mol wt. In conclusion, granulosa cells within the membrana granulosa die by apoptosis, with phagocytosis by a neighboring cell preceding any potential budding of the nucleus or cell itself. Granulosa cells near the antrum are sloughed off into the antrum, and their death has features more consistent with that of other cell types that undergo death as a result of terminal differentiation.
Distribution of AMPA receptor subunits GluR1-4 in the dorsal vagal complex of the rat: a light and electron microscope immunocytochemical study.

PubMed

Kessler, J P; Baude, A

1999-10-01

The dorsal vagal complex, localized in the dorsomedial medulla, includes the nucleus tractus solitarii (NTS), the dorsal motor nucleus of the vagus nerve (DMN) and the area postrema (AP). The distribution of AMPA-preferring glutamate receptors (AMPA receptors) within this region was investigated using immunohistochemistry and antibodies recognizing either one (GluR1 or GluR4) or two (GluR2 and GluR3) AMPA receptors subunits. The distribution of GluR1 immunoreactivity showed high contrast of staining between strongly and lightly labeled areas. Labeling was intense in the AP and weak in the NTS, except for its medial and dorsalmost parts which exhibited moderate staining. Almost no GluR1 immunoreactivity was found in the DMN. GluR2/3 immunolabeling was present in the entire dorsal vagal complex. This labeling was strong in the AP, the DMN and the medial half of the NTS and moderate in the lateral half of the NTS, except for the interstitial subdivision which exhibited intense staining. Labeling induced by the GluR4 antibody was very weak throughout the dorsal vagal complex. Ultrastructural examination showed that GluR1 and GluR2/3 immunoreactivity was localized in neuronal cell bodies and dendrites. No labeled axon terminal or glial cell body was found. Immunoperoxidase staining in labeled cell bodies and dendrites was associated with intracellular organelles (microtubules, mitochondria, cisternae of the endoplasmic reticulum,.) and/or parts of the plasma membrane. Plasma membrane labeling was often associated with asymmetrical synaptic differentiations. No labeled symmetrical synapse was found using either GluR1 or GluR2/3 antibody. The present results show that AMPA receptors have a widespread distribution in neuronal perikarya and dendrites of the rat dorsal vagal complex. They suggest differences in subunit composition between AMPA receptors localized in the NTS, the DMN and the AP. Ultrastructural data are consistent with the fact that AMPA receptors associated with the plasma membrane are mostly synaptic receptors. However, they also suggest the existence of a large intracellular pool of receptor subunits in neuronal soma and dendrites. Copyright 1999 Wiley-Liss, Inc.
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.
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.
Chronological changes of radiofrequency ablation zone in rabbit liver: an in vivo correlation between gross pathology and histopathology

PubMed Central

Song, Kyoung D; Rhim, Hyunchul; Kang, Tae Wook; Cha, Dong Ik; Yang, Jehoon

2017-01-01

Objective: To examine the gross pathology and histopathology of ablation zones created from radiofrequency (RF) ablation and to correlate their chronological changes. Methods: A total of 48 in vivo ablation zones (16 rabbit livers) were obtained immediately after and also 30 min, 1 h and 2 h after RF ablation and were subjected to haematoxylin and eosin (H&E) staining, nicotinamide adenine dinucleotide (NADH) diaphorase staining, terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) staining. Chronological changes in gross pathology and histopathology were evaluated and correlated with each other. Results: Peripheral red zones on gross pathology correlated with peripheral zones on H&E staining, lightly stained peripheral zones on NADH staining and peripheral positive zones on TUNEL staining. Central white zones on gross pathology correlated with combined central and border zones on H&E staining, central negative zones on NADH staining and combined central-positive and middle-negative zones on TUNEL staining. Boundary visibility between central white and peripheral red zones on gross pathology was significantly higher at 1 and 2 h than immediately after RF ablation. As time increased after RF ablation, visibility of the border zone on H&E staining and the grade of positively stained hepatocytes in the peripheral zone on TUNEL staining increased. Conclusion: Chronological changes in gross pathology of RF ablation zones correlated well with histopathology. The boundary between the central white and peripheral red zones tended to become clear at 1 h after RF ablation. Advances in knowledge: (1) RF ablation zones show chronological changes on gross pathology and histopathology. (2) Gross pathology and histopathology correlate well with each other. PMID:28139942
Comparison of effectiveness of abrasive and enzymatic action of whitening toothpastes in removal of extrinsic stains - a clinical trial.

PubMed

Patil, P A; Ankola, A V; Hebbal, M I; Patil, A C

2015-02-01

To compare the effectiveness of abrasive component (perlite/calcium carbonate) and enzymatic component (papain and bromelain) of whitening toothpaste in removal of extrinsic stains. This study is a randomized, triple blind and parallel group study in which 90 subjects aged 18-40 years were included. At baseline, stains scores were assessed by Macpherson's modification of Lobene Stain Index and subjects were randomly assigned to two groups with 45 subjects in each. Group 1 used whitening toothpaste with enzymatic action and group 2 with abrasive action. After 1 month, stain scores were assessed for the effectiveness of the two toothpastes and 2 months later to check the stain prevention efficacy. Wilcoxson's test was used to compare between baseline 1 and 2 months stain scores, and Mann-Witney U-test was applied for intragroup comparison. The mean baseline total stain score for the subjects allocated to the enzymatic toothpaste was 37.24 ± 2.11 which reduced to 30.77 ± 2.48 in 1 month, and for the abrasive paste, total stain reduced from 35.08 ± 2.96 to 32.89 ± 1.95. The reductions in total stain scores with both the pastes were significant compared with baseline stain scores (at 1 month Group 1, P = 0.0233 and Group 2, P = 0.0324; at 2 months, Group 1 P = 0.0356). Both the toothpastes proved to be equally good in removal of extrinsic stains; however, the enzymatic paste showed better results as compared to abrasive toothpaste. Whitening toothpaste with abrasive action and enzymatic action are equally effective in removal of extrinsic stains; however, whitening toothpaste with abrasive action needs to be used with caution. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Diagnosing periprosthetic infection: false-positive intraoperative Gram stains.

PubMed

Oethinger, Margret; Warner, Debra K; Schindler, Susan A; Kobayashi, Hideo; Bauer, Thomas W

2011-04-01

Intraoperative Gram stains have a reported low sensitivity but high specificity when used to help diagnose periprosthetic infections. In early 2008, we recognized an unexpectedly high frequency of apparent false-positive Gram stains from revision arthroplasties. The purpose of this report is to describe the cause of these false-positive test results. We calculated the sensitivity and specificity of all intraoperative Gram stains submitted from revision arthroplasty cases during a 3-month interval using microbiologic cultures of the same samples as the gold standard. Methods of specimen harvesting, handling, transport, distribution, specimen processing including tissue grinding/macerating, Gram staining, and interpretation were studied. After a test modification, results of specimens were prospectively collected for a second 3-month interval, and the sensitivity and specificity of intraoperative Gram stains were calculated. The retrospective review of 269 Gram stains submitted from revision arthroplasties indicated historic sensitivity and specificity values of 23% and 92%, respectively. Systematic analysis of all steps of the procedure identified Gram-stained but nonviable bacteria in commercial broth reagents used as diluents for maceration of periprosthetic membranes before Gram staining and culture. Polymerase chain reaction and sequencing showed mixed bacterial DNA. Evaluation of 390 specimens after initiating standardized Millipore filtering of diluent fluid revealed a reduced number of positive Gram stains, yielding 9% sensitivity and 99% specificity. Clusters of false-positive Gram stains have been reported in other clinical conditions. They are apparently rare related to diagnosing periprosthetic infections but have severe consequences if used to guide treatment. Even occasional false-positive Gram stains should prompt review of laboratory methods. Our observations implicate dead bacteria in microbiologic reagents as potential sources of false-positive Gram stains.
Clinical utility of an automated instrument for gram staining single slides.

PubMed

Baron, Ellen Jo; Mix, Samantha; Moradi, Wais

2010-06-01

Gram stains of 87 different clinical samples were prepared by the laboratory's conventional methods (automated or manual) and by a new single-slide-type automated staining instrument, GG&B AGS-1000. Gram stains from either heat- or methanol-fixed slides stained with the new instrument were easy to interpret, and results were essentially the same as those from the methanol-fixed slides prepared as a part of the routine workflow. This instrument is well suited to a rapid-response laboratory where Gram stain requests are commonly received on a stat basis.
Differential staining of bacteria: gram stain.

PubMed

Moyes, Rita B; Reynolds, Jackie; Breakwell, Donald P

2009-11-01

In 1884, Hans Christian Gram, a Danish doctor, developed a differential staining technique that is still the cornerstone of bacterial identification and taxonomic division. This multistep, sequential staining protocol separates bacteria into four groups based on cell morphology and cell wall structure: Gram-positive cocci, Gram-negative cocci, Gram-positive rods, and Gram-negative rods. The Gram stain is useful for assessing bacterial contamination of tissue culture samples or for examining the Gram stain status and morphological features of bacteria isolated from mixed or isolated bacterial cultures. (c) 2009 by John Wiley & Sons, Inc.
Identification criteria of the rare multi-flagellate Lophomonas blattarum: comparison of different staining techniques.

PubMed

Alam-Eldin, Yosra Hussein; Abdulaziz, Amany Mamdouh

2015-09-01

Bronchopulmonary lophomoniasis (BPL) is an emerging disease of potential importance. BPL is presented by non-specific clinical picture and is usually accompanied by immunosuppression. Culture of Lophomonas blattarum is difficult and its molecular diagnosis has not yet been developed. Therefore, microscopic examination of respiratory samples, e.g., bronchoalveolar lavage (BAL) or sputum, is the mainstay of BPL diagnosis. Creola bodies and ciliocytophthoria are two forms of bronchial cells which occur in chest diseases with non-specific clinical picture like that of BPL. Both forms could be misrecognized as multi-flagellates because of their motile cilia in the wet mounts and due to shape variability of L. blattarum in stained smears. The aim of the study is to compare different staining techniques for visualizing L. blattarum to improve the recognition and diagnosis of BPL, to distinguish respiratory epithelial cells from L. blattarum and to decide which stain is recommended in suspected cases of BPL. BAL samples from patients which contain L. blattarum, creola bodies, and ciliocytophthoria were collected then wet mounts were examined. The BAL samples were also stained by Papanicolaou (PAP), Giemsa, hematoxylin and eosin (H & E), trichrome, Gram, and Diff-Quik (DQ) stains. The different staining techniques were compared regarding the stain quality. In wet mounts, the ciliary movement was coordinate and synchronous while the flagellar movement was wavy and leaded to active swimming of L. blattarum. In stained slides, bronchial cells were characterized by the presence of basal nucleus and the terminal bar from which the cilia arise. Trichrome was the best stain in demonstration of cellular details of L. blattarum. H & E, PAP, and Giemsa stains showed good quality of stains. Gram and DQ stains showed only pale hues of L. blattarum. We recommended adding Wheatley's trichrome staining to the differential diagnosis workup of cases of non-specific chest infections, especially when BPL is suspected, to avoid overdiagnosis or underdiagnosis of it.
Sudan stain of fecal fat: new insight into an old test.

PubMed

Khouri, M R; Huang, G; Shiau, Y F

1989-02-01

The 72-h fecal fat determination is used as the gold standard to document the presence of steatorrhea. Although the Sudan stain for fecal fat is advocated as a sensitive screening test, a quantitative correlation between the 72-h fecal fat quantitation and the fecal Sudan stain is lacking. This study was designed to examine the staining properties of different classes of purified lipids in an experimentally defined artificial matrix, and to elucidate the reasons for the lack of quantitative correlation between these two tests. Our results indicate that the "neutral fat" stain without acidification or heating identifies triglyceride; and at an appropriate pH, the "neutral stain" also identifies fatty acid. The "split fat" stain with acidification and heating identifies both triglyceride and fatty acid. After acidification, fatty acid soaps are converted to the nonionized fatty acid. Thus, fatty acid soaps can be identified indirectly as fat droplets that are stained by the split fat stain. Although cholesterol is stained with Sudan stain after heating, upon cooling, cholesterol forms crystals of anhydrous cholesterol, making its staining pattern distinct. Neither the neutral fat nor the split fat stain can detect phospholipid or cholesteryl ester. The 72-h fecal fat determination is a measure of the total fatty acid content after a specimen is saponified. The resulting fatty acids are derived from a variety of endogenous and exogenous sources, including free fatty acids, soaps of fatty acids, triglycerides, cholesterol esters, and phospholipids. Therefore, the 72-h fecal fat quantitation does not differentiate between the primary sources of the measured fatty acid. It is concluded that the 72-h fecal fat determination is not specific for documenting triglyceride (fat) malabsorption. Until new methods are developed that specifically measure fecal triglyceride and fatty acid, the Sudan stain of fecal fat appears to be a more specific method for detecting the presence of triglyceride and fatty acid in a matrix.
Gram stain of tissue biopsy

MedlinePlus

... biopsy To use the sharing features on this page, please enable JavaScript. Gram stain of tissue biopsy test involves using crystal violet stain to test a sample of tissue taken from a biopsy . The Gram stain method can ...
DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Deok Hee; Hwang, Jae Cheol; Lim, Soo Mee

Purpose: To describe the findings of pleural and pulmonary staining of the inferior phrenic artery, which can be confused with tumor staining during transarterial chemoembolization (TACE) of hepatoma.Methods: Fifteen patients who showed pleural and pulmonary staining without relationship to hepatic masses at inferior phrenic arteriography were enrolled. The staining was noted at initial TACE (n = 8), at successive TACE (n = 5), and after hepatic surgery (n = 2). The angiographic pattern, the presence of pleural change on computed tomography (CT), and clinical history were evaluated.Results: Draining pulmonary veins were seen in all cases. The lower margin of themore » staining corresponded to the lower margin of the pleura in 10 patients. CT showed pleural and/or pulmonary abnormalities in all cases. After embolization of the inferior phrenic artery, the accumulation of iodized oil in the lung was noted.Conclusion: Understanding the CT and angiographic findings of pleural and pulmonary staining during TACE may help differentiate benign staining from tumor staining.« less
A review of curcumin as a biological stain and as a self-visualizing pharmaceutical agent.

PubMed

Hope-Roberts, M; Horobin, R W

2017-01-01

Curcumin has been widely used to color textiles but, unlike other natural dyes such as hematoxylin or saffron, it rarely has been discussed as a biological stain. Aspects of the physicochemistry of curcumin relevant to biological staining and self-visualization, i.e., its acidic properties, lipophilicity, metal and pseudometal complexes, and optical properties, are summarized briefly here. Reports of staining of non-living biological specimens in sections and smears, both fixed and unfixed, including specimens embedded in resin, are summarized here. Staining of amyloid, boron and chromatin are outlined and possible reaction mechanisms discussed. Use of curcumin as a vital stain also is described, both in cultured monolayers and in whole organisms. Staining mechanisms are considered especially for the selective uptake of curcumin into cancer cells. Staining with curcumin labeled nanoparticles is discussed. Toxicity and safety issues associated with the dye also are presented.
Prototype Slide Stainer

NASA Technical Reports Server (NTRS)

1971-01-01

The prototype slide staining system capable of performing both one-component Wright's staining of blood smears and eight-step Gram staining of heat fixed slides of microorganisms is described. Attention was given to liquid containment, waste handling, absence of contamination from previous staining, and stability of the staining reagents. The unit is self-contained, capable of independent operation under one- or zero-g conditions, and compatible with Skylab A.
Comparison of staining of mitotic figures by haematoxylin and eosin-and crystal violet stains, in oral epithelial dysplasia and squamous cell carcinoma.

PubMed

Ankle, Madhuri R; Kale, Alka D; Charantimath, Seema

2007-01-01

Mitosis of cells gives rise to tissue integrity. Defects during mitosis bring about abnormalities. Excessive proliferation of cells due to increased mitosis is one such outcome, which is the hallmark in precancer and cancer. The localization of proliferating cells or their precursors may not be obvious and easy. Establishing an easy way to distinguish these mitotic cells will help in grading and understanding their biological potential. Although immunohistochemistry is an advanced method in use, the cost and time factor makes it less feasible for many laboratories. Selective histochemical stains like toluidine blue, giemsa and crystal violet have been used in tissues including the developing brain, neural tissue and skin. 1) To compare the staining of mitotic cells in haematoxylin and eosin with that in crystal violet. 2) To compare the number of mitotic figures present in normal oral mucosa, epithelial dysplasia and oral squamous cell carcinoma in crystal violet-stained sections with that in H and E-stained sections. Ten tissues of normal oral mucosa and 15 tissues each of oral epithelial dysplasia seen in tobacco-associated leukoplakia and squamous cell carcinoma were studied to evaluate the selectivity of 1% crystal violet for mitotic figures. The staining was compared with standard H and E staining. Statistical analysis was done using Mann-Whitney U test. A statistically significant increase in the mean mitotic count was observed in crystal violet-stained sections of epithelial dysplasia as compared to the H and E-stained sections (p=0.0327). A similar increase in the mitotic counts was noted in crystal violet-stained sections of oral squamous cell carcinoma as compared to the H and E-stained sections.(p=0.0443). No significant difference was found in the mitotic counts determined in dysplasia or carcinoma by either the crystal violet (p=0.4429) or the H and E-staining techniques (p=0.2717). One per cent crystal violet provides a definite advantage over the H and E-stained sections in selectively staining the mitotic figures.

Primary epidural hemangiopericytoma in the lumbar spine: a case report.

PubMed

Ijiri, Kosei; Yuasa, Shinya; Yone, Kazunori; Matsunaga, Shunji; Ryoki, Yoshihiro; Taniguchi, Noboru; Yonezawa, Suguru; Komiya, Setsuro

2002-04-01

A case report of primary epidural hemangiopericytoma in the lumbar spine and a review of the literature are presented. To present the result of pathologic diagnosis using immunohistochemical staining and the treatment of spinal hemangiopericytoma. Spinal hemangiopericytoma is a very rare soft tissue tumor with specific pathologic features and a clinical course featuring high rates of recurrence and metastasis. A 39-year-old woman reported numbness in both legs. Neither sensory abnormalities nor muscle weakness was present in her lower extremities. Magnetic resonance imaging showed a tumor dorsal to the thecal sac at L1-L2. After L1 and L2 laminectomy, the tumor with its dural base was resected en bloc. The patient's clinical and neurologic symptoms disappeared after surgery. Microscopic examination showed oval- or spindle-shaped cells with slightly acidic cytoplasm and oval nuclei. Silver staining emphasized fibers around tumor cells. The test results for the tumor cells were positive for vimentin staining, but negative for alpha-TM staining using thrombomodulin, a marker for endothelial cells. On the basis of these pathologic findings, the tumor was diagnosed as a hemangiopericytoma, a type of tumor composed of mesenchymal hemangiopericytes. Neither recurrence nor metastasis of the tumor was found during the 2-year follow-up period after surgery. Soft tissue hemangiopericytoma is a well-recognized entity considered to be an aggressive neoplasm with a high rate of recurrence and a propensity to metastasize. Immunohistochemical investigation was essential for the diagnosis of this tumor. Although hemangiopericytoma very rarely occurs in the spine, surgeons treating patients with this tumor should be aware of its metastatic potential.
Muscle-specific kinase (MuSK) autoantibodies suppress the MuSK pathway and ACh receptor retention at the mouse neuromuscular junction

PubMed Central

Ghazanfari, Nazanin; Morsch, Marco; Reddel, Stephen W; Liang, Simon X; Phillips, William D

2014-01-01

Muscle-specific kinase (MuSK) autoantibodies from myasthenia gravis patients can block the activation of MuSK in vitro and/or reduce the postsynaptic localization of MuSK. Here we use a mouse model to examine the effects of MuSK autoantibodies upon some key components of the postsynaptic MuSK pathway and upon the regulation of junctional ACh receptor (AChR) numbers. Mice became weak after 14 daily injections of anti-MuSK-positive patient IgG. The intensity and area of AChR staining at the motor endplate was markedly reduced. Pulse-labelling of AChRs revealed an accelerated loss of pre-existing AChRs from postsynaptic AChR clusters without a compensatory increase in incorporation of (newly synthesized) replacement AChRs. Large, postsynaptic AChR clusters were replaced by a constellation of tiny AChR microaggregates. Puncta of AChR staining also appeared in the cytoplasm beneath the endplate. Endplate staining for MuSK, activated Src, rapsyn and AChR were all reduced in intensity. In the tibialis anterior muscle there was also evidence that phosphorylation of the AChR β-subunit-Y390 was reduced at endplates. In contrast, endplate staining for β-dystroglycan (through which rapsyn couples AChR to the synaptic basement membrane) remained intense. The results suggest that anti-MuSK IgG suppresses the endplate density of MuSK, thereby down-regulating MuSK signalling activity and the retention of junctional AChRs locally within the postsynaptic membrane scaffold. PMID:24860174
Characterization of Uterine Granular Cell Tumors in B6C3F1 Mice: A Histomorphologic, Immunohistochemical, and Ultrastructural Study

PubMed Central

Veit, A. C.; Painter, J. T.; Miller, R. A.; Hardisty, J. F.; Dixon, D.

2009-01-01

The granular cell tumor is most often a benign neoplasm of uncertain origin. Four uterine granular cell tumors in control and treated female B6C3F1 mice were identified in chronic studies at the National Toxicology Program. Two tumors occurred in untreated control animals and 2 in treated animals receiving different compounds. Tissue sections were evaluated histologically and stained with hematoxylin and eosin, periodic acid–Schiff with diastase resistance, Masson’s trichrome, toluidine blue, phosphotungstic acid–hematoxylin, and stained immunohistochemically with a panel of antibodies to muscle (desmin, alpha smooth muscle actin), neural (S-100, neuron specific enolase), epithelial (wide-spectrum cytokeratin), and macrophage (F4/80) markers. The main histomorphologic feature of tumor cells was the presence of abundant cytoplasmic eosinophilic granules that stained positive for periodic acid–Schiff with diastase resistance. Tumors varied in appearance and were comprised of sheets and nests of round to polygonal cells with distinct borders. Nuclei were hyperchromatic, pleomorphic, and centrally to eccentrically located and often contained single nucleoli. Occasional multinucleated giant cells were observed. Tumors were pale pink and homogeneous with trichrome stain and negative with toluidine blue. Three tumors had positive to weakly positive immunoreactivity for desmin, and 1 was positive for alpha smooth muscle actin. Expression of S-100, wide-spectrum cytokeratin, and neuron-specific enolase was negative for all tumors. Ultrastructurally, prominent electron-dense cytoplasmic granules were abundant and contained secondary lysosomes with heterogeneous lysosomal contents. The characteristics of these uterine granular cell tumors were suggestive of a myogenic origin. PMID:18725470
Background-free, high sensitivity staining of proteins in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gels using a luminescent ruthenium complex.

PubMed

Berggren, K; Chernokalskaya, E; Steinberg, T H; Kemper, C; Lopez, M F; Diwu, Z; Haugland, R P; Patton, W F

2000-07-01

SYPRO Ruby dye is a permanent stain comprised of ruthenium as part of an organic complex that interacts noncovalently with proteins. SYPRO Ruby Protein Gel Stain provides a sensitive, gentle, fluorescence-based method for detecting proteins in one-dimensional and two-dimensional sodium dodecyl sulfate-polyacrylamide gels. Proteins are fixed, stained from 3h to overnight and then rinsed in deionized water or dilute methanol/acetic acid solution for 30 min. The stain can be visualized using a wide range of excitation sources commonly used in image analysis systems including a 302 nm UV-B transilluminator, 473 nm second harmonic generation (SHG) laser, 488 nm argon-ion laser, 532 nm yttrium-aluminum-garnet (YAG) laser, xenon arc lamp, blue fluorescent light bulb or blue light-emitting diode (LED). The sensitivity of SYPRO Ruby Protein Gel Stain is superior to colloidal Coomassie Brilliant Blue (CBB) stain or monobromobimane labeling and comparable with the highest sensitivity silver or zinc-imidazole staining procedures available. The linear dynamic range of SYPRO Ruby Protein Gel stain extends over three orders of magnitude, which is vastly superior to silver, zinc-imidazole, monobromobimane and CBB stain. The fluorescent stain does not contain superfluous chemicals (formaldehyde, glutaraldehyde, Tween-20) that frequently interfere with peptide identification in mass spectrometry. While peptide mass profiles are severely altered in protein samples prelabeled with monobromobimane, successful identification of proteins by peptide mass profiling using matrix-assisted laser desorption/ionization mass spectrometry was easily performed after protein detection with SYPRO Ruby Protein Gel stain.
Contact lens-induced circumlimbal staining in silicone hydrogel contact lenses worn on a daily wear basis.

PubMed

Maïssa, Cécile; Guillon, Michel; Garofalo, Renee J

2012-01-01

The principal objective of the study was to measure the conjunctival staining produced in the circumlimbal region by silicone hydrogel contact lenses with different edge designs. The secondary objective was to investigate the association between circumlimbal staining and comfort. Four silicone hydrogel contact lenses: ACUVUE OASYS (knife edge design), AIR OPTIX, Biofinity (chisel edge rounded edge combination), and PureVision (rounded edge design), and 1 hydrogel contact lens, ACUVUE 2 (knife edge design), were tested. The study was conducted on a cohort population of 27 established soft contact lens wearers, who wore each contact lens type, in a random order, for a period of 10 (±2) days. Circumlimbal staining was measured in a double-masked fashion through image analysis of digital photographs of lissamine green taken under controlled experimental conditions. The results obtained showed that contact lens edge design was the primary factor controlling circumlimbal staining for silicone hydrogel lenses: a rounded edge away from the ocular surface produced the lowest staining (average, 0.19%) and a knife edge in close apposition to the ocular surface produced the highest staining (average, 1.34%). Contact lens material rigidity was also identified to affect circumlimbal staining and an inverse association between circumlimbal staining and contact lens comfort was demonstrated: the rounded edge design produced the lowest comfort (72 of 100) and the knife edge design produced the highest (87 out of 100). Soft contact lens wear induces circumlimbal staining, the level of staining being influenced by the contact lens edge design. However, high level of circumlimbal staining is not associated with decreased comfort.
A comparative study for selectivity of micronuclei in oral exfoliated epithelial cells

PubMed Central

Grover, S; Mujib, ABR; Jahagirdar, A; Telagi, N; Kulkarni, PG

2012-01-01

Background: Micronucleus (MN) represents small, additional nuclei formed by the exclusion of chromosome fragments or whole chromosomes lagging at mitosis. MN rates, therefore, indirectly reflect chromosome breakage or impairment of the mitotic apparatus. During the last few decades, micronuclei (“MNi”) in oral exfoliated epithelial cells are widely used as biomarkers of chromosomal damage, genome instability and cancer risk in humans. However, until now only little attention has been given to the effect of different staining procedures on the results of these MN assays. Aim: To compare the MNi frequencies in oral exfoliated epithelial cells using three different stains, i.e.,Feulgen stain, Papanicolaou stain (Pap) and hemotoxylin and eosin stain (H and E). Materials and Methods: Oral exfoliated cells from 45 cases of potentially malignant disorders (15 oral submucous fibrosis, 15 lichen planus and 15 leukoplakia) and 15 controls with healthy mucosa, were taken and MNi frequencies (No. of MNi/1000 cells) were compared using three different stains. Results: Mean MNi frequency in cases was found to be 3.8 with Feulgen stain, 16.8 with PAP and 25.9 with H and E. In controls, mean MNi frequency was 1.6 with Feulgen stain, 7.7 with PAP and 9.6 with H and E stain. Statistically significant value (P < 0.01) were observed when the three stains were compared together using Kruskal Walli's ANOVA test. Conclusion: Feulgen being a DNA-specific stain gave the least counts, although statistically significant results from the comparison of MNi frequency between cases and controls were obtained with all the three stains. PMID:23326025
A comparative study for selectivity of micronuclei in oral exfoliated epithelial cells.

PubMed

Grover, S; Mujib, Abr; Jahagirdar, A; Telagi, N; Kulkarni, Pg

2012-10-01

Micronucleus (MN) represents small, additional nuclei formed by the exclusion of chromosome fragments or whole chromosomes lagging at mitosis. MN rates, therefore, indirectly reflect chromosome breakage or impairment of the mitotic apparatus. During the last few decades, micronuclei ("MNi") in oral exfoliated epithelial cells are widely used as biomarkers of chromosomal damage, genome instability and cancer risk in humans. However, until now only little attention has been given to the effect of different staining procedures on the results of these MN assays. To compare the MNi frequencies in oral exfoliated epithelial cells using three different stains, i.e.,Feulgen stain, Papanicolaou stain (Pap) and hemotoxylin and eosin stain (H and E). Oral exfoliated cells from 45 cases of potentially malignant disorders (15 oral submucous fibrosis, 15 lichen planus and 15 leukoplakia) and 15 controls with healthy mucosa, were taken and MNi frequencies (No. of MNi/1000 cells) were compared using three different stains. Mean MNi frequency in cases was found to be 3.8 with Feulgen stain, 16.8 with PAP and 25.9 with H and E. In controls, mean MNi frequency was 1.6 with Feulgen stain, 7.7 with PAP and 9.6 with H and E stain. Statistically significant value (P < 0.01) were observed when the three stains were compared together using Kruskal Walli's ANOVA test. Feulgen being a DNA-specific stain gave the least counts, although statistically significant results from the comparison of MNi frequency between cases and controls were obtained with all the three stains.
Standardization of stain used for diagnosing erythrocytic inclusion body syndrome (EIBS)

USGS Publications Warehouse

1987-01-01

Erythrocytic inclusion body syndrome (EIBS), a viral erythrocytic necrosis (VEN)-like disease, has been observed in several areas in the Northwest. This virus disease is clinically diagnosed by microscopic examination of blood smears for intracytoplasmic erythrocytic inclusion bodies. Fish biologists involved in EIBS diagnostic work have been using several types of hematological stains. It became apparent that standardization of the staining procedure was needed. Comparative tests were conducted on blood smears and kidney imprints with the following commonly used blood stains: (1) Leishman-Giesma, (2) Pinacyanol chloride, (3) Powell 's Giemsa, (4) Harleco's Giemsa, (5) Diff Quik differential stain, (6) Wright's.Pinacyanol chloride stain was found to be the most consistent. The following staining procedure is recommended.
Techniques for controlling variability in gram staining of obligate anaerobes.

PubMed Central

Johnson, M J; Thatcher, E; Cox, M E

1995-01-01

Identification of anaerobes recovered from clinical samples is complicated by the fact that certain gram-positive anaerobes routinely stain gram negative; Peptostreptococcus asaccharolyticus, Eubacterium plautii, Clostridium ramosum, Clostridium symbiosum, and Clostridium clostridiiforme are among the nonconformists with regard to conventional Gram-staining procedures. Accurate Gram staining of American Type Culture Collection strains of these anaerobic bacteria is possible by implementing fixing and staining techniques within a gloveless anaerobic chamber. Under anaerobic conditions, gram-positive staining occurred in all test organisms with "quick" fixing techniques with both absolute methanol and formalin. The results support the hypothesis that, when anaerobic bacteria are exposed to oxygen, a breakdown of the physical integrity of the cell wall occurs, introducing Gram stain variability in gram-positive anaerobes. PMID:7538512
Black stains in the mixed dentition: a PCR microbiological study of the etiopathogenic bacteria.

PubMed

Saba, C; Solidani, M; Berlutti, F; Vestri, A; Ottolenghi, L; Polimeni, A

2006-01-01

The aim of this work is to emphasize that particular stains on the third cervical of the buccal and lingual surfaces in mixed dentition, called "black stain." Previous research showed the microbiological etiology of this discoloration by chromogen bacterias. Our study shows bacteria spp involved in stains by means of PCR process and electrophoresis gel on the agarose medium. Sample was formed by 100 subject with black stain and 100 control subjects stain-free. A statistical analysis (SPSS 10.0) using X2 was performed in this study. Porphyromonas gingivalis and Prevotella melaninogenica, were not involved in both in black stain subjects and in the control. On the contrary, Actinomyces could be involved in the pigmentation process.
Comparative stain removal properties of four commercially available denture cleaning products: an in vitro study.

PubMed

Alam, M; Jagger, R; Vowles, R; Moran, J

2011-02-01

Formulations of commercially available denture cleaners vary widely. Unfortunately, comparative data to suggest which products are the most effective can become invalid as newer products are introduced or formulations are changed. The aim of this in vitro study was to measure the stain removal properties of four currently available denture cleaners. Stain was deposited on multi-well polystyrene saliva coated microplates using multiple chlorhexidine and tea solutions. Following drying, each stained well was exposed to a solution of denture cleaner, dried again and the amount of stain remaining measured using a microplate reader. The cleaning procedure was repeated with further multiple exposures of the wells to solutions of the denture cleaners. All denture cleaners removed stain better than water used as a control. At five cleaning cycles only one of the cleaners (Superdrug Cleaning Powder) had removed 100% of the stain. At 30 cycles three of the cleaners had removed 100% of the stain. All the commercial denture cleaners removed stain. Superdrug Cleaning Powder, which contains sodium percarbonate and sodium lauryl sulphate, was particularly effective. © 2010 John Wiley & Sons A/S.
[Automated analysis of bacterial preparations manufactured on automatic heat fixation and staining equipment].

PubMed

2012-01-01

Heat fixation of preparations was made in the fixation bath designed by EMKO (Russia). Programmable "Emkosteiner" (EMKO, Russia) was used for trial staining. Reagents set Micko-GRAM-NITsF was applied for Gram's method of staining. It was demostrated that automatic smear fixation equipment and programmable staining ensure high-quality imaging (1% chromaticity variation) good enough for standardization of Gram's staining of microbial preparations.
Neural Plasticity and Neurorehabilitation Following Traumatic Brain Injury

DTIC Science & Technology

2010-10-01

for sectioning and staining . To date, the brains have been sectioned and one set stained for Nissl . Using the Nissl stained sections, Dorothy...all behavioral data. • Brains have been harvested and sent to Dr. Jones’ lab • Dr. Jones’ lab has sliced the brains and stained one set with Nissl ...remaining sets of brain sections are currently being stained with markers of plasticity using immunohistochemistry. We have completed immunohistochemical
Mass spectrometry-compatible silver staining of histones resolved on acetic acid-urea-Triton PAGE.

PubMed

Pramod, Khare Satyajeet; Bharat, Khade; Sanjay, Gupta

2009-05-01

Acetic acid-Urea-Triton (AUT) PAGE is commonly used method to separate histone variants and their post-translationally modified forms. Coomassie staining is the preferred method for protein visualization; however, its sensitivity is less than that of silver staining. Though silver staining of histones in AUT-PAGE has been reported, the method is time-consuming, dependent on prior staining by Amido black and has not been reported suitable for mass spectrometry. Here, we propose 'SDS-Silver' method for rapid, sensitive and mass spectrometry-compatible staining of histones resolved on AUT-PAGE.
Clinical Utility of an Automated Instrument for Gram Staining Single Slides ▿

PubMed Central

Baron, Ellen Jo; Mix, Samantha; Moradi, Wais

2010-01-01

Gram stains of 87 different clinical samples were prepared by the laboratory's conventional methods (automated or manual) and by a new single-slide-type automated staining instrument, GG&B AGS-1000. Gram stains from either heat- or methanol-fixed slides stained with the new instrument were easy to interpret, and results were essentially the same as those from the methanol-fixed slides prepared as a part of the routine workflow. This instrument is well suited to a rapid-response laboratory where Gram stain requests are commonly received on a stat basis. PMID:20410348
Image analysis assisted study of mitotic figures in oral epithelial dysplasia and squamous cell carcinoma using differential stains.

PubMed

Tandon, Ankita; Singh, Narendra Nath; Brave, V R; Sreedhar, Gadiputi

2016-11-01

Mitosis is a process of cell division resulting in two genetically equivalent daughter cells. Excessive proliferation of cells due to mitosis is the hallmark in pre cancer and cancer. This study was conducted to count the number of mitotic figures in normal oral mucosa, oral epithelial dysplasia and squamous cell carcinoma in both Hematoxylin and Eosin (H&E) and Crystal Violet stained sections. Also the overall number of mitotic figures with both stains were compared along with the evaluation of staining efficacy of both the stains. The present study was conducted on 20 specimens each of the three categories. These were further divided into two groups for staining with H&E and with 1% Crystal Violet respectively. Images were captured and analyzed using image analysis software Dewinter Biowizard 4.1. Comparison of mitotic figure count in three categories in sections stained with both stains showed statistically significant difference ( p < 0.001). The mean number of mitotic figures seen in Crystal Violet reagent were significantly higher as seen in H&E stain ( p < 0.001). The overall diagnostic efficacy of Crystal Violet was 87.6%. Crystal Violet scored over H&E stain and also helped to better appreciate metaphases in Squamous cell carcinoma and telophases in dysplasia. Number of mitotic figures progressively increase with the advancement of the pathology. Use of 1% Crystal Violet provides better appreciation of mitotic figures and can be employed as a selective stain in routine histopathology.
Single step modified ink staining for Tzanck test: quick detection of herpetic giant cells in Tzanck smear.

PubMed

Mizutani, Hitoshi; Akeda, Tomoko; Yamanaka, Kei-Ichi; Isoda, Kenichi; Gabazza, Esteban C

2012-02-01

Tzanck test has been recently re-evaluated as a method for the diagnosis of herpes virus infection. Giemsa staining for the Tzanck test is time-consuming and laborious. There is a need to develop simple and quick staining methods for bedside diagnosis of this disease. We report a single step and quick method for staining herpes giant cells in Tzanck smears using routinely available inks and physiological saline. A keratinocyte cell line (HaCaT) was cultured on a slide glass and stained with various commercially available blue, blue-black and black inks serially diluted with physiological saline. Clinical smear samples from herpes lesions were also stained with these solutions without specific pretreatment. The nuclei of HaCaT were clearly stained showing high contrast with the cytoplasm using 5% Parker-Quink blue-black ink saline solution. Concentration of ink solution higher or lower than 5% resulted in less contrast. Blue or black inks or other manufacturers' inks can also be used, but staining of the cultured keratinocytes was less clear. Smear of clinical samples from herpes lesions were also stained with 5% ink solution. The nuclei of the multinucleated giant cells were clearly stained, and the sample could be immediately used for microscopic examination. One step staining of Tzanck smear using this diluted ink solution is an inexpensive and a convenient bedside diagnostic tool for the dermatologist. © 2011 Japanese Dermatological Association.
Sperm viability assessment in marine invertebrates by fluorescent staining and spectrofluorimetry: A promising tool for assessing marine pollution impact.

PubMed

Gallo, Alessandra; Boni, Raffaele; Tosti, Elisabetta

2018-01-01

The viability of spermatozoa is a crucial parameter to evaluate their quality that is an important issue in ecotoxicological studies. Here, a new method has been developed to rapidly determine the viability of spermatozoa in three marine invertebrates: the ascidian Ciona intestinalis, the sea urchin Paracentrotus lividus and the mollusc Mytilus galloprovincialis. This method employed the dual DNA fluorescent staining coupled with spectrofluorimetric analysis. The dual fluorescent staining used the SYBR-14 stained live spermatozoa and propidium iodide stained degenerated cells that had lost membrane integrity. Stain uptake was assessed by confocal microscopy and then the percentage of live and dead spermatozoa was quantified by spectrofluorimetric analysis. The microscopic examination revealed three populations of spermatozoa: living-SYBR-14 stained, dead-PI stained, and dying-doubly stained spermatozoa. The fluorescence emission peak values recorded in a spectrofluorimeter provide the portion of live and dead spermatozoa showing a significant negative correlation. The stain combination was further validated using known ratios of live and dead spermatozoa. The present study demonstrated that the dual DNA staining with SYBR-14 and propidium iodide was effective in assessing viability of spermatozoa in marine invertebrates and that spectrofluorimetric analysis can be successfully employed to evaluate the percentage of live and dead spermatozoa. The method develop herein is simple, accurate, rapid, sensitive, and cost-effective, so it could be a useful tool by which marine pollutants may be screened for spermiotoxicity. Copyright © 2017 Elsevier Inc. All rights reserved.
Control of brown stain: in Eastern white pine

Treesearch

Robert E. Stutz; Peter Koch; Millard L. Oldham

1961-01-01

Degrade caused by brown stain and blue stain in eastern white pine was virtually eliminated by the use of sap stain chemicals and sodium azide. Combinations of buffered sodium azide with both sodium pentachlorophenate plus borax and buffered ethyl mercury phosphate were effective.
Unsupervised color normalisation for H and E stained histopathology image analysis

NASA Astrophysics Data System (ADS)

Celis, Raúl; Romero, Eduardo

2015-12-01

In histology, each dye component attempts to specifically characterise different microscopic structures. In the case of the Hematoxylin-Eosin (H&E) stain, universally used for routine examination, quantitative analysis may often require the inspection of different morphological signatures related mainly to nuclei patterns, but also to stroma distribution. Nevertheless, computer systems for automatic diagnosis are often fraught by color variations ranging from the capturing device to the laboratory specific staining protocol and stains. This paper presents a novel colour normalisation method for H&E stained histopathology images. This method is based upon the opponent process theory and blindly estimates the best color basis for the Hematoxylin and Eosin stains without relying on prior knowledge. Stain Normalisation and Color Separation are transversal to any Framework of Histopathology Image Analysis.

COMPARISON OF PERMANENT STAINING METHODS FOR THE LABORATORY DIAGNOSIS OF TRICHOMONIASIS

PubMed Central

MENEZES, Camila Braz; MELLO, Mariana dos Santos; TASCA, Tiana

2016-01-01

Trichomonas vaginalis is the etiologic agent of trichomoniasis, the most common non-viral sexually transmitted disease (STD) in the world. The diagnosis is based on wet mount preparation and direct microscopy on fixed and stained clinical specimens. The aim of this study was to compare the performance of different fixing and staining techniques used in the detection of T. vaginalis in urine. The smears were fixed and submitted to different methods of permanent staining and then, the morphological aspects of the parasites were analyzed and compared. The Papanicolaou staining with ethanol as the fixative solution showed to be the best method of permanent staining. Our data suggest that staining techniques in association with wet mount examination of fresh specimens contribute to increase the sensitivity in the diagnosis of trichomoniasis. PMID:26910452
Double-Staining Method for Differentiation of Morphological Changes and Membrane Integrity of Campylobacter coli Cells

PubMed Central

Alonso, Jose L.; Mascellaro, Salvatore; Moreno, Yolanda; Ferrús, María A.; Hernández, Javier

2002-01-01

We developed a double-staining procedure involving NanoOrange dye (Molecular Probes, Eugene, Oreg.) and membrane integrity stains (LIVE/DEAD BacLight kit; Molecular Probes) to show the morphological and membrane integrity changes of Campylobacter coli cells during growth. The conversion from a spiral to a coccoid morphology via intermediary forms and the membrane integrity changes of the C. coli cells can be detected with the double-staining procedure. Our data indicate that young or actively growing cells are mainly spiral shaped (green-stained cells), but older cells undergo a degenerative change to coccoid forms (red-stained cells). Club-shaped transition cell forms were observed with NanoOrange stain. Chlorinated drinking water affected the viability but not the morphology of C. coli cells. PMID:12324366
Monoclonal antibody against human ovarian tumor-associated antigens.

PubMed

Poels, L G; Peters, D; van Megen, Y; Vooijs, G P; Verheyen, R N; Willemen, A; van Niekerk, C C; Jap, P H; Mungyer, G; Kenemans, P

1986-05-01

Mouse monoclonal antibodies (OV-TL 3) were raised against human ovarian tumor-associated antigens for diagnostic purposes. A cloned hybridoma cell line was obtained by fusion of murine myeloma cells with spleen lymphocytes from BALB/c mice immunized with a tumor cell suspension prepared from an ovarian endometrioid carcinoma. The antibodies were initially screened for their ability to bind on frozen sections of human ovarian carcinoma tissue and a negative reaction on gastric carcinoma tissue by indirect immunofluorescence. The reactivity of the selected OV-TL 3 clone (IgG1 subclass) was studied on normal and neoplastic tissues as well as on a cell line derived from the original tumor cell suspension used for immunization. OV-TL 3 antibodies stained frozen sections of human ovarian carcinomas of the following histological types: serous, mucinous, endometrioid, and clear cell. No reaction was found with breast cancers or other nongynecological tumors. No differences in staining pattern were observed between primary and metastatic ovarian carcinomas. OV-TL 3 antibodies brightly stained ovarian carcinoma cell clusters in ascitic fluids and left unstained mesothelial cells and peripheral blood cells. The OV-TL 3-defined antigen also remained strongly expressed on a cell line derived from the endometrioid ovarian carcinoma originally used for generation of OV-TL 3 clone. Reactivity was weak and irregular in a few ovarian cysts, while traces of fluorescence were sometimes detected in epithelial cells lining the female genital tract. In only 3 specimens of 15 endometrium carcinomas was weak focal reactivity with OV-TL 3 antibodies observed. The results of the immunofluorescence study were confirmed by the more sensitive avidin-biotin method and by 125I-labeled OV-TL 3 antibodies. Thus OV-TL 3 recognizes a common antigen for most ovarian carcinomas and may be a useful tool for rapid diagnosis of ovarian carcinomas.
DOE Office of Scientific and Technical Information (OSTI.GOV)

Poels, L.G.; Peters, D.; van Megen, Y.

Mouse monoclonal antibodies (OV-TL 3) were raised against human ovarian tumor-associated antigens for diagnostic purposes. A cloned hybridoma cell line was obtained by fusion of murine myeloma cells with spleen lymphocytes from BALB/c mice immunized with a tumor cell suspension prepared from an ovarian endometrioid carcinoma. The antibodies were initially screened for their ability to bind on frozen sections of human ovarian carcinoma tissue and a negative reaction on gastric carcinoma tissue by indirect immunofluorescence. The reactivity of the selected OV-TL 3 clone (IgG1 subclass) was studied on normal and neoplastic tissues as well as on a cell line derivedmore » from the original tumor cell suspension used for immunization. OV-TL 3 antibodies stained frozen sections of human ovarian carcinomas of the following histological types: serous, mucinous, endometrioid, and clear cell. No reaction was found with breast cancers or other nongynecological tumors. No differences in staining pattern were observed between primary and metastatic ovarian carcinomas. OV-TL 3 antibodies brightly stained ovarian carcinoma cell clusters in ascitic fluids and left unstained mesothelial cells and peripheral blood cells. The OV-TL 3-defined antigen also remained strongly expressed on a cell line derived from the endometrioid ovarian carcinoma originally used for generation of OV-TL 3 clone. Reactivity was weak and irregular in a few ovarian cysts, while traces of fluorescence were sometimes detected in epithelial cells lining the female genital tract. In only 3 specimens of 15 endometrium carcinomas was weak focal reactivity with OV-TL 3 antibodies observed. The results of the immunofluorescence study were confirmed by the more sensitive avidin-biotin method and by /sup 125/I-labeled OV-TL 3 antibodies.« less
Tissue preparation for immunocytochemistry.

PubMed Central

Williams, J H; Mepham, B L; Wright, D H

1997-01-01

AIMS: To investigate the effect of tissue preparation on immunostaining and to establish whether there is a standard tissue preparation schedule that allows optimal demonstration of all antigens. METHODS: Blocks of tonsil were subjected to variations to a standard fixation, processing, and section preparation schedule. The sections were stained with five antibodies-L26 (CD20), UCHL1 (CD45RO), CD3, vimentin, and anti-kappa light chain--using the streptavidinbiotin immunostaining technique. When further investigation was necessary, other tissues and antibodies were used and where weak immunostaining was obtained the use of microwave pretreatment to improve staining was tested. RESULTS: Several factors involved in fixation were found to affect immunoreactivity. These included the duration, pH, and type of fixative used. In tissue processing only temperature and the duration of the dehydration and wax infiltration steps affected immunoreactivity. Of all the factors investigated, the temperature and duration of the section drying had the greatest effect. In contrast, long term storage of cut sections before immunostaining had no effect on the reactivity of the antibodies tested. Antibodies were found to be affected by alterations to tissue preparation by varying degrees, UCHL1 and vimentin being the most susceptible to changes in fixation and L26 to changes in processing. Where weak staining occurred, microwave pretreatment was generally found to eliminate the problem. CONCLUSIONS: There is no standard tissue preparation schedule for the optimal demonstration of all antigens. Factors involved in all aspects of tissue preparation can affect immunoreactivity, so it is important that precise details of the preparation schedule are given when reporting immunocytochemical studies, rather than using the general term "routinely fixed and processed". Images PMID:9215127
Platinum blue as an alternative to uranyl acetate for staining in transmission electron microscopy.

PubMed

Inaga, Sumire; Katsumoto, Tetsuo; Tanaka, Keiichi; Kameie, Toshio; Nakane, Hironobu; Naguro, Tomonori

2007-04-01

This paper introduces an aqueous solution of platinum blue (Pt-blue) as an alternative to uranyl acetate (UA) for staining in transmission electron microscopy (TEM). Pt-blue was prepared from a reaction of cis-dichlorodiamine-platinum (II) (cis-platin) with thymidine. When Pt-blue was dried on a microgrid and observed by TEM it showed a uniform appearance with tiny particles less than 1 nm in diameter. The effect of Pt-blue as an electron stain was then examined not only for positive staining of conventional ultrathin resin sections and counterstaining of post-embedding immuno-electron microscopy but also for negative staining. In ultrathin sections of the rat liver and renal glomerulus, Pt-blue provided good contrast images, especially in double staining combined with a lead stain (Pb). Almost all cell organelles were clearly observed with high contrast in these sections. Glycogen granules in the hepatic parenchymal cells were particularly electron dense in Pt-blue stained sections compared with those treated with UA. In longitudinal and transverse sections of budding influenza A viruses, a specific arrangement of rod-like structures, which correspond to the ribonucleoprotein complexes, was clearly shown in each virion stained with Pt-blue and Pb. When post-embedding immunoelectron microscopy was performed in ultrathin sections of HeLa cells embedded in Lowicryl K4M, the localization of Ki-67 protein was sufficiently detected even after Pt-blue and Pb staining. The present study also revealed that Pt-blue could be used for the negative staining of E. coli, allowing the visualization of a flagellum. These findings indicate that Pt-blue is a useful, safe, and easily obtainable electron stain that is an alternative to UA for TEM preparations.
A double blind randomised controlled clinical trial comparing a novel anti-stain and calculus reducing dentifrice with a standard fluoride dentifrice.

PubMed

Jowett, Adrian K; Marlow, Ian; Rawlinson, Andrew

2013-04-01

This clinical trial tested the anti-stain efficacy at 3 and 6 months of a novel, sodium polyaspartate-containing, anti-stain dentifrice. In addition, the efficacy of the new dentifrice in controlling gingival inflammation and inhibition of calculus deposition was tested. Participants were recruited to this double blind randomised control clinical trial, and allocated to either test or control groups. The presence of stain and calculus were entry criteria. Measurements of stain, calculus and gingival inflammation were recorded using the Shaw and Murray Stain score, Volpe-Manhold Calculus score and the Modified Gingival Index respectively. Measurements were made at baseline, prior to the removal of stain and calculus, and after 3 and 6 months. Missing data were imputed by and the outcomes were analysed using univariate analysis. At three months, toothpaste containing sodium polyaspartate was better (difference of mean 1.13 with SEM 0.57) than control for the control of dental stain (p<0.05). Stain scores also showed a trend in favour of the test product (difference of mean 1.03 with SEM 0.78) at six months (p>0.05). There was no difference between toothpastes with respect to calculus deposition or gingival inflammation. Toothpaste containing sodium polyaspartate was more effective than a control toothpaste at preventing deposition of dental stain for 3 months after professional tooth cleaning but showed no significant effect at 6 months. Sodium polyaspartate toothpaste was more effective than a control toothpaste at preventing dental stain formation and maybe helpful in controlling staining between episodes of scaling and polishing. Copyright © 2012 Elsevier Ltd. All rights reserved.
[Comparison of four different staining methods for ear cytology of dogs with otitis externa].

PubMed

Bouassiba, C; Osthold, W; Mueller, R S

2013-01-01

Cytological examination is crucial for the diagnosis and classification of canine otitis externa. Staining should reveal micro-organisms as perpetuating factors of otitis externa. The aim of the study was to compare four different staining methods (Diff-Quik®, Diff-Quik® after dipping in acetone, Gram Quick stain® and a commercial rapid stain for otitis externa) for ear cytology of dogs with otitis externa and to investigate the agreement of cytology and culture. In a study evaluating dogs with otitis externa, five ear swabs (one for culture and four for cytology) were taken from the horizontal part of the external auditory canal of 224 affected ears and compared semi-quantitatively. Diff-Quik® with and without prior dipping in acetone as well as the Gram Quick stain® displayed a high degree of agreement in the detection of micro-organisms (cocci p = 0.2366; rods p = 0.4832; yeasts p = 0.1574), while the commercial otitis rapid stain revealed significantly less micro-organisms (p < 0.001 for all comparisons). The results of the first three stains corresponded to the culture results by > 70%; the agreement was lower with the commercial otitis rapid stain. The quickest and easiest method was staining with Diff-Quik®. Diff-Quik® with or without prior dipping in acetone and the Gram Quick stain® had a high agreement in the detection of microorganisms and can thus be considered nearly equivalent for the diagnosis of otitis externa infectiosa. The commercial otitis rapid stain is less reliable. Based on this study Diff-Quik® can be recommended for the routine cytology of ear swabs. Additionally, a culture may be indicated and must be interpreted in the context of the cytology.
A comparative study to evaluate liquid dish washing soap as an alternative to xylene and alcohol in deparaffinization and hematoxylin and eosin staining.

PubMed

Pandey, Pinki; Dixit, Alok; Tanwar, Aparna; Sharma, Anuradha; Mittal, Sanjeev

2014-07-01

Our study presents a new deparaffinizing and hematoxylin and eosin (H and E) staining method that involves the use of easily available, nontoxic and eco-friendly liquid diluted dish washing soap (DWS) by completely eliminating expensive and hazardous xylene and alcohol from deparaffinizing and rehydration prior to staining, staining and from dehydration prior to mounting. The aim was to evaluate and compare the quality of liquid DWS treated xylene and alcohol free (XAF) sections with that of the conventional H and E sections. A total of 100 paraffin embedded tissue blocks from different tissues were included. From each tissue block, one section was stained with conventional H and E (normal sections) and the other with XAF H and E (soapy sections) staining method. Slides were scored using five parameters: Nuclear, cytoplasmic, clarity, uniformity, and crispness of staining. Z-test was used for statistical analysis. Soapy sections scored better for cytoplasmic (90%) and crisp staining (95%) with a statistically significant difference. Whereas for uniformity of staining, normal sections (88%) scored over soapy sections (72%) (Z = 2.82, P < 0.05). For nuclear (90%) and clarity of staining (90%) total scored favored soapy sections, but the difference was not statistically significant. About 84% normal sections stained adequately for diagnosis when compared with 86% in soapy sections (Z = 0.396, P > 0.05). Liquid DWS is a safe and efficient alternative to xylene and alcohol in deparaffinization and routine H and E staining procedure. We are documenting this project that can be used as a model for other histology laboratories.
A comparative study to evaluate liquid dish washing soap as an alternative to xylene and alcohol in deparaffinization and hematoxylin and eosin staining

PubMed Central

Pandey, Pinki; Dixit, Alok; Tanwar, Aparna; Sharma, Anuradha; Mittal, Sanjeev

2014-01-01

Introduction: Our study presents a new deparaffinizing and hematoxylin and eosin (H and E) staining method that involves the use of easily available, nontoxic and eco-friendly liquid diluted dish washing soap (DWS) by completely eliminating expensive and hazardous xylene and alcohol from deparaffinizing and rehydration prior to staining, staining and from dehydration prior to mounting. The aim was to evaluate and compare the quality of liquid DWS treated xylene and alcohol free (XAF) sections with that of the conventional H and E sections. Materials and Methods: A total of 100 paraffin embedded tissue blocks from different tissues were included. From each tissue block, one section was stained with conventional H and E (normal sections) and the other with XAF H and E (soapy sections) staining method. Slides were scored using five parameters: Nuclear, cytoplasmic, clarity, uniformity, and crispness of staining. Z-test was used for statistical analysis. Results: Soapy sections scored better for cytoplasmic (90%) and crisp staining (95%) with a statistically significant difference. Whereas for uniformity of staining, normal sections (88%) scored over soapy sections (72%) (Z = 2.82, P < 0.05). For nuclear (90%) and clarity of staining (90%) total scored favored soapy sections, but the difference was not statistically significant. About 84% normal sections stained adequately for diagnosis when compared with 86% in soapy sections (Z = 0.396, P > 0.05). Conclusion: Liquid DWS is a safe and efficient alternative to xylene and alcohol in deparaffinization and routine H and E staining procedure. We are documenting this project that can be used as a model for other histology laboratories. PMID:25328332
Thermal Inactivation of Bacillus Anthracis using Laser Irradiation of Micro-Etched Platforms

DTIC Science & Technology

2009-03-01

Application of Malachite Green Staining protocol for greater clarity of spore population; fluorescent staining techniques. Further assess toxicity or...to normal staining procedures. The primary stain in the endospore stain procedure, malachite green, is driven into the cells with heat. Malachite ...dimethyl-aniline is a toxic chemical primarily used as a dye. Since malachite green is water-soluble and does not adhere well to the cell, and since the
Differential fluorescent staining method for detection of bacteria in blood cultures, cerebrospinal fluid and other clinical specimens.

PubMed

Fazii, P; Ciancaglini, E; Riario Sforza, G

2002-05-01

The aim of this study was to evaluate a differential staining method to distinguish gram-positive from gram-negative bacteria in fluorescence. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein. With this method, gram-positive bacteria appear yellow and gram-negative bacteria appear green. In view of the importance of a rapid aetiological diagnosis in cases of septicaemia, the differential staining method in fluorescence was compared with Gram stain for the detection of bacteria in blood. Of 5,820 blood cultures entered into the study and identified by the Bactec 9120 fluorescent series instrument (Becton Dickinson Europe, France), 774 were positive. Of the 774 positive cultures, 689 yielded only a single organism. The differential staining method in fluorescence detected 626 of the 689 cultures, while Gram stain detected 468. On the basis of these results, the sensitivity of the differential staining method in fluorescence was 90.9%, while that of Gram stain was 67.9%. The difference between the two methods was statistically significant ( P<0.001). The differential fluorescent staining method was more sensitive than Gram stain in the detection of bacteria in blood cultures during the incubation period. This technique provides a rapid, simple and highly sensitive staining method that can be used in conjunction with subculture methods. Whereas subculture requires an incubation period of 18-24 h, the fluorescent staining technique can detect bacteria on the same day that smears are prepared and examined. The differential fluorescent staining method was also evaluated for its ability to detect microorganisms in cerebrospinal fluid and other clinical specimens. The microorganisms were easily detected, even when bacterial counts in the specimens were low.
Near-UV laser treatment of extrinsic dental enamel stains.

PubMed

Schoenly, J E; Seka, W; Featherstone, J D B; Rechmann, P

2012-04-01

The selective ablation of extrinsic dental enamel stains using a 400-nm laser is evaluated at several fluences for completely removing stains with minimal damage to the underlying enamel. A frequency-doubled Ti:sapphire laser (400-nm wavelength, 60-nanosecond pulse duration, 10-Hz repetition rate) was used to treat 10 extracted human teeth with extrinsic enamel staining. Each tooth was irradiated perpendicular to the surface in a back-and-forth motion over a 1-mm length using an ∼300-µm-diam 10th-order super-Gaussian beam with fluences ranging from 0.8 to 6.4 J/cm(2) . Laser triangulation determined stain depth and volume removed by measuring 3D surface images before and after irradiation. Scanning electron microscopy evaluated the surface roughness of enamel following stain removal. Fluorescence spectroscopy measured spectra of unbleached and photobleached stains in the spectral range of 600-800 nm. Extrinsic enamel stains are removed with laser fluences between 0.8 and 6.4 J/cm(2) . Stains removed on sound enamel leave behind a smooth enamel surface. Stain removal in areas with signs of earlier cariogenic acid attacks resulted in isolated and randomly located laser-induced, 50-µm-diam enamel pits. These pits contain 0.5-µm diam, smooth craters indicative of heat transfer from the stain to the enamel and subsequent melting and water droplet ejection. Ablation stalling of enamel stains is typically observed at low fluences (<3 J/cm(2) ) and is accompanied by a drastic reduction in porphyrin fluorescence from the Soret band. Laser ablation of extrinsic enamel stains at 400 nm is observed to be most efficient above 3 J/cm(2) with minimal damage to the underlying enamel. Unsound underlying enamel is also observed to be selectively removed after irradiation. Copyright © 2012 Wiley Periodicals, Inc.
Microscopic quantification of bacterial invasion by a novel antibody-independent staining method.

PubMed

Agerer, Franziska; Waeckerle, Stephanie; Hauck, Christof R

2004-10-01

Microscopic discrimination between extracellular and invasive, intracellular bacteria is a valuable technique in microbiology and immunology. We describe a novel fluorescence staining protocol, called FITC-biotin-avidin (FBA) staining, which allows the differentiation between extracellular and intracellular bacteria and is independent of specific antibodies directed against the microorganisms. FBA staining of eukaryotic cells infected with Gram-negative bacteria of the genus Neisseria or the Gram-positive pathogen Staphylococcus aureus are employed to validate the novel technique. The quantitative evaluation of intracellular pathogens by the FBA staining protocol yields identical results compared to parallel samples stained with conventional, antibody-dependent methods. FBA staining eliminates the need for cell permeabilization resulting in robust and rapid detection of invasive microbes. Taken together, FBA staining provides a reliable and convenient alternative for the differential detection of intracellular and extracellular bacteria and should be a valuable technical tool for the quantitative analysis of the invasive properties of pathogenic bacteria and other microorganisms.
An optimized staining technique for the detection of Gram positive and Gram negative bacteria within tissue.

PubMed

Becerra, Sandra C; Roy, Daniel C; Sanchez, Carlos J; Christy, Robert J; Burmeister, David M

2016-04-12

Bacterial infections are a common clinical problem in both acute and chronic wounds. With growing concerns over antibiotic resistance, treatment of bacterial infections should only occur after positive diagnosis. Currently, diagnosis is delayed due to lengthy culturing methods which may also fail to identify the presence of bacteria. While newer costly bacterial identification methods are being explored, a simple and inexpensive diagnostic tool would aid in immediate and accurate treatments for bacterial infections. Histologically, hematoxylin and eosin (H&E) and Gram stains have been employed, but are far from optimal when analyzing tissue samples due to non-specific staining. The goal of the current study was to develop a modification of the Gram stain that enhances the contrast between bacteria and host tissue. A modified Gram stain was developed and tested as an alternative to Gram stain that improves the contrast between Gram positive bacteria, Gram negative bacteria and host tissue. Initially, clinically relevant strains of Pseudomonas aeruginosa and Staphylococcus aureus were visualized in vitro and in biopsies of infected, porcine burns using routine Gram stain, and immunohistochemistry techniques involving bacterial strain-specific fluorescent antibodies as validation tools. H&E and Gram stain of serial biopsy sections were then compared to a modification of the Gram stain incorporating a counterstain that highlights collagen found in tissue. The modified Gram stain clearly identified both Gram positive and Gram negative bacteria, and when compared to H&E or Gram stain alone provided excellent contrast between bacteria and non-viable burn eschar. Moreover, when applied to surgical biopsies from patients that underwent burn debridement this technique was able to clearly detect bacterial morphology within host tissue. We describe a modification of the Gram stain that provides improved contrast of Gram positive and Gram negative microorganisms within host tissue. The samples used in this study demonstrate that this staining technique has laboratory and clinical applicability. This modification only adds minutes to traditional Gram stain with reusable reagents, and results in a cost- and time-efficient technique for identifying bacteria in any clinical biopsy containing connective tissue.
Recognition of Pneumocystis carinii by gram stain in impression smears of lung tissue.

PubMed Central

Felegie, T P; Pasculle, A W; Dekker, A

1984-01-01

In 12 of 20 (60%) biopsy-proven cases of Pneumocystis carinii pneumonia, the diagnosis was first suggested by examination of routine Gram stains of impression smears made from infected lung tissue and later confirmed by methenamine-silver staining. The cysts appeared as 5- to 7-microns unstained spheres, each containing six to eight intracystic gram-negative bodies (sporozoites). Although the Gram stain does not appear to be as sensitive as more traditional staining techniques for the detection of P. carinii, clinical microbiologists should be aware of the morphology of this organism in gram-stained specimens because this relatively simple procedure gives quick results. Images PMID:6084017
Research on fluorescence detection method of Microcystis aeruginosa

NASA Astrophysics Data System (ADS)

Wang, Xiao-xiong

2017-07-01

The paper studied the viability determination of Microcystis aeruginosa by FDA and PI staining. The staining results were measured by fluorescence microscopy. The results indicated that viable and dead cells were stained as bright green and red fluorescent respectively by FDA and PI. Through PI-FDA dual color fluorescence staining, the color of green and red distinct obviously by fluorescence microscope. The staining rate has relation with the cell density. If the cell density of M. aeruginosa was 1.0×107-1.0×109 cell·mL-1, the staining rate would be 100.0% or 98.0% by PI and of FDA respectively.
The effect of dietary pigmentation on the esthetic appearance of clear orthodontic elastomeric modules

PubMed Central

Talic, Nabeel F; Almudhi, Abdullazez A

2016-01-01

Objective: To compare the stain resistance of three types of clear elastomeric modules exposed to several common dietary substances through the assessment of the perception of a group of dentists to discoloration using visual analog scale (VAS). Materials and Methods: Elastomeric modules from Unitek (AU), Ormco (OR), and dentaurum (DE) were immersed in the following food substances: Coffee, black tea, chocolate, energy drink, ketchup, and Coca-Cola for 72 h. VAS was used to reflect the module staining severity. Results: Significant difference was found among the three types of modules examined in this study. OR modules showed the least mean staining ratings by the examiners. There was no statistical difference in the staining properties between AU and DE modules. Coffee and tea showed higher staining potential as compared to all staining media. Furthermore, there was no difference in the staining characteristics of coffee and black tea. Conclusions: Coffee and tea are strong staining media that should be avoided by patients who opted to have esthetic appliances for their orthodontic treatment. Elastomeric modules manufactured by AU showed higher staining optical properties as compared to the other two companies, which could be related to the manufacturing processing of these modules. PMID:27127754
Confocal mosaicing microscopy of human skin ex vivo: spectral analysis for digital staining to simulate histology-like appearance

NASA Astrophysics Data System (ADS)

Bini, Jason; Spain, James; Nehal, Kishwer; Hazelwood, Vikki; Dimarzio, Charles; Rajadhyaksha, Milind

2011-07-01

Confocal mosaicing microscopy enables rapid imaging of large areas of fresh tissue, without the processing that is necessary for conventional histology. Mosaicing may offer a means to perform rapid histology at the bedside. A possible barrier toward clinical acceptance is that the mosaics are based on a single mode of grayscale contrast and appear black and white, whereas histology is based on two stains (hematoxylin for nuclei, eosin for cellular cytoplasm and dermis) and appears purple and pink. Toward addressing this barrier, we report advances in digital staining: fluorescence mosaics that show only nuclei, are digitally stained purple and overlaid on reflectance mosaics, which show only cellular cytoplasm and dermis, and are digitally stained pink. With digital staining, the appearance of confocal mosaics mimics the appearance of histology. Using multispectral analysis and color matching functions, red, green, and blue (RGB) components of hematoxylin and eosin stains in tissue were determined. The resulting RGB components were then applied in a linear algorithm to transform fluorescence and reflectance contrast in confocal mosaics to the absorbance contrast seen in pathology. Optimization of staining with acridine orange showed improved quality of digitally stained mosaics, with good correlation to the corresponding histology.
Bone marrow cells stained by azide-conjugated Alexa fluors in the absence of an alkyne label.

PubMed

Lin, Guiting; Ning, Hongxiu; Banie, Lia; Qiu, Xuefeng; Zhang, Haiyang; Lue, Tom F; Lin, Ching-Shwun

2012-09-01

Thymidine analog 5-ethynyl-2'-deoxyuridine (EdU) has recently been introduced as an alternative to 5-bromo-2-deoxyuridine (BrdU) for cell labeling and tracking. Incorporation of EdU into replicating DNA can be detected by azide-conjugated fluors (eg, Alexa-azide) through a Cu(i)-catalyzed click reaction between EdU's alkyne moiety and azide. While this cell labeling method has proven to be valuable for tracking transplanted stem cells in various tissues, we have found that some bone marrow cells could be stained by Alexa-azide in the absence of EdU label. In intact rat femoral bone marrow, ~3% of nucleated cells were false-positively stained, and in isolated bone marrow cells, ~13%. In contrast to true-positive stains, which localize in the nucleus, the false-positive stains were cytoplasmic. Furthermore, while true-positive staining requires Cu(i), false-positive staining does not. Reducing the click reaction time or reducing the Alexa-azide concentration failed to improve the distinction between true- and false-positive staining. Hematopoietic and mesenchymal stem cell markers CD34 and Stro-1 did not co-localize with the false-positively stained cells, and these cells' identity remains unknown.

Accurate counting of neurons in frozen sections: some necessary precautions.

PubMed Central

Cooper, J D; Payne, J N; Horobin, R W

1988-01-01

In 30 microns frozen sections of rat midbrain the retrograde axonal transport of diamidino yellow, a fluorescent tracer, was used to demonstrate a population of neurons in the substantia nigra. However, when visualisation was carried out using the routine Nissl method a significant proportion of neurons failed to stain. As the presence of the retrograde tracer did not affect Nissl staining of such cells, such incomplete staining, with consequent underestimation of neuronal populations, is probably a common error in similar material. Further investigation revealed that the proportion of such unstained neurons was greater when the staining time was short, when stain concentration was low, or when section thickness was increased. Some stains were worse in this respect than others. Cresyl fast violet resulted in the highest proportion of unstained neurons, thionin resulted in the lowest proportion. It was concluded that the rate of diffusion of the stain into the section was the main factor limiting the staining of neurons present. Staining with pure thionin at 0.1% concentration for at least 3 minutes and with sections no thicker than 30 microns is one regime which would avoid this problem. Images Fig. 1 PMID:2461923
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... Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper than that of Strict Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] ...
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... Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper than that of Strict Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] ...
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... Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper than that of Strict Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] ...
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... Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper than that of Strict Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] ...
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... Strict Middling Yellow Stained Color. Strict Middling Yellow Stained Color is color which is deeper than that of Strict Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] ...
Evaluation of efficacy of 1% Crystal Violet & Nuclear Fast Red stain compared to Haematoxyline & Eosin stain for assessing mitotic figures in oral premalignant and malignant lesions.

PubMed

Motiwale, Gauri; Jaiswal, Shradha; Vikey, Ashok; Motiwale, Tejas; Bagulkar, Bhupesh; Bhat, Atul; Kapoor, Prakhar

2016-07-01

Various chromosomal arrangements in cells undergoing division are referred to as Mitotic figure (MF). The abnormal excess of mitotic figures is commonly seen in oral epithelial dysplasia (ED) and oral squamous cell carcinoma (OSCC). In present study, we compared the number of mitotic figures in normal oral mucosa, epithelial dysplasia & OSCC sections with haematoxyline & eosine (H&E) and 1%Crystal Violet & Nuclear Fast Red (CV&NFR) stain, also the efficacy of the CV&NFR stain as compared to H & E stain. We investigated the correlation between the number of mitotic figures & grades of OSCC. Study sample comprised of two serial sections of archival blocks of normal oral mucosa & diagnosed cases of epithelial dysplasia & OSCC. One slide stained with H& E & the other one with 1% CV & NFR. Mitotic figures were counted with the grid eyepiece. There was significant increase in number of MFs in oral ED and OSCC in comparison with normal oral mucosa. There was a highly significant increase in number of MFs in CV&NFR stained tissue sections when compared with H & E stain. Metaphase is the most commonly observed phase of mitosis. In summary, our study proposes the use of Crystal violet & Nuclear fast red stain as a selective stain for better contrast & easy identification MFs. © 2016 Old City Publishing, Inc.
Staining for factor VIII related antigen and Ulex europaeus agglutinin I (UEA-I) in 230 tumours. An assessment of their specificity for angiosarcoma and Kaposi's sarcoma.

PubMed

Leader, M; Collins, M; Patel, J; Henry, K

1986-11-01

In this study we examined the staining reactivity of commercially available antisera to factor VIII related antigen (F VIII RAg) and Ulex europaeus agglutinin I (UEA-I) on sections from 230 formalin fixed paraffin embedded tumours. These included 196 sarcomas, 20 carcinomas and 14 angiomas. All angiomas showed positive staining for F VIII RAg; all carcinomas showed negative staining; the vasoformative areas of all angiosarcomas stained positively but only four of six angiosarcomas showed positive staining of their solid areas; of seven Kaposi's sarcomas, all showed positive staining of vessels and six showed positive staining of the spindle cell component. In the remaining 181 non-vascular sarcomas there was a false positive result in four tumours (2.2%), three of which had a history of irradiation. Pre-radiotherapy biopsies of these three tumours stained negatively with anti-F VIII RAg. UEA-I was demonstrated in all the angiomas studied, in all angiosarcomas (including the solid components) and in well-formed vessels of all Kaposi's sarcomas, but only in the spindle cell component of 3/6. However, there was an unacceptably high rate of false positive staining amongst the carcinomas and non-vascular sarcomas. In conclusion, F VIII RAg is a specific but not a sensitive marker of angiosarcomas; UEA-I is a sensitive but not a specific marker of angiosarcomas.
Transmission Electron Microscopy as a Tool to Image Bio-Inorganic Nanohybrids: The Case of Phage-Gold Nanocomposites

PubMed Central

Cao, Binrui; Xu, Hong; Mao, Chuanbin

2011-01-01

In recent years, bio-inorganic nanohybrids composed of biological macromolecules and functional inorganic nanomaterials have revealed many unique properties that show promise for the future. Transmission electron microscopy (TEM) is a popular and relatively simple tool that can offer a direct visualization of the nanomaterials with high resolutions. When TEM is applied to visualize bio-inorganic nanohybrids, a treatment of negative staining is necessary due to the presence of biological molecules in the nanohybrids except for those with densely packed inorganic materials. However, the conventional negative-staining procedure for regular biological samples cannot be directly applied to such bio-inorganic nanohybrids. To image a specific bio-inorganic nanohybrid, negative-staining factors such as negative stain type, working pH, staining time, and drying method, should be identified. Currently, no detailed studies have been done to investigate how to adjust negative-staining factors based on specific bio-inorganic nanohybrids. In this study, bacteriophage-gold nanoparticle hybrids were chosen as a model to systematically study the effects of each factor on the negative staining of the nanohybrids. The best staining conditions for gold nanoparticle-phage nanohybrids were obtained and the effects of each factor on the negative staining of general nanohybrids were discussed. This work indicates that with proper staining it is possible to use TEM to directly visualize both biological and inorganic components without introducing any artifact. PMID:21678527
The use of an oxidising mouthwash to reduce staining associated with chlorhexidine. Studies in vitro and in vivo.

PubMed

Addy, M; al-Arrayed, F; Moran, J

1991-04-01

Discolouration of the teeth, tongue and dental restorative materials associated with the use of cationic antiseptics is still the limiting factor in long-term usage of antiplaque agents such as chlorhexidine. Oxidising agents have been used successfully to remove dental stain in animals, but products and regimens have not been evaluated in humans. These pilot studies in vitro and in vivo assessed a long-established oxidising mouthwash, based on peroxyborate, for stain removal from teeth and acrylic. The laboratory studies measured chlorhexidine tea stain removal from tooth and acrylic specimens. The oxidising agent markedly reduced staining on specimens. In a short-term clinical experiment, dental staining was forced over a 2 1/2-day period by reciprocal chlorhexidine and tea rinsing. 5 rinses with the oxidising mouthwash during the next 1 1/2 days dramatically reduced staining compared with water after rinses. Additionally, a single rinse with peroxyborate following the 5 water after rinses, again considerably reduced the residual staining. Effects on tongue staining were less impressive. Both experimental models produced findings consistent with those from the animal model. The results suggest proprietary oxidising mouthwashes could have a place in the control of dental stain associated with the use of chlorhexidine. Further studies to evaluate such products in a more conventional chlorhexidine regimen are indicated.
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.
Utility of Acridine Orange staining for detection of bacteria from positive blood cultures.

PubMed

Neeraja, M; Lakshmi, V; Padmasri, C; Padmaja, K

2017-08-01

The diagnostic performance of AO stain was evaluated for the detection of bacteria and or fungi from positive blood cultures. The sensitivity of Gram stain (GS) was 98.26% while Acridine Orange (AO) stain proved to be more sensitive (100%) with a Positive and Negative Predictive Value of 100% each. The specificity of both the stains was 100%. Overall agreement between the two stains was 98.23% (688/700). The organisms that were missed by GS and positive by AO were Candida species (Sutton, 2006) and Gram negative bacilli (GNB) (Sutton, 2006). Sensitivity of GS was 82.35% and AO was 100% among mixed cultures. Immediate reporting of the results of AO stain would have a significant impact on clinical management of patients with serious blood stream infections. Copyright © 2017 Elsevier B.V. All rights reserved.
The development of fluorescence turn-on probe for Al(III) sensing and live cell nucleus-nucleoli staining

NASA Astrophysics Data System (ADS)

Saini, Anoop Kumar; Sharma, Vinay; Mathur, Pradeep; Shaikh, Mobin M.

2016-10-01

The morphology of nucleus and nucleolus is powerful indicator of physiological and pathological conditions. The specific staining of nucleolus recently gained much attention due to the limited and expensive availability of the only existing stain “SYTO RNA-Select”. Here, a new multifunctional salen type ligand (L1) and its Al3+ complex (1) are designed and synthesized. L1 acts as a chemosensor for Al3+ whereas 1 demonstrates specific staining of nucleus as well as nucleoli. The binding of 1 with nucleic acid is probed by DNase and RNase digestion in stained cells. 1 shows an excellent photostability, which is a limitation for existing nucleus stains during long term observations. 1 is assumed to be a potential candidate as an alternative to expensive commercial dyes for nucleus and nucleoli staining.
The development of fluorescence turn-on probe for Al(III) sensing and live cell nucleus-nucleoli staining.

PubMed

Saini, Anoop Kumar; Sharma, Vinay; Mathur, Pradeep; Shaikh, Mobin M

2016-10-10

The morphology of nucleus and nucleolus is powerful indicator of physiological and pathological conditions. The specific staining of nucleolus recently gained much attention due to the limited and expensive availability of the only existing stain "SYTO RNA-Select". Here, a new multifunctional salen type ligand (L 1 ) and its Al 3+ complex (1) are designed and synthesized. L 1 acts as a chemosensor for Al 3+ whereas 1 demonstrates specific staining of nucleus as well as nucleoli. The binding of 1 with nucleic acid is probed by DNase and RNase digestion in stained cells. 1 shows an excellent photostability, which is a limitation for existing nucleus stains during long term observations. 1 is assumed to be a potential candidate as an alternative to expensive commercial dyes for nucleus and nucleoli staining.
Specimen block counter-staining for localization of GUS expression in transgenic arabidopsis and tobacco

NASA Technical Reports Server (NTRS)

Kim, M. K.; Choi, J-W; Jeon, J-H; Franceschi, V. R.; Davin, L. B.; Lewis, N. G.

2002-01-01

A simple counter-staining procedure has been developed for comparative beta-glucuronidase (GUS) expression and anatomical localization in transgenic herbaceous arabidopsis and tobacco. This protocol provides good anatomical visualization for monitoring chimeric gene expression at both the organ and tissue levels. It can be used with different histochemical stains and can be extended to the study of woody species. The specimens are paraffin-embedded, the block is trimmed to reveal internal structure, safranin-O staining solution is briefly applied to the surface of the block, then washed off and, after drying, a drop of immersion oil is placed on the stained surface for subsequent photographic work. This gives tissue counter-staining with good structural preservation without loss of GUS staining product; moreover, sample observation is rapid and efficient compared to existing procedures.
Blood culture gram stain, acridine orange stain and direct sensitivity-based antimicrobial therapy of bloodstream infection in patients with trauma.

PubMed

Behera, B; Mathur, P; Gupta, B

2010-01-01

The purpose of this study was to ascertain if the simple practice of Gram stain, acridine orange stain and direct sensitivity determination of positive blood culture bottles could be used to guide early and appropriate treatment in trauma patients with clinical suspicion of sepsis. The study also aimed to evaluate the error in interpreting antimicrobial sensitivity by direct method when compared to standard method and find out if specific antibiotic-organism combination had more discrepancies. Findings from consecutive episodes of blood stream infection at an Apex Trauma centre over a 12-month period are summarized. A total of 509 consecutive positive blood cultures were subjected to Gram staining. AO staining was done in BacT/ALERT-positive Gram-stain negative blood cultures. Direct sensitivity was performed from 369 blood culture broths, showing single type of growth in Gram and acridine orange staining. Results of direct sensitivity were compared to conventional sensitivity for errors. No 'very major' discrepancy was found in this study. About 5.2 and 1.8% minor error rates were noted in gram-positive and gram-negative bacteria, respectively, while comparing the two methods. Most of the discrepancies in gram-negative bacteria were noted in beta lactam - beta lactamase inhibitor combinations. Direct sensitivity testing was not reliable for reporting of methicillin and vancomycin resistance in Staphylococci. Gram stain result together with direct sensitivity testing is required for optimizing initial antimicrobial therapy in trauma patients with clinical suspicion of sepsis. Gram staining and AO staining proved particularly helpful in the early detection of candidaemia.
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... Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper than Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] below color grade cotton ...
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... Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper than Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] below color grade cotton ...
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... Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper than Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] below color grade cotton ...
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... Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper than Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] below color grade cotton ...

Aureobasidium pullulans morphology: two adapted polysaccharide stains.

PubMed

Oller, Anna R

2005-12-01

Morphological stages of Aureobasidium pullulans were investigated utilizing different media ingredients and were visualized by bright-field microscopy. A polysaccharide stain was developed to stain chlamydospores, cell walls, hyphae, and conidia, since current staining techniques do not reveal subcellular details to identify fungi, especially those that exhibit polysaccharide secretions.
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... Stained Color. Middling Yellow Stained Color is American Upland cotton which in color is deeper than Middling Tinged Color. [57 FR 34498, Aug. 5, 1992] below color grade cotton ...
Fabrics coated with lubricated nanostructures display robust omniphobicity

DOE PAGES

Shillingford, Cicely; MacCallum, Noah; Wong, Tak -Sing; ...

2013-12-11

The development of a stain-resistant and pressure-stable textile is desirable for consumer and industrial applications alike, yet it remains a challenge that current technologies have been unable to fully address. Traditional superhydrophobic surfaces, inspired by the lotus plant, are characterized by two main components: hydrophobic chemical functionalization and surface roughness. While this approach produces water-resistant surfaces, these materials have critical weaknesses that hinder their practical utility, in particular as robust stain-free fabrics. For example, traditional superhydrophobic surfaces fail (i.e., become stained) when exposed to low-surface-tension liquids, under pressure when impacted by a high-velocity stream of water (e.g., rain), and whenmore » exposed to physical forces such as abrasion and twisting. We have recently introduced slippery lubricant-infused porous surfaces (SLIPS), a self-healing, pressure-tolerant and omniphobic surface, to address these issues. However we present the rational design and optimization of nanostructured lubricant-infused fabrics and demonstrate markedly improved performance over traditional superhydrophobic textile treatments: SLIPS-functionalized cotton and polyester fabrics exhibit decreased contact angle hysteresis and sliding angles, omni-repellent properties against various fluids including polar and nonpolar liquids, pressure tolerance and mechanical robustness, all of which are not readily achievable with the state-of-the-art superhydrophobic coatings.« less
Fabrics coated with lubricated nanostructures display robust omniphobicity

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

Shillingford, C; MacCallum, N; Wong, TS

2013-12-11

The development of a stain-resistant and pressure-stable textile is desirable for consumer and industrial applications alike, yet it remains a challenge that current technologies have been unable to fully address. Traditional superhydrophobic surfaces, inspired by the lotus plant, are characterized by two main components: hydrophobic chemical functionalization and surface roughness. While this approach produces water-resistant surfaces, these materials have critical weaknesses that hinder their practical utility, in particular as robust stain-free fabrics. For example, traditional superhydrophobic surfaces fail (i.e., become stained) when exposed to low-surface-tension liquids, under pressure when impacted by a high-velocity stream of water (e. g., rain), andmore » when exposed to physical forces such as abrasion and twisting. We have recently introduced slippery lubricant-infused porous surfaces (SLIPS), a self-healing, pressure-tolerant and omniphobic surface, to address these issues. Herein we present the rational design and optimization of nanostructured lubricant-infused fabrics and demonstrate markedly improved performance over traditional superhydrophobic textile treatments: SLIPS-functionalized cotton and polyester fabrics exhibit decreased contact angle hysteresis and sliding angles, omni-repellent properties against various fluids including polar and nonpolar liquids, pressure tolerance and mechanical robustness, all of which are not readily achievable with the state-of-the-art superhydrophobic coatings.« less
[Pneumocystis pneumonia biological diagnosis at Fann Teaching Hospital in Dakar, Senegal].

PubMed

Dieng, Y; Dieng, T; Sow, D; Wlouhou, S; Sylla, K; Tine, R; Ndiaye, M; Ndiaye, J L; Faye, B; Faye, O; Gaye, O

2016-03-01

Data relative to Pneumocystis pneumonia in sub-Saharan Africa are not well known. Weakness of the technical material and use of little sensitive biological tools of diagnosis are among the evoked reasons. The objective of this study is to update the data of the disease at the Fann Teaching Hospital in Dakar and to estimate biological methods used in diagnosis. A descriptive longitudinal study was carried out from January 5th, 2009 to October 31st, 2011 in the parasitology and mycology laboratory of the Fann Teaching Hospital in Dakar. The bronchoalveolar lavages received in the laboratory were examined microscopically for Pneumocystis jirovecii by indirect fluorescent assay or after Giemsa or toluidine blue O staining. One hundred and eighty-three bronchoalveolar lavages withdrawn from 183 patients were received in the laboratory. Sixteen were positive for P. jirovecii at 9% frequency. Four among these patients were HIV positive. Indirect fluorescent assay allowed finding of P. jirovecii among 16 patients while Giemsa staining discovered P. jirovecii only in a single patient. No case was diagnosed by toluidine blue O staining. Pneumocystis pneumonia in Parasitology and Mycology Laboratory of Fann Teaching Hospital at Dakar was mainly diagnosed among HIV patients. Copyright © 2015 Elsevier Masson SAS. All rights reserved.
Fabrics coated with lubricated nanostructures display robust omniphobicity.

PubMed

Shillingford, Cicely; MacCallum, Noah; Wong, Tak-Sing; Kim, Philseok; Aizenberg, Joanna

2014-01-10

The development of a stain-resistant and pressure-stable textile is desirable for consumer and industrial applications alike, yet it remains a challenge that current technologies have been unable to fully address. Traditional superhydrophobic surfaces, inspired by the lotus plant, are characterized by two main components: hydrophobic chemical functionalization and surface roughness. While this approach produces water-resistant surfaces, these materials have critical weaknesses that hinder their practical utility, in particular as robust stain-free fabrics. For example, traditional superhydrophobic surfaces fail (i.e., become stained) when exposed to low-surface-tension liquids, under pressure when impacted by a high-velocity stream of water (e.g., rain), and when exposed to physical forces such as abrasion and twisting. We have recently introduced slippery lubricant-infused porous surfaces (SLIPS), a self-healing, pressure-tolerant and omniphobic surface, to address these issues. Herein we present the rational design and optimization of nanostructured lubricant-infused fabrics and demonstrate markedly improved performance over traditional superhydrophobic textile treatments: SLIPS-functionalized cotton and polyester fabrics exhibit decreased contact angle hysteresis and sliding angles, omni-repellent properties against various fluids including polar and nonpolar liquids, pressure tolerance and mechanical robustness, all of which are not readily achievable with the state-of-the-art superhydrophobic coatings.
Fabrics coated with lubricated nanostructures display robust omniphobicity

NASA Astrophysics Data System (ADS)

Shillingford, Cicely; MacCallum, Noah; Wong, Tak-Sing; Kim, Philseok; Aizenberg, Joanna

2014-01-01

The development of a stain-resistant and pressure-stable textile is desirable for consumer and industrial applications alike, yet it remains a challenge that current technologies have been unable to fully address. Traditional superhydrophobic surfaces, inspired by the lotus plant, are characterized by two main components: hydrophobic chemical functionalization and surface roughness. While this approach produces water-resistant surfaces, these materials have critical weaknesses that hinder their practical utility, in particular as robust stain-free fabrics. For example, traditional superhydrophobic surfaces fail (i.e., become stained) when exposed to low-surface-tension liquids, under pressure when impacted by a high-velocity stream of water (e.g., rain), and when exposed to physical forces such as abrasion and twisting. We have recently introduced slippery lubricant-infused porous surfaces (SLIPS), a self-healing, pressure-tolerant and omniphobic surface, to address these issues. Herein we present the rational design and optimization of nanostructured lubricant-infused fabrics and demonstrate markedly improved performance over traditional superhydrophobic textile treatments: SLIPS-functionalized cotton and polyester fabrics exhibit decreased contact angle hysteresis and sliding angles, omni-repellent properties against various fluids including polar and nonpolar liquids, pressure tolerance and mechanical robustness, all of which are not readily achievable with the state-of-the-art superhydrophobic coatings.
Spinal cord compression secondary to hemangiosarcoma in a saddlebred stallion.

PubMed Central

Berry, S

1999-01-01

Hemangiosarcoma in the spinal canal was diagnosed in a 25-year-old stallion showing progressive and symmetrical 4-limb ataxia, proprioceptive deficits, and weakness. On necropsy, an extradural mass consisting of spindle-shaped cells and numerous free erythrocytes was found at the level of C7-T1. Immunohistochemical staining confirmed a neoplasm of endothelial origin. PMID:10646067
Reconnaissance geology of the Al Mukhul Quadrangle, sheet 26/42 B, Kingdom of Saudi Arabia

USGS Publications Warehouse

Du Bray, E.A.

1984-01-01

Mineral potential in the quadrangle is low. At a very small prospect pit in the north-central part of the quadrangle, massive, milky quartz veins cutting weakly metamorphosed volcanogenic sedimentary rocks are stained blue and green by copper minerals. A previously reported mine site in the southern part of the quadrangle was not relocated.
The role of different structural motifs in the ultrafast dynamics of second generation protein stains.

PubMed

Chatterjee, Soumit; Karuso, Peter; Boulangé, Agathe; Peixoto, Philippe A; Franck, Xavier; Datta, Anindya

2013-12-05

Engineering the properties of fluorescent probes through modifications of the fluorophore structure has become a subject of interest in recent times. By doing this, the photophysical and photochemical properties of the modified fluorophore can be understood and this can guide the design and synthesis of better fluorophores for use in biotechnology. In this work, the electronic spectra and fluorescence decay kinetics of four analogues of the fluorescent natural product epicocconone were investigated. Epicocconone is unique in that the native state is weakly green fluorescent, whereas the enamine formed reversibly with proteins is highly emissive in the red. It was found that the ultrafast dynamics of the analogues depends profoundly on the H-bonding effect of solvents and solvent viscosity though solvent polarity also plays a role. Comparing the steady state and time-resolved data, the weak fluorescence of epicocconone in its native state is most likely due to the photoisomerization of the hydrocarbon side chain, while the keto enol moiety also has a role to play in determining the fluorescence quantum yield. This understanding is expected to aid the design of better protein stains from the same family.
Bacillus Anthracis Spores of the bclA Mutant Exhibit Increased Adherence to Epithelial Cells, Fibroblasts, and Endothelial Cells but not to Macrophages

DTIC Science & Technology

2007-09-01

immunofluorescence (IFM) and light microscopy. Samples were fixed in forma- lin, stained with immunofluorescent dyes (as described below) or spore stain ( malachite ...BEC. Adherence was assessed by microscopic observation of the infected cells stained with malachite green and counterstaining of the BEC. For enzymatic...this significant difference, BEC infected with spores were stained with malachite green and counter- stained with Wright-Giemsa (Fig. 1B and C). This
Quantitative cytochemistry of nuclear and cytoplasmic proteins using the Naphthol Yellow S and dinitrofluorobenzene staining methods.

PubMed

Tas, J; James, J

1981-09-01

The 'total protein staining' of biological specimens with the electrostatically binding Naphthol Yellow S or the covalently binding dinitrofluorobenzene must be interpreted as methods which yield data on the specific amino acid pool of the proteins concerned. Both dyes bind to certain free amino-acid side-chains, giving different dye--protein ratios for various proteins. In the presence of DNA, dinitrofluorobenzene stains all proteins present in cell nuclei, whereas Naphthol Yellow S only stains the majority of the non-histone proteins. When protein staining methods are combined with the Feulgen--Pararosanile (SO2) procedure for DNA, decreased Feulgen--DNA contents were measured in dinitrofluorobenzene-stained isolated nuclei and lymphocytes.
Efficacy test of a toothpaste in reducing extrinsic dental stain

NASA Astrophysics Data System (ADS)

Agustanti, A.; Ramadhani, S. A.; Adiatman, M.; Rahardjo, A.; Callea, M.; Yavuz, I.; Maharani, D. A.

2017-08-01

This clinical trial compared the external dental stain reduction achieved by tested toothpaste versus placebo in adult patients. In this double-blind, parallel, randomised clinical trial, 45 female volunteers with a mean age of 20 years old were included. All study subjects front teeth were topically applicated with Silver Diamine Fluoride (SDF) to create external dental stains. Subjects were randomized into test (n=22) and control (n=23) groups. Toothpastes were used for two days to analyse the effects of removing external stains on the labial surfaces of all anterior teeth. VITA Easyshade Advance 4.0 was used to measure dental extrinsic stains changes. The analysis showed statistically significant efficacy of the tested toothpaste in reducing external dental stain caused by SDF, comparing to the placebo toothpaste, after one and two days of usage. The tested toothpaste was effective in reducing dental stain.
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.
Fluorogenic Ag+–Tetrazolate Aggregation Enables Efficient Fluorescent Biological Silver Staining

PubMed Central

Xie, Sheng; Wong, Alex Y. H.; Kwok, Ryan T. K.; Li, Ying; Su, Huifang; Lam, Jacky W. Y.

2018-01-01

Abstract Silver staining, which exploits the special bioaffinity and the chromogenic reduction of silver ions, is an indispensable visualization method in biology. It is a most popular method for in‐gel protein detection. However, it is limited by run‐to‐run variability, background staining, inability for protein quantification, and limited compatibility with mass spectroscopic (MS) analysis; limitations that are largely attributed to the tricky chromogenic visualization. Herein, we reported a novel water‐soluble fluorogenic Ag+ probe, the sensing mechanism of which is based on an aggregation‐induced emission (AIE) process driven by tetrazolate‐Ag+ interactions. The fluorogenic sensing can substitute the chromogenic reaction, leading to a new fluorescence silver staining method. This new staining method offers sensitive detection of total proteins in polyacrylamide gels with a broad linear dynamic range and robust operations that rival the silver nitrate stain and the best fluorescent stains. PMID:29575702
Use of the gram stain in microbiology.

PubMed

Beveridge, T J

2001-05-01

The Gram stain differentiates bacteria into two fundamental varieties of cells. Bacteria that retain the initial crystal violet stain (purple) are said to be "gram-positive," whereas those that are decolorized and stain red with carbol fuchsin (or safranin) are said to be "gram-negative." This staining response is based on the chemical and structural makeup of the cell walls of both varieties of bacteria. Gram-positives have a thick, relatively impermeable wall that resists decolorization and is composed of peptidoglycan and secondary polymers. Gram-negatives have a thin peptidoglycan layer plus an overlying lipid-protein bilayer known as the outer membrane, which can be disrupted by decolorization. Some bacteria have walls of intermediate structure and, although they are officially classified as gram-positives because of their linage, they stain in a variable manner. One prokaryote domain, the Archaea, have such variability of wall structure that the Gram stain is not a useful differentiating tool.
Fluorescent staining for leukocyte chemotaxis. Eosinophil-specific fluorescence with aniline blue.

PubMed

McCrone, E L; Lucey, D R; Weller, P F

1988-11-10

To overcome problems associated with the quantitation of human eosinophil chemotaxis in micropore filters, we have developed a fluorescent method of specifically staining eosinophils in chemotactic filters. A neutral solution of aniline blue yielded bright green fluorescent staining of the cytoplasmic granules of eosinophils. Other leukocytes and contaminating neutrophils potentially present with eosinophils did not fluoresce with aniline blue. The fluorescent staining eosinophils within filters provided bright, non-fading images that facilitated visual microscopic counting and were of sufficiently high contrast, unlike those with conventional eosinophil stains, to allow image analyzer based enumeration of eosinophil chemotactic responses at levels through the filters. Although not cell type-specific, congo red and ethidium bromide also provided high contrast, fluorescent images of all leukocyte types within chemotactic filters. Fluorescent staining with aniline blue constitutes a rapid, stable and eosinophil-specific stain that facilitates the visual or image analyzer-based quantitation of eosinophil chemotaxis.
Histochemical discrimination of fibers in regenerating rat infraorbital nerve

NASA Technical Reports Server (NTRS)

Wilke, R. A.; Riley, D. A.; Sanger, J. R.

1992-01-01

In rat dorsal root ganglia, histochemical staining of carbonic anhydrase (CA) and cholinesterase (CE) yields a reciprocal pattern of activity: Sensory processes are CA positive and CE negative, whereas motor processes are CA negative and CE positive. In rat infraorbital nerve (a sensory peripheral nerve), we saw extensive CA staining of nearly 100% of the myelinated axons. Although CE reactivity in myelinated axons was extremely rare, we did observe CE staining of unmyelinated autonomic fibers. Four weeks after transection of infraorbital nerves, CA-stained longitudinal sections of the proximal stump demonstrated 3 distinct morphological zones. A fraction of the viable axons retained CA activity to within 2 mm of the distal extent of the stump, and the stain is capable of resolving growth sprouts being regenerated from these fibers. Staining of unmyelinated autonomic fibers in serial sections shows that CE activity was not retained as far distally as is the CA sensory staining.
Gram staining in the diagnosis of acute septic arthritis.

PubMed

Faraj, A A; Omonbude, O D; Godwin, P

2002-10-01

This study aimed at determining the sensitivity and specificity of Gram staining of synovial fluid as a diagnostic tool in acute septic arthritis. A retrospective study was made of 22 patients who had arthroscopic lavage following a provisional diagnosis of acute septic arthritis of the knee joint. Gram stains and cultures of the knee aspirates were compared with the clinical and laboratory parameters, to evaluate their usefulness in diagnosing acute arthritis. All patients who had septic arthritis had pain, swelling and limitation of movement. CRP was elevated in 90% of patients. The incidence of elevated white blood cell count was higher in the group of patients with a positive Gram stain study (60%) as compared to patients with a negative Gram stain study (33%). Gram staining sensitivity was 45%. Its specificity was however 100%. Gram staining is an unreliable tool in early decision making in patients requiring urgent surgical drainage and washout.
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…

How to Guarantee Long Life for your Carpeting.

ERIC Educational Resources Information Center

Gilliland, John W.

1968-01-01

Carpeting must be maintained through a proper maintenance program so as to extend the carpet's life and allow for continued sound control. Four types of common soils or stains are discussed--(1) dry soils, (2) water soluble stains, (3) petroleum soluble stains, and (4) other stains. Various cleaning methods, such as, vacuuming, spot removal, wet…
Tracking living decapod larvae: mass staining of eggs with neutral red prior to hatching.

PubMed

Øresland, V; Horobin, R W

2012-04-01

Mass staining of decapod females carrying eggs, with subsequent identification of hatched larvae in the environment, is a research tool with great potential for field ecologists wishing to track the movements of larvae. For this to be achieved, however, numerous requirements must be met. These include adequate dye solubility, short staining time, dye penetration through different tissues, dye retention within the organism, absence of toxic and behavioral effects, low visibility to predators of stained larvae, no loss of staining owing to preservatives and low cost. The dye, neutral red, appears to meet most of these requirements. This dye was used in aliquots of 0.7 g/770 ml seawater applied to the females of Norway lobster (Nephrops norvegicus) and European lobster (Homarus gammarus) for 10 min. This procedure stained lobster eggs and embryos so that hatched larvae could be distinguished easily by fluorescence microscopy from larvae that hatched from unstained eggs. Stained larvae that were preserved in 4% formaldehyde in seawater were still stained after 1 year. Larvae should not come in contact with ethanol, because it extracts the dye rapidly.
DNA comet Giemsa staining for conventional bright-field microscopy.

PubMed

Osipov, Andreyan; Arkhangelskaya, Ekaterina; Vinokurov, Alexei; Smetaninа, Nadezhda; Zhavoronkov, Alex; Klokov, Dmitry

2014-04-10

This study was undertaken to evaluate the compatibility of Giemsa staining protocol with the comet assay. We showed, for the first time, that DNA comets can be visualized and analyzed using Giemsa staining. We generated DNA damage dose response curves for human peripheral blood lymphocytes exposed to X-ray radiation using the comet assay with either SybrGreen I or Giemsa stain. The dose response curves were fitted by linear regressions (R2>0.977). The SybrGreen I results showed only ~1.2-fold higher slope coefficient (method sensitivity) compared to the Giemsa results. The unexpectedly high sensitivity of Giemsa staining for the comet assay is due to the Romanowsky-Giemsa effect, the stain photo-stability and the higher resolution of bright-field imaging compared to fluorescence imaging. Our results demonstrate that Giemsa staining can effectively be used for measuring DNA damage by the comet assay. The low cost and availability of Giemsa stain makes this method affordable for any low budget research and will facilitate new applications of the comet assay in biology and medicine.
A procedure for Alcian blue staining of mucins on polyvinylidene difluoride membranes.

PubMed

Dong, Weijie; Matsuno, Yu-ki; Kameyama, Akihiko

2012-10-16

The isolation and characterization of mucins are critically important for obtaining insight into the molecular pathology of various diseases, including cancers and cystic fibrosis. Recently, we developed a novel membrane electrophoretic method, supported molecular matrix electrophoresis (SMME), which separates mucins on a polyvinylidene difluoride (PVDF) membrane impregnated with a hydrophilic polymer. Alcian blue staining is widely used to visualize mucopolysaccharides and acidic mucins on both blotted membranes and SMME membranes; however, this method cannot be used to stain mucins with a low acidic glycan content. Meanwhile, periodic acid-Schiff staining can selectively visualize glycoproteins, including mucins, but is incompatible with glycan analysis, which is indispensable for mucin characterizations. Here we describe a novel staining method, designated succinylation-Alcian blue staining, for visualizing mucins on a PVDF membrane. This method can visualize mucins regardless of the acidic residue content and shows a sensitivity 2-fold higher than that of Pro-Q Emerald 488, a fluorescent periodate Schiff-base stain. Furthermore, we demonstrate the compatibility of this novel staining procedure with glycan analysis using porcine gastric mucin as a model mucin.
Loss of Fumarate Hydratase and Aberrant Protein Succination Detected With S-(2-Succino)-Cysteine Staining to Identify Patients With Multiple Cutaneous and Uterine Leiomyomatosis and Hereditary Leiomyomatosis and Renal Cell Cancer Syndrome.

PubMed

Llamas-Velasco, Mar; Requena, Luis; Adam, Julie; Frizzell, Norma; Hartmann, Arndt; Mentzel, Thomas

2016-12-01

Hereditary leiomyomatosis and renal cell cancer (HLRCC) syndrome is an autosomal dominant disorder caused by heterozygotic germline mutations in fumarate hydratase (FH) with incomplete penetrance, and clinically challenging to diagnose. Immunohistochemical stainings may favor an earlier diagnosis. The authors have tested 31 smooth muscle neoplasms. Ten of the 13 lesions from patients with HLRCC syndrome showed negative FH staining. Most sporadic piloleiomyomas presented strongly positive FH staining although 5 cases were negative. Sensitivity of FH staining in our series is 83.3% but specificity is 75%. Anti-S-(2-succino)-cysteine (2SC) showed the opposite intensity staining pattern and showed great correlation with anti-FH (rho spearman = -0.797). Anti-2SC staining increased the diagnostic accuracy in 19% of the cases. The main limitation of this study is the lack additional clinical data to further classify the cases as the case inclusion was histopathological. Negative FH staining could indicate a high risk of HLRCC but it could also suggest the presence of a syndrome in up to 25% of sporadic cases. Thus, when there is a doubtful case, anti-2SC may be added to exclude the syndrome if a negative staining is found.
Confocal mosaicing microscopy of human skin ex vivo: spectral analysis for digital staining to simulate histology-like appearance

PubMed Central

Bini, Jason; Spain, James; Nehal, Kishwer; Hazelwood, Vikki; DiMarzio, Charles; Rajadhyaksha, Milind

2011-01-01

Confocal mosaicing microscopy enables rapid imaging of large areas of fresh tissue, without the processing that is necessary for conventional histology. Mosaicing may offer a means to perform rapid histology at the bedside. A possible barrier toward clinical acceptance is that the mosaics are based on a single mode of grayscale contrast and appear black and white, whereas histology is based on two stains (hematoxylin for nuclei, eosin for cellular cytoplasm and dermis) and appears purple and pink. Toward addressing this barrier, we report advances in digital staining: fluorescence mosaics that show only nuclei, are digitally stained purple and overlaid on reflectance mosaics, which show only cellular cytoplasm and dermis, and are digitally stained pink. With digital staining, the appearance of confocal mosaics mimics the appearance of histology. Using multispectral analysis and color matching functions, red, green, and blue (RGB) components of hematoxylin and eosin stains in tissue were determined. The resulting RGB components were then applied in a linear algorithm to transform fluorescence and reflectance contrast in confocal mosaics to the absorbance contrast seen in pathology. Optimization of staining with acridine orange showed improved quality of digitally stained mosaics, with good correlation to the corresponding histology. PMID:21806269
Rapid contrast evaluation method based on affinity beads and backscattered electron imaging for the screening of electron stains.

PubMed

Kaku, Hiroki; Inoue, Kanako; Muranaka, Yoshinori; Park, Pyoyun; Ikeda, Kenichi

2015-10-01

Uranyl salts are toxic and radioactive; therefore, several studies have been conducted to screen for substitutes of electron stains. In this regard, the contrast evaluation process is time consuming and the results obtained are inconsistent. In this study, we developed a novel contrast evaluation method using affinity beads and a backscattered electron image (BSEI), obtained using scanning electron microscopy. The contrast ratios of BSEI in each electron stain treatment were correlated with those of transmission electron microscopic images. The affinity beads bound to cell components independently. Protein and DNA samples were enhanced by image contrast treated with electron stains; however, this was not observed for sugars. Protein-conjugated beads showed an additive effect of image contrast when double-stained with lead. However, additive effect of double staining was not observed in DNA-conjugated beads. The varying chemical properties of oligopeptides showed differences in image contrast when treated with each electron stain. This BSEI-based evaluation method not only enables screening for alternate electron stains, but also helps analyze the underlying mechanisms of electron staining of cellular structures. © The Author 2015. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
ALK-positive large B-cell lymphoma: identification of EML4-ALK and a review of the literature focusing on the ALK immunohistochemical staining pattern.

PubMed

Sakamoto, Kana; Nakasone, Hideki; Togashi, Yuki; Sakata, Seiji; Tsuyama, Naoko; Baba, Satoko; Dobashi, Akito; Asaka, Reimi; Tsai, Chien-Chen; Chuang, Shih-Sung; Izutsu, Koji; Kanda, Yoshinobu; Takeuchi, Kengo

2016-04-01

Anaplastic lymphoma kinase-positive large B-cell lymphoma (ALK+LBCL) is a rare, aggressive B-cell lymphoma with ALK fusion genes. Histopathologically, the ALK immunohistochemical staining pattern is suggestive of the fusion partner of ALK. Here, we examined an ALK+LBCL case showing a unique diffuse cytoplasmic ALK staining pattern and identified EML4-ALK, which has not previously been reported in ALK+LBCL. Furthermore, to clarify whether the prognosis differs depending on the staining pattern, we reviewed 112 previously reported cases, and analyzed immunohistochemical markers and clinical data stratified by the staining pattern. We found that ALK staining can be classified into a granular cytoplasmic staining (GCS) or a non-GCS patterns. Sixty-four adult cases for which both the ALK staining pattern and survival time were reported were further analyzed for survival trends. The non-GCS pattern was significantly associated with inferior overall survival (P = 0.031). This difference remained significant after adjusting for age and clinical stage (hazard ratio 5.08, 95 % CI 1.88-13.7, P = 0.0013). Given that the ALK immunohistochemical staining pattern is associated with the ALK fusion partner, the present results suggest that the prognosis for ALK+LBCL differs depending on the ALK fusion partner.
Histopathological findings in immunohistological staining of the granulomatous tissue reaction associated with tuberculosis.

PubMed

Karimi, Shirin; Shamaei, Masoud; Pourabdollah, Mihan; Sadr, Makan; Karbasi, Mehrdad; Kiani, Arda; Bahadori, Moslem

2014-01-01

Purpose. The histological diagnosis of Mycobacterium tuberculosis (MTB) remains a diagnostic challenge despite different methods. Immunohistochemistry (IHC) not only could confirm granulomatous tissue involvement but also can demonstrate MTB antigen immunolocalization. This study tries to clarify the details of immunohistochemical staining for MTB with pAbBCG. Materials/Methods. Twenty-three confirmed TB granulomatous tissue samples were studied by Ziehl-Neelsen and immunohistochemistry (IHC) staining with pAbBCG. Samples were selected from the archive of the Department of Pathology, National Research Institute of Tuberculosis and Lung Disease, Tehran, Iran. Results. IHC staining was positive in all samples, whereas Ziehl-Neelsen was positive in 9 cases out of 23 (39.1%). Tissue types used were pleural tissue, lymph nodes, and lung tissue. IHC showed positive coarse granular cytoplasmic and round, fragmented bacillary staining. In this study, epithelioid cells clearly showed more positive staining at the periphery of the granuloma rather than the center of granuloma. There is also positive staining in endothelial cells, fibroblasts, plasma cells, lymphocytes, and macrophages outside the granuloma. Conclusion. Considering the criteria of positive immunohistochemical staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize its distribution in different cells.
Histopathological Findings in Immunohistological Staining of the Granulomatous Tissue Reaction Associated with Tuberculosis

PubMed Central

Karimi, Shirin; Pourabdollah, Mihan; Sadr, Makan; Karbasi, Mehrdad; Bahadori, Moslem

2014-01-01

Purpose. The histological diagnosis of Mycobacterium tuberculosis (MTB) remains a diagnostic challenge despite different methods. Immunohistochemistry (IHC) not only could confirm granulomatous tissue involvement but also can demonstrate MTB antigen immunolocalization. This study tries to clarify the details of immunohistochemical staining for MTB with pAbBCG. Materials/Methods. Twenty-three confirmed TB granulomatous tissue samples were studied by Ziehl-Neelsen and immunohistochemistry (IHC) staining with pAbBCG. Samples were selected from the archive of the Department of Pathology, National Research Institute of Tuberculosis and Lung Disease, Tehran, Iran. Results. IHC staining was positive in all samples, whereas Ziehl-Neelsen was positive in 9 cases out of 23 (39.1%). Tissue types used were pleural tissue, lymph nodes, and lung tissue. IHC showed positive coarse granular cytoplasmic and round, fragmented bacillary staining. In this study, epithelioid cells clearly showed more positive staining at the periphery of the granuloma rather than the center of granuloma. There is also positive staining in endothelial cells, fibroblasts, plasma cells, lymphocytes, and macrophages outside the granuloma. Conclusion. Considering the criteria of positive immunohistochemical staining of TB granulomatous reactions, this stain not only highlights the presence of mycobacterial antigens for tissue diagnosis, but also could morphologically localize its distribution in different cells. PMID:24511393
Immunohistochemical expression of p53 proteins in Wilms' tumour: a possible association with the histological prognostic parameter of anaplasia.

PubMed

Cheah, P L; Looi, L M; Chan, L L

1996-01-01

Wilms' tumour (nephroblastoma) has been associated with chromosomal abnormalities at the 11p13, 11p15 and 16q regions. A study into the possibility of mutations occurring within p53, the ubiquitous adult tumour suppressor gene, in Wilms' tumour was carried out. Thirty-eight cases were studied. Of these 36 were categorised into the favourable histology group and two into the unfavourable histology group based on the National Wilms' Tumour Study criteria. Archival formalin-fixed, paraffin-embedded tissue sections from each case were stained with a polyclonal (AB565:Chemicon) and a monoclonal (DO7:Dako) antibody raised against p53 protein using a peroxidase-labelled streptavidin biotin kit (Dako). 'Cure' (disease-free survival of 60 months or longer) was documented in 39% of cases with favourable histology tumours. Eleven percent in this group succumbed to the disease. Both cases with unfavourable histology died. Four out of 36 (11%) tumours with favourable histology demonstrated weak to moderate staining with both AB565 and DO7 in more than 75% of tumour cells. In contrast, p53 protein expression in unfavourable histology tumours was significantly increased compared with the favourable histology group (P = 0.021) with both cases demonstrating immunopositivity in > 75% of tumour cells when stained with AB565 and DO7. The intensity of staining ranged from moderate to strong in both cases. It appears from this preliminary study that the immunohistochemical expression of p53 protein in Wilms' tumour, presumably a result of mutation in the p53 tumour suppressor gene, correlates with histological classification, histological categorisation being one of the useful features in the prognostic assessment of Wilms' tumours.
Osteogenic properties of PBLG-g-HA/PLLA nanocomposites.

PubMed

Liao, Lan; Yang, Shuang; Miron, Richard J; Wei, Junchao; Zhang, Yufeng; Zhang, Meng

2014-01-01

New development of biomaterial scaffolds remains a prominent issue for the regeneration of lost or fractured bone. Of these scaffolds, a number of bioactive polymers have been synthesized and fabricated for diverse biological roles. Although recent evidence has demonstrated that composite scaffolds such as HA/PLLA have improved properties when compared to either HA or PLLA alone, recent investigations have demonstrated that the phase compatibility between HA and PLLA layers is weak preventing optimal enhancement of the mechanical properties and making the composites prone to breakdown. In the present study, poly (γ-benzyl-L-glutamate) modified hydroxyapatite/(poly (L-lactic acid)) (PBLG-g-HA/PLLA) composite scaffolds were fabricated with improved phase compatibility and tested for their osteogenic properties in 18 Wistar female rats by analyzing new bone formation in 3 mm bilateral femur defects in vivo. At time points, 2, 4 and 8 weeks post surgery, bone formation was evaluated by µ-CT and histological analysis by comparing 4 treatment groups; 1) blank defect, 2) PLLA, 3) HA/PLLA and 4) PBLG-g-HA/PLLA scaffolds. The in vivo analysis demonstrated that new bone formation was much more prominent in HA/PLLA and PBLG-g-HA/PLLA groups as depicted by µ-CT, H&E staining and immunohistochemistry for collagen I. TRAP staining was also utilized to determine the influence of osteoclast cell number and staining intensity to the various scaffolds. No significant differences in either staining intensity or osteoclast numbers between all treatment modalities was observed, however blank defects did contain a higher number of osteoclast-like cells. The results from the present study illustrate the potential of PBLG-g-HA/PLLA scaffolds for bone tissue engineering applications by demonstrating favorable osteogenic properties.
Muscle-specific kinase (MuSK) autoantibodies suppress the MuSK pathway and ACh receptor retention at the mouse neuromuscular junction.

PubMed

Ghazanfari, Nazanin; Morsch, Marco; Reddel, Stephen W; Liang, Simon X; Phillips, William D

2014-07-01

Muscle-specific kinase (MuSK) autoantibodies from myasthenia gravis patients can block the activation of MuSK in vitro and/or reduce the postsynaptic localization of MuSK. Here we use a mouse model to examine the effects of MuSK autoantibodies upon some key components of the postsynaptic MuSK pathway and upon the regulation of junctional ACh receptor (AChR) numbers. Mice became weak after 14 daily injections of anti-MuSK-positive patient IgG. The intensity and area of AChR staining at the motor endplate was markedly reduced. Pulse-labelling of AChRs revealed an accelerated loss of pre-existing AChRs from postsynaptic AChR clusters without a compensatory increase in incorporation of (newly synthesized) replacement AChRs. Large, postsynaptic AChR clusters were replaced by a constellation of tiny AChR microaggregates. Puncta of AChR staining also appeared in the cytoplasm beneath the endplate. Endplate staining for MuSK, activated Src, rapsyn and AChR were all reduced in intensity. In the tibialis anterior muscle there was also evidence that phosphorylation of the AChR β-subunit-Y390 was reduced at endplates. In contrast, endplate staining for β-dystroglycan (through which rapsyn couples AChR to the synaptic basement membrane) remained intense. The results suggest that anti-MuSK IgG suppresses the endplate density of MuSK, thereby down-regulating MuSK signalling activity and the retention of junctional AChRs locally within the postsynaptic membrane scaffold. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.
Increased expression of tyrosine phosphatase SHP-2 in Helicobacter pylori-infected gastric cancer

PubMed Central

Jiang, Jing; Jin, Mei-Shan; Kong, Fei; Wang, Yin-Ping; Jia, Zhi-Fang; Cao, Dong-Hui; Ma, Hong-Xi; Suo, Jian; Cao, Xue-Yuan

2013-01-01

AIM: To explore the alteration of tyrosine phosphatase SHP-2 protein expression in gastric cancer and to assess its prognostic values. METHODS: Three hundred and five consecutive cases of gastric cancer were enrolled into this study. SHP-2 expression was carried out in 305 gastric cancer specimens, of which 83 were paired adjacent normal gastric mucus samples, using a tissue microarray immunohistochemical method. Correlations were analyzed between expression levels of SHP-2 protein and tumor parameters or clinical outcomes. Serum anti-Helicobacter pylori (H. pylori) immunoglobulin G was detected with enzyme-linked immunosorbent assay. Cox proportional hazards model was used to evaluate prognostic values by compassion of the expression levels of SHP-2 and disease-specific survivals in patients. RESULTS: SHP-2 staining was found diffuse mainly in the cytoplasm and the weak staining was also observed in the nucleus in gastric mucosa cells. Thirty-two point five percent of normal epithelial specimen and 62.6% of gastric cancer specimen were identified to stain with SHP-2 antibody positively (P < 0.001). Though SHP-2 staining intensities were stronger in the H. pylori (+) group than in the H. pylori (-) group, no statistically significant difference was found in the expression levels of SHP-2 between H. pylori (+) and H. pylori (-) gastric cancer (P = 0.40). The SHP-2 expression in gastric cancer was not significantly associated with cancer stages, lymph node metastases, and distant metastasis of the tumors (P = 0.34, P = 0.17, P = 0.52). Multivariate analysis demonstrated no correlation between SHP-2 expression and disease-free survival (P = 0.86). CONCLUSION: Increased expression of SHP-2 protein in gastric cancer specimen suggesting the aberrant up-regulation of SHP-2 protein might play an important role in the gastric carcinogenesis. PMID:23382639
Localization of Alpha-Keratin and Beta-Keratin (Corneous Beta Protein) in the Epithelium on the Ventral Surface of the Lingual Apex and Its Lingual Nail in the Domestic Goose (Anser Anser f. domestica) by Using Immunohistochemistry and Raman Microspectroscopy Analysis.

PubMed

Skieresz-Szewczyk, Kinga; Jackowiak, Hanna; Buchwald, Tomasz; Szybowicz, Mirosław

2017-08-01

The epithelium of the ventral surface of the apex of the tongue in most birds is specified by the presence of the special superficial layer called lingual nail. The aim of the present study is to determine the localization of the alpha-keratin and beta-keratin (corneous beta protein) in this special epithelium in the domestic goose by using immunohistochemistry staining and the Raman spectroscopy analysis. Due to lack of commercially available antibodies to detect beta-keratin (corneous beta protein), the Raman spectroscopy was used as a specific tool to detect and describe the secondary structure of proteins. The immunohistochemical (IHC) detections reveal the presence of alpha-keratin in all layers of the epithelium, but significant differences in the distribution of the alpha-keratin in the epithelial layers appear. The staining reaction is stronger from the basal layer to the upper zone of the intermediate layer. The unique result is weak staining for the alpha-keratin in the lingual nail. Applications of the Raman spectroscopy as a complementary method not only confirmed results of IHC staining for alpha-keratin, but showed that this technique could be used to demonstrate the presence of beta-keratin (corneous beta protein). Functionally, the localization of alpha-keratin in the epithelium of the ventral surface of the lingual apex provides a proper scaffold for epithelial cells and promotes structural integrity, whereas the presence of beta-keratin (corneous beta protein) in the lingual nail, described also as exoskeleton of the ventral surface of the apex, endures mechanical stress. Anat Rec, 300:1361-1368, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
[Application of Warthin-Starry stain, immunohistochemistry and transmission electron microscopy in diagnosis of cat scratch disease].

PubMed

Huang, Juan; Dai, Lin; Lei, Song; Liao, Dian-ying; Wang, Xiao-qing; Luo, Tian-you; Chen, Yu; Hang, Zhen-biao; Li, Gan-di; Dong, Dan-dan; Xu, Gang; Gu, Zheng-ce; Hao, Ji-ling; Hua, Ping; He, Lei; Duan, Fang-lei

2010-04-01

To evaluate the diagnostic utility of Warthin-Starry silver stain, immunohistochemistry and transmission electron microscopy in the detection of human Bartonella henselae infection and pathologic diagnosis of cat scratch disease (CSD). The paraffin-embedded lymph node tissues of 77 histologically-defined cases of cat scratch disease collected during the period from January, 1998 to December, 2008 were retrieved and studied using Warthin-Starry silver stain (WS stain) and mouse monoclonal antibody against Bartonella henselae (BhmAB stain). Five cases rich in bacteria were selected for transmission electron microscopy. Under electron microscope, the organisms Bartonella henselae appeared polymorphic, round, elliptical, short rod or bacilliform shapes, ranged from 0.489 to 1.110 microm by 0.333 to 0.534 microm and often clustered together. Black short rod-shaped bacilli arranged in chains or clumps were demonstrated in 61.0% (47/77) of CSD by WS stain. The organisms were located outside the cells and lie mainly in the necrotic debris, especially near the nodal capsule. In 72.7% (56/77) of the cases, dot-like, granular as well as few linear positive signals were observed using BhmAB immunostain and showed similar localization. Positive results for both stains were identified in 59.7% (46/77) of the cases. When applying both stains together, Bartonella henselae was observed in 74.0% (57/77) of the case. The difference between the results obtained by WS stain and BhmAB immunostain was of statistical significance (P < 0.05). Bartonella henselae is the causative pathogen of cat scratch disease. WS stain, BhmAB immunostain and transmission electron microscopy are helpful in confirming the histologic diagnosis. Immunostaining using BhmAB can be a better alternative than WS stain in demonstrating the organisms.
Diagnostic accuracy of intracellular mycobacterium tuberculosis detection for tuberculous meningitis.

PubMed

Feng, Guo-dong; Shi, Ming; Ma, Lei; Chen, Ping; Wang, Bing-ju; Zhang, Min; Chang, Xiao-lin; Su, Xiu-chu; Yang, Yi-ning; Fan, Xin-hong; Dai, Wen; Liu, Ting-ting; He, Ying; Bian, Ting; Duan, Li-xin; Li, Jin-ge; Hao, Xiao-ke; Liu, Jia-yun; Xue, Xin; Song, Yun-zhang; Wu, Hai-qin; Niu, Guo-qiang; Zhang, Li; Han, Cui-juan; Lin, Hong; Lin, Zhi-hui; Liu, Jian-jun; Jian, Qian; Zhang, Jin-she; Tian, Ye; Zhou, Bai-yu; Wang, Jing; Xue, Chang-hu; Han, Xiao-fang; Wang, Jian-feng; Wang, Shou-lian; Thwaites, Guy E; Zhao, Gang

2014-02-15

Early diagnosis and treatment of tuberculous meningitis saves lives, but current laboratory diagnostic tests lack sensitivity. We investigated whether the detection of intracellular bacteria by a modified Ziehl-Neelsen stain and early secretory antigen target (ESAT)-6 in cerebrospinal fluid leukocytes improves tuberculous meningitis diagnosis. Cerebrospinal fluid specimens from patients with suspected tuberculous meningitis were stained by conventional Ziehl-Neelsen stain, a modified Ziehl-Neelsen stain involving cytospin slides with Triton processing, and an ESAT-6 immunocytochemical stain. Acid-fast bacteria and ESAT-6-expressing leukocytes were detected by microscopy. All tests were performed prospectively in a central laboratory by experienced technicians masked to the patients' final diagnosis. Two hundred and eighty patients with suspected tuberculous meningitis were enrolled. Thirty-seven had Mycobacterium tuberculosis cultured from cerebrospinal fluid; 40 had a microbiologically confirmed alternative diagnosis; the rest had probable or possible tuberculous meningitis according to published criteria. Against a clinical diagnostic gold standard the sensitivity of conventional Ziehl-Neelsen stain was 3.3% (95% confidence interval, 1.6-6.7%), compared with 82.9% (95% confidence interval, 77.4-87.3%) for modified Ziehl-Neelsen stain and 75.1% (95% confidence interval, 68.8-80.6%) for ESAT-6 immunostain. Intracellular bacteria were seen in 87.8% of the slides positive by the modified Ziehl-Neelsen stain. The specificity of modified Ziehl-Neelsen and ESAT-6 stain was 85.0% (95% confidence interval, 69.4-93.8%) and 90.0% (95% confidence interval, 75.4-96.7%), respectively. Enhanced bacterial detection by simple modification of the Ziehl-Neelsen stain and an ESAT-6 intracellular stain improve the laboratory diagnosis of tuberculous meningitis.
Investigation of the Application of miR10b and miR135b in the Identification of Semen Stains

PubMed Central

Xue, Tianyu; Ma, Xiaoyan; Zhang, Jinxiang; Ou, Xueling; Cheng, Jianding; Sun, Hongyu

2015-01-01

To evaluate the identification method using the microRNA markers miR10b and miR135b to distinguish semen stains from menstrual blood, peripheral blood, vaginal fluid and so on body fluid stains. The expression levels of miR10b and miR35b in semen stains and menstrual blood and so on were detected utilizing a real-time quantitative PCR technique with a specific fluorescence-labeled TaqMan probe. RNU6b was used as the internal reference gene; the difference in their expression was analyzed, and the specificity, sensitivity, and detection capability of the techniques were evaluated. The expression of miR10b and miR135b in semen stains was significantly higher than that of other body fluid stains, with a mean value of ΔCт from-6 to-7. However, it ranged from-2 to-4 for other body fluid stains. The initial criteria for judging which semen stains can be identified were determined by analyzing the research results. When the threshold value was set to 0.04, the CT value could be detected in the target genes miR10b, miR135b and in the internal reference gene RNU6b, and CT values are<40, ΔCT[10b-U6]<-5.5, and ΔCT[135b-U6]<-6, respectively, and the semen stain could be identified. The expression levels of miR10b and miR135b are higher in semen with strong tissue specificity; thus, they can be used to differentiate semen stains from other body fluid stains in forensic science. PMID:26355456
[Effect of staining method and sintering temperature on the color of porcelain-fused-to-metal restorations].

PubMed

Wang, Yan; Yan, Wei-hao; Zhang, Xin-chun; Huang, Yue; Shen, Lin-han

2004-12-01

To investigate the effect of staining method and sintering temperature on the color of porcelain-fused-metal restorations. 40 cylindrical stained porcelain-metal specimens of 15 mm in diameter and 6 mm in height were fabricated with customized mould, consisting of 2 mm Ni-Cr metal, 1 mm opaque porcelain, 2 mm dentine porcelain and 1 mm enamel porcelain. The specimens were prepared by 5 techniques, 8 for each group. Group A: internal staining, Group B: external staining, 900 degrees centigrade sintering temperature was used in both A and B group; Group C to E: external staining, with the sintering temperature of 880 degrees centigrade, 900 degrees centigrade and 920 degrees centigrade respectively. Sofu A2 porcelain and Sofu 44 stain system were used for the study. Using standard white plate as a reference, colors (L*,a*,b* coordinates) of the specimens were measured with a computerized colorimeter. Student's t test and one way ANOVA were used to analyze the data. DeltaE, b* and Delta C(ab)* of Group B (external staining) and group A (internal staining) were 43.72 +/- 2.99/26.51 +/- 1.64/31.31 +/- 2.48 and 39.71 +/- 1.78/23.69 +/- 0.36/26.55 +/- 2.16, respectively. The values of the former group were significantly higher than that of the latter (P < 0.05); For Group C to E, there were no significant differences in all the color parameters. Staining method has a significant effect on the color parameters of porcelain-fused-to-metal restorations; For external staining, within the clinically-used range, changing the sintering temperature does not have an obvious effect on the color.
Should gram stains have a role in diagnosing hip arthroplasty infections?

PubMed

Johnson, Aaron J; Zywiel, Michael G; Stroh, D Alex; Marker, David R; Mont, Michael A

2010-09-01

The utility of Gram stains in diagnosing periprosthetic infections following total hip arthroplasty has recently been questioned. Several studies report low sensitivity of the test, and its poor ability to either confirm or rule out infection in patients undergoing revision total hip arthroplasty. Despite this, many institutions including that of the senior author continue to perform Gram stains during revision total hip arthroplasty. We assessed the sensitivity, specificity, accuracy, and positive and negative predictive values of Gram stains from surgical-site samplings taken from procedures on patients with both infected and aseptic revision total hip arthroplasties. A review was performed on patients who underwent revision total hip arthroplasty between 2000 and 2007. Eighty-two Gram stains were performed on patients who had infected total hip arthroplasties and underwent revision procedures. Additionally, of the 410 revision total hip arthroplasties performed on patients who were confirmed infection-free, 120 Gram stains were performed. Patients were diagnosed as infected using multiple criteria at the time of surgery. Sensitivity, specificity, positive and negative predictive values, and accuracy were calculated from these Gram stain results. The Gram stain demonstrated a sensitivity and specificity of 9.8% and 100%, respectively. In this series, the Gram stain had a negative predictive value of 62%, a positive predictive value of 100%, and an accuracy of 63%. Gram stains obtained from surgical-site samples had poor sensitivity and poor negative predictive value. Based on these findings, as well as those of other authors, we believe that Gram stains should no longer be considered for diagnosing infections in revision total hip arthroplasty. Level III, diagnostic study. See Guidelines for Authors for a complete description of levels of evidence.

Quantification of histochemical stains using whole slide imaging: development of a method and demonstration of its usefulness in laboratory quality control.

PubMed

Gray, Allan; Wright, Alex; Jackson, Pete; Hale, Mike; Treanor, Darren

2015-03-01

Histochemical staining of tissue is a fundamental technique in tissue diagnosis and research, but it suffers from significant variability. Efforts to address this include laboratory quality controls and quality assurance schemes, but these rely on subjective interpretation of stain quality, are laborious and have low reproducibility. We aimed (1) to develop a method for histochemical stain quantification using whole slide imaging and image analysis and (2) to demonstrate its usefulness in measuring staining variation. A method to quantify the individual stain components of histochemical stains on virtual slides was developed. It was evaluated for repeatability and reproducibility, then applied to control sections of an appendix to quantify H&E staining (H/E intensities and H:E ratio) between automated staining machines and to measure differences between six regional diagnostic laboratories. The method was validated with <0.5% variation in H:E ratio measurement when using the same scanner for a batch of slides (ie, it was repeatable) but was not highly reproducible between scanners or over time, where variation of 7% was found. Application of the method showed H:E ratios between three staining machines varied from 0.69 to 0.93, H:E ratio variation over time was observed. Interlaboratory comparison demonstrated differences in H:E ratio between regional laboratories from 0.57 to 0.89. A simple method using whole slide imaging can be used to quantify and compare histochemical staining. This method could be deployed in routine quality assurance and quality control. Work is needed on whole slide imaging devices to improve reproducibility. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.
Putting vital stains in context.

PubMed

Efron, Nathan

2013-07-01

While vital staining remains a cornerstone in the diagnosis of ocular disease and contact lens complications, there are many misconceptions regarding the properties of commonly used dyes by eye-care practitioners and what is and what is not corneal staining after instillation of sodium fluorescein. Similarly, the proper use and diagnostic utility of rose Bengal and lissamine green B, the other two ophthalmic dyes commonly used for assessing ocular complications, have similarly remained unclear. Due to the limitations of vital stains for definitive diagnosis, concomitant signs and symptoms in addition to a complete patient history are required. Over the past decade, there have been many reports of a type of corneal staining--often referred to as solution-induced corneal staining (SICS)--that is observed with the use of multipurpose solutions in combination with soft lenses, more specifically silicone hydrogel lenses. Some authors believe that SICS is a sign of lens/solution incompatibility; however, new research shows that SICS may be neither a measure of lens/solution biocompatibility nor 'true' corneal staining, as that observed in pathological situations. A large component of SICS may be a benign phenomenon, known as preservative-associated transient hyperfluorescence (PATH). There is a lack of correlated signs and/or symptoms with SICS/PATH. Several properties of SICS/PATH, such as appearance and duration, differentiate it from pathological corneal staining. This paper reviews the properties of vital stains, their use and limitations in assessment of the ocular surface, the aetiology of corneal staining, characteristics of SICS/PATH that differentiate it from pathological corneal staining and what the SICS/PATH phenomenon means for contact lens-wearing patients. © 2012 The Author. Clinical and Experimental Optometry © 2012 Optometrists Association Australia.
A Randomized Controlled Clinical Trial to Evaluate Extrinsic Stain Removal of a Whitening Dentifrice.

PubMed

Terézhalmy, Géza; He, Tao; Anastasia, Mary Kay; Eusebio, Rachelle

2016-12-01

To evaluate the extrinsic stain removal efficacy of a new whitening dentifrice containing sodium hexametaphosphate (SHMP) over a two-week period. This study used a controlled and randomized, examiner-blind, single-center, two-treatment, parallel group design. Subjects with visible extrinsic dental stain on facial surfaces of their anterior teeth, and meeting all study criteria, were entered into the trial. The test group received the whitening dentifrice with sodium fluoride and SHMP and an ADA reference soft manual toothbrush. Subjects in the control group received a dental prophylaxis after the initial examination at Baseline and were instructed to use their usual oral hygiene products at home. Subjects returned at Day 3 and Week 2 for re-evaluation of extrinsic dental stain. Extrinsic stain was measured using the Interproximal Modified Lobene (IML) Stain Index; safety was assessed based on clinical examination. Fifty subjects (mean age 32.0 years) completed the study, with 25 in each group. Statistically significant reductions in composite stain for whole tooth, as well as interproximal, gingival, and body surfaces were observed for both groups at Day 3 and Week 2 (p < 0.0001) with no significant differences between the two groups (p > 0.3). At Day 3, median percent reductions in composite IML stain from Baseline were 98% for the prophylaxis group and 100% for the test dentifrice group. At Week 2, median percent reductions in composite IML stain were 100% compared to Baseline for both groups. No adverse events were reported for either group. The whitening dentifrice demonstrated a statistically significant reduction in IML stain after three days and two weeks of use relative to baseline. Stain reduction with the toothpaste was comparable to a dental prophylaxis.
[Technical recommendations and best practice guidelines for May-Grünwald-Giemsa staining: literature review and insights from the quality assurance].

PubMed

Piaton, Eric; Fabre, Monique; Goubin-Versini, Isabelle; Bretz-Grenier, Marie-Françoise; Courtade-Saïdi, Monique; Vincent, Serge; Belleannée, Geneviève; Thivolet, Françoise; Boutonnat, Jean; Debaque, Hervé; Fleury-Feith, Jocelyne; Vielh, Philippe; Cochand-Priollet, Béatrix; Egelé, Caroline; Bellocq, Jean-Pierre; Michiels, Jean-François

2015-08-01

May-Grünwald-Giemsa (MGG) stain is a Romanowsky-type, polychromatic stain as those of Giemsa, Leishman and Wright. Apart being the reference method of haematology, it has become a routine stain of diagnostic cytopathology for the study of air-dried preparations (lymph node imprints, centrifuged body fluids and fine needle aspirations). In the context of their actions of promoting the principles of quality assurance in cytopathology, the French Association for Quality Assurance in Anatomic and Cytologic Pathology (AFAQAP) and the French Society of Clinical Cytology (SFCC) conducted a proficiency test on MGG stain in 2013. Results from the test, together with the review of literature data allow pre-analytical and analytical steps of MGG stain to be updated. Recommendations include rapid air-drying of cell preparations/imprints, fixation using either methanol or May-Grünwald alone for 3-10minutes, two-step staining: 50% May-Grünwald in buffer pH 6.8 v/v for 3-5minutes, followed by 10% buffered Giemsa solution for 10-30minutes, and running water for 1-3minutes. Quality evaluation must be performed on red blood cells (RBCs) and leukocytes, not on tumour cells. Under correct pH conditions, RBCs must appear pink-orange (acidophilic) or buff-coloured, neither green nor blue. Leukocyte cytoplasm must be almost transparent, with clearly delineated granules. However, staining may vary somewhat and testing is recommended for automated methods (slide stainers) which remain the standard for reproducibility. Though MGG stain remains the reference stain, Diff-Quik(®) stain can be used for the rapid evaluation of cell samples. Copyright © 2015 Elsevier Masson SAS. All rights reserved.
Comparison of algorithms for blood stain detection applied to forensic hyperspectral imagery

NASA Astrophysics Data System (ADS)

Yang, Jie; Messinger, David W.; Mathew, Jobin J.; Dube, Roger R.

2016-05-01

Blood stains are among the most important types of evidence for forensic investigation. They contain valuable DNA information, and the pattern of the stains can suggest specifics about the nature of the violence that transpired at the scene. Early detection of blood stains is particularly important since the blood reacts physically and chemically with air and materials over time. Accurate identification of blood remnants, including regions that might have been intentionally cleaned, is an important aspect of forensic investigation. Hyperspectral imaging might be a potential method to detect blood stains because it is non-contact and provides substantial spectral information that can be used to identify regions in a scene with trace amounts of blood. The potential complexity of scenes in which such vast violence occurs can be high when the range of scene material types and conditions containing blood stains at a crime scene are considered. Some stains are hard to detect by the unaided eye, especially if a conscious effort to clean the scene has occurred (we refer to these as "latent" blood stains). In this paper we present the initial results of a study of the use of hyperspectral imaging algorithms for blood detection in complex scenes. We describe a hyperspectral imaging system which generates images covering 400 nm - 700 nm visible range with a spectral resolution of 10 nm. Three image sets of 31 wavelength bands were generated using this camera for a simulated indoor crime scene in which blood stains were placed on a T-shirt and walls. To detect blood stains in the scene, Principal Component Analysis (PCA), Subspace Reed Xiaoli Detection (SRXD), and Topological Anomaly Detection (TAD) algorithms were used. Comparison of the three hyperspectral image analysis techniques shows that TAD is most suitable for detecting blood stains and discovering latent blood stains.
Mechanisms of superficial micropunctate corneal staining with sodium fluorescein: the contribution of pooling.

PubMed

Bandamwar, Kalika L; Garrett, Qian; Papas, Eric B

2012-04-01

To establish if sodium fluorescein (SFL) dye accumulation within intercellular spaces on the ocular surface contributes to the appearance of superficial punctate corneal staining. Thirteen subjects bilaterally wore PureVision™ lenses that had been pre-soaked in ReNu MultiPlus® multipurpose solution. After 1h of lens wear, corneal staining with SFL was assessed using a standard slit-lamp technique. Participants who presented with bilateral, corneal staining were selected for further evaluation. A randomly selected eye was rinsed with saline three times. Fellow eyes (control) received no rinsing. After each rinse, the appearance of SFL staining was recorded without any further instillation of the dye. To eliminate any confounding effects of staining due to residual fluorescein in the tear menisci, corneal staining was induced in freshly excised, isolated, rabbit eyes by topical administration of 0.001% PHMB and staining, rinsing and grading were performed as above. Nine out of 13 subjects presented with bilateral diffuse corneal staining (mean grade±SD: 2.4±0.7). The mean staining grades in test and control eyes respectively after each of the three rinses were (1) 2.41±0.41, 2.25±0.69 (p=0.9); (2) 2.34±0.79, 2.1±0.83 (p=0.8); and (3) 1.71±0.65, 1.60±0.79 (p=0.6) there was no significant reduction in staining with rinsing (p>0.05) and no difference was observed between test and control eyes at any sampling-point. Similar observations made in ex vivo rabbit eyes replicated these results. Pooling or accumulation of SFL solution within intercellular spaces does not appear to contribute to the appearance of superficial micropunctate corneal staining. Copyright Â© 2011 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.
A simple and rapid microwave-assisted hematoxylin and eosin staining method using 1,1,1 trichloroethane as a dewaxing and a clearing agent.

PubMed

Temel, S G; Noyan, S; Cavusoglu, I; Kahveci, Z

2005-01-01

The use and practicability of microwave-assisted staining procedures in routine histopathology has been well established for more than 17 years. In the study reported here, we aimed to examine an alternative approach that would shorten the duration of dewaxing and clearing steps of hematoxylin and eosin (H & E) staining of paraffin sections by using a microwave oven. Although xylene is one of the most popular dewaxing and clearing agents, its flammability restricts its use in a microwave oven; thus we preferred 1,1,1 trichloroethane, which is not flammable, as the dewaxing and clearing agent in the present study. In Group I and Group II (control groups), intestine was processed with xylene and 1,1,1 trichloroethane, respectively. The sections were then stained with H & E according to the conventional staining protocol at room temperature and subdivided into two groups according to the duration of dewaxing and clearing in xylene. In Groups III and IV (experimental groups) similar tissues were processed with xylene and 1,1,1 trichloroethane, respectively; however, sections from these groups were divided into four subgroups to study the period required for dewaxing and clearing in 1,1,1 trichloroethane, then stained with H & E in the microwave oven at 360 W for 30 sec. Our conventional H & E staining procedure, which includes dewaxing, staining and clearing of sections, requires approximately 90 min, while our method using 1,1,1 trichloroethane and microwave heating required only 2 min. Our alternative method for H & E staining not only reduced the procedure time significantly, but also yielded staining quality equal or superior to those stained the conventional way. Our results suggest that 1,1,1 trichloroethane can be used effectively and safely as a dewaxing and clearing agent for H & E staining in a microwave oven.
Fluorescence detection of proteins in sodium dodecyl sulfate-polyacrylamide gels using environmentally benign, nonfixative, saline solution.

PubMed

Steinberg, T H; Lauber, W M; Berggren, K; Kemper, C; Yue, S; Patton, W F

2000-02-01

SYPRO Tangerine stain is an environmentally benign alternative to conventional protein stains that does not require solvents such as methanol or acetic acid for effective protein visualization. Instead, proteins can be stained in a wide range of buffers, including phosphate-buffered saline or simply 150 mM NaCl using an easy, one-step procedure that does not require destaining. Stained proteins can be excited by ultraviolet light of about 300 nm or with visible light of about 490 nm. The fluorescence emission maximum of the dye is approximately 640 nm. Noncovalent binding of SYPRO Tangerine dye is mediated by sodium dodecyl sulfate (SDS) and to a lesser extent by hydrophobic amino acid residues in proteins. This is in stark contrast to acidic silver nitrate staining, which interacts predominantly with lysine residues or Coomassie Blue R, which in turn interacts primarily with arginine and lysine residues. The sensitivity of SYPRO Tangerine stain is similar to that of the SYPRO Red and SYPRO Orange stains - about 4-10 ng per protein band. This detection sensitivity is comparable to colloidal Coomassie blue staining and rapid silver staining procedures. Since proteins stained with SYPRO Tangerine dye are not fixed, they can easily be eluted from gels or utilized in zymographic assays, provided that SDS does not inactivate the protein of interest. This is demonstrated with in-gel detection of rabbit liver esterase activity using alpha-naphthyl acetate and Fast Blue BB dye as well as Escherichia coli beta-glucuronidase activity using ELF-97 beta-D-glucuronide. The dye is also suitable for staining proteins in gels prior to their transfer to membranes by electroblotting. Gentle staining conditions are expected to improve protein recovery after electroelution and to reduce the potential for artifactual protein modifications such as the alkylation of lysine and esterification of glutamate residues, which complicate interpretation of peptide fragment profiles generated by mass spectrometry.
A Novel Contrast Stain for the Rapid Diagnosis of Pityriasis Versicolor: A Comparison of Chicago Sky Blue 6B Stain, Potassium Hydroxide Mount and Culture

PubMed Central

Lodha, Nikita; Poojary, Shital Amin

2015-01-01

Background: The mycological study of pityriasis versicolor is usually done by potassium hydroxide (KOH) mount and culture. However, KOH mount lacks a color contrast and requires a trained eye to interpret, while culture is difficult to perform, time consuming and has low sensitivity. Chicago Sky Blue 6B (CSB) is a new contrast stain that highlights the fungal hyphae and spores, blue against a purplish background. Aims and Objectives: This study was done to compare the utility of a novel contrast stain (CSB stain) with KOH mount and culture. Materials and Methods: Skin scrapings from the lesions of 100 clinically diagnosed cases of P. versicolor were subjected to (1) KOH mount and CSB stain for direct microscopic examination and (2) culture using Sabouraud's dextrose agar. The statistical analysis of CSB stain and culture was done using KOH mount as the reference method, as it is the most commonly performed and practical diagnostic test available for P. versicolor. An interrater reliability analysis using the Cohen's Kappa statistic was performed to determine consistency (agreement) among the different modalities. Observations and Results: Direct microscopy with CSB stain, KOH mount and mycological culture showed positive results in 98 (98%), 92 (92%) and 56 (56%) patients, respectively. Using KOH mount as the reference method, CSB stain had a sensitivity of 100% which was significantly higher than culture (60.9%). Statistically significant fair agreement was found between CSB stain and KOH mount (94% with κ=0.38, P < 0.001). Negligible agreement was found between CSB stain and culture (66%, κ=0.199, P = 0.001) as well as between KOH mount and culture (64%, κ=0.051, P = 0.107). Conclusion: CSB staining of skin scrapings is the most sensitive method for the diagnosis of pityriasis versicolor. Due to the distinct contrast provided by CSB, it is easy to perform, rapid and qualitatively superior to KOH mount. PMID:26288400
A Novel Contrast Stain for the Rapid Diagnosis of Pityriasis Versicolor: A Comparison of Chicago Sky Blue 6B Stain, Potassium Hydroxide Mount and Culture.

PubMed

Lodha, Nikita; Poojary, Shital Amin

2015-01-01

The mycological study of pityriasis versicolor is usually done by potassium hydroxide (KOH) mount and culture. However, KOH mount lacks a color contrast and requires a trained eye to interpret, while culture is difficult to perform, time consuming and has low sensitivity. Chicago Sky Blue 6B (CSB) is a new contrast stain that highlights the fungal hyphae and spores, blue against a purplish background. This study was done to compare the utility of a novel contrast stain (CSB stain) with KOH mount and culture. Skin scrapings from the lesions of 100 clinically diagnosed cases of P. versicolor were subjected to (1) KOH mount and CSB stain for direct microscopic examination and (2) culture using Sabouraud's dextrose agar. The statistical analysis of CSB stain and culture was done using KOH mount as the reference method, as it is the most commonly performed and practical diagnostic test available for P. versicolor. An interrater reliability analysis using the Cohen's Kappa statistic was performed to determine consistency (agreement) among the different modalities. Direct microscopy with CSB stain, KOH mount and mycological culture showed positive results in 98 (98%), 92 (92%) and 56 (56%) patients, respectively. Using KOH mount as the reference method, CSB stain had a sensitivity of 100% which was significantly higher than culture (60.9%). Statistically significant fair agreement was found between CSB stain and KOH mount (94% with κ=0.38, P < 0.001). Negligible agreement was found between CSB stain and culture (66%, κ=0.199, P = 0.001) as well as between KOH mount and culture (64%, κ=0.051, P = 0.107). CSB staining of skin scrapings is the most sensitive method for the diagnosis of pityriasis versicolor. Due to the distinct contrast provided by CSB, it is easy to perform, rapid and qualitatively superior to KOH mount.
Drip bloodstain appearance on inclined apparel fabrics: Effect of prior-laundering, fibre content and fabric structure.

PubMed

de Castro, Therese C; Carr, Debra J; Taylor, Michael C; Kieser, Jules A; Duncan, Warwick

2016-09-01

The interaction of blood and fabrics is currently a 'hot topic', since the understanding and interpretation of these stains is still in its infancy. A recent simplified perpendicular impact experimental programme considering bloodstains generated on fabrics laid the foundations for understanding more complex scenarios. Blood rarely impacts apparel fabrics perpendicular; therefore a systematic study was conducted to characterise the appearance of drip stains on inclined fabrics. The final drip stain appearance for 45° and 15° impact angles on torso apparel fabrics (100% cotton plain woven, 100% polyester plain woven, a blend of polyester and cotton plain woven and 100% cotton single jersey knit) that had been laundered for six, 26 and 52 cycles prior to testing was investigated. The relationship between drop parameters (height and volume), angle and the stain characteristics (parent stain area, axis 1 and 2 and number of satellite stains) for each fabric was examined using analysis of variance. The appearance of the drip stains on these fabrics was distorted, in comparison to drip stains on hard-smooth surface. Examining the parent stain allowed for classification of stains occurring at an angle, however the same could not be said for the satellite stains produced. All of the dried stains visible on the surface of the fabric were larger than just after the impacting event, indicating within fabric spreading of blood due to capillary force (wicking). The cotton-containing fabrics spread the blood within the fabrics in all directions along the stain's circumference, while spreading within the polyester plain woven fabric occurred in only the weft (width of the fabric) and warp (length) directions. Laundering affected the formation of bloodstain on the blend plain woven fabric at both impact angles, although not all characteristics were significantly affected for the three impact conditions considered. The bloodstain characteristics varied due to the fibre content and fabric structure for both impact angles investigated. It is therefore necessary to consider the age of the fabric (which is fabric specific), the fibre type (including blends) and the fabric structure, before interpreting bloodstain patterns. An understanding of this simplified inclined drip stain interaction has been investigated to generate a basis for more complex interactions, such as spatter bloodstains. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.
[Isolation and identification of dog periodontal ligament stem cells].

PubMed

Chang, Xiu-Mei; Liu, Hong-Wei; Jin, Yan; Liu, Yuan; He, Hui-Xia

2009-02-01

To isolate, culture and identify a dog periodontal ligament stem cells (PDLSC) line in vitro. The adult dog periodontal ligament cells were isolated by limited dilution of culture cell for single cell clone. Cells originated from one of these clones were assessed through colony-forming efficiency and immunocytochemistry assay and alkaline phosphatase stain was used to identify the source of adult dog periodontal stem cells, at the same time, PDLSC were induced with mineralizatin solution and was found to have long protrude like an osteoblast. Differentiation of PDLSC were assessed. Mineralized potential was studied by Von-Kossa staining. The dog PDLSC expressed STRO-1, which was the marker of mesenchymal stem cells. Also Vimentin, osteoblast-like marker alkaline phosphatase and Collagen-I expressed weakly. Cells were clonegenic, highly proliferative cells and capable of differentiating into osteoblasts/cementoblasts. The evidence suggests that the cultured cells were stem cells from adult dog periodontal ligament.
A Novel Mutation in DMD (c.10797+5G>A) Causes Becker Muscular Dystrophy Associated with Intellectual Disability.

PubMed

Banihani, Rudaina; Baskin, Berivan; Halliday, William; Kobayashi, Jeff; Kawamura, Anne; McAdam, Laura; Ray, Peter N; Yoon, Grace

2016-04-01

Severe intellectual disability has been reported in a subgroup of patients with Duchenne muscular dystrophy but is not typically associated with Becker muscular dystrophy. The authors report a 13-year-old boy, with severe intellectual disability (Wechsler Intelligence Scales for Children-IV, Full Scale IQ < 0.1 percentile), attention-deficit hyperactivity disorder, and mild muscle weakness. He had elevated serum creatine kinase and dystrophic changes on muscle biopsy. Dystrophin immunohistochemistry revealed decreased staining with the C-terminal and mid-rod antibodies and essentially absent staining of the N-terminal immunostain. Sequencing of muscle mRNA revealed aberrant splicing due to a c.10797+5G > A mutation in DMD. Dystrophinopathy may be associated with predominantly cognitive impairment and neurobehavioral disorder, and should be considered in the differential diagnosis of unexplained cognitive or psychiatric disturbance in males.
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 under...
Lignin staining ...a limited success in identifying koa growth rings

Treesearch

Herbert L. Wick

1970-01-01

Among the lignin stains tested in trying to identify growth rings in koa (Acacia koa Gray), phloroglucinol was the most effective. The light colored sapwood of mature trees stained readily, with growth rings apparent. But staining failed to emphasize rings in the dark colored heartwood. Growth rings were not apparent on samples from young fast...
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 under...
Preparation of colloidal gold for staining proteins electrotransferred onto nitrocellulose membranes.

PubMed

Yamaguchi, K; Asakawa, H

1988-07-01

This paper describes a simple method of preparing colloidal gold for staining protein blots. Colloidal gold was prepared from 0.005 or 0.01% HAuCl4 by the addition of formalin as a reductant and potassium hydroxide. Staining of small cell carcinoma tissue extract blotted onto nitrocellulose membranes with this colloidal gold solution resulted in the appearance of a large number of clear wine-red bands. The sensitivity of gold staining was 60 times higher than that of Coomassie brilliant blue staining and almost comparable to that of silver staining of proteins in polyacrylamide gel. The sensitivity of this method was also satisfactory in comparison with that of enzyme immunoblotting. The colloidal gold prepared by this method is usable for routine work.
Comparison and evaluation of mitotic figures in oral epithelial dysplasia using crystal violet and Feulgen stain.

PubMed

Rao, Roopa S; Patil, Shankargouda; Agarwal, Anveeta

2014-05-01

Routine staining procedures often pose a problem in differentiating a mitotic cell from an apoptotic cell, deteriorating the reliability of histology grading. Although various new methods have been recommended for identifying mitotic figures (MFs) in tissues, the time factor and cost makes them less feasible. Thus, an attempt was made to evaluate the efficacy of crystal violet and Feulgen reaction in identifying MFs and also to see for any variation in the number of MFs in various grades of Epithelial dysplasia. 1. Using crystal violet and Feulgen stain in the identification and counting of MFs on diagnosed cases of epithelial dysplasia and thereby to evaluate their efficacy. 2. To evaluate the variation in the number of MFs in various grades of epithelial dysplasia. The study sample includes retrieval of 30 formalin fixed paraffin embedded tissue sections diagnosed for different grades of epithelial dysplasia (WHO grading system, 2005) from the archives, Department of Oral Pathology, MSRDC, Bengaluru. Ten tissue sections each of mild, moderate and severe epithelial dysplasia were stained with H&E, Feulgen and 1% crystal violet stains and the number of MFs were counted. Five cases of cervical carcinoma were taken as control. Stained sections were compared, and data obtained was statistically analyzed using the Kruskal-Wallis test. A significant increase in the number of MFs (p = 0.02) was observed in Feulgen stained sections as compared to H&E stain. Feulgen stain can be considered as a simple, reliable, cost-effective and reproducible method of staining MFs.
Histochemical stains as promising means for the laser histochemical surgery of a number of pathologies

NASA Astrophysics Data System (ADS)

Piruzyan, L. A.; Mikhailovskiy, Ye. M.; Piruzyan, A. L.

1999-12-01

The directions of laboratory and clinical studies oriented to experimental confirmation of the priority concept of `laser histochemical surgery' are presented. The goal of the studies is reproduction on experimental model of a number of pathologies (in vivo and in vitro) of the `sensitization to laser radiation by staining' effect. Testing of the histochemical stains as sensitizers to laser irradiation of their `address substrates', i.e. vitally stained intracellular structures which participate in the pathologic processes evolution is under planning. The processes include: (a) metabolic disorders in the brain cells, i.e. disseminated sclerosis; (b) generalized metabolic disorders- -mucopolysaccharidosis and collagenosises (periarteritis nodosa, rheumatism, rheumatoid arthritis, sclerodermia); (3) metabolic disorders in individual organs--vessel atherosclerosis, hypercholesterolemia, myocardial infarction, cardiosclerosis, caries and parodontosis. The conditions of the studies are detailed in the recommendations along the positions: (1) disease name; (2) disease characteristics: (a) pathomorphologic, (b) biochemical; (3) stains revealing the disease signs and recommended for testing; (4) `address substrates' of the stains that are targets for laser radiation; (5) lasers recommended for the testing after the cells staining in vivo in the corresponding pathology; (6) experimental models of the pathologies suggested for the testing; (7) criteria of the stain efficiency as target sensitizer to the laser light (criteria of the `laser sensitization by staining' efficiency). Possible perspectives for the experimental clinical medicine are indicated of common histochemical stains and lasers use and of practice introduction of the `laser histochemical surgery' in the case the described concept is confirmed in experiments and clinically.
Comparative study of efficacy, rapidity of detection, and cost-effectiveness of potassium hydroxide, calcofluor white, and Chicago sky blue stains in the diagnosis of dermatophytoses.

PubMed

Prakash, R; Prashanth, Hosakere Veerappa; Ragunatha, Shivanna; Kapoor, Meenakshi; Anitha, T K; Krishnamurthy, Veena

2016-04-01

The diagnosis of superficial mycosis such as dermatophytosis is often done clinically. However, in difficult cases, a rapid test with high sensitivity and specificity helps in the immediate confirmation and administration of treatment. The efficacy, rapidity of detection, and cost-effectiveness of KOH preparation, calcofluor white (CW) stain, and Chicago sky blue (CSB) stain in the identification of fungal elements were assessed in patients with dermatophytoses attending the dermatology clinic of a tertiary care hospital. All three tests were performed on each sample collected from 73 patients according to standard procedure. The slides were examined after 5 and 30 minutes in × 10 and × 40 magnifications. The sensitivity and specificity of CW and CSB at 5 and 30 minutes were calculated using KOH preparation as the standard test. CSB stain showed highest positivity (94.5%) within 5 minutes when compared to KOH (75.3%) and CW (83.5%). After 30 minutes, positivity increased in KOH (84.9%) and CW stains (89%), but it remained the same in CSB stain. Both CW and CSB stains when compared to 10% KOH are equally sensitive (100%), but CW was more specific (72.7%), particularly at 30 minutes. When cost of performing tests on 100 specimens is considered, KOH, CW, and CSB stains cost Rs 5, 100, and 15, respectively. CSB stain is a better stain for rapid diagnosis of dermatophytoses because of ease of performance, rapidity of detection, better appreciation of morphology of fungal elements, and cost effectiveness. © 2016 The International Society of Dermatology.

Lack of clinical utility of urine gram stain for suspected urinary tract infection in pediatric patients.

PubMed

Cantey, Joseph B; Gaviria-Agudelo, Claudia; McElvania TeKippe, Erin; Doern, Christopher D

2015-04-01

Urinary tract infection (UTI) is one of the most common infections in children. Urine culture remains the gold standard for diagnosis, but the utility of urine Gram stain relative to urinalysis (UA) is unclear. We reviewed 312 pediatric patients with suspected UTI who had urine culture, UA, and urine Gram stain performed from a single urine specimen. UA was considered positive if ≥10 leukocytes per oil immersion field were seen or if either nitrates or leukocyte esterase testing was positive. Urine Gram stain was considered positive if any organisms were seen. Sensitivity, specificity, and positive and negative predictive values were calculated using urine culture as the gold standard. Thirty-seven (12%) patients had a culture-proven UTI. Compared to urine Gram stain, UA had equal sensitivity (97.3% versus 97.5%) and higher specificity (85% versus 74%). Empirical therapy was prescribed before the Gram stain result was known in 40 (49%) patients and after in 42 (51%) patients. The antibiotics chosen did not differ between the two groups (P=0.81), nor did they differ for patients with Gram-negative rods on urine Gram stain compared to those with Gram-positive cocci (P=0.67). From these data, we conclude that UA has excellent negative predictive value that is not enhanced by urine Gram stain and that antibiotic selection did not vary based on the urine Gram stain result. In conclusion, the clinical utility of urine Gram stain does not warrant the time or cost it requires. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
Lack of Clinical Utility of Urine Gram Stain for Suspected Urinary Tract Infection in Pediatric Patients

PubMed Central

Gaviria-Agudelo, Claudia; McElvania TeKippe, Erin; Doern, Christopher D.

2015-01-01

Urinary tract infection (UTI) is one of the most common infections in children. Urine culture remains the gold standard for diagnosis, but the utility of urine Gram stain relative to urinalysis (UA) is unclear. We reviewed 312 pediatric patients with suspected UTI who had urine culture, UA, and urine Gram stain performed from a single urine specimen. UA was considered positive if ≥10 leukocytes per oil immersion field were seen or if either nitrates or leukocyte esterase testing was positive. Urine Gram stain was considered positive if any organisms were seen. Sensitivity, specificity, and positive and negative predictive values were calculated using urine culture as the gold standard. Thirty-seven (12%) patients had a culture-proven UTI. Compared to urine Gram stain, UA had equal sensitivity (97.3% versus 97.5%) and higher specificity (85% versus 74%). Empirical therapy was prescribed before the Gram stain result was known in 40 (49%) patients and after in 42 (51%) patients. The antibiotics chosen did not differ between the two groups (P = 0.81), nor did they differ for patients with Gram-negative rods on urine Gram stain compared to those with Gram-positive cocci (P = 0.67). From these data, we conclude that UA has excellent negative predictive value that is not enhanced by urine Gram stain and that antibiotic selection did not vary based on the urine Gram stain result. In conclusion, the clinical utility of urine Gram stain does not warrant the time or cost it requires. PMID:25653411
The role of serum amyloid A staining of granulomatous tissues for the diagnosis of sarcoidosis.

PubMed

Huho, Albert; Foulke, Llewellyn; Jennings, Timothy; Koutroumpakis, Efstratios; Dalvi, Siddhartha; Chaudhry, Haroon; Chopra, Amit; Modi, Aakash; Rane, Neha; Prezant, David J; Sheehan, Christine; Yucel, Recai; Patel, Mehul; Judson, Marc A

2017-05-01

Previous studies demonstrated that SAA staining of sarcoidosis granulomas was qualitatively and quantitatively different from other granulomatous diseases. These data suggest that positive SAA staining of granulomatous tissue may have adequate specificity to establish a diagnosis of sarcoidosis. Our objective was to determine the diagnostic specificity of SAA staining for sarcoidosis relative to other granulomatous disorders. Pathological specimens demonstrating granulomatous inflammation were retrospectively identified at one institution, plus 4 specimens were obtained from New York City firefighters with biopsy-confirmed World Trade Center "sarcoidosis-like" pulmonary disease. Specimens were analyzed if specific diagnoses related to the granulomatous inflammation were confirmed through medical record review. SAA staining was performed using previously developed methods. Two pathologists, blinded to each other and the diagnoses, determined if the stained material was SAA positive or negative. Discordant results were adjudicated by the two pathologists. 106 specimens were analyzed from 100 patients, with 36 biopsies (34%) from sarcoidosis tissues and 70 (66%) from other granulomatous disorders. The Cohen Kappa correlation between the two pathologists for SAA staining positivity was excellent (0.85, 0.73-0.98). The overall specificity of SAA staining for the diagnosis of sarcoidosis was 84% (59/70). The sensitivity was 44% (16/36). Although SAA staining of various granulomatous tissues was fairly specific for the diagnosis of sarcoidosis, the specificity was inadequate for SAA staining to be used as a diagnostic test for sarcoidosis in isolation. These data suggest that SAA production may not be a universal mechanism in the development of sarcoidosis. Copyright © 2017 Elsevier Ltd. All rights reserved.
Immunohistochemical Analysis of Histone H3 Modifications in Germ Cells during Mouse Spermatogenesis

PubMed Central

Song, Ning; Liu, Jie; An, Shucai; Nishino, Tomoya; Hishikawa, Yoshitaka; Koji, Takehiko

2011-01-01

Histone modification has been implicated in the regulation of mammalian spermatogenesis. However, the association of differently modified histone H3 with a specific stage of germ cells during spermatogenesis is not fully understood. In this study, we examined the localization of variously modified histone H3 in paraffin-embedded sections of adult mouse testis immunohistochemically, focusing on acetylation at lysine 9 (H3K9ac), lysine 18 (H3K18ac), and lysine 23 (H3K23ac); tri-methylation at lysine 4 (H3K4me3) and lysine 27 (H3K27me3); and phosphorylation at serine 10 (H3S10phos). As a result, we found that there was a significant fluctuation in the modifications; in spermatogonia, the stainings for H3K9ac, H3K18ac, and H3K23ac were strong while that for H3K4me3 was weak. In spermatocytes, the stainings for H3K9ac, H3K18ac, H3K23ac, and H3K4me3 were reduced in the preleptotene to pachytene stage, but in diplotene stage the stainings for H3K18ac, H3K23ac, and H3K4me3 seemed to become intense again. The staining for H3K27me3 was nearly constant throughout these stages. In the ensuing spermiogenesis, a dramatic acetylation and methylation of histone H3 was found in the early elongated spermatids and then almost all signals disappeared in the late elongated spermatids, in parallel with the replacement from histones to protamines. In addition, we confirmed that the staining of histone H3S10phos was exclusively associated with mitotic and meiotic cell division. Based upon the above results, we indicated that the modification pattern of histone H3 is subject to dynamic change and specific to a certain stage of germ cell differentiation during mouse spermatogenesis. PMID:21927517
Immunohistochemical Analysis of Histone H3 Modifications in Germ Cells during Mouse Spermatogenesis.

PubMed

Song, Ning; Liu, Jie; An, Shucai; Nishino, Tomoya; Hishikawa, Yoshitaka; Koji, Takehiko

2011-08-27

Histone modification has been implicated in the regulation of mammalian spermatogenesis. However, the association of differently modified histone H3 with a specific stage of germ cells during spermatogenesis is not fully understood. In this study, we examined the localization of variously modified histone H3 in paraffin-embedded sections of adult mouse testis immunohistochemically, focusing on acetylation at lysine 9 (H3K9ac), lysine 18 (H3K18ac), and lysine 23 (H3K23ac); tri-methylation at lysine 4 (H3K4me3) and lysine 27 (H3K27me3); and phosphorylation at serine 10 (H3S10phos). As a result, we found that there was a significant fluctuation in the modifications; in spermatogonia, the stainings for H3K9ac, H3K18ac, and H3K23ac were strong while that for H3K4me3 was weak. In spermatocytes, the stainings for H3K9ac, H3K18ac, H3K23ac, and H3K4me3 were reduced in the preleptotene to pachytene stage, but in diplotene stage the stainings for H3K18ac, H3K23ac, and H3K4me3 seemed to become intense again. The staining for H3K27me3 was nearly constant throughout these stages. In the ensuing spermiogenesis, a dramatic acetylation and methylation of histone H3 was found in the early elongated spermatids and then almost all signals disappeared in the late elongated spermatids, in parallel with the replacement from histones to protamines. In addition, we confirmed that the staining of histone H3S10phos was exclusively associated with mitotic and meiotic cell division. Based upon the above results, we indicated that the modification pattern of histone H3 is subject to dynamic change and specific to a certain stage of germ cell differentiation during mouse spermatogenesis.
Impact of changing from staining to culture techniques on detection rates of Campylobacter spp. in routine stool samples in Chile.

PubMed

Porte, Lorena; Varela, Carmen; Haecker, Thomas; Morales, Sara; Weitzel, Thomas

2016-05-13

Campylobacter is a leading cause of bacterial gastroenteritis, but sensitive diagnostic methods such as culture are expensive and often not available in resource limited settings. Therefore, direct staining techniques have been developed as a practical and economical alternative. We analyzed the impact of replacing Campylobacter staining with culture for routine stool examinations in a private hospital in Chile. From January to April 2014, a total of 750 consecutive stool samples were examined in parallel by Hucker stain and Campylobacter culture. Isolation rates of Campylobacter were determined and the performance of staining was evaluated against culture as the gold standard. Besides, isolation rates of Campylobacter and other enteric pathogens were compared to those of past years. Campylobacter was isolated by culture in 46 of 750 (6.1 %) stool samples. Direct staining only identified three samples as Campylobacter positive and reached sensitivity and specificity values of 6.5 and 100 %, respectively. In comparison to staining-based detection rates of previous years, we observed a significant increase of Campylobacter cases in our patients. Direct staining technique for Campylobacter had a very low sensitivity compared to culture. Staining methods might lead to a high rate of false negative results and an underestimation of the importance of campylobacteriosis. With the inclusion of Campylobacter culture, this pathogen became a leading cause of intestinal infection in our patient population.
Negative Stains Containing Trehalose: Application to Tubular and Filamentous Structures

NASA Astrophysics Data System (ADS)

Harris, J. Robin; Gerber, Max; Gebauer, Wolfgang; Wernicke, Wolfgang; Markl, Jürgen

1996-02-01

Several examples are presented that show the successful application of uranyl acetate and ammonium molybdate negative staining in the presence of trehalose for TEM studies of filamentous and tubular structures. The principal benefit to be gained from the inclusion of trehalose stems from the considerably reduced flattening of the large tubular structures and the greater orientational freedom of single molecules due to an increased depth of the negative stain in the presence of trehalose. Trehalose is likely to provide considerable protection to protein molecules and their assemblies during the drying of negatively stained specimens. Some reduction in the excessive density imparted by uranyl acetate around large assemblies is also achieved. Nevertheless, in the presence of 1% (w/v) trehalose, it is desirable to increase the concentration of negative stain to 5% (w/v) for ammonium molybdate and to 4% for uranyl acetate to produce satisfactory image contrast. In general, the ammonium molybdate-trehalose negative stain is more satisfactory than the uranyl acetate-trehalose combination, because of the greater electron beam sensitivity of the uranyl negative stain. Reassembled taxol-stabilized pig brain microtubules, together with collagen fibrils, sperm tails, helical filaments, and reassociated hemocyanin (KLH2), all from the giant keyhole limpet Megathura crenulata, have been studied by negative staining in the presence of trehalose. In all cases satisfactory TEM imaging conditions were readily obtained on the specimens, as long as regions of excessively deep stain were avoided.
Portal vein territory identification using indocyanine green fluorescence imaging: Technical details and short-term outcomes.

PubMed

Kobayashi, Yuta; Kawaguchi, Yoshikuni; Kobayashi, Kosuke; Mori, Kazuhiro; Arita, Junichi; Sakamoto, Yoshihiro; Hasegawa, Kiyoshi; Kokudo, Norihiro

2017-12-01

Portal vein (PV) territory identification during liver resection may be performed using indocyanine green (ICG) fluorescence imaging technique. However, the technical details of the fluorescence staining technique have not been fully elucidated. This study was performed to demonstrate the technical details of PV territory identification using fluorescence imaging and evaluates the short-term outcomes. From 2011 to 2015, 105 underwent liver resection at the University of Tokyo Hospital with one of the following fluorescence staining techniques by transhepatic PV injection or intravenous injection of ICG: single staining (n = 36), multiple staining (n = 31), counterstaining (n = 22), negative staining (n = 13), or paradoxical negative staining (n = 3). The PV territory was identified as a region with fluorescence or a defect of fluorescence using one of the five staining techniques. ICG was administered by transhepatic PV injection in all but the negative staining technique, which employed intravenous injection. No adverse events associated with the ICG administration occurred. The mortality, postoperative total morbidity, and the major complication (Clavien-Dindo grade ≥III) rates were 0.0%, 14.3%, and 7.6%. We have demonstrated the technical details of five types of fluorescence staining techniques. These techniques are safe to perform and facilitate clear visualization of the PV territory in real time, enhancing the efficacy of anatomical removal of such territories. © 2017 Wiley Periodicals, Inc.
Feasibility of a tetracycline-binding method for detecting synovial fluid basic calcium phosphate crystals.

PubMed

Rosenthal, Ann K; Fahey, Mark; Gohr, Claudia; Burner, Todd; Konon, Irina; Daft, Laureen; Mattson, Eric; Hirschmugl, Carol; Ryan, Lawrence M; Simkin, Peter

2008-10-01

Basic calcium phosphate (BCP) crystals are common components of osteoarthritis (OA) synovial fluid. Progress in understanding the role of these bioactive particles in clinical OA has been hampered by difficulties in their identification. Tetracyclines stain calcium phosphate mineral in bone. The aim of this study was to investigate whether tetracycline staining might be an additional or alternative method for identifying BCP crystals in synovial fluid. A drop of oxytetracycline was mixed with a drop of fluid containing synthetic or native BCP, calcium pyrophosphate dihydrate (CPPD), or monosodium urate (MSU) crystals and placed on a microscope slide. Stained and unstained crystals were examined by light microscopy, with and without a portable broad-spectrum ultraviolet (UV) pen light. A small set of characterized synovial fluid samples were compared by staining with alizarin red S and oxytetracycline. Synthetic BCP crystals in synovial fluid were quantified fluorimetrically using oxytetracycline. After oxytetracycline staining, synthetic and native BCP crystals appeared as fluorescent amorphous aggregates under UV light. Oxytetracycline did not stain CPPD or MSU crystals or other particulates. Oxytetracycline staining had fewer false-positive test results than did alizarin red S staining and could provide estimates of the quantities of synthetic BCP crystals in synovial fluid. With further validation, oxytetracycline staining may prove to be a useful adjunct or alternative to currently available methods for identifying BCP crystals in synovial fluid.
Light microscopy with differential staining techniques for the characterisation and discrimination of insects versus marine arthropods processed animal proteins.

PubMed

Ottoboni, Matteo; Tretola, Marco; Cheli, Federica; Marchis, Daniela; Veys, Pascal; Baeten, Vincent; Pinotti, Luciano

2017-08-01

The aim of this study was to evaluate the use of light microscopy with differential staining techniques for the discrimination of insect material from marine arthropods - classified as fishmeal. Specifically, three samples of single-species insect material, Hermetia illucens (HI), Bombyx mori (BM) and Tenebrio molitor (TM), and two samples of marine arthropods, shrimp material and krill, were analysed and compared after staining by two reagents to enhance fragment identification. Alizarin Red (AR) and Chlorazol Black (CB), which react respectively with calcium salts and chitin, were tested for their potential efficacy in distinguishing between insect and marine materials. Results indicated that AR failed to stain HI, BM and TM materials. By contrast, the three insect species materials tested were stained by CB. When shrimp fragments and krill were considered, AR and CB stained marine materials reddish-pink and light blue to black, respectively. By combining these results, it can be suggested that CB staining may efficiently be used to mark insect materials; AR does stain shrimp fragments but does not stain the tested insect material, indicating a possible approach for discriminating between insects and marine arthropods. However, since the present study was performed on pure materials and a small set of samples, possible implementation of this technique still needs to be confirmed in complex matrices such as compound feed.
Exploring methods for prevention of oxidative stain in soft maple

Treesearch

Michael C. Wiemann; Richard D. Bergman; Mark Knaebe; Scott A. Bowe

2009-01-01

Interior gray enzymatic oxidative stain for white woods such as maple has plagued the wood industry for many years because methods that have been found to reduce stain are hard to scale up to industrial levels. We examined possible alternative treatments to eliminate stain in soft maple (Acer rubrum L.), and found that exposure to sulfur dioxide gas eliminates interior...
Test Characteristics of Acridine Orange, Gram, and May-Grünwald-Giemsa Stains for Enumeration of Intracellular Organisms in Bronchoalveolar Lavage Fluid

PubMed Central

De Brauwer, Els; Jacobs, Jan; Nieman, Fred; Bruggeman, Cathrien; Drent, Marjolein

1999-01-01

For enumeration of intracellular organisms (ICO) in bronchoalveolar lavage fluid samples, the May-Grünwald-Giemsa (MGG) stain displayed higher interobserver agreement than the acridine orange and Gram stains. The MGG stain offered a reliable enumeration of ICO when 200 cells were counted by one observer. PMID:9889233
Comparative Tissue Stainability of Lawsonia inermis (Henna) and Eosin as Counterstains to Hematoxylin in Brain Tissues.

PubMed

Alawa, Judith N; Gideon, Gbenga O; Adetiba, Bamidele; Alawa, Clement B

2015-04-01

We hyposthesized that henna staining could provide an alternative to eosin when used as a counterstain to hematoxylin for understanding basic neurohistological principles. Therefore, this study was aimed at investigating the suitability of henna as counterstain to hematoxylin for the demonstration of the layer stratification and cellular distribution in the brain tissue. Henna stained nervous tissue by reacting with the basic elements in proteins via its amino groups. It stained the neuropil and connective tissue membranes brown and effectively outlined the perikarya of neurons with no visible nuclei demonstrating that it is an acidic dye. Henna as a counterstain to hematoxylin demonstrated reliability as a new neurohistological stain. It facilitated identification of cortical layer stratification and cellular distribution in brain tissue sections from Wistar rats. This was comparable to standard hematoxylin and eosin staining as morphological and morphometrical analyses of stained cells did not show significant differences in size or number. This study presents a method for staining with henna and demonstrates that although henna and eosin belong to different dye groups (anthraquinone and xanthenes, respectively) based on their chromophores, they share similar staining techniques and thus could be used interchangeably in neurohistology.
Measuring the Level of Agreement Between Cloacal Gram's Stains and Bacterial Cultures in Hispaniolan Amazon Parrots ( Amazona ventralis ).

PubMed

Evans, Erika E; Mitchell, Mark A; Whittington, Julia K; Roy, Alma; Tully, Thomas N

2014-12-01

Cloacal or fecal Gram's stains and bacterial cultures are routinely performed during avian physical examinations to assess the microbial flora of the gastrointestinal tract. Although cloacal or fecal Gram's stains and bacterial cultures are considered routine diagnostic procedures, the level of agreement between the individual tests has not been determined. To investigate the level of agreement between results from Gram's stain and bacterial culture when used to assess cloacal or fecal samples from psittacine birds, samples were taken from 21 clinically healthy Hispaniolan Amazon parrots ( Amazona ventralis ) and tested by Gram's stain cytology and bacterial culture. Most bacteria (97.2%) identified by Gram's stain were gram positive. However, gram-negative organisms were identified in 7 of 21 (33.3%; 95% confidence interval: 13.3%-53.3%) birds. Escherichia coli was the only gram-negative organism identified on culture. Agreement between results of Gram's stain and culture was fair (weighted κ = 0.27). The results of this study suggest that Gram's stains and bacterial culture may need to be performed with a parallel testing strategy to limit the likelihood of misclassifying the microbial flora of psittacine patients.
Improving Gram stain proficiency in hospital and satellite laboratories that do not have microbiology.

PubMed

Guarner, Jeannette; Street, Cassandra; Matlock, Margaret; Cole, Lisa; Brierre, Francoise

2017-03-01

Consolidation of laboratories has left many hospitals and satellite laboratories with minimal microbiologic testing. In many hospitals and satellite laboratories, Gram stains on primary specimens are still performed despite difficultly in maintaining proficiency. To maintain Gram stain proficiency at a community 450-bed hospital with an active emergency room we designed bimonthly challenges that require reporting Gram staining and morphology of different organisms. The challenges consist of five specimens prepared by the reference microbiology laboratory from cultures and primary specimens. Twenty to 23 medical laboratory scientists participate reading the challenges. Results from the challenges are discussed with each medical laboratory scientists. In addition, printed images from the challenges are presented at huddle to add microbiology knowledge. On the first three challenges, Gram staining was read correctly in 71%-77% of the time while morphology 53%-66%. In the last six challenges correct answers for Gram stain were 77%-99% while morphology 73%-96%. We observed statistically significant improvement when reading Gram stains by providing frequent challenges to medical laboratory scientists. The clinical importance of Gram stain results is emphasized during huddle presentations increasing knowledge and motivation to perform the test for patients.
Comparison of different diagnostic techniques for the detection of cryptosporidiosis in bovines

PubMed Central

Rekha, K. M. H.; Puttalakshmamma, G. C.; D’Souza, Placid E.

2016-01-01

Aim: Aim of the present study was to compare different methods, viz., Sheather's sugar flotation (SSF), Ziehl-Neelsen (ZN), Kinyoun's acid-fast method (KAF), safranin-methylene blue staining (SMB), and negative staining techniques such as nigrosin staining, light green staining, and malachite green staining for the detection of Cryptosporidium spp. oocysts in bovines. Materials and Methods: A total of 455 fecal samples from bovines were collected from private, government farms and from the clinical cases presented to Department of Medicine, Veterinary College, Bengaluru. They were subjected for SSF, ZN, KAF, SMB and negative staining methods. Results: Out of 455 animal fecal samples screened 5.71% were found positive for Cryptosporidium spp. oocysts. The species were identified as Cryptosporidium parvum in calves and Cryptosporidium andersoni in adults based on the morphological characterization and micrometry of the oocysts. Conclusions: Of all the techniques, fecal flotation with sheather's was found to be more specific and sensitive method for the detection of Cryptosporidium spp. oocysts. Among the conventional staining methods, the SMB gives better differentiation between oocysts and yeast. Among the three negative staining methods, malachite green was found sensitive over the other methods. PMID:27051211
A novel washing algorithm for underarm stain removal

NASA Astrophysics Data System (ADS)

Acikgoz Tufan, H.; Gocek, I.; Sahin, U. K.; Erdem, I.

2017-10-01

After contacting with human sweat which comprise around 27% sebum, anti-perspirants comprising aluminium chloride or its compounds form a jel-like structure whose solubility in water is very poor. In daily use, this jel-like structure closes sweat pores and hinders wetting of skin by sweat. However, when in contact with garments, they form yellowish stains at the underarm of the garments. These stains are very hard to remove with regular machine washing. In this study, first of all, we focused on understanding and simulating such stain formation on the garments. Two alternative procedures are offered to form jel-like structures. On both procedures, commercially available spray or deo-stick type anti-perspirants, standard acidic and basic sweat solutions and artificial sebum are used to form jel-like structures, and they are applied on fabric in order to get hard stains. Secondly, after simulation of the stain on the fabric, we put our efforts on developing a washing algorithm specifically designed for removal of underarm stains. Eight alternative washing algorithms are offered with varying washing temperature, amounts of detergent, and pre-stain removal procedures. Better algorithm is selected by comparison of Tristimulus Y values after washing.
A comparative clinical study evaluating stain removal efficacy of a new sensitivity whitening dentifrice compared to commercially available whitening dentifrices.

PubMed

Hughes, Nathan; Maggio, Brenda; Sufi, Farzana; Mason, Stephen; Kleber, Carl J

2009-01-01

To assess the extrinsic stain removal efficacy of a new sensitivity dentifrice containing sodium tripolyphosphate (STP) to marketed whitening toothpastes after six weeks of product use. This was a single-center, double-blind, stratified, six-week clinical study comparing the reduction in stain area and intensity of Sensodyne Extra Whitening to Crest Maximum Strength Sensitivity Protection Whitening plus Scope and Colgate Tartar Control Plus Whitening Mint dentifrice, as measured by MacPherson's Modification of the Lobene Stain Index (MMLSI) in a forced stain model. Two-hundred and ninety-five subjects completed the study. Tooth stain MMSLI scores showed significant differences between Sensodyne and Crest dentifrices in favor of Sensodyne for all surface sites (p = 0.014), and individually for facial (p = 0.023), lingual (p = 0.027), and interproximal (p = 0.014) surfaces. No significant statistical differences between Sensodyne and Colgate dentifrices were observed for any of the surfaces. Results from this stain removal clinical study demonstrate significant extrinsic stain removal efficacy for all dentifrices relative to baseline. Significant differences between the two marketed sensitivity whitening dentifrices were demonstrated in favor of the new Sensodyne Sensitivity Whitening dentifrice.
[Usefulness of sputum Gram staining in community-acquired pneumonia].

PubMed

Sato, Tadashi; Aoshima, Masahiro; Ohmagari, Norio; Tada, Hiroshi; Chohnabayashi, Naohiko

2002-07-01

To evaluate the usefulness of sputum gram staining in community-acquired pneumonia (CAP), we reviewed 144 cases requiring hospitalization in the last 4 years. The sensitivity was 75.5%, specificity 68.2%, positive predictive value 74.1%, negative predictive value 69.8%, positive likelihood ratio 2.37, negative likelihood ratio 0.36 and accuracy 72.2% in 97 cases. Both sputum gram staining and culture were performed. Concerning bacterial pneumonia (65 cases), we compared the Gram staining group (n = 33), which received initial antibiotic treatment, based on sputum gram staining with the Empiric group (n = 32) that received antibiotics empirically. The success rates of the initial antibiotic treatment were 87.9% vs. 78.1% (P = 0.473); mean hospitalization periods were 9.67 vs. 11.75 days (P = 0.053); and periods of intravenous therapy were 6.73 vs. 7.91 days (P = 0.044), respectively. As for initial treatment, penicillins were used in the Gram staining group more frequently (P < 0.01). We conclude that sputum gram staining is useful for the shortening of the treatment period and the appropriate selection of initial antibiotics in bacterial pneumonia. We believe, therefore, that sputum gram staining is indispensable as a diagnostic tool CAP.
Distribution of calmodulin in corn seedlings - Immunocytochemical localization in coleoptiles and root apices

NASA Technical Reports Server (NTRS)

Dauwalder, M.; Roux, S. J.

1986-01-01

Immunofluorescence techniques have been used to study the distribution of calmodulin in several tissues in etiolated corn (Zea mays, var. Bear Hybrid) seedlings. Uniform staining was seen in the background cytoplasm of most cell types. Cell walls and vacuoles were not stained. In coleoptile mesophyll cells the nucleoplasm of most nuclei was stained as was the stroma of most amyloplasts. The lumen border of mature tracheary elements in coleoptiles also stained. In the rootcap the most intensely stained regions were the cytoplasms of columella cells and of the outermost cells enmeshed in the layer of secreted slime. Nuclei in the rootcap cells did not stain distinctly, but those in all cell types of the root meristem did. Also in the root meristem, the cytoplasm of metaxylem elements stained brightly. These results are compared and contrasted with previous data on the localization of calmodulin in pea root apices and epicotyls and discussed in relation to current hypotheses on mechanisms of gravitropism.

A new procedure for fast soft staining of BN-PAGEs on photosynthetic complexes.

PubMed

Farci, Domenica; Kirkpatrick, Joanna; Piano, Dario

2017-02-01

We report a fast and sensitive procedure for blue native PAGE staining, in which the conventional staining step with CBB is avoided. After running, a short exposure to a mix of polar protic solvents (ethanol and acetic acid) leads to a fast and selective removal of the dye from the migration front and a specific binding to the protein bands, while the rest undergo a selective and complete background removal, leading to an intense contrast. This single-step staining-destaining technique is useful in protein samples that bind colored cofactors such as photosystems, which can be selectively discerned by their characteristic green color. After the staining of such samples, the green color persists, while the other unpigmented protein complexes and the molecular standard remain CBB stained, creating a useful reference system for the assignment of the bands. The advantages and chemical basis of this staining procedure are discussed. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Optimal staining methods for delineation of cortical areas and neuron counts in human brains.

PubMed

Uylings, H B; Zilles, K; Rajkowska, G

1999-04-01

For cytoarchitectonic delineation of cortical areas in human brain, the Gallyas staining for somata with its sharp contrast between cell bodies and neuropil is preferable to the classical Nissl staining, the more so when an image analysis system is used. This Gallyas staining, however, does not appear to be appropriate for counting neuron numbers in pertinent brain areas, due to the lack of distinct cytological features between small neurons and glial cells. For cell counting Nissl is preferable. In an optimal design for cell counting at least both the Gallyas and the Nissl staining must be applied, the former staining for cytoarchitectural delineaton of cortical areas and the latter for counting the number of neurons in the pertinent cortical areas. Copyright 1999 Academic Press.
CGP stain: An inexpensive, odorless, rapid, sensitive, and in principle in vitro methylation-free Coomassie Brilliant Blue stain.

PubMed

Yasumitsu, Hidetaro; Ozeki, Yasuhiro; Kawsar, Sarkar M A; Toda, Tosifusa; Kanaly, Robert

2010-11-01

Coomassie Brilliant Blue (CBB) protein stains are inexpensive but detect proteins at only at microgram levels. Because of acetic acid and methanol, they cause skin irritation and reduce work motivation by malodor. Recent mass spectrometric (MS) analyses demonstrated that nanogram-sensitive colloidal CBB staining resulted in in vitro methylations of proteins. We propose a rapid, inexpensive, sensitive, odorless, less harsh, and in vitro methylation-free CBB stain. CGP uses three components: citric acid, CBB G-250, and polyvinylpyrrolidone. CGP detects proteins at 12ng within 45min, and because it is nonalcohol, in principle in vitro methylation would be eliminated. Indeed, MS analysis of CGP-stained bands confirmed a lack of methylation. 2010 Elsevier Inc. All rights reserved.
[Histochemical stains for minerals by hematoxylin-lake method].

PubMed

Miyagawa, Makoto

2013-04-01

The present study was undertaken to establish the experimental animal model by histological staining methods for minerals. After intraperitoneal injections of minerals, precipitates deposited on the surface of the liver. Liver tissues were fixed in paraformaldehyde, embedded in paraffin and cut into thin sections which were used as minerals containing standard section. Several reagents for histological stains and spectrophotometry for minerals were applied in both test-tube experiments and stainings of tissue sections to test for minerals. Hematoxylin-lake was found of capable of staining minerals in tissue. A simple technique used was described for light microscopic detection of minerals.
A combined Bodian-Nissl stain for improved network analysis in neuronal cell culture.

PubMed

Hightower, M; Gross, G W

1985-11-01

Bodian and Nissl procedures were combined to stain dissociated mouse spinal cord cells cultured on coverslips. The Bodian technique stains fine neuronal processes in great detail as well as an intracellular fibrillar network concentrated around the nucleus and in proximal neurites. The Nissl stain clearly delimits neuronal cytoplasm in somata and in large dendrites. A combination of these techniques allows the simultaneous depiction of neuronal perikarya and all afferent and efferent processes. Costaining with little background staining by either procedure suggests high specificity for neurons. This procedure could be exploited for routine network analysis of cultured neurons.
Quantitative proteome analysis of barley seeds using ruthenium(II)-tris-(bathophenanthroline-disulphonate) staining.

PubMed

Witzel, Katja; Surabhi, Giridara-Kumar; Jyothsnakumari, Gottimukkala; Sudhakar, Chinta; Matros, Andrea; Mock, Hans-Peter

2007-04-01

This paper describes the application of the recently introduced fluorescence stain Ruthenium(II)-tris-(bathophenanthroline-disulphonate) (RuBP) on a comparative proteome analysis of two phenotypically different barley lines. We carried out an analysis of protein patterns from 2-D gels of the parental lines of the Oregon Wolfe Barley mapping population DOM and REC and stained with either the conventional colloidal Coomassie Brilliant Blue (cCBB) or with the novel RuBP solution. We wished to experimentally verify the usefulness of such a stain in evaluating the complex pattern of a seed proteome, in comparison to the previously used cCBB staining technique. To validate the efficiency of visualization by both stains, we first compared the overall number of detected protein spots. On average, 790 spots were visible by cCBB staining and 1200 spots by RuBP staining. Then, the intensity of a set of spots was assessed, and changes in relative abundance were determined using image analysis software. As expected, staining with RuBP performed better in quantitation in terms of sensitivity and dynamic range. Furthermore, spots from a cultivar-specific region in the protein map were chosen for identification to asses the gain of biological information due to the staining procedure. From this particular region, eight spots were visualized exclusively by RuBP and identification was successful for all spots, proving the ability to identify even very low abundant proteins. Performance in MS analysis was comparable for both protein stains. Proteins were identified by MALDI-TOF MS peptide mass fingerprinting. This approach was not successful for all spots, due to the restricted entry number for barley in the database. Therefore, we subsequently used LC-ESI-Q-TOF MS/MS and de novo sequencing for identification. Because only an insufficient number of proteins from barley is annotated, an EST-based identification strategy was chosen for our experiment. We wished to test whether under these limitations the application of a more sensitive stain would lead to a more advanced proteome approach. In summary, we demonstrate here that the application of RuBP as an economical but reliable and sensitive fluorescence stain is highly suitable for quantitative proteome analysis of plant seeds.
Determining the relationship between "microvessel density" and different grades of astrocytoma based on immunohistochemistry for "factor VIII-related antigen" (von Willebrand factor) expression in tumor microvessels.

PubMed

Mahzouni, Parvin; Mohammadizadeh, Fereshteh; Mougouei, Kourosh; Moghaddam, Noushin Afshar; Chehrei, Ali; Mesbah, Alireza

2010-01-01

Astrocytic brain tumors are the most common primary central nervous system tumors, which are classified into four grades. One of the most important pathologic criteria for the diagnosis of higher-grade astrocytomas (especially glioblastoma multiforme) is microvessel proliferation, particularly in the form of glomeruloid complex. Because tumor angiogenesis is a necessary factor for growth and invasiveness of malignancies, microvessel density (MVD) and intensity of angiogenesis may be used to determine the grade of astrocytomas and plan therapy accordingly. We have planned this study to evaluate the relationship between vwf expression in microvessels and different grades of astrocytoma. Sixty-four formalin-fixed and paraffin-embedded blocks of surgical specimens with diagnosis of astrocytoma (grades I to IV, each of them 16 blocks) were selected in a simple-nonrandom sampling. Thin sections of tissue blocks underwent immunohistochemical staining for vwf. The stained slides were examined using a light microscope at low (100) and high (400) magnifications. MVD was estimated by calculating the mean number of stained microvessels in three areas of highest vascularization in the high-power field (400). The intensity of staining was determined based on a 3 scale model, in which scores 0, 1, 2, and 3 mean no detectable stain, trace staining, moderate amount of diffuse stain, and strong diffuse staining, respectively. Thirty-six (56%) patients were male and 28 (44%) were female. Scores 0 and 1 of microvessel staining intensity were not observed in any grades studied, but severe staining intensity (score 3) was observed in 18.8%, 37.5%, 56.3%, and 87.5% of grades I, II, III, and IV astrocytomas, respectively. "Vwf vessel index" (MVD staining intensity of microvessels) was 23.84, 25.62, 31.62, and 62.43 in grades I, II, III, and IV astrocytomas, respectively. We found a significant relationship between staining intensity of vwf in microvessels and different grades of astrocytomas. The intensity of microvessel stain increases in parallel with increasing tumor grade. Regarding "microvessel density" and "vwf vessel index," the difference is predominantly between grade IV and all other grades. However, there is no other statistically meaningful difference between grades I, II and III.
Usefulness of Leifson Staining Method in Diagnosis of Helicobacter pylori Infection

PubMed Central

Piccolomini, Raffaele; Di Bonaventura, Giovanni; Neri, Matteo; Di Girolamo, Arturo; Catamo, Giovanni; Pizzigallo, Eligio

1999-01-01

The Leifson staining method was used to diagnose Helicobacter pylori infection and was compared to histology, culture, and the rapid urease test (RUT). Histology gave the best sensitivity (98%), compared to Leifson staining (97%), culture (92%), and RUT (85%) (P < 0.005). Leifson staining is a sensitive, rapid, economical method for diagnosis of H. pylori infection in dyspeptic patients. PMID:9854090
Micronucleus Assay: An Early Diagnostic Tool to Assess Genotoxic Changes in Patients with Tobacco Use, Oral Leukoplakia and Oral Submucous Fibrosis

PubMed Central

Ahuja, Puneet; Mehendiratta, Monica; Sharma, Mohit; Dutta, Jahnobi

2017-01-01

Introduction Micronuclei (MNi) are acentric chromatid or chromosome fragments produced via genetic damage through genotoxic agents contained in tobacco and betel nut. Evidently, the various Oral Potentially Malignant Disorders (OPMDs) like oral lichen Planus, oral leukoplakia and Oral Submucous Fibrosis (OSMF) demonstrate MNi, as a substantiation of genetic damage. As these changes can be easily appreciated in oral exfoliated cells, an exfoliated cell based MNi assay might be utilized as handy and non invasive biomonitoring tool for gauging the genetic damage and hence the propensity for malignant transformation in OPMDs. To this end, MNi are definitely easier to evaluate when compared to chromosome aberrations. Aim To compare the MNi frequency in normal mucosa, in individuals using various tobacco forms without oral leukoplakia, individuals using various tobacco forms with oral leukoplakia, and areca nut chewers with OSMF, using three different stains. Materials and Methods Oral exfoliated cells from 50 cases of normal mucosa (Group I), 50 cases of tobacco chewing people without Oral Leukoplakia (Group II), 50 cases of people with Oral Leukoplakia (Group III) and 50 cases of areca nut chewers with OSMF (Group IV) were taken. MNi frequencies were compared in these groups using three different stains i.e., Papanicolaou (PAP) stain, May Grunwald Giemsa (MGG) stain and Feulgen stain. The data between cases (Group II, III and IV) and control groups (Group I) was analyzed by Kruskal-Wallis Test. The comparison between two independent groups was done by Mann-Whitney U test and interstain comparison between cases and control was done by Wilcoxon Signed Rank Test and the individual p-value was obtained. Results A significant increase in the count was observed during transition of normal mucosa to OPMDs. The best stain for detecting MNi was PAP stain followed by MGG stain and Feulgen stain. Conclusion The higher mean MNi count for PAP stain and MGG stain could be attributed to nonspecific staining. Further study using a larger sample size on quantitative assessment of MNi count in various OPMDs is warranted. PMID:29207828
Propidium iodide staining: a new application in fluorescence microscopy for analysis of cytoarchitecture in adult and developing rodent brain.

PubMed

Hezel, Marcus; Ebrahimi, Fahim; Koch, Marco; Dehghani, Faramarz

2012-10-01

Immunohistochemical visualization of antigens in specimen has evolved to an indispensable technique in biomedical research for investigations of cell morphology and pathology both in bright field and fluorescence microscopy. While there are couple of staining methods that reveal entire cytoarchitecture in bright field microscopy such as Nissl or hemalaun-eosin, there are still limitations in visualizations of cytoarchitecture in fluorescence microscopy. The present study reports a simple staining method that provides the required illustration of cell allocations and cellular composition in fluorescence microscopy in adult and in developing rodent central nervous system using the fluorophore propidium iodide (PI, 5μg/mL). PI is a well-accepted marker for degenerating cells when applied prior to fixation (pre-fixation PI staining). Here, PI was added to the sections after the fixation (post-fixation PI staining). This revised labeling procedure led to similar cytoarchitectural staining patterns in fluorescence microscopy as observed with hemalaun in bright field microscopy. This finding was proven in organotypic hippocampal slice cultures (OHSC) and brain sections obtained from different postnatal developmental stages. Excitotoxically lesioned OHSC subjected to pre-fixation PI staining merely showed brightly labeled condensed nuclei of degenerating neurons. In contrast, post-fixation PI staining additionally revealed extensive labeling of neuronal cell bodies and glial cells within the OHSC, thus allowing visualization of stratification of neuronal layers and cell morphology. Furthermore, post-fixation PI staining was combined with NeuN, calbindin, calretinin, glial fibrillary acidic protein or Griffonia simplicifolia isolectin B4 (IB(4)) in post natal (p1 and p9) and adult rats. In early post-natal brain sections almost all mentioned cellular markers led to an incomplete staining of the native cell organization and resulted in an inaccurate estimation of cell morphology when compared to adult brains. In contrast, post-fixation PI staining allowed investigation of the whole cytoarchitecture independent of the developmental stage. Taken together, post-fixation PI staining provides a detailed insight in the morphology of both developing and adult brain tissues in fluorescence microscopy. Copyright © 2012 Elsevier Ltd. All rights reserved.
Audit and internal quality control in immunohistochemistry

PubMed Central

Maxwell, P; McCluggage, W

2000-01-01

Aims—Although positive and negative controls are performed and checked in surgical pathology cases undergoing immunohistochemistry, internal quality control procedures for immunohistochemistry are not well described. This study, comprising a retrospective audit, aims to describe a method of internal quality control for immunohistochemistry. A scoring system that allows comparison between cases is described. Methods—Two positive tissue controls for each month over a three year period (1996–1998) of the 10 antibodies used most frequently were evaluated. All test cases undergoing immunohistochemistry in the months of April in this three year period were also studied. When the test case was completely negative for a given antibody, the corresponding positive tissue control from that day was examined. A marking system was devised whereby each immunohistochemical slide was assessed out of a possible score of 8 to take account of staining intensity, uniformity, specificity, background, and counterstaining. Using this scoring system, cases were classified as showing optimal (7–8), borderline (5–6), or unacceptable (0–4) staining. Results—Most positive tissue controls showed either optimal or borderline staining with the exception of neurone specific enolase (NSE), where most slides were unacceptable or borderline as a result of a combination of low intensity, poor specificity, and excessive background staining. All test cases showed either optimal or borderline staining with the exception of a single case stained for NSE, which was unacceptable. Conclusions—This retrospective audit shows that immunohistochemically stained slides can be assessed using this scoring system. With most antibodies, acceptable staining was achieved in most cases. However, there were problems with staining for NSE, which needs to be reviewed. Laboratories should use a system such as this to evaluate which antibodies regularly result in poor staining so that they can be excluded from panels. Routine evaluation of immunohistochemical staining should become part of everyday internal quality control procedures. Key Words: immunohistochemistry • audit • internal quality control PMID:11265178
Porcine Intestinal Mast Cells. Evaluation of Different Fixatives for Histochemical Staining Techniques Considering Tissue Shrinkage

PubMed Central

Rieger, J.; Twardziok, S.; Huenigen, H.; Hirschberg, R.M.; Plendl, J.

2013-01-01

Staining of mast cells (MCs), including porcine ones, is critically dependent upon the fixation and staining technique. In the pig, mucosal and submucosal MCs do not stain or stain only faintly after formalin fixation. Some fixation methods are particularly recommended for MC staining, for example the fixation with Carnoy or lead salts. Zinc salt fixation (ZSF) has been reported to work excellently for the preservation of fixation-sensitive antigens. The aim of this study was to establish a reliable histological method for counting of MCs in the porcine intestinum. For this purpose, different tissue fixation and staining methods that also allow potential subsequent immunohistochemical investigations were evaluated in the porcine mucosa, as well as submucosa of small and large intestine. Tissues were fixed in Carnoy, lead acetate, lead nitrate, Zamboni and ZSF and stained subsequently with either polychromatic methylene blue, alcian blue or toluidine blue. For the first time our study reveals that ZSF, a heavy metal fixative, preserves metachromatic staining of porcine MCs. Zamboni fixation was not suitable for histochemical visualization of MCs in the pig intestine. All other tested fixatives were suitable. Alcian blue and toluidine blue co-stained intestinal goblet cells which made a prima facie identification of MCs difficult. The polychromatic methylene blue proved to be the optimal staining. In order to compare MC counting results of the different fixation methods, tissue shrinkage was taken into account. As even the same fixation caused shrinkagedifferences between tissue from small and large intestine, different factors for each single fixation and intestinal localization had to be calculated. Tissue shrinkage varied between 19% and 57%, the highest tissue shrinkage was found after fixation with ZSF in the large intestine, the lowest one in the small intestine after lead acetate fixation. Our study emphasizes that MC counting results from data using different fixation techniques can only be compared if the respective studyimmanent shrinkage factor has been determined and quantification results are adjusted accordingly. PMID:24085270
NMDAR-1 staining in the lateral geniculate nucleus of normal and visually deprived cats.

PubMed

Ziburkus, J; Bickford, M E; Guido, W

2000-01-01

In normal adult cats, a monoclonal antibody directed toward the NR-1 subunit of the N-methyl-D-aspartate (NMDA) receptor (Pharmingen, clone 54.1) produced dense cellular and neuropil labeling throughout all layers of the lateral geniculate nucleus (LGN) and adjacent thalamic nuclei, including the thalamic reticular, perigeniculate, medial intralaminar, and ventral lateral geniculate nuclei. Cellular staining revealed well-defined somata, and in some cases proximal dendrites. NMDAR-1 cell labeling was also evident in the LGN of early postnatal kittens, suggesting that developing LGN cells possess this receptor subunit at or before eye opening. Within the A-layers of the adult LGN, staining encompassed a wide range of soma sizes. Soma size comparisons of NMDAR-1 stained cells with those stained with an antibody directed toward a nonphosphorylated neurofilament protein (SMI-32), which selectively stains Y-relay cells (Bickford et al., 1998), or an antibody to glutamic acid decarboxylase (GAD), which stains for GABAergic interneurons, suggested that NMDA receptors are utilized by relay cells and interneurons. NMDAR-1 staining was also observed in the LGN of cats with early monocular lid suture. Although labeling was apparent in both deprived and nondeprived A-layers of LGN, the distribution of soma sizes was significantly different. In the deprived A-layers of LGN, staining was limited to small- and medium-sized cells. Cells with relatively large soma were lacking. However, cell density measurements as well as soma size comparisons with cells stained for Nissl substance suggested these differences were due to deprivation-induced cell shrinkage and not to a loss of NMDAR-1 staining in Y-cells. Taken together, these results suggest that NMDA receptors are utilized by both relay cells and interneurons in LGN and that alterations in early visual experience do not necessarily affect the expression of NMDA receptors in the LGN.
An in vitro screening assay for dental stain cleaning.

PubMed

Wang, Changxiang; Lucas, Robert; Smith, Anthony J; Cooper, Paul R

2017-01-09

The present study aimed to develop an in vitro model for stain removal from natural enamel for the assessment and comparison of oral hygiene products. Bovine teeth (n = 8 per group) were ground/polished to provide flat enamel specimens and ferric-tannate deposits were precipitated onto the enamel surfaces. The ferric-tannate stained enamel specimens were brushed using an in vitro tooth-brushing simulator with slurries containing commercially available toothpaste products, dental abrasive particles, and sodium tripolyphosphate (STP) solutions of different concentrations. The colour of the enamel surfaces was measured using a spectrophotometer before and after stain application as well as after the brushing treatments. Differences in stain removal efficacy were found between the toothpastes categorised as whitening and non-whitening comprising of different types of dental abrasives (hydrated silica and alumina). A mean value of 27% for stain removal was detected for the three non-whitening toothpastes and 59% of stain removal was detected for the three whitening toothpastes after 1000 strokes. Compared with the slurry with Zeodent 113 abrasive alone, the addition of STP provided better performance for stain removal under the same brushing conditions (mean value of 62% for Zeodent 113 abrasive alone and 72% with the addition of 5% (w/w) STP after 1000 strokes). No difference was evident between the STP concentration of 5% (w/w) and 10% (w/w). The ferric-tannate/bovine enamel model reported here provides good stain retention, is rapidly and easily prepared, and is shown to be progressively and reproducibly sensitive to toothbrushing using different toothpastes and surfactant/chelating agent solutions. Importantly, it provides good discrimination between various oral hygiene products. The stain removal assay reported here has considerable potential to enable comparative assessments of different toothpaste types in terms of their cleaning capabilities.
Multicenter Assessment of Gram Stain Error Rates.

PubMed

Samuel, Linoj P; Balada-Llasat, Joan-Miquel; Harrington, Amanda; Cavagnolo, Robert

2016-06-01

Gram stains remain the cornerstone of diagnostic testing in the microbiology laboratory for the guidance of empirical treatment prior to availability of culture results. Incorrectly interpreted Gram stains may adversely impact patient care, and yet there are no comprehensive studies that have evaluated the reliability of the technique and there are no established standards for performance. In this study, clinical microbiology laboratories at four major tertiary medical care centers evaluated Gram stain error rates across all nonblood specimen types by using standardized criteria. The study focused on several factors that primarily contribute to errors in the process, including poor specimen quality, smear preparation, and interpretation of the smears. The number of specimens during the evaluation period ranged from 976 to 1,864 specimens per site, and there were a total of 6,115 specimens. Gram stain results were discrepant from culture for 5% of all specimens. Fifty-eight percent of discrepant results were specimens with no organisms reported on Gram stain but significant growth on culture, while 42% of discrepant results had reported organisms on Gram stain that were not recovered in culture. Upon review of available slides, 24% (63/263) of discrepant results were due to reader error, which varied significantly based on site (9% to 45%). The Gram stain error rate also varied between sites, ranging from 0.4% to 2.7%. The data demonstrate a significant variability between laboratories in Gram stain performance and affirm the need for ongoing quality assessment by laboratories. Standardized monitoring of Gram stains is an essential quality control tool for laboratories and is necessary for the establishment of a quality benchmark across laboratories. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
Multicenter Assessment of Gram Stain Error Rates

PubMed Central

Balada-Llasat, Joan-Miquel; Harrington, Amanda; Cavagnolo, Robert

2016-01-01

Gram stains remain the cornerstone of diagnostic testing in the microbiology laboratory for the guidance of empirical treatment prior to availability of culture results. Incorrectly interpreted Gram stains may adversely impact patient care, and yet there are no comprehensive studies that have evaluated the reliability of the technique and there are no established standards for performance. In this study, clinical microbiology laboratories at four major tertiary medical care centers evaluated Gram stain error rates across all nonblood specimen types by using standardized criteria. The study focused on several factors that primarily contribute to errors in the process, including poor specimen quality, smear preparation, and interpretation of the smears. The number of specimens during the evaluation period ranged from 976 to 1,864 specimens per site, and there were a total of 6,115 specimens. Gram stain results were discrepant from culture for 5% of all specimens. Fifty-eight percent of discrepant results were specimens with no organisms reported on Gram stain but significant growth on culture, while 42% of discrepant results had reported organisms on Gram stain that were not recovered in culture. Upon review of available slides, 24% (63/263) of discrepant results were due to reader error, which varied significantly based on site (9% to 45%). The Gram stain error rate also varied between sites, ranging from 0.4% to 2.7%. The data demonstrate a significant variability between laboratories in Gram stain performance and affirm the need for ongoing quality assessment by laboratories. Standardized monitoring of Gram stains is an essential quality control tool for laboratories and is necessary for the establishment of a quality benchmark across laboratories. PMID:26888900
Different immunohistochemical and ultrastructural phenotypes of squamous differentiation in bladder cancer.

PubMed

Gaisa, Nadine T; Braunschweig, Till; Reimer, Nina; Bornemann, Jörg; Eltze, Elke; Siegert, Sabine; Toma, Marieta; Villa, Luigi; Hartmann, Arndt; Knuechel, Ruth

2011-03-01

Besides worse prognosis of bladder cancer with squamous differentiation (pure squamous cell carcinoma (SCC) or mixed urothelial carcinoma (UC/SCC)), high-grade non-keratinising squamous differentiation is difficult to identify in haematoxylin-eosin stainings. This study aims to validate routine immunohistochemical markers for squamous differentiation in a larger cohort of patients. Tissue microarrays of 89 pure SCCs and mixed UC/SCCs, 66 urothelial carcinomas (UC), precursor lesions and normal urothelium were stained for cytokeratin (CK) 5/6, CK 5/14, CK 7, CK 20 and uroplakin III. Electron microscopy was performed to confirm the differentiation. Pure SCCs displayed staining throughout the epithelium for CK 5/6 (76.6% (36/47)) and CK 5/14 (95.8% (46/48)), focal staining for CK 7 (28.9% (13/45)) and no staining for CK 20 and uroplakin III (both 0% (0/48)). UCs exhibited a basal or diffuse staining for CK 5/6 (30.2% (16/53)) and CK 5/14 (57.1% (32/56)), focal positivity for CK 7 (83.6% (46/55)), CK 20 (50.9% (29/57)) and uroplakin III (21.8% (12/55)). Each marker discriminated SCC and UC significantly (p < 0.01). A third subgroup rarely showed full epithelial staining for CK 5/6 (14.3% (1/7)) and CK 5/14 (28.6% (2/7)), focal staining for CK 7 (85.7% (6/7)) and no staining for CK 20 and uroplakin III (both 0% (0/7)). Electron microscopy could prove both, SCC and UC characteristics, revealing a transient type. A staining pattern with CK 5/6- and CK 5/14-positivity plus CK 20- and uroplakin III-negativity identified squamous differentiation in bladder tumours and revealed a third type of squamous transdifferentiation.
Methylene blue intra-arterial staining of resected colorectal cancer specimens improves accuracy of nodal staging: A randomized controlled trial.

PubMed

Reima, H; Saar, H; Innos, K; Soplepmann, J

2016-11-01

Metastatic involvement of regional lymph nodes is a major prognostic factor of colorectal cancer, which influences also its treatment strategy. International consensus foresees retrieval of ≥12 lymph nodes from colorectal specimens. The aim of the study was to assess the effect of intra-arterial staining of colorectal specimens with methylene blue on lymph node harvest. A total of 266 radically operated colorectal cancer patients were randomized into the methylene blue staining and non-staining groups. In the staining group, methylene blue solution was injected into the colorectal specimen's artery after its removal. The specimens were analysed for lymph node count, diameter and metastatic involvement. The median number of lymph nodes was higher in the staining group, 27 (95% CI 23-31%), compared with the control group, 16 (95% CI 14-19, p < 0.001). The number of examined lymph-nodes was ≥12 in 86% of the cases in the staining group and in 69% of the cases in the control group (p = 0.001). In the staining group more small-diameter (≤4 mm) lymph nodes were examined (median number 20.5 vs. 10, p < 0.001). The proportion of patients with metatatic lymph nodes was 42% in the staining group and 43% in the control group (NS). Methylene blue staining improves significantly staging accuracy through finding more small-diameter lymph nodes. It enables to detect ≥12 lymph nodes in the majority of cases. We recommend routine use of this technique in all colorectal resections with curative intent. Copyright © 2016 Elsevier Ltd, BASO ~ the Association for Cancer Surgery, and the European Society of Surgical Oncology. All rights reserved.
Stain susceptibility of composite and ceramic CAD/CAM blocks versus direct resin composites with different resinous matrices.

PubMed

Alharbi, Amal; Ardu, Stefano; Bortolotto, Tissiana; Krejci, Ivo

2017-04-01

To evaluate the stain susceptibility of CAD/CAM blocks and direct composite after long term exposure to various staining agents. 40 disk-shaped samples were fabricated from each of nine materials; six CAD/CAM (Vitablocs Mark II, Paradigm MZ100, Experimental Vita Hybrid Ceramic, Vita Enamic, Experimental Kerr and Lava Ultimate) and three direct composites (Filtek Supreme, Venus Diamond and Filtek Silorane). Samples were randomly divided into five groups (n = 8) according to different staining solutions (distilled water, tea, red wine, coffee and artificial saliva). Initial L*a*b* values were assessed using a calibrated digital spectrophotometer. Specimens were immersed in staining solutions and stored in an incubator at 37 °C for 120 days. L*a*b* values were assessed again and color change (∆E) was calculated as difference between recorded L*a*b* values. ANOVA, and Duncan test were used to identify differences between groups (α = 0.05). Significant differences in ∆E values were detected between materials (p = 0.000). Among all staining solutions, the highest ∆E value was observed with red wine. The new CAD/CAM blocks (Vita Enamic, Vita Hybrid Ceramic and Lava Ultimate) showed the highest resistance to staining compared to the MZ100 composite resin blocks. Filtek Silorane, a direct composite, showed high stain resistance values compared to CAD/CAM materials and other direct composites. Ceramic and composite CAD/CAM blocks had lower staining susceptibility than methacrylate based direct composite. Staining susceptibility of the new resin based CAD/CAM materials Vita Enamic and Lava Ultimate was comparable to feldspathic ceramic blocks (Vitablocs Mark II). Filtek Silorane showed promising results that were comparable to some CAD/CAM blocks.
Histological staining can enhance the performance of spectroscopic microscopy on sensing nanoarchitectural alterations of biological cells (Conference Presentation)

NASA Astrophysics Data System (ADS)

Zhang, Di; Cherkezyan, Lusik; Li, Yue; Capoglu, Ilker; Subramanian, Hariharan; Taflove, Allen; Backman, Vadim

2017-02-01

Our group had previously established that nanoscale three-dimensional refractive index (RI) fluctuations of a linear, dielectric, label-free medium can be sensed in the far field through spectroscopic microscopy, regardless of the diffraction limit of optical microscopy. Adopting this technique, Partial Wave Spectroscopic (PWS) Microscopy was able to sense nanoarchitectural alterations in early-stage cancers. With the success of PWS on detecting cancer from healthy clinical samples, we further investigated whether and how histological staining can enhance the performance of PWS by both finite difference time domain (FDTD) simulations and experiments. In this investigation, the dispersion models of hematoxylin and eosin were extracted from the absorption spectra of H and E stained cells. Using these models, the effect of staining were studied via FDTD simulations of unstained versus stained samples with various nanostructures. We observed that, the spectral variance was increased and the spectral variance difference between two samples with distinct nanostructures was enhanced in stained samples by over 200%. Furthermore, we investigated with FDTD whether molecule-specific staining can be used to enhance signals from a medium composing of the desired molecule. Samples with two mixed random media were created and the desired medium was either stained or unstained. Our results showed that the difference between the nanostructures of only the desired medium was enhanced in stained samples. We concluded that, with molecule-specific staining, PWS can selectively target the nanoarchitecture of a desired molecule. Lastly, these results were validated by experiments using human buccal cells from healthy or lung cancer patients. This study has significant impact in improving the performance of PWS on quantifying nanoarchitectural alterations during cancer.

Immunohistochemical localization of beta-amyloid precursor protein sequences in Alzheimer and normal brain tissue by light and electron microscopy.

PubMed

McGeer, P L; Akiyama, H; Kawamata, T; Yamada, T; Walker, D G; Ishii, T

1992-03-01

Immunohistochemical staining with antibodies directed against four segments of the amyloid precursor protein (APP) was studied by light and electron microscopy in normal and Alzheimer (AD) brain tissue. The segments according to the Kang et al. sequence were: 18-38 (T97); 527-540 (R36); 597-620 (1-24 of beta-amyloid protein [BAP], R17); and 681-695 (R37) (Kang et al. [1987]: Nature 325:733-736). The antibodies recognized full length APP in Western blots of extracts of APP transfected cells. They stained cytoplasmic granules in some pyramidal neurons in normal appearing tissue from control and AD cases. In AD affected tissue, the antibodies to amino terminal sections of APP stained tangled neurons and neuropil threads, and intensely stained dystrophic neurites in senile plaques. By electron microscopy, this staining was localized to abnormal filaments. The antibody to the carboxy terminal segment failed to stain neurofibrillary tangles or neuropil threads; it did stain some neurites with globular swellings. It also stained globular and elongated deposits in senile plaque areas. The antibody against the BAP intensely stained extracellular material in senile plaques and diffuse deposits. By electron microscopy, the antibodies all stained intramicroglial deposits. Some of the extracellular and intracellular BAP-positive deposits were fibrillary. Communication between intramicroglial and extracellular fibrils was detected in plaque areas. These data suggest the following sequence of events. APP is normally concentrated in intraneuronal granules. In AD, it accumulates in damaged neuronal fibers. The amino terminal portion binds to abnormal neurofilaments. Major fragments of APP are phagocytosed and processed by microglia with the BAP portion being preserved. The preserved BAP is then extruded and accumulates in extracellular tissue.
Secondary mutation in a coding mononucleotide tract in MSH6 causes loss of immunoexpression of MSH6 in colorectal carcinomas with MLH1/PMS2 deficiency.

PubMed

Shia, Jinru; Zhang, Liying; Shike, Moshe; Guo, Min; Stadler, Zsofia; Xiong, Xiaoling; Tang, Laura H; Vakiani, Efsevia; Katabi, Nora; Wang, Hangjun; Bacares, Ruben; Ruggeri, Jeanine; Boland, C Richard; Ladanyi, Marc; Klimstra, David S

2013-01-01

Immunohistochemical staining for DNA mismatch repair proteins may be affected by various biological and technical factors. Staining variations that could potentially lead to erroneous interpretations have been recognized. A recently recognized staining variation is the significant reduction of staining for MSH6 in some colorectal carcinomas. The frequency and specific characteristics of this aberrant MSH6 staining pattern, however, have not been well analyzed. In this study of 420 colorectal carcinoma samples obtained from patients fulfilling the Revised Bethesda Guidelines, we detected 9 tumors (2%) showing extremely limited staining for MSH6 with positive staining present in <5% of the tumor cells. Our analyses showed that these tumors belonged to two distinct categories: (1) MLH1 and/or PMS2 protein-deficient carcinomas (n=5, including 1 with a pathogenic mutation in PMS2); and (2) MLH1, PMS2 and MSH2 normal but with chemotherapy or chemoradiation therapy before surgery (n=4). To test our hypothesis that somatic mutation in the coding region microsatellite of the MSH6 gene might be a potential underlying mechanism for such limited MSH6 staining, we evaluated frameshift mutation in a (C)(8) tract in exon 5 of the MSH6 gene in seven tumors that had sufficient DNA for analysis, and detected mutation in four; all four tumors belonged to the MLH1/PMS2-deficient group. In conclusion, our data outline the main scenarios where significant reduction of MSH6 staining is more likely to occur in colorectal carcinoma, and suggest that somatic mutations of the coding region microsatellites of the MSH6 gene is an underlying mechanism for this staining phenomenon in MLH1/PMS2-deficient carcinomas.
Histological staining methods preparatory to laser capture microdissection significantly affect the integrity of the cellular RNA.

PubMed

Wang, Hongyang; Owens, James D; Shih, Joanna H; Li, Ming-Chung; Bonner, Robert F; Mushinski, J Frederic

2006-04-27

Gene expression profiling by microarray analysis of cells enriched by laser capture microdissection (LCM) faces several technical challenges. Frozen sections yield higher quality RNA than paraffin-imbedded sections, but even with frozen sections, the staining methods used for histological identification of cells of interest could still damage the mRNA in the cells. To study the contribution of staining methods to degradation of results from gene expression profiling of LCM samples, we subjected pellets of the mouse plasma cell tumor cell line TEPC 1165 to direct RNA extraction and to parallel frozen sectioning for LCM and subsequent RNA extraction. We used microarray hybridization analysis to compare gene expression profiles of RNA from cell pellets with gene expression profiles of RNA from frozen sections that had been stained with hematoxylin and eosin (H&E), Nissl Stain (NS), and for immunofluorescence (IF) as well as with the plasma cell-revealing methyl green pyronin (MGP) stain. All RNAs were amplified with two rounds of T7-based in vitro transcription and analyzed by two-color expression analysis on 10-K cDNA microarrays. The MGP-stained samples showed the least introduction of mRNA loss, followed by H&E and immunofluorescence. Nissl staining was significantly more detrimental to gene expression profiles, presumably owing to an aqueous step in which RNA may have been damaged by endogenous or exogenous RNAases. RNA damage can occur during the staining steps preparatory to laser capture microdissection, with the consequence of loss of representation of certain genes in microarray hybridization analysis. Inclusion of RNAase inhibitor in aqueous staining solutions appears to be important in protecting RNA from loss of gene transcripts.
Histological staining methods preparatory to laser capture microdissection significantly affect the integrity of the cellular RNA

PubMed Central

Wang, Hongyang; Owens, James D; Shih, Joanna H; Li, Ming-Chung; Bonner, Robert F; Mushinski, J Frederic

2006-01-01

Background Gene expression profiling by microarray analysis of cells enriched by laser capture microdissection (LCM) faces several technical challenges. Frozen sections yield higher quality RNA than paraffin-imbedded sections, but even with frozen sections, the staining methods used for histological identification of cells of interest could still damage the mRNA in the cells. To study the contribution of staining methods to degradation of results from gene expression profiling of LCM samples, we subjected pellets of the mouse plasma cell tumor cell line TEPC 1165 to direct RNA extraction and to parallel frozen sectioning for LCM and subsequent RNA extraction. We used microarray hybridization analysis to compare gene expression profiles of RNA from cell pellets with gene expression profiles of RNA from frozen sections that had been stained with hematoxylin and eosin (H&E), Nissl Stain (NS), and for immunofluorescence (IF) as well as with the plasma cell-revealing methyl green pyronin (MGP) stain. All RNAs were amplified with two rounds of T7-based in vitro transcription and analyzed by two-color expression analysis on 10-K cDNA microarrays. Results The MGP-stained samples showed the least introduction of mRNA loss, followed by H&E and immunofluorescence. Nissl staining was significantly more detrimental to gene expression profiles, presumably owing to an aqueous step in which RNA may have been damaged by endogenous or exogenous RNAases. Conclusion RNA damage can occur during the staining steps preparatory to laser capture microdissection, with the consequence of loss of representation of certain genes in microarray hybridization analysis. Inclusion of RNAase inhibitor in aqueous staining solutions appears to be important in protecting RNA from loss of gene transcripts. PMID:16643667
Organic staining on bone from exposure to wood and other plant materials.

PubMed

Pollock, Corey R; Pokines, James T; Bethard, Jonathan D

2018-02-01

Determining the depositional environment and the postmortem alterations to a set of remains are necessary aspects of a forensic investigation to explain the circumstances surrounding the death of an individual. The present study examines organic staining as a method for reconstructing the depositional environment of skeletal remains and the taphonomic agents with which they came into contact. Organic staining results largely from tannins leaching from plant materials and therefore can be seen on bone deposited in wooden coffin environments or on terrestrial surfaces. The present study examines the hypothesis that the degree of staining observed on skeletal elements would increase as the length of exposure to the organic matter increased and that different plant materials and environments would leave different patterns or colorations of staining. The sample consisted of 165 pig (Sus scrofa) femora divided into four groups exposed to differing experimental conditions, including burial in direct contact with soil or burial in a simulated coffin environment, immersion in water with wood samples, and surface deposition with plant matter contact. The bones were removed once a month from their experimental environments and the level of staining was recorded qualitatively using the Munsell Soil Color Chart. In all of the experimental environments, staining was present after two months of exposure, and the color darkened across the bone surface with each episode of data collection. The results from the present study indicate that staining can manifest on bone within a relatively short time frame once skeletonization occurs and a variety of colorations or patterns of staining can manifest based on the plant material. The present research also demonstrates the potential of organic staining to aid in estimations of the postmortem interval as well as a depositional environmental reconstruction through plant species identification. Copyright © 2017 Elsevier B.V. All rights reserved.
Microwave-Assisted Tissue Preparation for Rapid Fixation, Decalcification, Antigen Retrieval, Cryosectioning, and Immunostaining

PubMed Central

2016-01-01

Microwave irradiation of tissue during fixation and subsequent histochemical staining procedures significantly reduces the time required for incubation in fixation and staining solutions. Minimizing the incubation time in fixative reduces disruption of tissue morphology, and reducing the incubation time in staining solution or antibody solution decreases nonspecific labeling. Reduction of incubation time in staining solution also decreases the level of background noise. Microwave-assisted tissue preparation is applicable for tissue fixation, decalcification of bone tissues, treatment of adipose tissues, antigen retrieval, and other special staining of tissues. Microwave-assisted tissue fixation and staining are useful tools for histological analyses. This review describes the protocols using microwave irradiation for several essential procedures in histochemical studies, and these techniques are applicable to other protocols for tissue fixation and immunostaining in the field of cell biology. PMID:27840640
Identification of a silver binding protein associated with the cytological silver staining of actively transcribing nucleolar regions.

PubMed

Hubbell, H R; Rothblum, L I; Hsu, T C

1979-10-01

Nucleoli isolated from Novikoff hepatoma cells were stained with AgNO3 to demonstrate the typical staining of active ribosomal cistrons. Pre-treatment of the nucleoli with 80 mM Tris-HCl (pH 7.5) -- 2.0 M NaCl did not interfere with silver staining. Treatment of the nucleoli with 80 mM Tris-HCl (pH 7.5) -- 0.15 M NaCl did, however, eliminate silver binding. Serial extraction of nucleoli with 2.0 M NaCl buffer followed by 0.15 M NaCl buffer also abolished silver staining. Analysis of the supernatant fraction of these extracts by polyacrylamide gel electrophoresis indicates that, although more than one nucleolar protein can bind silver, only one protein is associated with the staining of active ribosomal cistrons.
Photometric Application of the Gram Stain Method To Characterize Natural Bacterial Populations in Aquatic Environments

PubMed Central

Saida, H.; Ytow, N.; Seki, H.

1998-01-01

The Gram stain method was applied to the photometric characterization of aquatic bacterial populations with a charge-coupled device camera and an image analyzer. Escherichia coli and Bacillus subtilis were used as standards of typical gram-negative and gram-positive bacteria, respectively. A mounting agent to obtain clear images of Gram-stained bacteria on Nuclepore membrane filters was developed. The bacterial stainability by the Gram stain was indicated by the Gram stain index (GSI), which was applicable not only to the dichotomous classification of bacteria but also to the characterization of cell wall structure. The GSI spectra of natural bacterial populations in water with various levels of eutrophication showed a distinct profile, suggesting possible staining specificity that indicates the presence of a particular bacterial population in the aquatic environment. PMID:9464416
Visualizing hydrophobic domains in silicone hydrogel lenses with Sudan IV.

PubMed

Jacob, Jean T; Levet, Jacques; Edwards, Tamika A; Dassanayake, Nissanke; Ketelson, Howard

2012-06-08

A lipophilic dye is used to investigate the degree to which the surface and bulk hydrophobic domains of the lenses can be imaged and to identify specific changes in the availability of those domains after in vitro wear and cleaning conditions. The effect of a multipurpose solution (MPS), OPTI-FREE RepleniSH, on lens hydrophobic domains was also investigated. Hydrophobic domains were determined using a saturated solution of Sudan IV. Staining periods of 30 minutes and 16 hours were used to determine surface versus bulk hydrophobic domains. Four types of silicone hydrogel lens materials were tested. The degree of staining was visually documented by photography and quantitatively determined by extraction and analysis of the total amount of dye adsorbed. Specific differences in staining were found for all control lenses. Exposure to in vitro wear conditions significantly decreased the staining response for all lens types as compared with unworn lenses (P = 0.001). However, the trend of staining remained the same: balafilcon A > galyfilcon A > senofilcon A > lotrafilcon B. MPS decreased the extent of staining; the degree of its effect varied with lens type. Hydrophobic staining with Sudan IV visualized domains on and within silicone hydrogel lenses. Differences in staining response after exposure to wear and cleaning conditions indicate the potential for protein and lipid deposition on the different lens types and the ability of MPS to affect that deposition. Hydrophobic staining may be useful for determining differences in surface modification and lipophilicity of silicone hydrogel lenses.
Gram stain method shows better sensitivity than clinical criteria for detection of bacterial vaginosis in surveillance of pregnant, low-income women in a clinical setting.

PubMed Central

Tam, M T; Yungbluth, M; Myles, T

1998-01-01

OBJECTIVE: The purpose of the study is to determine whether the Gram stain method is superior to the clinical criteria for the diagnosis of bacterial vaginosis in low-income pregnant women seen in a resident clinic setting. The clinical criteria is the current diagnostic method employed to diagnose bacterial vaginosis. STUDY DESIGN: In this study, 51 pregnant women with vaginal discharge were prospectively evaluated. All were screened using the clinical criteria, Gram stain method, and culture of the discharge. The modified scoring system instituted by Nugent et al. (J Clin Microbiol 29:297-301, 1991) was employed in reading the Gram stain smears. The clinical criteria were then compared with the Gram stain method. Isolation of moderate to many Gardnerella vaginalis growth by culture was used as the confirmatory finding. RESULTS: Sensitivity of the Gram stain method (91%) was significantly higher than that of the clinical criteria (46%), (sign test P = 0.0023, < 0.01). The Gram stain method also has both a low false-negative (4%) and high negative predictive value (96%), making it an ideal diagnostic test. CONCLUSION: The Gram stain method is a rapid and cost-effective test that is also highly reproducible and readily available in many laboratories. These features make the Gram stain method a more desirable screening procedure for bacterial vaginosis in a clinic population. PMID:9894174
Gram-stain-based antimicrobial selection reduces cost and overuse compared with Japanese guidelines.

PubMed

Taniguchi, Tomohiro; Tsuha, Sanefumi; Shiiki, Soichi; Narita, Masashi

2015-10-26

The Gram stain has been used as an essential tool for antimicrobial stewardship in our hospital since the 1970s. The objective of this study was to clarify the difference in the targeted therapies selected based on the Gram stain and simulated empirical therapies based on the antimicrobial guidelines used in Japan. A referral-hospital-based prospective descriptive study was undertaken between May 2013 and April 2014 in Okinawa, Japan. All enrolled patients were adults who had been admitted to the Division of Infectious Diseases through the emergency room with suspected bacterial infection at one of three sites: respiratory system, urinary tract, or skin and soft tissues. The study outcomes were the types and effectiveness of the antibiotics initially selected, and their total costs. Two hundred eight patients were enrolled in the study. The median age was 80 years. A significantly narrower spectrum of antibiotics was selected based on the Gram stain than was selected based on the Japanese guidelines. The treatments based on the Gram stain and on the guidelines were estimated to be equally highly effective. The total cost of antimicrobials after Gram-stain testing was less than half the cost after the guidelines were followed. Compared with the Japanese guidelines, the Gram stain dramatically reduced the overuse of broad-spectrum antimicrobials without affecting the effectiveness of the treatment. Drug costs were reduced by half when the Gram stain was used. The Gram stain should be included in all antimicrobial stewardship programs.
Utility of Gram stain for the microbiological analysis of burn wound surfaces.

PubMed

Elsayed, Sameer; Gregson, Daniel B; Lloyd, Tracie; Crichton, Marilyn; Church, Deirdre L

2003-11-01

Surface swab cultures have attracted attention as a potential alternative to biopsy histology or quantitative culture methods for microbiological burn wound monitoring. To our knowledge, the utility of adding a Gram-stained slide in this context has not been evaluated previously. To determine the degree of correlation of Gram stain with culture for the microbiological analysis of burn wound surfaces. Prospective laboratory analysis. Urban health region/centralized diagnostic microbiology laboratory. Burn patients hospitalized in any Calgary Health Region burn center from November 2000 to September 2001. Gram stain plus culture of burn wound surface swab specimens obtained during routine dressing changes or based on clinical signs of infection. Degree of correlation (complete, high, partial, none), including weighted kappa statistic (kappa(w)), of Gram stain with culture based on quantitative microscopy and degree of culture growth. A total of 375 specimens from 50 burn patients were evaluated. Of these, 239 were negative by culture and Gram stain, 7 were positive by Gram stain only, 89 were positive by culture only, and 40 were positive by both methods. The degree of complete, high, partial, and no correlation of Gram stain with culture was 70.9% (266/375), 1.1% (4/375), 2.4% (9/375), and 25.6% (96/375), respectively. The degree of correlation for all 375 specimens, as expressed by the weighted kappa statistic, was found to be fair (kappa(w) = 0.32).Conclusion.-The Gram stain is not suitable for the microbiological analysis of burn wound surfaces.
Alternative to xylene as a clearing agent in histopathology

PubMed Central

Alwahaibi, Nasar; Aljaradi, Shaima; Alazri, Horiyah

2018-01-01

INTRODUCTION: Clearing is an essential step in processing tissue for light microscopy. Xylene is the clearing agent used most commonly worldwide. Xylene is toxic and therefore a threat to personnel working in histopathology laboratories. We evaluated a safer alternative clearing agent for use in the histopathology laboratory. MATERIALS AND METHODS: We used 230 formalin-fixed, paraffin-embedded tissue blocks from 19 different tissues. Half of the specimens were processed using xylene and half were processed using UltraClear™. Tissues were evaluated for eight parameters: sectioning, nuclear staining, cytoplasmic staining, overall cell morphology, clarity of staining, uniformity of staining, quality of immunohistochemistry (IHC), and cost. RESULTS: Both UltraClear™ and xylene processed sections scored 100% for IHC. Sections processed using UltraClear™ were easy to cut (81.7%) as were xylene processed sections (96.5%). UltraClear™ processed sections showed 67%, 60.9%, 52.2%, 63.5%, and 67% for nuclear staining, cytoplasmic staining, cell morphology, clarity of staining, and uniformity of staining, respectively. UltraClear™ is twice as expensive as xylene. We found that tissues processed using UltraClear™ were easy to cut and worked well for both hematoxylin and eosin and IHC staining. CONCLUSION: UltraClear™ is less toxic, less flammable, friendlier to the environment, and easy to handle, but it is two times expensive than xylene. The findings of this study recommend the use of UltraClear™ solution as a routine clearing agent in histopathology laboratories. However, further studies are required. PMID:29692586
Combined histochemical staining, RNA amplification, regional, and single cell cDNA analysis within the hippocampus.

PubMed

Ginsberg, Stephen D; Che, Shaoli

2004-08-01

The use of five histochemical stains (cresyl violet, thionin, hematoxylin & eosin, silver stain, and acridine orange) was evaluated in combination with an expression profiling paradigm that included regional and single cell analyses within the hippocampus of post-mortem human brains and adult mice. Adjacent serial sections of human and mouse hippocampus were labeled by histochemistry or neurofilament immunocytochemistry. These tissue sections were used as starting material for regional and single cell microdissection followed by a newly developed RNA amplification procedure (terminal continuation (TC) RNA amplification) and subsequent hybridization to custom-designed cDNA arrays. Results indicated equivalent levels of global hybridization signal intensity and relative expression levels for individual genes for hippocampi stained by cresyl violet, thionin, and hematoxylin & eosin, and neurofilament immunocytochemistry. Moreover, no significant differences existed between the Nissl stains and neurofilament immunocytochemistry for individual CA1 neurons obtained via laser capture microdissection. In contrast, a marked decrement was observed in adjacent hippocampal sections stained for silver stain and acridine orange, both at the level of the regional dissection and at the CA1 neuron population level. Observations made on the cDNA array platform were validated by real-time qPCR using primers directed against beta-actin and glyceraldehyde-3 phosphate dehydrogenase. Thus, this report demonstrated the utility of using specific Nissl stains, but not stains that bind RNA species directly, in both human and mouse brain tissues at the regional and cellular level for state-of-the-art molecular fingerprinting studies.
Orientation specific deposition of mesoporous particles

NASA Astrophysics Data System (ADS)

Kjellman, Tomas; Bodén, Niklas; Wennerström, Hâkan; Edler, Karen J.; Alfredsson, Viveka

2014-11-01

We present a protocol for a facile orientation specific deposition of plate-like mesoporous SBA-15 silica particles onto a surface (mesopores oriented normal to surface). A drop of an aqueous dispersion of particles is placed on the surface and water vaporizes under controlled relative humidity. Three requirements are essential for uniform coverage: particle dispersion should not contain aggregates, a weak attraction between particles and surface is needed, and evaporation rate should be low. Aggregates are removed by stirring/sonication. Weak attraction is realized by introducing cationic groups to the surface. Insight into the mechanisms of the so-called coffee stain effect is also provided.
Comparison of modified Chicago sky blue stain and potassium hydroxide mount for the diagnosis of dermatomycoses and onychomycoses.

PubMed

Liu, Zhong; Sheng, Ping; Yang, Yan-Ping; Li, Wen; Huang, Wen-Ming; Wang, Jie-Di; Fan, Yi-Ming

2015-05-01

The diagnostic value of modified Chicago sky blue (CSB) stain and potassium hydroxide (KOH) mount for superficial mycoses was compared using fungal culture as gold standard. The sensitivity and screening time of the CSB stain were superior to the KOH mount. The CBS stain is simple, quick and reliable for diagnosing superficial mycoses. Copyright © 2015. Published by Elsevier B.V.
Rapid alkaline methylene blue supravital staining for assessment of anterior segment infections.

PubMed

Kiuchi, Katsuji

2016-01-01

To present the Löffler's alkaline methylene blue technique of staining eye discharges in eyes with anterior segment infections. The Löffler's alkaline methylene blue staining method is a simple staining technique that can be used to differentiate bacterial, viral, and fungal infections. It is a cationic dye that stains cells blue because the positively charged dye is attracted to negatively charged particles such as polyphosphates, DNAs, and RNAs. Specimens collected from patients by swabbing are smeared onto microscope slides and the methylene blue solution is dropped on the slide. The slide is covered with a glass cover slip and examined under a microscope. The entire time from the collection to the viewing is about 30 seconds. Histopathological images of the conjunctival epithelial cells and neutrophils in eye discharges were dyed blue and the nuclei were stained more intensely blue. Bacterial infections consisted mainly of neutrophils, and viral infections consisted mainly of lymphocytes. Löffler's alkaline methylene blue staining can be done in about 30 seconds for diagnosis. Even though this is a one color stain, it is possible to infer the cause of the infection by detection of the absence of bacteria and/or fungi in context of the differential distribution of neutrophils and lymphocytes.
The application of visible wavelength reflectance hyperspectral imaging for the detection and identification of blood stains.

PubMed

Li, Bo; Beveridge, Peter; O'Hare, William T; Islam, Meez

2014-12-01

Current methods of detection and identification of blood stains rely largely on visual examination followed by presumptive tests such as Kastle-Meyer, Leuco-malachite green or luminol. Although these tests are useful, they can produce false positives and can also have a negative impact on subsequent DNA tests. A novel application of visible wavelength reflectance hyperspectral imaging has been used for the detection and positive identification of blood stains in a non contact and non destructive manner on a range of coloured substrates. The identification of blood staining was based on the unique visible absorption spectrum of haemoglobin between 400 and 500 nm. Images illustrating successful discrimination of blood stains from nine red substances are included. It has also been possible to distinguish between blood and approximately 40 other reddish stains. The technique was also successfully used to detect latent blood stains deposited on white filter paper at dilutions of up to 1 in 512 folds and on red tissue at dilutions of up to 1 in 32 folds. Finally, in a blind trial, the method successfully detected and identified a total of 9 blood stains on a red T-shirt. Copyright © 2014 Forensic Science Society. Published by Elsevier Ireland Ltd. All rights reserved.
Inadvertent trypan blue posterior capsule staining during cataract surgery.

PubMed

Burkholder, Bryn M; Srikumaran, Divya; Nanji, Afshan; Lee, Bryan; Weinberg, Robert S

2013-04-01

To report 5 cases of inadvertent posterior capsule staining with trypan blue during phacoemulsification. Retrospective, observational case series. Five cases of posterior capsule staining with trypan blue were identified from cataract surgeries performed at an academic institution. All 5 eyes underwent phacoemulsification with use of iris retractors. The surgical videos from each case were reviewed to better understand the mechanisms and risk factors for posterior capsule staining with trypan blue and techniques to avoid this complication. No eyes had clinical evidence of zonular pathology during their preoperative examination. Only 1 patient reported a possible childhood history of trauma to both eyes. One eye had uveitis, requiring posterior synechialysis. All 5 cases involved the use of iris retractors. No other intraoperative complications occurred, and the intraocular lens was successfully placed in the capsular bag in all cases. All eyes had resolution of posterior capsule staining by postoperative day 8. Inadvertent posterior capsule staining with trypan blue can occur in eyes that appear structurally normal. The use of iris retractors may facilitate posterior capsule staining by allowing the posterior flow of trypan blue under the iris and through the zonules to the posterior capsule. Surgeons should consider techniques to minimize the risk of posterior capsule staining, particularly in cases involving the use of iris retractors. Copyright © 2013 Elsevier Inc. All rights reserved.
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.

Whole blood staining in suspension for nonspecific esterase and alkaline phosphatase analyzed with a Technicon H-1.

PubMed

Ross, D W; Bishop, C; Henderson, A; Kaplow, L

1990-01-01

We adapted previously published methods for nonspecific esterase and alkaline phosphatase staining of white blood cells in suspension for use on a Technicon H-1 hematology analyzer. The objective was to develop a semiautomated method using whole blood that could be employed on a large scale for hematology laboratory applications, including toxicology studies, measurement of neutrophil left shift, and cytochemical classification of myeloid leukemias. The nonspecific esterase method uses the pararosaniline stain, generating the unstable substrate from two stable precursors. Whole blood is added to the substrate plus dye mix. Next, acid lysis and fixation steps destroy red cells and stabilize the monocyte staining. The alkaline phosphatase stain employs a stable naphthyl phosphate substrate and fast blue B coupling dye. The red cells are lysed with a pH 10.3 propanediol buffer, and the white blood cells are then stabilized with formalin fixation. For both methods the staining is performed off-line, and the sample is then diluted with propanediol to match the refractive index of the sheath on the H-1 analyzer, before aspiration into the direct cytometry port. A cytogram of scattered versus absorbed light is obtained. The number of cells staining and the intensity of the stain can be quantified from the cytogram.
Inhibin may be involved in negative feedback in the prairie dog (Cynomys ludovicianus).

PubMed

Foreman, Darhl

2007-02-01

The changes in inhibin immunostaining in the gonads during the annual reproductive cycle of both sexes of the prairie dog are described. No inhibin immunostaining was found in primary or secondary follicles of the ovary. Theca and granulosa cells of preovulatory Graafian follicles found in January and February stained for inhibin. Corpora lutea of both pregnant and non-pregnant females stain more densely for inhibin than follicles. Inhibin staining is present in luteal cells for at least 4 months during regression, longer than detectable progesterone is secreted. Sertoli cells in the testes do not have inhibin immunostaining during recrudescence. These cells show light immunostain for inhibin during peak spermatogenic activity in January and February but stain more deeply during early regression of the testis. Stain is gradually lost in the next 4-5 months as the tubules close. Leydig cells and germ cells do not stain for inhibin at any stage of the annual cycle but interstitial cells and tunic cells stain during the breeding phase. The presence of immunochemical staining for inhibin in prairie dog gonads during regression suggests that inhibin is part of a negative feedback complex that includes progesterone in the female and testosterone or another androgen in the male. Negative feedback during regression may also cause gonadal inactivity.
Calbindin-D28k is a more reliable marker of human Purkinje cells than standard Nissl stains: a stereological experiment.

PubMed

Whitney, Elizabeth R; Kemper, Thomas L; Rosene, Douglas L; Bauman, Margaret L; Blatt, Gene J

2008-02-15

In a study of human Purkinje cell (PC) number, a striking mismatch between the number of PCs observed with the Nissl stain and the number of PCs immunopositive for calbindin-D28k (CB) was identified in 2 of the 10 brains examined. In the remaining eight brains this mismatch was not observed. Further, in these eight brains, analysis of CB immunostained sections counterstained with the Nissl stain revealed that more than 99% Nissl stained PCs were also immunopositive for CB. In contrast, in the two discordant brains, only 10-20% of CB immunopositive PCs were also identified with the Nissl stain. Although this finding was unexpected, a historical survey of the literature revealed that Spielmeyer [Spielmeyer W. Histopathologie des nervensystems. Julius Springer: Berlin; 1922. p. 56-79] described human cases with PCs that lacked the expected Nissl staining intensity, an important historical finding and critical issue when studying postmortem human brains. The reason for this failure in Nissl staining is not entirely clear, but it may result from premortem circumstances since it is not accounted for by postmortem delay or processing variables. Regardless of the exact cause, these observations suggest that Nissl staining may not be a reliable marker for PCs and that CB is an excellent alternative marker.
[Expression of EV71-VP1, PSGL-1 and SCARB2 in Tissues of Infants with Brain Stem Encephalitis].

PubMed

Li, Ming; Kong, Xiao-ping; Liu, Hong; Cheng, Ling-xi; Huang, Jing-lu; Quan, Li; Wu, Fang-yu; Hao, Bo; Liu, Chao; Luo, Bin

2015-04-01

To understand the correlation of enterovirus 71 (EV71), P-selectin glycoprotein ligand-1 (PSGL-1), and scavenger receptor B2 (SCARB2) and to explore the possible pathway and mechanism of EV71 infection by observing the expression of EV71, PSGL-1 and SCARB2 in tissues of infants with brain stem encephalitis. The organs and tissues of infants with EV71-VP1 positivity in their brain stems were chosen. Expression and distribution of EV71-VP1, PSGL-1, and SCARB2 were detected and compared by immunohistochemistry. Strong staining of EV71 -VP1 was observed in the neuron, glial cells, the inflammatory cells of perivascular cuffing, parietal cells of the gastric fundus gland while alveolar macrophages, intestinal gland epithelium cells, mucosa lymphoid nodule and lymphocyte of palatine tonsil showed moderate staining and weak staining were displayed in mesenteric lymph nodes and lymphocyte of spleen. PSGL-1 expression was detected in parietal cells of the gastric fundus gland, tonsillar crypt squamous epithelium, alveolar macrophages and leukocytes in each tissue. SCARB2 expression was observed in all the above tissues except the intestines and spleen. The distribution of EV71 correlates with SCARB2 expression. SCARB2 plays an important role in virus infection and replication. Stomach may be an important site for EV71 replication.
c-KIT positive schistosomal urinary bladder carcinoma are frequent but lack KIT gene mutations.

PubMed

Shams, Tahany M; Metawea, Mokhtar; Salim, Elsayed I

2013-01-01

Urinary bladder squamous cell carcinoma (SCC), one of the most common neoplasms in Egypt, is attributed to chronic urinary infection with Schistosoma haematobium (Schistosomiasis). The proto-oncogene c-KIT, encoding a tyrosine kinase receptor and implicated in the development of a number of human malignancies, has not been studied so far in schistosomal urinary bladder SCCs. We therefore determined immunohistochemical (IHC) expression of c-KIT in paraffin sections from 120 radical cystectomies of SCCs originally obtained from the Pathology Department of Suez Canal University (Ismailia, Egypt). Each slide was evaluated for staining intensity where the staining extent of >10% of cells was considered positive. c-KIT overexpression was detected in 78.3% (94/120) of the patients, the staining extents in the tumor cells were 11-50% and >50% in 40 (42.6%) and 54 (57.4%) respectively. The positive cases had 14.9%, 63.8%, 21.3% as weak, moderate and strong intensity respectively. Patients with positive bilharzial ova had significantly higher c-KIT expression than patients without (95.2% vs. 38.9%, P=0.000). Mutation analysis of exons 9-13 was negative in thirty KIT positive cases. The high rate of positivity in SBSCC was one of the striking findings; However, CD117 may be a potential target for site specific immunotherapy to improve the outcome of this tumor.
The evaluation and validation of Phadebas® paper as a presumptive screening tool for saliva on forensic exhibits.

PubMed

Wornes, Danielle J; Speers, Samuel J; Murakami, Julie A

2018-07-01

The Phadebas ® Forensic Press Test is routinely used for the detection of saliva. However, assessment of the use of Phadebas ® paper for this purpose has not been studied extensively. The suitability of Phadebas ® paper as a presumptive screening tool for saliva on forensic exhibits, was investigated by analysing the following: (1) sensitivity, (2) specificity, (3) effects of temperature on sensitivity and specificity, (4) detection of saliva in mixed body fluid samples, and (5) influence of substrate porosity. The results of this study demonstrated that Phadebas ® paper is more sensitive to α-amylase activity and less specific for saliva than previously reported. The use of an examination temperature of 37°C had no effect on sensitivity, but increased the incidence of cross-reactivity with other forensically relevant body fluid stains. Blood, urine and vaginal secretions can inhibit the detection of α-amylase activity with Phadebas ® paper in mixed stains of saliva and body fluid. Substrate porosity is a weak predictor for the time taken for a saliva stain to achieve a strong positive result on Phadebas ® paper. Overall, this study demonstrated that the Phadebas ® Forensic Press Test has limitations as a presumptive test for the accurate identification of saliva. Copyright © 2018 Elsevier B.V. All rights reserved.
PSCA expression is associated with favorable tumor features and reduced PSA recurrence in operated prostate cancer.

PubMed

Heinrich, Marie-Christine; Göbel, Cosima; Kluth, Martina; Bernreuther, Christian; Sauer, Charlotte; Schroeder, Cornelia; Möller-Koop, Christina; Hube-Magg, Claudia; Lebok, Patrick; Burandt, Eike; Sauter, Guido; Simon, Ronald; Huland, Hartwig; Graefen, Markus; Heinzer, Hans; Schlomm, Thorsten; Heumann, Asmus

2018-05-31

Prostate Stem Cell Antigen (PSCA) is frequently expressed in prostate cancer but its exact function is unclear. To clarify contradictory findings on the prognostic role of PSCA expression, a tissue microarray containing 13,665 prostate cancers was analyzed by immunohistochemistry. PSCA staining was absent in normal epithelial and stromal cells of the prostate. Membranous and cytoplasmic PSCA staining was seen in 53.7% of 9642 interpretable tumors. Staining was weak in 22.4%, moderate in 24.5% and strong in 6.8% of tumors. PSCA expression was associated with favorable pathological and clinical tumor features: Early pathological tumor stage (p < 0.0001), low Gleason grade (p < 0.0001), absence of lymph node metastasis (p < 0.0001), low pre-operative PSA level (p = 0.0118), negative surgical margin (p < 0.0001) and reduced PSA recurrence (p < 0.0001). PSCA expression was an independent predictor of prognosis in multivariate analysis (hazard ratio 0.84, p < 0.0001). The absence of statistical relationship to TMPRSS2:ERG fusion status, chromosomal deletion or high tumor cell proliferation argues against a major role of PSCA for regulation of cell cycle or genomic integrity. PSCA expression is linked to favorable prognosis. PSCA measurement is a candidate for inclusion in multi-parametric prognostic prostate cancer tests.
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.
Utility of Gram staining for diagnosis of Malassezia folliculitis.

PubMed

Tu, Wei-Ting; Chin, Szu-Ying; Chou, Chia-Lun; Hsu, Che-Yuan; Chen, Yu-Tsung; Liu, Donald; Lee, Woan-Ruoh; Shih, Yi-Hsien

2018-02-01

Malassezia folliculitis (MalF) mimics acne vulgaris and bacterial folliculitis in clinical presentations. The role of Gram staining in rapid diagnosis of MalF has not been well studied. In our study, 32 patients were included to investigate the utility of Gram staining for MalF diagnosis. The final diagnoses of MalF were determined according to clinical presentation, pathological result and treatment response to antifungal agents. Our results show that the sensitivity and specificity of Gram staining are 84.6% and 100%, respectively. In conclusion, Gram staining is a rapid, non-invasive, sensitive and specific method for MalF diagnosis. © 2017 Japanese Dermatological Association.
Ghost mycobacteria on Gram stain.

PubMed Central

Trifiro, S; Bourgault, A M; Lebel, F; René, P

1990-01-01

The Gram stain is a key tool in diagnostic microbiology. Its usefulness with respect to mycobacteria is undefined. The neutrality of mycobacteria other than Mycobacterium tuberculosis on Gram staining of various clinical specimens is described. Images PMID:1688872
Concentric scheme of monkey auditory cortex

NASA Astrophysics Data System (ADS)

Kosaki, Hiroko; Saunders, Richard C.; Mishkin, Mortimer

2003-04-01

The cytoarchitecture of the rhesus monkey's auditory cortex was examined using immunocytochemical staining with parvalbumin, calbindin-D28K, and SMI32, as well as staining for cytochrome oxidase (CO). The results suggest that Kaas and Hackett's scheme of the auditory cortices can be extended to include five concentric rings surrounding an inner core. The inner core, containing areas A1 and R, is the most densely stained with parvalbumin and CO and can be separated on the basis of laminar patterns of SMI32 staining into lateral and medial subdivisions. From the inner core to the fifth (outermost) ring, parvalbumin staining gradually decreases and calbindin staining gradually increases. The first ring corresponds to Kaas and Hackett's auditory belt, and the second, to their parabelt. SMI32 staining revealed a clear border between these two. Rings 2 through 5 extend laterally into the dorsal bank of the superior temporal sulcus. The results also suggest that the rostral tip of the outermost ring adjoins the rostroventral part of the insula (area Pro) and the temporal pole, while the caudal tip adjoins the ventral part of area 7a.
Issues in using whole slide imaging for diagnostic pathology: "routine" stains, immunohistochemistry and predictive markers.

PubMed

Taylor, C R

2014-08-01

The traditional microscope, together with the "routine" hematoxylin and eosin (H & E) stain, remains the "gold standard" for diagnosis of cancer and other diseases; remarkably, it and the majority of associated biological stains are more than 150 years old. Immunohistochemistry has added to the repertoire of "stains" available. Because of the need for specific identification and even measurement of "biomarkers," immunohistochemistry has increased the demand for consistency of performance and interpretation of staining results. Rapid advances in the capabilities of digital imaging hardware and software now offer a realistic route to improved reproducibility, accuracy and quantification by utilizing whole slide digital images for diagnosis, education and research. There also are potential efficiencies in work flow and the promise of powerful new analytical methods; however, there also are challenges with respect to validation of the quality and fidelity of digital images, including the standard H & E stain, so that diagnostic performance by pathologists is not compromised when they rely on whole slide images instead of traditional stained tissues on glass slides.
Present and future technologies of tooth whitening.

PubMed

Viscio, D; Gaffar, A; Fakhry-Smith, S; Xu, T

2000-01-01

Dental stains can be broadly classified as intrinsic or extrinsic. Intrinsic stains are a result of defects in tooth development, fluorosis, or acquired through the use of tetracycline. Extrinsic stains are localized mainly in the pellicle and are generated by the reaction between sugars and amino acids or acquired from the retention of exogenous chromophores in the pellicle. Three clinical methods are currently used for measuring stain removal and tooth whitening in the development of new whitening technologies: Lobene Stain Index, Shade Guide Color Change, and Minolta ChromaMeter. Professional tooth whitening products rely on proven technologies--35% hydrogen peroxide for in-office power bleaching or 10% to 15% carbamide peroxide for at-home bleaching--to reduce intrinsic stain and change the inherent tooth color. Over-the-counter tooth whitening products use a combination of surfactants, abrasives, anticalculus agents, and low levels of hydrogen peroxide to reduce extrinsic stain and help maintain tooth whiteness after professional treatment. Future technologies for whitening teeth could involve the use of activating agents to enhance the performance of hydrogen peroxide and natural enzymes.
Changes in the morphology and presumptive chemistry of impact and pooled bloodstain patterns by Lucilia sericata (Meigen) (Diptera: Calliphoridae).

PubMed

Fujikawa, Amanda; Barksdale, Larry; Higley, Leon G; Carter, David O

2011-09-01

Bloodstain pattern analysis can be critical to accurate crime scene reconstruction. However, bloodstain patterns can be altered in the presence of insects and can confound crime scene reconstruction. To address this problem, we conducted a series of controlled laboratory experiments to investigate the effect of Lucilia sericata (Meigen) on impact bloodstains and pooled bloodstains in association with three combinations of common surfaces (linoleum/painted drywall, wood floor/wallpaper, and carpet/wood paneling). L. sericata fed from the pooled bloodstains and added insect stains through regurgitation and defecation of consumed blood. L. sericata formed defecatory trails of insect stains that indicated directionality. Defecatory stains fluoresced when viewed at 465 nm with an orange filter. These observations differed from Calliphora vicina insect stains because feeding on blood spatter was not observed and trails of insect stains were formed by L. sericata. The fluorescence of defecatory stains can be used as a method to detect insect stains and discriminate them from real bloodstains. © 2011 American Academy of Forensic Sciences.
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 CFR 51.3210 - Serious damage.

Code of Federal Regulations, 2010 CFR

2010-01-01

...) Staining, dirt or foreign material when more than 25 percent of the onions in any lot are badly stained. Onions with adhering dirt or other foreign matter shall be judged on the same basis as stained onions; (e...
Overcoming Autophagy to Induce Apoptosis in Castration-Resistant Prostate Cancer

DTIC Science & Technology

2016-12-01

staining with antibodies against Ki67 or AR. The same analysis was done for the metformin group and got comparable results. Figure 8. Two sets of...were parafilm embedded and sectioned for H&E and immunohistochemical staining with antibodies against Ki67 or AR (Figure 9). H&E staining showed...combination group measured only 16% of the control. The immunohistochemical staining of representative samples clearly showed inhibition of pSrc by sara
An Analysis of Microbial Contamination in Military Aviation Fuel Systems

DTIC Science & Technology

2003-03-01

does not contact moisture. Corrosion occurs during the natural tendency of the elemental metals (except noble metals) to revert to a combined form...losing the primary stain). The Gram stain procedure used here consisted of staining a fixed smear with the primary dye crystal violet. An iodine...solution was applied as a mordant . The primary stain was next decolorized with acetone/alcohol and the smear was counterstained with safranin. The
Staining paraffin embedded sections of scald of barley before paraffin removal.

PubMed

Xi, K; Burnett, P A

1997-07-01

Staining of paraffin embedded sections with periodic acid-Schiff reagent and fast green before paraffin removal resulted in differentiation of barley seed and leaf tissue from fungal structures of Rhynchosporium secalis. Crystal violet, toluidine blue O and antiline blue also successfully stained fungal structures of R. secalis in barley leaf tissues. Staining of embedded sections before paraffin removal allows simple processing of a series of sections, saves time and reduces solvent consumption.
Detection of Proteins on Blot Membranes

PubMed Central

Goldman, Aaron; Harper, Sandra; Speicher, David W.

2017-01-01

Staining of blot membranes enables the visualization of bound proteins. Proteins are usually transferred to blot membranes by electroblotting, by direct spotting of protein solutions, or by contact blots. Staining allows the efficiency of transfer to the membrane to be monitored. This unit describes protocols for staining proteins after electroblotting from polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. The same methods can be used if proteins are directly spotted, either manually or using robotics. Protocols are included for seven general protein stains (amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, India ink, and MemCode) and three fluorescent protein stains (fluorescamine, IAEDANS, and SYPRO Ruby). Also included is an in-depth discussion of the different blot membrane types and the compatibility of different protein stains with downstream applications, such as immunoblotting or N-terminal Edman sequencing. PMID:27801518

Detection of Proteins on Blot Membranes.

PubMed

Goldman, Aaron; Harper, Sandra; Speicher, David W

2016-11-01

Staining of blot membranes enables the visualization of bound proteins. Proteins are usually transferred to blot membranes by electroblotting, by direct spotting of protein solutions, or by contact blots. Staining allows the efficiency of transfer to the membrane to be monitored. This unit describes protocols for staining proteins after electroblotting from polyacrylamide gels to blot membranes such as polyvinylidene difluoride (PVDF), nitrocellulose, or nylon membranes. The same methods can be used if proteins are directly spotted, either manually or using robotics. Protocols are included for seven general protein stains (amido black, Coomassie blue, Ponceau S, colloidal gold, colloidal silver, India ink, and MemCode) and three fluorescent protein stains (fluorescamine, IAEDANS, and SYPRO Ruby). Also included is an in-depth discussion of the different blot membrane types and the compatibility of different protein stains with downstream applications, such as immunoblotting or N-terminal Edman sequencing. © 2016 by John Wiley & Sons, Inc. Copyright © 2016 John Wiley & Sons, Inc.
Comparison of routine urinalysis and urine Gram stain for detection of bacteriuria in dogs.

PubMed

Way, Leilani Ireland; Sullivan, Lauren A; Johnson, Valerie; Morley, Paul S

2013-01-01

To determine the utility of performing urine Gram stain for detection of bacteriuria compared to routine urine sediment examination and bacterial aerobic urine culture. Prospective, observational study. University teaching hospital. Urine samples acquired via cystocentesis through convenience sampling from 103 dogs presenting to a tertiary referral institution. All samples underwent routine urinalysis, including sediment examination, as well as urine Gram stain and quantitative bacterial aerobic urine culture. The urine Gram stain demonstrated improved sensitivity (96% versus 76%), specificity (100% versus 77%), positive predictive value (100% versus 83%), and negative predictive value (93% versus 69%) when identifying bacteriuria, compared to routine urine sediment examination. The urine Gram stain is highly sensitive and specific when detecting the presence of bacteria in canine urine samples. Gram staining should be considered when bacteriuria is highly suspected and requires rapid identification while bacterial culture is pending. © Veterinary Emergency and Critical Care Society 2013.
Multimodal confocal mosaicing microscopy: an emphasis on squamous cell carcinoma

NASA Astrophysics Data System (ADS)

Chen, Nathaniel W.; Sensibaugh, Jordan; Ardeshiri, Ardaland; Blanchard, Adam; Jacques, Steven; Gareau, Daniel

2010-02-01

Our previous study reported a sensitivity of 96.6% and a specificity of 89.2% in rapidly detecting Basal Cell Carcinomas (BCCs) when nuclei were stained with acridine orange. Squamous Cell Carcinomas (SCCs) and infiltrative BCCs remain difficult to detect. More complete screening can be achieved utilizing both acridine orange for nuclei staining and eosin for cytoplasmic contrast, using two lasers to excite the two stains independently. Nuclear fluorescence is achieved by staining with acridine orange (0.5mM, 60 s), and cytoplasmic fluorescence is achieved by staining with eosin working solution (30 s). This work shows good morphological contrast of SCC and infiltrative BCC with eosin, acridine orange, and reflectance, and presents a means for rapid SCC and infiltrative BCC detection in fresh skin excisions using multimodal confocal microscopy. In addition, digital staining is shown to effectively simulate hematoxylin and eosin (H&E) histology with confocal mosaics.
Proposals for best-quality immunohistochemical staining of paraffin-embedded brain tissue slides in forensics.

PubMed

Trautz, Florian; Dreßler, Jan; Stassart, Ruth; Müller, Wolf; Ondruschka, Benjamin

2018-01-03

Immunohistochemistry (IHC) has become an integral part in forensic histopathology over the last decades. However, the underlying methods for IHC vary greatly depending on the institution, creating a lack of comparability. The aim of this study was to assess the optimal approach for different technical aspects of IHC, in order to improve and standardize this procedure. Therefore, qualitative results from manual and automatic IHC staining of brain samples were compared, as well as potential differences in suitability of common IHC glass slides. Further, possibilities of image digitalization and connected issues were investigated. In our study, automatic staining showed more consistent staining results, compared to manual staining procedures. Digitalization and digital post-processing facilitated direct analysis and analysis for reproducibility considerably. No differences were found for different commercially available microscopic glass slides regarding suitability of IHC brain researches, but a certain rate of tissue loss should be expected during the staining process.
Identifying motor and sensory myelinated axons in rabbit peripheral nerves by histochemical staining for carbonic anhydrase and cholinesterase activities

NASA Technical Reports Server (NTRS)

Riley, Danny A.; Sanger, James R.; Matloub, Hani S.; Yousif, N. John; Bain, James L. W.

1988-01-01

Carbonic anhydrase (CA) and cholinesterase (CE) histochemical staining of rabbit spinal nerve roots and dorsal root ganglia demonstrated that among the reactive myeliated axons, with minor exceptions, sensory axons were CA positive and CE negative whereas motor axons were CA negative and CE positive. The high specificity was achieved by adjusting reaction conditions to stain subpopulations of myelinated axons selectively while leaving 50 percent or so unstained. Fixation with glutaraldehyde appeared necessary for achieving selectivity. Following sciatic nerve transection, the reciprocal staining pattern persisted in damaged axons and their regenerating processes which formed neuromas within the proximal nerve stump. Within the neuromas, CA-stained sensory processes were elaborated earlier and in greater numbers than CE-stained regenerating motor processes. The present results indicate that histochemical axon typing can be exploited to reveal heterogeneous responses of motor and sensory axons to injury.
Effect of Light-Activated Tooth Whitening on Color Change Relative to Color of Artificially Stained Teeth.

PubMed

Kwon, So Ran; Kurti, Steven R; Oyoyo, Udochukwu; Li, Yiming

2015-01-01

There is still controversy as to the efficacy of light activation used in tooth whitening. The purpose of this study was to evaluate the effect of light activation on tooth color change relative to the artificial dye color. Extracted human third molars (160) were randomly distributed into eight groups of 20 specimens each based on artificial staining and use of light activation. All groups received three 45-minute sessions of in-office whitening at 3-day intervals. Color measurements were performed with an intraoral spectrophotometer at baseline prior to staining (T0), after artificial staining (T1), 1-day--(T2), and 1-week--(T3) post-whitening. Color differences were calculated relative to after artificial staining color parameters (L*1, a*1, b*1) with the use of a software analysis program enabling synchronization of two images. Within the same staining groups, the light-activated samples exhibited a greater color change than their nonlight-activated counterparts. However, only in the case of the yellow-stained samples at 1-day post-whitening was there a significant difference between the nonlight-activated and light-activated groups (Tukey's post hoc multiple comparison test for pairwise comparisons, p < 0.05). Light activation is a valid method for enhancing the efficacy of tooth whitening with respect to overall color change and works best with yellow stains. Light activation is a valid method for enhancing the efficacy of tooth whitening with respect to overall color change and works best with yellow stains.
Saccharomyces cerevisiae samples stained with FUN-1 dye can be stored at -20 degrees C for later observation.

PubMed

Eggleston, M D; Marshall, P A

2007-01-01

FUN-1, a fluorescent vital dye, has been observed to form cylindrical intravacuolar structures within the vacuoles of metabolically active yeast cells. FUN-1 staining, which begins as a diffuse pool of fluorescent cytoplasmic stain, uses an unknown endogenous biochemical processing mechanism to compact and form orange-red cylindrical intravacuolar structures within the cell vacuole. In the clinical setting, FUN-1 is primarily used for identification of fungal infection. FUN-1 is utilized in the laboratory to distinguish between metabolically active and dead fungal cells. Although this stain is useful for distinguishing between live and dead fungal dead cells, few studies have utilized this chemical. This lack of use in the scientific community may be due to the requirement that cells are visualized directly after staining. Thus, it would be of interest to be able to stain cells and store them for later use. Our lab examined the longevity of cylindrical intravacuolar structures in two strains of Saccharomyces cerevisiae stained with FUN-1 and stored at -20 degrees C. We found that cylindrical intravacuolar structures could be reliably observed and imaged utilizing differential interference contrast microscopy and fluorescence microscopy for 21 days. We also observed that cells stained with FUN-1 would resume propagation on yeast extract, peptone, dextrose (YPD) plates after being frozen at -20 degrees C for 21 days. These modifications to the published procedure for FUN-1 dye staining should allow for a more prevalent and less time sensitive use of this important biological tool.
The Immunohistochemistry and Toluidine Blue Roles for Helicobacter pylori Detection in Patients with Gastritis

PubMed Central

Tajalli, Raziye; Nobakht, Maliheh; Mohammadi-Barzelighi, Hajar; Agah, Shahram; Rastegar-Lari, Abdolaziz; Sadeghipour, Alireza

2013-01-01

Background: Helicobacter pylori, which is associated with many upper gastrointestinal diseases, is found in half of the population of the world. Several special stains and immunohistochemistry stain for H. pylori are available. The need for and usefulness of immunohistochemical (IHC) technique has been debated for years. Toluidine blue is a simple stain for microbiological studies and is easily available in laboratories. Therefore, this study was conducted to compare hematoxylin and eosin (H&E), Giemsa and toluidine blue staining with immunehistochemistry for detection of H. pylori in patients with gastritis and also to correlate the results of these staining methods with pathological grading. Methods: We reviewed 54 consecutive gastric biopsy specimens stained by H&E and Giemsa as well as by toluidine blue and immunohistochemistry stains for H. pylori. Results: H. pylori was positively identified by IHC in 43 (79.63%) patients, while positive samples were found in 18 (33.33%), 24 (44.44%) and 33 (61.11%) patients using H&E, Giemsa and toluidine blue staining methods. Our results showed that classical histological staining methods are not sensitive enough to identify low numbers or coccoid forms of organism, while toluidine blue and immunohistochemistry play an important role in detection of H. pylori infection. Conclusion: Toluidine blue has been proved to be much more reliable than H&E and Giemsa in detection of H. pylori. In addition, in post treatment biopsies and in biopsies with unexplained chronic active gastritis without histological evidence of H. pylori should have immunohistochemistry done to detect possible low density or coccoid form of organisms. PMID:23279833
A Digital Staining Algorithm for Optical Coherence Tomography Images of the Optic Nerve Head

PubMed Central

Mari, Jean-Martial; Aung, Tin; Cheng, Ching-Yu; Strouthidis, Nicholas G.; Girard, Michaël J. A.

2017-01-01

Purpose To digitally stain spectral-domain optical coherence tomography (OCT) images of the optic nerve head (ONH), and highlight either connective or neural tissues. Methods OCT volumes of the ONH were acquired from one eye of 10 healthy subjects. We processed all volumes with adaptive compensation to remove shadows and enhance deep tissue visibility. For each ONH, we identified the four most dissimilar pixel-intensity histograms, each of which was assumed to represent a tissue group. These four histograms formed a vector basis on which we ‘projected' each OCT volume in order to generate four digitally stained volumes P1 to P4. Digital staining was also verified using a digital phantom, and compared with k-means clustering for three and four clusters. Results Digital staining was able to isolate three regions of interest from the proposed phantom. For the ONH, the digitally stained images P1 highlighted mostly connective tissues, as demonstrated through an excellent contrast increase across the anterior lamina cribrosa boundary (3.6 ± 0.6 times). P2 highlighted the nerve fiber layer and the prelamina, P3 the remaining layers of the retina, and P4 the image background. Further, digital staining was able to separate ONH tissue layers that were not well separated by k-means clustering. Conclusion We have described an algorithm that can digitally stain connective and neural tissues in OCT images of the ONH. Translational Relevance Because connective and neural tissues are considerably altered in glaucoma, digital staining of the ONH tissues may be of interest in the clinical management of this pathology. PMID:28174676
Hue-saturation-density (HSD) model for stain recognition in digital images from transmitted light microscopy.

PubMed

van Der Laak, J A; Pahlplatz, M M; Hanselaar, A G; de Wilde, P C

2000-04-01

Transmitted light microscopy is used in pathology to examine stained tissues. Digital image analysis is gaining importance as a means to quantify alterations in tissues. A prerequisite for accurate and reproducible quantification is the possibility to recognise stains in a standardised manner, independently of variations in the staining density. The usefulness of three colour models was studied using data from computer simulations and experimental data from an immuno-doublestained tissue section. Direct use of the three intensities obtained by a colour camera results in the red-green-blue (RGB) model. By decoupling the intensity from the RGB data, the hue-saturation-intensity (HSI) model is obtained. However, the major part of the variation in perceived intensities in transmitted light microscopy is caused by variations in staining density. Therefore, the hue-saturation-density (HSD) transform was defined as the RGB to HSI transform, applied to optical density values rather than intensities for the individual RGB channels. In the RGB model, the mixture of chromatic and intensity information hampers standardisation of stain recognition. In the HSI model, mixtures of stains that could be distinguished from other stains in the RGB model could not be separated. The HSD model enabled all possible distinctions in a two-dimensional, standardised data space. In the RGB model, standardised recognition is only possible by using complex and time-consuming algorithms. The HSI model is not suitable for stain recognition in transmitted light microscopy. The newly derived HSD model was found superior to the existing models for this purpose. Copyright 2000 Wiley-Liss, Inc.
A memory-efficient staining algorithm in 3D seismic modelling and imaging

NASA Astrophysics Data System (ADS)

Jia, Xiaofeng; Yang, Lu

2017-08-01

The staining algorithm has been proven to generate high signal-to-noise ratio (S/N) images in poorly illuminated areas in two-dimensional cases. In the staining algorithm, the stained wavefield relevant to the target area and the regular source wavefield forward propagate synchronously. Cross-correlating these two wavefields with the backward propagated receiver wavefield separately, we obtain two images: the local image of the target area and the conventional reverse time migration (RTM) image. This imaging process costs massive computer memory for wavefield storage, especially in large scale three-dimensional cases. To make the staining algorithm applicable to three-dimensional RTM, we develop a method to implement the staining algorithm in three-dimensional acoustic modelling in a standard staggered grid finite difference (FD) scheme. The implementation is adaptive to the order of spatial accuracy of the FD operator. The method can be applied to elastic, electromagnetic, and other wave equations. Taking the memory requirement into account, we adopt a random boundary condition (RBC) to backward extrapolate the receiver wavefield and reconstruct it by reverse propagation using the final wavefield snapshot only. Meanwhile, we forward simulate the stained wavefield and source wavefield simultaneously using the nearly perfectly matched layer (NPML) boundary condition. Experiments on a complex geologic model indicate that the RBC-NPML collaborative strategy not only minimizes the memory consumption but also guarantees high quality imaging results. We apply the staining algorithm to three-dimensional RTM via the proposed strategy. Numerical results show that our staining algorithm can produce high S/N images in the target areas with other structures effectively muted.
Identification of metastatic papillary thyroid carcinoma in FNA specimens using thyroid peroxidase immunohistochemistry.

PubMed

Shield, P W; Crouch, S J; Papadimos, D J; Walsh, M D

2018-06-01

We evaluated immunohistochemical staining for thyroid peroxidase (TPO), a glycoprotein found in the apical plasma membrane of thyroid follicular cells, as a marker for metastatic PTC in FNA samples and compared results with thyroglobulin (Tg) and thyroid transcription factor 1 (TTF1) staining. Cell block sections prepared from 100 FNA specimens were stained with a rabbit monoclonal antibody to TPO (EP159). The FNAs included 64 metastatic malignancies from non-thyroid primary sites, including 18 lung, and 36 cases of thyroid tumours (29 PTC, six cases of medullary thyroid carcinoma and one thyroid anaplastic carcinoma). Thyroid tumours were stained with TTF1 and Tg in addition to TPO. All cases of metastatic lung carcinoma also had TTF-1 staining results. TPO staining was negative in all non-thyroid malignancies. Ninety percent (26/29) of PTC were positive. All positive cases showed strong cytoplasmic staining, although 54% (14/26) showed positivity in less than half of the cells. By comparison, Tg staining of TPC cases was present in 62% and TTF-1 in 100%. In addition to showing higher sensitivity, interpretation of staining results with TPO was generally easier with than Tg. All metastatic lung adenocarcinomas were positive for TTF-1 and TPO negative. The six medullary cancers showed positivity in 17%, 0% and 83% with TPO, Tg and TTF-1, respectively. TPO (mAb EP159) may be a useful addition to immunohistochemical panels for FNA specimens where metastatic PTC is a consideration, particularly in cases where metastatic lung carcinoma features in the differential diagnosis. © 2018 John Wiley & Sons Ltd.
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.
42 CFR 493.913 - Mycobacteriology.

Code of Federal Regulations, 2012 CFR

2012-10-01

... purposes: (1) Those that interpret acid-fast stains and refer specimen to another laboratory appropriately certified in the subspecialty of mycobacteriology; (2) Those that interpret acid-fast stains, perform... of mycobacteriology for identification; (3) Those that interpret acid-fast stains, isolate and...
42 CFR 493.913 - Mycobacteriology.

Code of Federal Regulations, 2014 CFR

2014-10-01

... purposes: (1) Those that interpret acid-fast stains and refer specimen to another laboratory appropriately certified in the subspecialty of mycobacteriology; (2) Those that interpret acid-fast stains, perform... of mycobacteriology for identification; (3) Those that interpret acid-fast stains, isolate and...
42 CFR 493.913 - Mycobacteriology.

Code of Federal Regulations, 2013 CFR

2013-10-01

... purposes: (1) Those that interpret acid-fast stains and refer specimen to another laboratory appropriately certified in the subspecialty of mycobacteriology; (2) Those that interpret acid-fast stains, perform... of mycobacteriology for identification; (3) Those that interpret acid-fast stains, isolate and...
42 CFR 493.913 - Mycobacteriology.

Code of Federal Regulations, 2010 CFR

2010-10-01

... purposes: (1) Those that interpret acid-fast stains and refer specimen to another laboratory appropriately certified in the subspecialty of mycobacteriology; (2) Those that interpret acid-fast stains, perform... of mycobacteriology for identification; (3) Those that interpret acid-fast stains, isolate and...
42 CFR 493.913 - Mycobacteriology.

Code of Federal Regulations, 2011 CFR

2011-10-01

... purposes: (1) Those that interpret acid-fast stains and refer specimen to another laboratory appropriately certified in the subspecialty of mycobacteriology; (2) Those that interpret acid-fast stains, perform... of mycobacteriology for identification; (3) Those that interpret acid-fast stains, isolate and...
Improved Nissl method to stain formaldehyde or glutaraldehyde-fixed material.

PubMed

Böck, P

1979-05-15

Nissl staining of paraffin sections from formaldehyde- or glutaraldehyde-fixed specimens is significantly intensified when sections are kept in a 50% (w/v) aqueous solution of potassium metabisulfite before being stained by a conventional Nissl method.
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.

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
The value of intraoperative Gram stain in revision spine surgery.

PubMed

Shifflett, Grant D; Nwachukwu, Benedict U; Bjerke-Kroll, Benjamin T; Kueper, Janina; Koltsov, Jayme B; Sama, Andrew A; Girardi, Federico P; Cammisa, Frank P; Hughes, Alexander P

2015-10-01

Intraoperative cultures and Gram stains are often obtained in cases of revision spine surgery even when clinical signs of infection are not present. The clinical utility and cost-effectiveness of this behavior remain unproven. The aim was to evaluate the clinical utility and cost-effectiveness of routine intraoperative Gram stains in revision spine surgery. This was a retrospective clinical review performed at an academic center in an urban setting. One hundred twenty-nine consecutive adult revision spine surgeries were performed. The outcome measures included intraoperative Gram stains. We retrospectively reviewed the records of 594 consecutive revision spine surgeries performed by four senior surgeons between 2008 and 2013 to identify patients who had operative cultures and Gram stains performed. All revision cases including cervical, thoracic, and lumbar fusion and non-fusion, with and without instrumentation were reviewed. One hundred twenty-nine (21.7%) patients had operative cultures obtained and were included in the study. The most common primary diagnosis code at the time of revision surgery was pseudarthrosis, which was present in 41.9% of cases (54 of 129). Infection was the primary diagnosis in 10.1% (13 of 129) of cases. Operative cultures were obtained in 129 of 595 (21.7%) cases, and 47.3% (61 of 129) were positive. Gram stains were performed in 98 of 129 (76.0%) cases and were positive in 5 of 98 (5.1%) cases. Overall, there was no correlation between revision diagnosis and whether or not a Gram stain was obtained (p=.697). Patients with a history of prior instrumentation were more likely to have a positive Gram stain (p<.0444). Intraoperative Gram staining was found to have a sensitivity of 10.9% (confidence interval [CI] 3.9%-23.6%) and specificity of 100% (CI 93.1%-100%). The positive and negative predictive values were 100% (CI 48.0%-100%) and 57.3% (CI 45.2%-66.2%), respectively. Kappa coefficient was calculated to be 0.1172 (CI 0.0194-0.2151). The cost per discrepant diagnosis (total cost/number discrepant) was $172.10. This study demonstrates that while very specific for infection, the sensitivity of intraoperative Gram staining is low, and agreement between positive cultures and Gram stains is very poor. Gram staining demonstrated limited cost-effectiveness because of the low prevalence of findings that altered patient management. Copyright © 2015 Elsevier Inc. All rights reserved.
One Small Step for the Gram Stain, One Giant Leap for Clinical Microbiology

PubMed Central

2016-01-01

The Gram stain is one of the most commonly performed tests in the clinical microbiology laboratory, yet it is poorly controlled and lacks standardization. It was once the best rapid test in microbiology, but it is no longer trusted by many clinicians. The publication by Samuel et al. (J. Clin. Microbiol. 54:1442–1447, 2016, http://dx.doi.org/10.1128/JCM.03066-15) is a start for those who want to evaluate and improve Gram stain performance. In an age of emerging rapid molecular results, is the Gram stain still relevant? How should clinical microbiologists respond to the call to reduce Gram stain error rates? PMID:27008876
Microbiology of liver abscesses and the predictive value of abscess gram stain and associated blood cultures.

PubMed

Chemaly, Roy F; Hall, Gerri S; Keys, Thomas F; Procop, Gary W

2003-08-01

Although rare, pyogenic liver abscesses are potentially fatal. We evaluated the predictive value of Gram stain of liver abscess aspirates and temporally associated blood cultures. Gram stains detected bacteria in 79% of the liver abscesses tested. The sensitivity and specificity of Gram stain of the liver abscesses were 90% and 100% for Gram-positive cocci (GPC) and 52% and 94% for Gram-negative bacilli (GNB). The sensitivities of the blood cultures for any GPC and GNB present in the liver abscess were 30% and 39%, respectively. Although, Gram stains and blood cultures offer incomplete detection of the microbial contents of pyogenic liver abscesses, both tests should always accompany liver abscess cultures.
Gram staining apparatus for space station applications.

PubMed Central

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. Images PMID:1690529
Reduction of Oxidative Stain in Ochoó (Hura crepitans L.) = Reducción de la Mancha de Oxidación en Ochoó (Hura crepitans L.)

Treesearch

Michael C. Wiemann; Mark Knaebe

2007-01-01

The sapwood of ochoÃ³ (Hura crepitans) suffers from an oxidative stain that results in substantial loss in value, making it one of the most important utilization problems encountered among Bolivian timber species. Initiating kiln drying within 6 days of felling did not reduce staining of ochoÃ³, although many boards with extensive interior stain had bright surfaces, so...
Staining of Tissue Sections for Electron Microscopy with Heavy Metals

PubMed Central

Watson, Michael L.

1958-01-01

Descriptions of three heavy metal stains and methods of application to tissue sections for electron microscopy are presented. Lead hydroxide stains rather selectively two types of particles in liver: those associated with the endoplasmic reticulum and containing ribonucleic acid and other somewhat larger particles. Barium hydroxide emphasizes certain bodies within vesicles of the Golgi region of hepatic cells. Alkalized lead acetate is useful as a general stain, as are also lead and barium hydroxides. PMID:13610936
Impedance Measurements Could Accelerate Phage-Based Identification of Bacillus anthracis and Other Bacteria

DTIC Science & Technology

2016-09-01

The Bacillus-inoculated NSM agar plates were incubated at 35°C for at least 48 h until Gram stains revealed the presence of > 90% Bacillus spores in...longer visible in Gram stained samples. Finally, centrifugation was used to remove soluble debris from the preparation and spore concentrations were...minutes post treatment. Gram Stains . Gram stains were used to track the emergence of vegetative Bacillus cells from spores. In this assay, bacterial
Acridine orange staining reaction as an index of physiological activity in Escherichia coli

NASA Technical Reports Server (NTRS)

McFeters, G. A.; Singh, A.; Byun, S.; Callis, P. R.; Williams, S.

1991-01-01

The assumption that the acridine orange (AO) color reaction may be used as an index of physiological activity was investigated in laboratory grown Escherichia coli. Spectrofluorometric observations of purified nucleic acids, ribosomes and the microscopic color of bacteriophage-infected cells stained with AO confirmed the theory that single-stranded nucleic acids emit orange to red fluorescence while those that are double-stranded fluoresce green in vivo. Bacteria growing actively in a rich medium could be distinguished from cells in stationary phase by the AO reaction. Cells from log phase appeared red, whereas those in stationary phase were green. However, this differentiation was not seen when the bacteria were grown in a minimal medium or when a variation of the staining method was used. Also, shifting bacteria in stationary phase to starvation conditions rapidly changed their AO staining reaction. Boiling and exposure to lethal concentrations of azide and formalin resulted in stationary-phase cells that appeared red after staining but bacteria killed with chlorine remained green. These findings indicate that the AO staining reaction may be suggestive of physiological activity under defined conditions. However, variables in staining and fixation procedures as well as uncertainties associated with mixed bacterial populations in environmental samples may produce results that are not consistent with the classical interpretation of this reaction. The importance of validating the putative physiological implications of this staining reaction is stressed.
Immunocytochemistry of cell surface heparan sulfate proteoglycan in mouse tissues. A light and electron microscopic study.

PubMed

Hayashi, K; Hayashi, M; Jalkanen, M; Firestone, J H; Trelstad, R L; Bernfield, M

1987-10-01

The core protein of the proteoglycan at the cell surface of NMuMG mouse mammary epithelial cells bears both heparan and chondroitin sulfate chains and is recognized by the monoclonal antibody 281-2. Using this antibody and the peroxidase-antiperoxidase staining technique in adult mouse tissues, we found that the antibody recognizes the antigen in a highly restricted distribution, staining a variety of epithelial cells but no cells derived from embryonic mesoderm or neural crest. The antibody fails to stain any stromal (mesenchymal) or neuronal cells, with the exception of plasma cells and Leydig cells. Squamous and transitional epithelia stain intensely over their entire surfaces, whereas cuboidal and columnar epithelia stain moderately and only at the lateral surface of the basal cells. Within squamous and transitional epithelial tissues that undergo physiological regeneration (e.g., epidermis), the most superficial and differentiated cell types fail to stain. Within glandular and branched epithelia (e.g., pancreas), the secretory alveolar cells fail to stain. When evaluated by electron microscopy, granular deposits of stain are seen on the plasma membrane, especially on lateral surfaces, but none are noted within the cells or the basement membrane. These results indicate that in adult tissues the core protein of this heparan sulfate-rich proteoglycan is expressed almost exclusively at epithelial cell surfaces. Expression appears to be lost as the cells become either mature or highly differentiated.
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.
A colourimetric evaluation of the effect of bacterial contamination on teeth stained with blood in vitro: Evaluation of the efficacy of two different bleaching regimes.

PubMed

Wang, S; Cathro, P; Heithersay, G; Briggs, N; Ratnayake, J; Zilm, P

2018-02-27

Tooth discolouration could occur due to bacterial contamination in traumatized teeth. Hydrogen peroxide is the commonly used bleaching agent. However, due to concerns over safety, alternative bleaching regimes such as sodium perborate (S) and thiourea-hydrogen peroxide (T) have been investigated. Apices resected and pulp extirpated 99 premolars were divided into two groups. Group 1 and 2 was injected with blood and blood/bacteria, stored anaerobically for 35 days. The two groups were treated by bleaching with water, S or T. Teeth were rebleached after 7 days. Colourimetric evaluation was assessed using digital imaging, CasMatch standardization and CIE L*a*b colour system preoperatively, 35 days of staining and 7 and 14 of bleaching. A linear mixed model with fixed effects of time, group and bleach was used to examine colour difference. Blood-stained teeth were significantly redder and darker on day 35 compared with blood/bacteria-stained teeth. After bleaching, blood-stained teeth retained significant redness compared with blood/bacteria-stained teeth using either S or T. T produced a significantly whiter shade in both the groups after 14 days. Blood-stained teeth were significantly darker and red compared with blood/bacteria-stained teeth. T bleaching regime was more effective than S. © 2018 Australian Dental Association.
Analysis of the utility of stone gram stain in urolithiasis treated with percutaneous nephrolithotomy.

PubMed

Cockerill, Patrick A; Rivera, Marcelino E; Krambeck, Amy E

2014-06-01

To define the sensitivity and specificity of stone gram stain for infected urolithiasis treated with percutaneous nephrolithotomy (PCNL). PCNL procedures performed at our institution were analyzed between January 2009 and May 2013. Stone fragments were sent in a sterile fashion for aerobic and fungal cultures. A gram stain and fungal smear were performed on the stones and reported within 24 hours of collection. A total of 228 patients underwent 248 PCNLs. Of the 248 stones, 81 (33%) had a positive stone culture. Stone gram stain was positive in 31 cases and negative in 50. There were 167 negative stone cultures, and in these cases, gram stain was positive in 5 and negative in 162. The calculated sensitivity and specificity of stone gram stain were 38% and 97%. The positive and negative predictive values were 86% and 76%, respectively. In the subset of 16 patients with positive stone fungal cultures, fungal smear was performed in 12 and was positive in 4, giving fungal smear a sensitivity of 33%. The results of this study suggest that stone gram stain cannot be relied on to detect a positive stone culture and may fail to detect up to 62% of infected stones. However, when positive, gram stain accurately predicts a positive stone culture in 86% of cases. Copyright © 2014 Elsevier Inc. All rights reserved.
Comparative efficacy of cedarwood oil and xylene in hematoxylin and eosin staining procedures: An experimental study.

PubMed

Indu, Sudip; Ramesh, V; Indu, Priyanka Chakravarty; Prashad, Karthikshree V; Premalatha, B; Ramadoss, K

2014-07-01

Xylene is used as a clearing agent in hematoxylin and eosin (H and E) staining of tissue sections in routine histopathology based diagnosis. However, the hazards associated with exposure to xylene are of concern. Numerous solutions mainly essential oils have been evaluated in the past as clearing agents, which can possibly be substituted for xylene during the routine tissue processing. The aim of this study is to compare the efficacy of essential oil (cedarwood oil), as a possible replacement for xylene in H and E staining procedures. The study was carried out in the Department of Oral Pathology and Microbiology. Thirty paraffin blocks of the routine biopsy specimen were retrieved from the department archives. The cedarwood oil was procured from organic and essential oil dealer in the local market. Two to three paraffin sections of four micron thickness were cut from each of the 30 paraffin blocks of processed tissue specimens, were subjected to different clearing agents: Essential oil (8% cedarwood oil) or xylene and stained with H and E stain. The stained sections were scored based on nuclear and cytoplasmic details, clarity and uniformity of staining. Significant correlation was observed between cedarwood oil and xylene in terms of the three staining quality parameters assessed. We conclude that cedarwood oil can be an effective, eco-friendly and safe alternative to xylene as a clearing agent in the histopathological laboratory.
Histologic evaluation of post-implantation immediate C4d deposition in 13 intestinal grafts: correlation with cell-based crossmatching, cold ischemia time, and preservation injury.

PubMed

López-García, P; Calvo Pulido, J; Colina, F; Ballestin Carcavilla, C; Jiménez-Romero, C; Martinez González, M A; Ibarrola de Andrés, C; López-Alonso, G; Cambra Molero, F; Justo Alonso, I; Moreno-González, E

2014-01-01

C4d deposits are predictive of humoral rejection in kidney and heart transplantation. The aim of this study was to identify C4d deposit patterns in intestinal mucosa of the grafts on biopsy specimens obtained immediately after implantation and to detect if it could be a valuable tool to predict humoral or acute rejection. A second objective was to search for a statistically significant relationship between positive C4d deposition and other collected variables. Thirteen immediately post-transplantation mucosal graft biopsy specimens, formalin fixed, underwent immunohistochemical stain for C4d deposits. Diffuse intense staining of capillary endothelium was considered positive and absent, focal or weak stains as negative. Preservation injury grade and cold ischemia times were registered for each case. Donor-specific preformed antibodies were detected by complement dependent cytotoxicity serologic technique (crossmatching). Another 19 endoscopic follow-up biopsy specimens from days 2 to 6 were also evaluated. Statistical studies were made using the index of correlation ρ (Spearman's test). Diffuse intense C4d deposits were observed in 2 grafts, focal and weak in 5, and completely negative in 6. The mean cold ischemia time was 327 ± 101 minutes. Two cases showed diffuse positive deposits, 1 had a positive crossmatch and the cold ischemia time was 360 minutes whereas the other had not preformed antibodies and its cold ischemia time was 475 minutes. Humoral or acute rejection was not observed in follow-up mucosal biopsy specimens. There was no statistically significant relationship between the C4d deposition, cold ischemia time, crossmatching results, and preservation injury degree. In conclusion, C4d deposition was not a helpful tool for diagnosis of humoral rejection and prediction of acute rejection during the early post-transplantation period. Copyright © 2014 Elsevier Inc. All rights reserved.
Different cytokeratin and neuronal cell adhesion molecule staining patterns in focal nodular hyperplasia and hepatic adenoma and their significance

PubMed Central

Iyer, Anita; Robert, Marie E.; Bifulco, Carlo B.; Salem, Ronald R.; Jain, Dhanpat

2013-01-01

Summary Differentiating focal nodular hyperplasia from hepatic adenoma can be challenging. Cytokeratin 7, neuronal cell adhesion molecule, and cytokeratin 19 are differentially expressed in hepatocytes, biliary epithelium, and possibly hepatic progenitor/stem cells. CD34 is known to have altered expression patterns in the hepatic endothelium in conditions associated with abnormal perfusion and in hepatocellular carcinoma. The purpose of this study was to examine the expression pattern of these markers in focal nodular hyperplasia and hepatic adenoma and assess their diagnostic use. Ten resection specimens each of hepatic adenoma and focal nodular hyperplasia (including a case of telangiectatic focal nodular hyperplasia) were selected for the study. Immunohistochemical analysis was performed using antibodies against cytokeratin 7, cytokeratin 19, neuronal cell adhesion molecule, and CD34 on formalin-fixed, paraffin-embedded sections from each case. The staining patterns and intensity for each marker were analyzed. In hepatic adenoma, the cytokeratin 7 stain revealed strong positivity in hepatocytes in patches, with a gradual decrease in the staining intensity as the cells differentiated towards mature hepatocytes. Although bile ducts were typically absent in hepatic adenoma, occasional ductules could be identified with cytokeratin 7 stain. In focal nodular hyperplasia, cytokeratin 7 showed strong staining of the biliary epithelium within the fibrous septa and staining of the peripheral hepatocytes of most lobules that was focal and weaker than hepatic adenoma. Cytokeratin 19 and neuronal cell adhesion molecule showed patchy and moderate staining in the biliary epithelium of the ductules in focal nodular hyperplasia. While in the hepatic adenoma, cytokeratin 19 showed only rare positivity in occasional cells within ductules, and neuronal cell adhesion molecule marked occasional isolated cells in the lesion. CD34 showed staining of sinusoids in the inflow areas (periportal areas) in both focal nodular hyperplasia and hepatic adenoma. One case of telangiectatic focal nodular hyperplasia revealed both hepatic adenoma–like and focal nodular hyperplasia–like staining patterns. Distinct cytokeratin 7, cytokeratin 19, and neuronal cell adhesion molecule staining patterns are seen in hepatic adenoma and focal nodular hyperplasia possibly suggest activation of different subsets of hepatic progenitor/stem cell and can be diagnostically useful. PMID:18602664
Glutamine synthetase, heat shock protein-70, and glypican-3 in intrahepatic cholangiocarcinoma and tumors metastatic to liver.

PubMed

Lagana, Stephen M; Moreira, Roger K; Remotti, Helen E; Bao, Fei

2013-05-01

Glutamine synthetase (GS), heat shock protein-70 (HSP-70), and glypican-3 (GPC-3) are markers best characterized in hepatocellular lesions, where they are useful in distinguishing hepatocellular carcinoma from dysplastic nodules. Their staining patterns in intrahepatic cholangiocarcinoma (IH-ChCa) and metastatic tumors in liver are not well described. Tissue microarrays containing 41 IH-ChCa and 24 metastatic tumors in liver were stained with commercially available antibodies to GS, HSP-70, and GPC-3. Five percent staining of tumor cells was considered positive for HSP-70 and GPC-3. For GS, 50% was the cut-off. GS reactivity was present in 31 of 41 IH-ChCa (76%), with the median amount of staining being 65% of tumor cells. HSP-70 reactivity was present in 36 of 41 IH-ChCa (88%) with the median amount of staining being 75% of tumor cells. GPC-3 reactivity was absent from all IH-ChCa. Twenty-seven of 41 IH-ChCa cases were positive for both GS and HSP-70 (66%). GS reactivity was present in 17 of 24 tumors metastatic to liver (71%), with the median amount of staining being 50% of tumor cells. HSP-70 reactivity was present in 21 of 24 tumors metastatic to liver (88%) with the median amount of staining being 80% of tumor cells. GPC-3 reactivity was present in 2 of 24 tumors metastatic to liver (8%) with one showing 5% staining and the other showing 50% staining of tumor cells. Fifteen of 24 cases were positive for both GS and HSP-70 (63%), and 2 cases were positive for all 3 markers (8%). Of the panel of immunostains currently commonly used to distinguish hepatocellular carcinoma from dysplastic hepatocytic nodules, only GPC-3 did not react frequently with metastatic tumors and IH-ChCa, although there was staining in 2 metastatic tumors. GS and HSP-70 are typically positive in IH-ChCa and metastatic tumors. Nothing should be inferred about the histogenesis of a tumor based on positive staining with either of these 2 markers, which currently have no role in tumor of unknown origin panels.
p16/Ki-67 Dual Stain Cytology for Detection of Cervical Precancer in HPV-Positive Women.

PubMed

Wentzensen, Nicolas; Fetterman, Barbara; Castle, Philip E; Schiffman, Mark; Wood, Shannon N; Stiemerling, Eric; Tokugawa, Diane; Bodelon, Clara; Poitras, Nancy; Lorey, Thomas; Kinney, Walter

2015-12-01

Human papillomavirus (HPV)-based cervical cancer screening requires triage markers to decide who should be referred to colposcopy. p16/Ki-67 dual stain cytology has been proposed as a biomarker for cervical precancers. We evaluated the dual stain in a large population of HPV-positive women. One thousand five hundred and nine HPV-positive women screened with HPV/cytology cotesting at Kaiser Permanente California were enrolled into a prospective observational study in 2012. Dual stain cytology was performed on residual Surepath material, and slides were evaluated for dual stain-positive cells. Disease endpoints were ascertained from the clinical database at KPNC. We evaluated the clinical performance of the assay among all HPV-positive women and among HPV-positive, cytology-negative women. We used internal benchmarks for clinical management to evaluate the clinical relevance of the dual stain assay. We evaluated sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the dual stain compared with Pap cytology. All statistical tests were two-sided. The dual stain had lower positivity (45.9%) compared with cytology at an ASC-US threshold (53.4%). For detection of CIN2+, the dual stain had similar sensitivity (83.4% vs 76.6%, P = .1), and statistically higher specificity (58.9% vs 49.6%, P < .001), PPV (21.0% vs 16.6%, P < .001), and NPV (96.4% vs 94.2%, P = .01) compared with cytology. Similar patterns were observed for CIN3+. Women with a positive test had high enough risk for referral to colposcopy, while the risk for women with negative tests was below a one-year return threshold based on current US management guidelines. Dual stain cytology showed good risk stratification for all HPV-positive women and for HPV-positive women with normal cytology. Additional follow-up is needed to determine how long dual stain negative women remain at low risk of precancer. Published by Oxford University Press 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.
Port-wine stain

MedlinePlus

... Laser therapy is most successful in removing port-wine stains. It is the only method that can destroy the tiny blood vessels in the skin without causing much damage to the skin. The exact type ... on the person's age, skin type, and particular port-wine stain.
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.

Negative Staining for COL4A5 Correlates With Worse Prognosis and More Severe Ultrastructural Alterations in Males With Alport Syndrome.

PubMed

Said, Samar M; Fidler, Mary E; Valeri, Anthony M; McCann, Brooke; Fiedler, Wade; Cornell, Lynn D; Alexander, Mariam Priya; Alkhunaizi, Ahmed M; Sullivan, Anne; Cramer, Carl H; Hogan, Marie C; Nasr, Samih H

2017-01-01

Alport syndrome (AS) is a genetic disorder characterized by progressive hematuric nephropathy with or without sensorineural hearing loss and ocular lesions. Previous studies on AS included mostly children. To determine the prognostic value of loss of staining for collagen type IV alpha 5 (COL4A5) and its relationship with the ultrastructural glomerular basement membrane alterations, we performed direct immunofluorescence using a mixture of fluorescein isothiocyanate-conjugated and Texas-red conjugated antibodies against COL4A5 and COL4A2, respectively, on renal biopsies of 25 males with AS (including 16 who were diagnosed in adulthood). All patients showed normal positive staining of glomerular basement membranes and tubular basement membranes for COL4A2. Of the 25 patients, 10 (40%) patients showed loss of staining for COL4A5 (including 89% of children and 13% of adults) and the remaining 15 (60%) had intact staining for COL4A5. Compared with patients with intact staining for COL4A5, those with loss of staining had more prominent ultrastructural glomerular basement membrane alterations and were younger at the time of biopsy. By Kaplan-Meier survival analysis and Cox regression analysis, loss of staining for COL4A5 predicted earlier progression to overt proteinuria and stage 2 chronic kidney disease or worse. By multivariate Cox regression analysis, loss of staining for COL4A5 was an independent predictor of the development of overt proteinuria and stage 2 chronic kidney disease or worse. Thus, the COL4A5 expression pattern has an important prognostic value and it correlates with the severity of ultrastructural glomerular basement membrane alterations in males with AS. Loss of COL4A5 staining is uncommon in patients with AS diagnosed in their adulthood.
Color normalization of histology slides using graph regularized sparse NMF

NASA Astrophysics Data System (ADS)

Sha, Lingdao; Schonfeld, Dan; Sethi, Amit

2017-03-01

Computer based automatic medical image processing and quantification are becoming popular in digital pathology. However, preparation of histology slides can vary widely due to differences in staining equipment, procedures and reagents, which can reduce the accuracy of algorithms that analyze their color and texture information. To re- duce the unwanted color variations, various supervised and unsupervised color normalization methods have been proposed. Compared with supervised color normalization methods, unsupervised color normalization methods have advantages of time and cost efficient and universal applicability. Most of the unsupervised color normaliza- tion methods for histology are based on stain separation. Based on the fact that stain concentration cannot be negative and different parts of the tissue absorb different stains, nonnegative matrix factorization (NMF), and particular its sparse version (SNMF), are good candidates for stain separation. However, most of the existing unsupervised color normalization method like PCA, ICA, NMF and SNMF fail to consider important information about sparse manifolds that its pixels occupy, which could potentially result in loss of texture information during color normalization. Manifold learning methods like Graph Laplacian have proven to be very effective in interpreting high-dimensional data. In this paper, we propose a novel unsupervised stain separation method called graph regularized sparse nonnegative matrix factorization (GSNMF). By considering the sparse prior of stain concentration together with manifold information from high-dimensional image data, our method shows better performance in stain color deconvolution than existing unsupervised color deconvolution methods, especially in keeping connected texture information. To utilized the texture information, we construct a nearest neighbor graph between pixels within a spatial area of an image based on their distances using heat kernal in lαβ space. The representation of a pixel in the stain density space is constrained to follow the feature distance of the pixel to pixels in the neighborhood graph. Utilizing color matrix transfer method with the stain concentrations found using our GSNMF method, the color normalization performance was also better than existing methods.
Stain removal and whitening by baking soda dentifrice: A review of literature.

PubMed

Li, Yiming

2017-11-01

Tooth discoloration may be caused by intrinsic or extrinsic stains or a combination of both. There are 2 major approaches to removing the stains, including the chemical mechanism using peroxides for tooth bleaching and the mechanical mechanism using abrasives in prophylactic pastes and dentifrices to remove stains, resulting in a whitening effect. Attempts have also been made to add a low concentration of peroxides to dentifrices to enhance their abrasive cleaning to remove tooth stains. This article provides a review of both in vitro and clinical studies on stain removal and whitening effect of dentifrices containing sodium bicarbonate (baking soda). In recent years, whitening dentifrices have become popular because of little additional effort for use, ease of availability, low cost, and accumulated evidence of clinical efficacy and safety in the literature. Advances in research and technology have led to innovative formulations of dentifrices using baking soda as the sole abrasive or a component of an abrasive system. Baking soda is biologically compatible with acid-buffering capacities, antibacterial at high concentrations, and has a relatively lower abrasivity. The evidence available in the literature indicates that baking soda-based dentifrices are effective and safe for tooth stain removal and consequently whitening. A number of clinical studies have also shown that baking soda-based dentifrices are more effective in stain removal and whitening than some non-baking soda-containing dentifrices with a higher abrasivity. So far, research efforts have mainly focused on stain removal and tooth-whitening efficacy and clinical safety of baking soda dentifrices used with manual toothbrushes, with only a few studies investigating their effects using powered toothbrushes, for which further research is encouraged. As part of a daily oral hygiene practice, baking soda-based dentifrice is a desirable, alternative or additional measure for tooth stain removal and whitening. Copyright © 2017 American Dental Association. Published by Elsevier Inc. All rights reserved.
Oxidative Stress Induced Age Dependent Meibomian Gland Dysfunction in Cu, Zn-Superoxide Dismutase-1 (Sod1) Knockout Mice

PubMed Central

Ibrahim, Osama M. A.; Dogru, Murat; Matsumoto, Yukihiro; Igarashi, Ayako; Kojima, Takashi; Wakamatsu, Tais Hitomi; Inaba, Takaaki; Shimizu, Takahiko; Shimazaki, Jun; Tsubota, Kazuo

2014-01-01

Purpose The purpose of our study was to investigate alterations in the meibomian gland (MG) in Cu, Zn-Superoxide Dismutase-1 knockout (Sod1 −/−) mouse. Methods Tear function tests [Break up time (BUT) and cotton thread] and ocular vital staining test were performed on Sod1 −/− male mice (n = 24) aged 10 and 50 weeks, and age and sex matched wild–type (+/+) mice (n = 25). Tear and serum samples were collected at sacrifice for inflammatory cytokine assays. MG specimens underwent Hematoxylin and Eosin staining, Mallory staining for fibrosis, Oil Red O lipid staining, TUNEL staining, immunohistochemistry stainings for 4HNE, 8-OHdG and CD45. Transmission electron microscopic examination (TEM) was also performed. Results Corneal vital staining scores in the Sod1 −/− mice were significantly higher compared with the wild type mice throughout the follow-up. Tear and serum IL-6 and TNF-α levels also showed significant elevations in the 10 to 50 week Sod1 −/− mice. Oil Red O staining showed an accumulation of large lipid droplets in the Sod1 −/− mice at 50 weeks. Immunohistochemistry revealed both increased TUNEL and oxidative stress marker stainings of the MG acinar epithelium in the Sod1 −/− mice compared to the wild type mice. Immunohistochemistry staining for CD45 showed increasing inflammatory cell infiltrates from 10 to 50 weeks in the Sod1 −/− mice compared to the wild type mice. TEM revealed prominent mitochondrial changes in 50 week Sod1 −/− mice. Conclusions Our results suggest that reactive oxygen species might play a vital role in the pathogensis of meibomian gland dysfunction. The Sod1 −/− mouse appears to be a promising model for the study of reactive oxygen species associated MG alterations. PMID:25036096
The evaluation of renal ischaemic damage: the value of CD10 monoclonal antibody staining and of biochemical assessments of tissue viability

PubMed Central

Tagboto, S; Griffiths, A Paul

2007-01-01

Background It is well recognised that there is often a disparity between the structural changes observed in the kidney following renal injury and the function of the organ. For this reason, we carried out studies to explore possible means of studying and quantifying the severity of renal ischaemic damage using a laboratory model. Methods To do this, freshly isolated rabbit kidney tissue was subjected to warm (37°C) or cold (1°C) ischaemia for 20 hours. Following this, the tissue was stained using Haematoxylin and Eosin (H+E), Periodic Schiff reagent (PAS) and the novel monoclonal antibody CD10 stain. Additionally, ischaemic damage to the kidneys was assessed by biochemical tests of tissue viability using formazan-based colorimetry. Results CD 10 antibody intensely stained the brush border of control kidney tissue with mild or no cytoplasmic staining. Cell injury was accompanied by a redistribution of CD10 into the lumen and cell cytoplasm. There was good correlation between a score of histological damage using the CD 10 monoclonal antibody stain and the biochemical assessment of viability. Similarly, a score of histological damage using traditional PAS staining correlated well with that using the CD10 antibody stain. In particular, the biochemical assay and the monoclonal antibody staining techniques were able to demonstrate the efficacy of Soltran (this solution is used cold to preserve freshly isolated human kidneys prior to transplantation) in preserving renal tissue at cold temperatures compared to other randomly selected solutions. Conclusion We conclude that the techniques described using the CD10 monoclonal antibody stain may be helpful in the diagnosis and assessment of ischaemic renal damage. In addition, biochemical tests of viability may have an important role in routine histopathological work by giving additional information about cellular viability which may have implications on the function of the organ. PMID:17531101
Gram staining of protected pulmonary specimens in the early diagnosis of ventilator-associated pneumonia.

PubMed

Mimoz, O; Karim, A; Mazoit, J X; Edouard, A; Leprince, S; Nordmann, P

2000-11-01

We evaluated prospectively the use of Gram staining of protected pulmonary specimens to allow the early diagnosis of ventilator-associated pneumonia (VAP), compared with the use of 60 bronchoscopic protected specimen brushes (PSB) and 126 blinded plugged telescopic catheters (PTC) obtained from 134 patients. Gram stains were from Cytospin slides; they were studied for the presence of microorganisms in 10 and 50 fields by two independent observers and classified according to their Gram stain morphology. Quantitative cultures were performed after serial dilution and plating on appropriate culture medium. A final diagnosis of VAP, based on a culture of > or = 10(3) c.f.u. ml-1, was established after 81 (44%) samplings. When 10 fields were analysed, a strong relationship was found between the presence of bacteria on Gram staining and the final diagnosis of VAP (for PSB and PTC respectively: sensitivity 74 and 81%, specificity 94 and 100%, positive predictive value 91 and 100%, negative predictive value 82 and 88%). The correlation was less when we compared the morphology of microorganisms observed on Gram staining with those of bacteria obtained from quantitative cultures (for PSB and PTC respectively: sensitivity 54 and 69%, specificity 86 and 89%, positive predictive value 72 and 78%, negative predictive value 74 and 84%). Increasing the number of fields read to 50 was associated with a slight decrease in specificity and positive predictive value of Gram staining, but with a small increase in its sensitivity and negative predictive value. The results obtained by the two observers were similar to each other for both numbers of fields analysed. Gram staining of protected pulmonary specimens performed on 10 fields predicted the presence of VAP and partially identified (using Gram stain morphology) the microorganisms growing at significant concentrations, and could help in the early choice of the treatment of VAP. Increasing the number of fields read or having the Gram stain analysed by two independent individuals did not improve the results.
p16/Ki-67 Dual Stain Cytology for Detection of Cervical Precancer in HPV-Positive Women

PubMed Central

Fetterman, Barbara; Castle, Philip E.; Schiffman, Mark; Wood, Shannon N.; Stiemerling, Eric; Tokugawa, Diane; Bodelon, Clara; Poitras, Nancy; Lorey, Thomas; Kinney, Walter

2015-01-01

Background: Human papillomavirus (HPV)–based cervical cancer screening requires triage markers to decide who should be referred to colposcopy. p16/Ki-67 dual stain cytology has been proposed as a biomarker for cervical precancers. We evaluated the dual stain in a large population of HPV-positive women. Methods: One thousand five hundred and nine HPV-positive women screened with HPV/cytology cotesting at Kaiser Permanente California were enrolled into a prospective observational study in 2012. Dual stain cytology was performed on residual Surepath material, and slides were evaluated for dual stain–positive cells. Disease endpoints were ascertained from the clinical database at KPNC. We evaluated the clinical performance of the assay among all HPV-positive women and among HPV-positive, cytology-negative women. We used internal benchmarks for clinical management to evaluate the clinical relevance of the dual stain assay. We evaluated sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the dual stain compared with Pap cytology. All statistical tests were two-sided. Results: The dual stain had lower positivity (45.9%) compared with cytology at an ASC-US threshold (53.4%). For detection of CIN2+, the dual stain had similar sensitivity (83.4% vs 76.6%, P = .1), and statistically higher specificity (58.9% vs 49.6%, P < .001), PPV (21.0% vs 16.6%, P < .001), and NPV (96.4% vs 94.2%, P = .01) compared with cytology. Similar patterns were observed for CIN3+. Women with a positive test had high enough risk for referral to colposcopy, while the risk for women with negative tests was below a one-year return threshold based on current US management guidelines. Conclusion: Dual stain cytology showed good risk stratification for all HPV-positive women and for HPV-positive women with normal cytology. Additional follow-up is needed to determine how long dual stain negative women remain at low risk of precancer. PMID:26376685
Statistical modeling, detection, and segmentation of stains in digitized fabric images

NASA Astrophysics Data System (ADS)

Gururajan, Arunkumar; Sari-Sarraf, Hamed; Hequet, Eric F.

2007-02-01

This paper will describe a novel and automated system based on a computer vision approach, for objective evaluation of stain release on cotton fabrics. Digitized color images of the stained fabrics are obtained, and the pixel values in the color and intensity planes of these images are probabilistically modeled as a Gaussian Mixture Model (GMM). Stain detection is posed as a decision theoretic problem, where the null hypothesis corresponds to absence of a stain. The null hypothesis and the alternate hypothesis mathematically translate into a first order GMM and a second order GMM respectively. The parameters of the GMM are estimated using a modified Expectation-Maximization (EM) algorithm. Minimum Description Length (MDL) is then used as the test statistic to decide the verity of the null hypothesis. The stain is then segmented by a decision rule based on the probability map generated by the EM algorithm. The proposed approach was tested on a dataset of 48 fabric images soiled with stains of ketchup, corn oil, mustard, ragu sauce, revlon makeup and grape juice. The decision theoretic part of the algorithm produced a correct detection rate (true positive) of 93% and a false alarm rate of 5% on these set of images.
Confocal mosaicing microscopy of basal-cell carcinomas ex vivo: progress in digital staining to simulate histology-like appearance

NASA Astrophysics Data System (ADS)

Bini, Jason; Spain, James; Nehal, Kishwer; Hazelwood, Vikki; DiMarzio, Charles; Rajadhyaksha, Milind

2011-03-01

Confocal mosaicing microscopy enables rapid imaging of large areas of fresh tissue, without the processing that is necessary for conventional histology. Using acridine orange (1 milliMolar, 20 seconds) to stain nuclei, basal cell carcinomas were detected in fluorescence confocal mosaics of Mohs surgical excisions with sensitivity of 96.6% and specificity of 89.2%. A possible barrier toward clinical acceptance is that confocal mosaics are based on a single mode of contrast and appear in grayscale, whereas histology is based on two (hematoxylin for nuclei, eosin for cellular cytoplasm and dermis) and appears purple-and-pink. Toward addressing this barrier, we report progress in developing a multispectral analytical model for digital staining: fluorescence confocal mosaics, which show only nuclei, are digitally stained purple and overlaid on reflectance confocal mosaics, which show only cellular cytoplasm and dermis, and digitally stained pink, to mimic the appearance of histology. Comparison of digitally stained confocal mosaics by our Mohs surgeon to the corresponding Mohs histology shows good correlation for normal and tumor detail. Digitally stained confocal mosaicing microscopy may allow direct examination of freshly excised tissue and serve as an adjunct for rapid pathology at-the-bedside.
Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells.

PubMed

Liesche, Johannes; Marek, Magdalena; Günther-Pomorski, Thomas

2015-01-01

Yeast cells are protected by a cell wall that plays an important role in the exchange of substances with the environment. The cell wall structure is dynamic and can adapt to different physiological states or environmental conditions. For the investigation of morphological changes, selective staining with fluorescent dyes is a valuable tool. Furthermore, cell wall staining is used to facilitate sub-cellular localization experiments with fluorescently-labeled proteins and the detection of yeast cells in non-fungal host tissues. Here, we report staining of Saccharomyces cerevisiae cell wall with Trypan Blue, which emits strong red fluorescence upon binding to chitin and yeast glucan; thereby, it facilitates cell wall analysis by confocal and super-resolution microscopy. The staining pattern of Trypan Blue was similar to that of the widely used UV-excitable, blue fluorescent cell wall stain Calcofluor White. Trypan Blue staining facilitated quantification of cell size and cell wall volume when utilizing the optical sectioning capacity of a confocal microscope. This enabled the quantification of morphological changes during growth under anaerobic conditions and in the presence of chemicals, demonstrating the potential of this approach for morphological investigations or screening assays.
Gram staining for the treatment of peritonsillar abscess.

PubMed

Takenaka, Yukinori; Takeda, Kazuya; Yoshii, Tadashi; Hashimoto, Michiko; Inohara, Hidenori

2012-01-01

Objective. To examine whether Gram staining can influence the choice of antibiotic for the treatment of peritonsillar abscess. Methods. Between 2005 and 2009, a total of 57 cases of peritonsillar abscess were analyzed with regard to cultured bacteria and Gram staining. Results. Only aerobes were cultured in 16% of cases, and only anaerobes were cultured in 51% of cases. Mixed growth of aerobes and anaerobes was observed in 21% of cases. The cultured bacteria were mainly aerobic Streptococcus, anaerobic Gram-positive cocci, and anaerobic Gram-negative rods. Phagocytosis of bacteria on Gram staining was observed in 9 cases. The bacteria cultured from these cases were aerobic Streptococcus, anaerobic Gram-positive cocci, and anaerobic Gram-negative rods. The sensitivity of Gram staining for the Gram-positive cocci and Gram-negative rods was 90% and 64%, respectively. The specificity of Gram staining for the Gram-positive cocci and Gram-negative rods was 62% and 76%, respectively. Most of the Gram-positive cocci were sensitive to penicillin, but some of anaerobic Gram-negative rods were resistant to penicillin. Conclusion. When Gram staining shows only Gram-positive cocci, penicillin is the treatment of choice. In other cases, antibiotics effective for the penicillin-resistant organisms should be used.
A Military-Relevant Model of Closed Concussive Head Injury: Longitudinal Studies Characterizing and Validating Single and Repetitive mTBI

DTIC Science & Technology

2013-10-01

processed at FD Neurotechnologies Inc. (Ellicott City, MD, USA). A series of coronal free floating brain sections (40 μm; 960 µm interval from +4.0...axonal damage. Another series of coronal sections was stained using FD NeuroSilverTM Kit II (FD Neurotechnologies Inc., Ellicott City, MD, USA) for... Neurotechnologies Inc for processing and staining. Tissue processing and staining protocols have been optimized for fluorescent staining of AQ4 co
An improved silver staining procedure for schizodeme analysis in polyacrylamide gradient gels.

PubMed

Gonçalves, A M; Nehme, N S; Morel, C M

1990-01-01

A simple protocol is described for the silver staining of polyacrylamide gradient gels used for the separation of restriction fragments of kinetoplast DNA [schizodeme analysis of trypanosomatids (Morel et al., 1980)]. The method overcomes the problems of non-uniform staining and strong background color which are frequently encountered when conventional protocols for silver staining of linear gels are applied to gradient gels. The method described has proven to be of general applicability for DNA, RNA and protein separations in gradient gels.
Staining of retinal neurons in the isolated eyecup by extracellular horseradish peroxidase injection.

PubMed

Amthor, F R; Jackson, C A

1986-01-01

A reliable method has been developed for the staining of mammalian retinal neurons in the isolated eyecup. Small injections of horseradish peroxidase are made at a number of places on the retinal surface while the hemisected eyecup preparation floats in Eagle's medium. After 30 min, the retinas are removed, fixed, and reacted with diaminobenzidine. Golgi-like staining of fine dendrites, spines and axons of ganglion cells is achieved. The method also stains amacrine, horizontal and bipolar cells.
Diagnosis of subclinical amniotic fluid infection prior to rescue cerclage using gram stain and glucose tests: an individual patient meta-analysis.

PubMed

Lisonkova, Sarka; Sabr, Yasser; Joseph, K S

2014-02-01

Microbial invasion of the amniotic cavity (MIAC) can affect outcomes following rescue cerclage. We carried out a study to compare the diagnostic performance of the Gram stain and glucose tests for detecting subclinical MIAC. We used individual-level data from published studies on Gram stain, glucose, and amniotic fluid culture among women with preterm labour. We calculated the sensitivity, specificity, area under the curve (AUC) and other indices, with amniotic fluid culture results used as the gold standard. The probability of infection using both tests as predictors was also estimated using logistic regression. The rate of culture-confirmed MIAC was 11.8% (34 of 288 women). The Gram stain test yielded a sensitivity of 65% (95% CI 46% to 78%) and a specificity of 99% (95% CI 98% to 100%). A positive Gram stain or glucose test had a sensitivity of 88% (95% CI 72% to 96%) and a specificity of 87% (95% CI 82% to 90%), while a positive Gram stain and a positive glucose test had a sensitivity of 62% (95% CI 44% to 77%) and a specificity of 100% (95% CI 98% to 100%). The AUC for the tests were Gram stain 0.82 (95% CI 0.74 to 0.90), glucose 0.86 (95% CI 0.80 to 0.93), and combined Gram stain and glucose 0.92 (95% CI 0.86 to 0.98). Using the tests, singly or in combination, provided greater clinically important calibration, risk-stratification, and classification accuracy than using no tests. Amniotic fluid Gram stain and/or glucose testing provides substantially improved performance for the diagnosis of subclinical MIAC compared with no testing.
Misinterpretation of Gram Stain from the Stationary Growth Phase of Positive Blood Cultures for Brucella and Acinetobacter Species.

PubMed

Bazzi, Ali M; Al-Tawfiq, Jaffar A; Rabaan, Ali A

2017-01-01

Acinetobacter baumannii and Brucella species are Gram-negative organisms that are vulnerable to misinterpretation as Gram-positive or Gram-variable in blood cultures. We assess the random errors in gram stain interpretation to reduce the likelihood of such errors and therefore patient harm. Aerobic and anaerobic blood cultures from two patients in an acute care facility in Saudi Arabia were subjected to preliminary Gram-staining. In case 1, VITEK-2 Anaerobe Identification, repeat Gram staining from a blood agar plate, Remel BactiDrop™ Oxidase test, Urea Agar urease test and real-time PCR were used to confirm presence of Brucella and absence of Coryneform species. In case 2, repeat Gram- staining from the plate and the vials, VITEK-2 Gram-Negative Identification, real-time PCR and subculture on to Columbia agar, blood agar, and MacConkey agar were carried out to identify A. baumannii . In case 1, initially pleomorphic Gram-positive bacteria were identified. Coryneform species were suspected. Tiny growth was observed after 24 h on blood agar plates, and good growth by 48 h. Presence of Brucella species was ultimately confirmed. In case 2, preliminary Gram-stain results suggested giant Gram-positive oval cocci. Further testing over 18-24 h identified A. baumannii . Oxidase test from the plate and urease test from the culture vial is recommended after apparent identification of pleomorphic Gram-positive bacilli from blood culture, once tiny growth is observed, to distinguish Brucella from Corynebacterium species. If giant Gram-positive oval cocci are indicated by preliminary Gram-staining, it is recommended that the Gram stain be repeated from the plate after 4-6 h, or culture should be tested in Triple Sugar Iron (TSI) medium and the Gram stain repeated after 2-4 h incubation.
Morphologic characterization of specific granules in Greyhound eosinophils.

PubMed

Iazbik, M C; Couto, C G

2005-06-01

"Vacuolated" eosinophils (ie, eosinophils with empty, nonstaining granules) have been described previously in normal Greyhounds. However, to our knowledge, detailed studies of granules in vacuolated and normal eosinophils in this breed have not been performed. The objective of this prospective study was to characterize some of the morphologic, ultrastructural, and cytochemical staining features of specific (primary) granules in both normal and vacuolated eosinophils in Greyhound blood. Morphologic features of eosinophils in Wright's- and Diff-Quik-stained peripheral blood smears from 49 Greyhounds were compared with 200 blood smears from non-Greyhound dogs. Transmission electron microscopy was done on blood from 3 Greyhounds with vacuolated eosinophils and 3 with normal eosinophil granules. Blood smears from 4 of these dogs also were stained cytochemically with alkaline phosphatase (AP), chloracetate esterase (CAE), and alpha naphthyl butyrate esterase (ANBE). The morphologic features and tinctorial properties of vacuolated and normal eosinophils were compared. Twenty-six Greyhounds (53%) had vacuolated eosinophils and 23 (47%) had normal granulated eosinophils in smears stained with Wright's stain. Only 1% of eosinophils were vacuolated in non-Greyhound dogs. Twenty of the 23 (85%) Greyhounds with normal granulated eosinophils on Wright's-stained smears had vacuolated eosinophils in smears stained with Diff-Quik. Ultrastructurally, no morphologic differences were observed between granules of vacuolated and normal eosinophils. Both vacuolated and normal eosinophils in Greyhounds were positive for AP and negative for CAE and ANBE, as expected for normal dogs. Vacuolated eosinophils in Greyhounds likely reflect, at least in part, differential staining properties of the specific granules with different hematologic stains. Ultrastuctural and cytochemical features of eosinophil granules were similar in normal and vacuolated eosinophils from Greyhounds.
Quantitative Analysis of Immunohistochemistry in Melanoma Tumors

PubMed Central

Lilyquist, Jenna; White, Kirsten Anne Meyer; Lee, Rebecca J.; Philips, Genevieve K.; Hughes, Christopher R.; Torres, Salina M.

2017-01-01

Abstract Identification of positive staining is often qualitative and subjective. This is particularly troublesome in pigmented melanoma lesions, because melanin is difficult to distinguish from the brown stain resulting from immunohistochemistry (IHC) using horse radish peroxidase developed with 3,3′-Diaminobenzidine (HRP-DAB). We sought to identify and quantify positive staining, particularly in melanoma lesions. We visualized G-protein coupled estrogen receptor (GPER) expression developed with HRP-DAB and counterstained with Azure B (stains melanin) in melanoma tissue sections (n = 3). Matched sections (n = 3), along with 22 unmatched sections, were stained only with Azure B as a control. Breast tissue (n = 1) was used as a positive HRP-DAB control. Images of the stained tissues were generated using a Nuance Spectral Imaging Camera. Analysis of the images was performed using the Nuance Spectral Imaging software and SlideBook. Data was analyzed using a Kruskal–Wallis one way analysis of variance (ANOVA). We showed that a pigmented melanoma tissue doubly stained with anti-GPER HRP-DAB and Azure B can be unmixed using spectra derived from a matched, Azure B-only section, and an anti-GPER HRP-DAB control. We unmixed each of the melanoma lesions using each of the Azure B spectra, evaluated the mean intensity of positive staining, and examined the distribution of the mean intensities (P = .73; Kruskal–Wallis). These results suggest that this method does not require a matched Azure B-only stained control tissue for every melanoma lesion, allowing precious tissues to be conserved for other studies. Importantly, this quantification method reduces the subjectivity of protein expression analysis, and provides a valuable tool for accurate evaluation, particularly for pigmented tissues. PMID:28403073
A baculovirus polyhedron envelope protein: immunogold localization in infected cells and mature polyhedra.

PubMed

Russell, R L; Rohrmann, G F

1990-01-01

A polyclonal antiserum against a trpE fusion protein containing the complete open reading frame of the polyhedron envelope (PE) protein from the nuclear polyhedrosis virus of Orgyia pseudotsugata (OpMNPV) was used for immunogold staining and electron microscopic examination of polyhedra, isolated polyhedron envelopes, and infected insect cells at selected times postinfection. The antiserum specifically stained the peripheral envelope of mature polyhedra and also stained the envelope structure which remained after polyhedra were dissolved in dilute alkaline solutions. In OpMNPV-infected Lymantria dispar cells, the PE protein was detected by 48 hr postinfection (hr p.i.) but specific localization and staining of developing polyhedra were not evident. However, by 72 hr p.i. substantial and preferential staining of the periphery of developing polyhedra was evident even though a distinct polyhedron envelope was not yet observed. In addition, the periphery of fibrillar structures was stained by the PE antiserum. By 96 hr p.i., mature envelopes surrounded polyhedra and these polyhedron envelopes were stained with the PE antibody. The progression of PE protein staining during polyhedron morphogenesis indicates that the PE protein accumulates and becomes associated with developing polyhedra in the nucleus between 48 and 72 hr p.i. Very late in infection the mature polyhedron envelope forms on the polyhedron surface. The apparent affinity of the PE protein for the surface of maturing polyhedra suggests that it may be a major component of the polyhedron envelope or may form the matrix for the deposition of other components which contribute to the mature envelope. Immunogold staining and protease digestion experiments indicate that protein is an essential component of the polyhedron envelope.
Immunoperoxidase staining for identification of Aspergillus species in routinely processed tissue sections.

PubMed Central

Verweij, P E; Smedts, F; Poot, T; Bult, P; Hoogkamp-Korstanje, J A; Meis, J F

1996-01-01

AIMS: To evaluate the performance of an immunoperoxidase stain using the monoclonal antibody EB-A1 to detect Aspergillus species in formalin fixed, paraffin wax embedded tissue. METHODS: The monoclonal antibody EB-A1 directed against galactomannan was used to detect Aspergillus species in 23 patients with suspected or confirmed invasive aspergillosis. Immunostaining was performed on formalin fixed, paraffin wax embedded tissue using the streptavidin-biotin method and compared with conventional haematoxylin and eosin, periodic acid-Schiff, and Gomori-Grocott stains. Results of immunostaining were semiquantitatively analysed with regard to both intensity of staining and number of positively staining micro-organisms. Tissue sections from 16 patients with confirmed invasive mycoses due to Candida species, Apophysomyces elegans, Rhizopus oryzae, Pseudallescheria boydii and Histoplasma capsulatum were used as controls. RESULTS: In 19 (83%) of 23 cases invasive aspergillosis was confirmed by both histological examination and culture (18 Aspergillus fumigatus and one A flavus). Immunoperoxidase stains were positive in 17 (89%) of 19 cases including one case of disseminated infection due to A flavus. Furthermore, the immunoperoxidase stain was positive in a culture negative tissue section with histological evidence of mycelial development, indicating the presence of Aspergillus species. Some cross-reactivity was observed with the highly related fungus P boydii, although the number of mycelial elements that stained was low. CONCLUSIONS: Immunoperoxidase staining using the monoclonal antibody EB-A1 performs well on routinely processed tissue sections and permits detection and generic identification of Aspergillus species, although it was no better than conventional histopathology in identifying the presence of an infection. An additional advantage is that the immunostain may help to provide an aetiological diagnosis when cultures remain negative. Images PMID:8943743

An analysis of the sensitivity and specificity of MHC-I and MHC-II immunohistochemical staining in muscle biopsies for the diagnosis of inflammatory myopathies.

PubMed

Rodríguez Cruz, Pedro M; Luo, Yue-Bei; Miller, James; Junckerstorff, Reimar C; Mastaglia, Frank L; Fabian, Victoria

2014-12-01

Although there have been several previous reports of immunohistochemical staining for MHC antigens in muscle biopsies, there appears to be a lack of consensus about its routine use in the diagnostic evaluation of biopsies from patients with suspected inflammatory myopathy. Positive MHC-I staining is nonspecific but is widely used as a marker for inflammatory myopathy, whilst the role of MHC-II staining is not clearly defined. We investigated the sensitivity and specificity of MHC-I and MHC-II immunostaining for the diagnosis of inflammatory myopathy in a large group of biopsies from a single reference laboratory. Positive staining for MHC-I was found to have a high sensitivity in biopsies from patients with inflammatory myopathy but a very low specificity, as it was also common in other non-inflammatory myopathies and neurogenic disorders. On the other hand, MHC-II positivity had a much higher specificity in all major subgroups of inflammatory myopathy, especially inclusion body myositis. The findings indicate that the combination of MHC-I and MHC-II staining results in a higher degree of specificity for the diagnosis of inflammatory myopathy and that in biopsies with inflammation, positive MHC-II staining strongly supports the diagnosis of an immune-mediated myopathy. We recommend that immunohistochemical staining for both MHC-I and MHC-II should be included routinely in the diagnostic evaluation of muscle biopsies from patients with suspected inflammatory myopathy. However, as the sensitivity and interpretation of MHC staining may depend on the technique used, further studies are needed to compare procedures in different centres and develop standardised protocols. Copyright © 2014 Elsevier B.V. All rights reserved.
The comparison of pyrosequencing molecular Gram stain, culture, and conventional Gram stain for diagnosing orthopaedic infections.

PubMed

Kobayashi, Naomi; Bauer, Thomas W; Tuohy, Marion J; Lieberman, Isador H; Krebs, Viktor; Togawa, Daisuke; Fujishiro, Takaaki; Procop, Gary W

2006-08-01

We have developed a combined real-time PCR and pyrosequencing assay that successfully differentiated the vast majority of gram-positive and gram-negative bacteria when bacterial isolates were tested. The purpose of this study was to evaluate this assay on clinical specimens obtained from orthopedic surgeries, and to prospectively compare the results of "molecular Gram stain" with culture and conventional direct Gram stain. Forty-five surgical specimens were obtained from patients who underwent orthopedic surgery procedures. The DNA was extracted and a set of broad-range PCR primers that targeted a part of the 16S rDNA gene was used for pan-bacterial PCR. The amplicons were submitted for pyrosequencing and the resulting molecular Gram stain characteristics were recorded. Culture and direct Gram staining were performed using standard methods for all cases. Surgical specimens were reviewed histologically for all cases that had a discrepancy between culture and molecular results. There was an 86.7% (39/45) agreement between the traditional and molecular methods. In 12/14 (85.7%) culture-proven cases of bacterial infection, molecular Gram stain characteristics were in agreement with the culture results, while the conventional Gram stain result was in agreement only for five cases (35.7%). In the 31 culture negative cases, 27 cases were also PCR negative, whereas 4 were PCR positive. Three of these were characterized as gram negative and one as gram positive by this molecular method. Molecular determination of the Gram stain characteristics of bacteria that cause orthopedic infections may be achieved, in most instances, by this method. Further studies are necessary to understand the clinical importance of PCR-positive/culture-negative results.
Touch cytology in diagnosing Helicobacter pylori: comparison of four staining methods.

PubMed

Hashemi, M R; Rahnavardi, M; Bikdeli, B; Dehghani Zahedani, M; Iranmanesh, F

2008-06-01

Helicobacter pylori (Hp), a major cause of peptic ulcer disease and an important risk factor for gastric malignancy, can be diagnosed by several methods. Touch cytology (TC) of the gastric mucosa has been noted to give good results and has been found to be very simple, inexpensive and rapid. However, evidence regarding the accuracy of different staining methods of TC is lacking. The present study aims at defining the diagnostic accuracy of four different staining methods of TC. Biopsy specimens were taken from the antral mucosa of one hundred consecutive patients referred for upper gastrointestinal endoscopy (UGIE) for various indications. TC slides were processed by four staining methods: Wright, Giemsa, Papanicolaou and Gram. Rapid urease test (RUT) and histological examination of specimens were also performed. The same experienced pathologist evaluated the coded samples. A patient's Hp status was established by minimum concordance of the three tests, including histology, RUT, and 'Touch mean'. The latter was defined positive when at least three of the four TC staining methods were positive. Forty-six patients (46%) were positive for Hp according to Hp status. TC stained by Wright had excellent agreement with both histology (kappa = 0.80, P < 0.001) and RUT (kappa = 0.84, P < 0.001). Regarding Hp status, histology was 100% sensitive and RUT was 100% specific. Wright-stained TC (88.89%) was significantly more specific than both Giemsa- (74.07%; P < 0.05) and Papanicolaou-stained (70.37%; P < 0.05) TC. RUT should still be acknowledged as the primary test in diagnosing Hp following UGIE. If RUT is negative and Hp detection is intended only, Wright-stained TC can safely substitute for histology. However, when assessment for severity of mucosal damage or cell atypias is meant, histology cannot be neglected.
Interobserver reproducibility and accuracy of p16/Ki-67 dual-stain cytology in cervical cancer screening.

PubMed

Wentzensen, Nicolas; Fetterman, Barbara; Tokugawa, Diane; Schiffman, Mark; Castle, Philip E; Wood, Shannon N; Stiemerling, Eric; Poitras, Nancy; Lorey, Thomas; Kinney, Walter

2014-12-01

Dual-stain cytology for p16 and Ki-67 has been proposed as a biomarker in cervical cancer screening. The authors evaluated the reproducibility and accuracy of dual-stain cytology among 10 newly trained evaluators. In total, 480 p16/Ki-67-stained slides from human papillomavirus-positive women were evaluated in masked fashion by 10 evaluators. None of the evaluators had previous experience with p16 or p16/Ki-67 cytology. All participants underwent p16/Ki-67 training and subsequent proficiency testing. Reproducibility of dual-stain cytology was measured using the percentage agreement, individual and aggregate κ values, as well as McNemar statistics. Clinical performance for the detection of cervical intraepithelial neoplasia grade 2 or greater (CIN2+) was evaluated for each individual evaluator and for all evaluators combined compared with the reference evaluation by a cytotechnologist who had extensive experience with dual-stain cytology. The percentage agreement of individual evaluators with the reference evaluation ranged from 83% to 91%, and the κ values ranged from 0.65 to 0.81. The combined κ value was 0.71 for all evaluators and 0.73 for cytotechnologists. The average sensitivity and specificity for the detection of CIN2+ among novice evaluators was 82% and 64%, respectively; whereas the reference evaluation had 84% sensitivity and 63% specificity, respectively. Agreement on dual-stain positivity increased with greater numbers of p16/Ki-67-positive cells on the slides. Good to excellent reproducibility of p16/Ki-67 dual-stain cytology was observed with almost identical clinical performance of novice evaluators compared with reference evaluations. The current findings suggest that p16/Ki-67 dual-stain evaluation can be implemented in routine cytology practice with limited training. © 2014 American Cancer Society.
(68)Ga-DOTATATE PET in juvenile angiofibroma.

PubMed

Gronkiewicz, Zuzanna; Kukwa, Wojciech; Krolicki, Leszek; Cyran-Chlebicka, Agata; Pawlak, Dariusz; Stankiewicz, Czeslaw; Krzeski, Antoni; Górnicka, Barbara; Wolosz, Dominika; Kunikowska, Jolanta

2016-06-01

As somatostatin receptors (SSTRs) may be overexpressed in rapidly growing vessels, the aim of this study was the analysis of in vivo and in vitro SSTR2A expression in juvenile angiofibroma (JA). A group of six male adolescents with a diagnosis of primary, recurrent/residual JA was enrolled in the study. All patients underwent (68)Ga-DOTATATE PET/computed tomography (CT) followed by immunohistochemical staining for SSTR expression. (68)Ga-DOTATATE PET/CT showed accumulation in areas matching the pathologic tissue in the nasopharynx of all patients studied with SUVmax of 5.1 ± 0.9 (ranging from 3.6 to 6.4). In all cases, the immunohistochemical examination showed a presence of SSTR2A with a high staining index. In vitro SSTR2A cytoplasm expression was found to be high in all tumor specimens. However, the uptake of (68)Ga-DOTATATE was weak in the PET/CT studies. We postulate that the intracellular localization of the SSTR2A in JA may cause this discrepancy.
Immunohistochemical analysis of EGFR and hyaluronan in tongue cancer and the development of regional recurrence in patients initially diagnosed N0.

PubMed

Lindell Jonsson, Eva; Nylander, Karin; Hallén, Lars; Laurell, Göran

2017-08-01

To investigate whether the extent of expression of hyaluronan (HA) and epidermal growth factor receptor (EGFR) in squamous cell carcinoma of the mobile tongue can predict the risk of cervical metastasis and survival. Retrospective histopathologic study. Surgical specimens from 64 patients who had undergone surgery for squamous cell carcinoma of the mobile tongue were assessed using immunohistochemistry to investigate the expression of HA and EGFR in the primary tumours, and the data were then correlated to cervical metastasis and survival. There was a significant correlation between the intensity of HA staining and patient survival (p .024), and a weak correlation between the staining proportion of EGFR and the risk for regional recurrence (AUC 66). This study indicates that immunoscoring using HA could be used to provide prognostic tools for tongue cancer, and that it might be of interest to study the prognostic properties of EGFR further concerning the risk for regional recurrence after the primary treatment.
HISTOLOGIC, IMMUNOHISTOCHEMICAL, AND ELECTRON MICROSCOPIC CHARACTERIZATION OF A MALIGNANT IRIDOPHOROMA IN A DWARF BEARDED DRAGON (POGONA HENRYLAWSONI).

PubMed

de Brot, Simone; Sydler, Titus; Nufer, Lisbeth; Ruetten, Maja

2015-09-01

A dwarf bearded dragon (Pogona henrylawsoni) was presented with a white subcutaneous mandibular mass and multiple nodules in the oral mucosa, heart, liver, kidney, intestine, and visceral fat. Histologically, the tumor consisted of densely packed spindle-shaped cells with brow intracytoplasmic pigment that exhibited white-blue birefringence with polarized light. Immunohistochemical staining was negative for S-100 and weakly positive with melan A. Electron microscopic examination revealed cytoplasmic irregular and oblong empty spaces, laminated and often arranged into short stacks, compatible with reflecting platelet profiles typically seen in iridophores. However, in unstained ultrathin sections, electron-dense crystalline material was present, which filled the empty spaces described for stained sections before. Based on histology, immunohistochemistry, and biologic behavior, a malignant iridophoroma was diagnosed. To the authors' knowledge, iridophoromas in lizards have rarely been characterized by using electronic microscopy. Moreover, this is the first description of an iridophoroma in a dwarf bearded dragon.
Unusual Differentiation to Pheochromocytoma-Like Cells in an Adrenal Neuroblastoma After Chemotherapy: A Case Report and Literature Review.

PubMed

Tatekawa, Yukihiro

2015-01-01

The authors present a case of 3-year-old female with Stage 4 neuroblastoma originating from the left adrenal gland. Biopsy of the left adrenal tumor showed neuroblastoma. After three courses of chemotherapy, the left adrenal gland including the left adrenal tumor was surgically removed. Pathological findings of the resected tumor revealed that most of the neuroblastoma tissues changed to pheochromocytoma-like cells. The tumor cells were arranged in well-defined nests surrounded by a delicate fibrovascular stroma and had granular eosinophilic cytoplasm, and round to oval nuclei. Immunohistological analysis of the biopsy samples showed strongly positive Ganglioside GD2-staining cells, whereas almost all of the tumor cells in the resected specimen were Ganglioside GD2-negative; cells were very weakly stained. The authors suggest that a part of the neuroblastoma in the left adrenal gland exhibited unusual differentiation toward pheochromocytic lineage Ganglioside GD2-negative neuroblastoma in a patient who had been treated with intensive chemotherapy.
Legionella micdadei prosthetic valve endocarditis complicated by brain abscess: case report and review of the literature.

PubMed

Fukuta, Yuriko; Yildiz-Aktas, Isil Z; William Pasculle, A; Veldkamp, Peter J

2012-06-01

Legionella endocarditis is extremely uncommon, and embolic phenomena have never been reported. We report the first case of Legionella micdadei prosthetic valve endocarditis complicated by brain abscess. A 57-y-old immunocompromised woman with a history of mitral valve replacement developed confusion and left-sided weakness. Brain magnetic resonance imaging showed a 3-cm peripheral-enhancing mass. Transoesophageal echocardiography suggested a perivalvular abscess. Blood cultures and valve cultures were negative. She was diagnosed with 16S rRNA polymerase chain reaction and silver stain, and was discharged with levofloxacin after a redo mitral valve replacement. Twelve cases of Legionella endocarditis were reviewed. Only one case had a native valve, and her endocarditis occurred after pneumonia. All cases were cured. The duration of antibiotic therapy was variable. Legionella species should be considered in the differential diagnosis of culture-negative endocarditis in both immunocompetent and immunocompromised patients. Molecular techniques and silver impregnation stains are useful, especially when cultures using buffered charcoal-yeast extract agar are negative.
Thrombospondin in colorectal disease.

PubMed

Behzad; Abdalla; Gardy; Battrick; Kumar; Haboubi

2000-05-01

To assess the presence of thrombospondin (TSP) in normal and colonic disease and correlate its presence to survival in colorectal cancers. A specific antibody was used to stain paraffin-embedded sections of human colorectal disease (103 carcinomas, 10 inflammatory bowel disease (IBD), 10 diverticulosis and 10 normal). The stained sections were viewed by light microscopy and the degree of staining was quantified using computer-assisted image analysis system. Seventy-three percent of carcinomas, 30% of IBD and 80% of the diverticulosis specimens stained for TSP. None of the 'normal' specimens examined showed any staining. No significant correlation could be detected between TSP expression and survival in colorectal cancer patients. The presence of TSP in the colon is not specific and merely indicates the presence of a disease process.
One Small Step for the Gram Stain, One Giant Leap for Clinical Microbiology.

PubMed

Thomson, Richard B

2016-06-01

The Gram stain is one of the most commonly performed tests in the clinical microbiology laboratory, yet it is poorly controlled and lacks standardization. It was once the best rapid test in microbiology, but it is no longer trusted by many clinicians. The publication by Samuel et al. (J. Clin. Microbiol. 54:1442-1447, 2016, http://dx.doi.org/10.1128/JCM.03066-15) is a start for those who want to evaluate and improve Gram stain performance. In an age of emerging rapid molecular results, is the Gram stain still relevant? How should clinical microbiologists respond to the call to reduce Gram stain error rates? Copyright © 2016, American Society for Microbiology. All Rights Reserved.
Curcuma longa extract as a histological dye for collagen fibres and red blood cells

PubMed Central

Avwioro, O G; Onwuka, S K; Moody, J O; Agbedahunsi, J M; Oduola, T; Ekpo, O E; Oladele, A A

2007-01-01

Crude ethanolic extract and column chromatographic fractions of the Allepey cultivar of Curcuma longa Roxb, commonly called turmeric (tumeric) in commerce, were used as a stain for tissue sections. Staining was carried out under basic, acidic and neutral media conditions. Inorganic and organic dissolution solvents were used. The stain was used as a counterstain after alum and iron haematoxylins. C. longa stained collagen fibres, cytoplasm, red blood cells and muscle cells yellow. It also stained in a fashion similar to eosin, except for its intense yellow colour. Preliminary phytochemical evaluation of the active column fraction revealed that it contained flavonoids, free anthraquinone and deoxy sugar. A cheap, natural dye can thus be obtained from C. longa. PMID:17451535
Rorschach inkblot port-wine stain.

PubMed

Coots, N V; Elston, D M

1997-01-01

We present an infant born with bilaterally symmetric, anterior and posterior port-wine stains. These lesions presented a striking resemblance to Rorschach inkblots, a phenomenon not previously reported. A discussion of the case as well as a discussion of syndromes associated with port-wine stains is provided.
42 CFR 493.911 - Bacteriology.

Code of Federal Regulations, 2012 CFR

2012-10-01

... laboratories: (1) Those that interpret Gram stains or perform primary inoculation, or both; and refer cultures...; (2) Those that use direct antigen techniques to detect an organism and may also interpret Gram stains... interpreting Gram stains, performing primary inoculations, and using direct antigen tests, also isolate and...
42 CFR 493.911 - Bacteriology.

Code of Federal Regulations, 2014 CFR

2014-10-01

... laboratories: (1) Those that interpret Gram stains or perform primary inoculation, or both; and refer cultures...; (2) Those that use direct antigen techniques to detect an organism and may also interpret Gram stains... interpreting Gram stains, performing primary inoculations, and using direct antigen tests, also isolate and...
42 CFR 493.911 - Bacteriology.

Code of Federal Regulations, 2013 CFR

2013-10-01

... laboratories: (1) Those that interpret Gram stains or perform primary inoculation, or both; and refer cultures...; (2) Those that use direct antigen techniques to detect an organism and may also interpret Gram stains... interpreting Gram stains, performing primary inoculations, and using direct antigen tests, also isolate and...
Ocular melanoma metastatic to skin: the value of HMB-45 staining.

PubMed

Schwartz, Robert A; Kist, Joseph M; Thomas, Isabelle; Fernández, Geover; Cruz, Manuel A; Koziorynska, Ewa I; Lambert, W Clark

2004-06-01

Cutaneous metastatic disease is an important finding that may represent the first sign of systemic cancer, or, if already known, that may change tumor staging and thus dramatically altered therapeutic plans. Although cutaneous metastases are relatively frequent in patients with cutaneous melanoma, they are less so from ocular melanoma. To demonstrate the value of HMB-45, staining in the detection of ocular melanoma metastatic to skin. The immunohistochemical stain HMB-45 a monoclonal antibody directed against intact human melanoma cells, was employed on a skin biopsy specimen from a cutaneous tumor. HMB-45 staining was positive in the atypical hyperchromatic cells of the deep dermis. HMB-45 may be of value in the detection of ocular melanoma metastatic to skin. Cutaneous metastatic disease is a somewhat common and extremely important diagnosis. Although cutaneous metastases from cutaneous melanoma are relatively frequent, those from ocular melanomas are less so. Use of histochemical staining, especially the HMB-45 stain, allows confirmation of the diagnosis.
Comparison of culture-based, vital stain and PMA-qPCR methods for the quantitative detection of viable hookworm ova.

PubMed

Gyawali, P; Sidhu, J P S; Ahmed, W; Jagals, P; Toze, S

2017-06-01

Accurate quantitative measurement of viable hookworm ova from environmental samples is the key to controlling hookworm re-infections in the endemic regions. In this study, the accuracy of three quantitative detection methods [culture-based, vital stain and propidium monoazide-quantitative polymerase chain reaction (PMA-qPCR)] was evaluated by enumerating 1,000 ± 50 Ancylostoma caninum ova in the laboratory. The culture-based method was able to quantify an average of 397 ± 59 viable hookworm ova. Similarly, vital stain and PMA-qPCR methods quantified 644 ± 87 and 587 ± 91 viable ova, respectively. The numbers of viable ova estimated by the culture-based method were significantly (P < 0.05) lower than vital stain and PMA-qPCR methods. Therefore, both PMA-qPCR and vital stain methods appear to be suitable for the quantitative detection of viable hookworm ova. However, PMA-qPCR would be preferable over the vital stain method in scenarios where ova speciation is needed.
Multiplication free neural network for cancer stem cell detection in H-and-E stained liver images

NASA Astrophysics Data System (ADS)

Badawi, Diaa; Akhan, Ece; Mallah, Ma'en; Üner, Ayşegül; ćetin-Atalay, Rengül; ćetin, A. Enis

2017-05-01

Markers such as CD13 and CD133 have been used to identify Cancer Stem Cells (CSC) in various tissue images. It is highly likely that CSC nuclei appear as brown in CD13 stained liver tissue images. We observe that there is a high correlation between the ratio of brown to blue colored nuclei in CD13 images and the ratio between the dark blue to blue colored nuclei in H&E stained liver images. Therefore, we recommend that a pathologist observing many dark blue nuclei in an H&E stained tissue image may also order CD13 staining to estimate the CSC ratio. In this paper, we describe a computer vision method based on a neural network estimating the ratio of dark blue to blue colored nuclei in an H&E stained liver tissue image. The neural network structure is based on a multiplication free operator using only additions and sign operations. Experimental results are presented.
Evaluation of gram stain as an alternative in the assessment of human spermatozoa quality.

PubMed

Mantas, D; Msaouel, P; Angelopoulou, R

2006-01-01

During spermiogenesis, protaminosis and sperm chromatin condensation are important prerequisites for the preservation of DNA integrity in spermatozoa. The aim of this study is to assess Gram stain as an alternative technique for the evaluation of human sperm chromatin condensation status. Aniline blue and Gram staining were applied to semen samples from 34 donors in order to determine the relationship between sperm chromatin condensation and infertility. In addition, the possible correlation between morphology and vitality (eosin-Y staining) of spermatozoa compared with their nuclear status (aniline blue and Gram staining) was studied. Chromatin condensation and sperm vitality were significantly higher in fertile men compared to the subfertile. A significant correlation was found between chromatin condensation and (a) sperm vitality (p < 0.01), and (b) nuclear protein status (p < 0.01). Gram staining may be used as a routine method in assisted reproduction laboratories and could assist in the evaluation of sperm quality as well as in the selection of the appropriate fertilization technique.

[Comparison of the quick Gram stain method to the B&M modified and favor methods].

PubMed

Osawa, Kayo; Kataoka, Nobumasa; Maruo, Toshio

2011-01-01

The Gram stain is an established method for bacterial identification, but the time needed to carry out this stain is 2-3 min. We attempted to shorten this time and stained a total of 70 clinical specimens isolated from using the Bartholomew & Mittwer (B&M) modified or Favor methods with a 3 s duration for washing and staining steps. Results were plotted and analyzed using a Hue Saturation Intensity (HSI) model. The range based on a plot of the two methods with the HSI model was presented as a reference interval. Our results indicated that 100% (35/35) of strains were Gram positive and 97.1% (34/35) were Gram negative for the quick B&M modified method. In the quick Favor method, 80.0% (28/35) were Gram positive and 68.6% (24/35) of strains were Gram negative. We propose that the quick B&M modified method is equivalent to the standard Gram staining method and is superior to the quick Favor method.
Effects of Microgravity on Embryonic Quail Eye Development

NASA Technical Reports Server (NTRS)

Barrett, Joyce E.; Wells, Diane C.; Paulsen, Avelina Q.; Conrad, Gary W.

1997-01-01

Immunohistochemical methods were used to stain neurofilament protein in corneal nerves of Embryonic Day 16 (E16) quail eyes that had been fixed in 4% paraformaldehyde at room temperature for several months. Fixation was according to the methods used by the Mir 21/NASA 2 Avian Developmental Biology Flight Experiments for quail embryos incubated on the Mir Space Station. After fixation, corneas were pretreated to improve immunohistochemical visualization of neurofilaments. A sequential combination of three pretreatments [microwave heating in saline G, followed by extraction with sodium dodecyl sulfate (SDS) at 37 C, followed by digestion with hyaluronidase at 37 C], produced increased antibody staining of corneal nerve neurofilament proteins, compared with corneas subjected to no prior pretreatments. Darker nerve staining and increased numbers of fine branches were observed, together with lower background staining after such pretreatments. In contrast, use of any single pretreatment or pair of pretreatments resulted in only slight and inconsistent enhancement of nerve staining. Only the sequential combination of all three pretreatments resulted in consistently better nerve staining.
A new fluorescent test for cell vitality using calcofluor white M2R.

PubMed

Fischer, J M; Peterson, C A; Bols, N C

1985-03-01

The fluorescent fabric-brightener dye, Calcofluor white M2R (CFW), can be used to distinguish between living and dead cells from a variety of animal and plant sources. CFW does not stain living mouse fibroblasts or trout red blood cells and stains only the cell walls in living cells from the epidermis of onion bulb scale, staminal hairs of Tradescantia, and longitudinal sections of broad bean stems and roots. Heat-killed plant or animal cells are recognized by their lightly stained cytoplasm and brightly stained nuclei. The optimum staining concentrations were very low (0.01% to 0.03%) and nontoxic. Using onion scale epidermis in which some cells had been killed by heating as a test system, and the plasmolysis-deplasmolysis rection as the ultimate test for cell vitality, results from CFW staining correctly predicted cell vitality for about 98% of the cells tested. This success rate was comparable to those for Evans blue, uranin or neutral red in this test system.
Determination of Protein Expression Level in Cultured Cells by Immunocytochemistry on Paraffin-embedded Cell Blocks.

PubMed

Poojan, Shiv; Kim, Han-Seong; Yoon, Ji-Woon; Sim, Hye Won; Hong, Kyeong-Man

2018-05-20

Immunofluorescent staining is currently the method of choice for determination of protein expression levels in cell-culture systems when morphological information is also necessary. The protocol of immunocytochemical staining on paraffin-embedded cell blocks, presented herein, is an excellent alternative to immunofluorescent staining on non-paraffin-embedded fixed cells. In this protocol, a paraffin cell block from HeLa cells was prepared using the thromboplastin-plasma method, and immunocytochemistry was performed for the evaluation of two proliferation markers, CKAP2 and Ki-67. The nuclei and cytoplasmic morphology of the HeLa cells were well preserved in the cell-block slides. At the same time, the CKAP2 and Ki-67 staining patterns in the immunocytochemistry were quite similar to those in immunohistochemical staining in paraffin cancer tissues. With modified cell-culture conditions, including pre-incubation of HeLa cells under serum-free conditions, the effect could be evaluated while preserving architectural information. In conclusion, immunocytochemistry on paraffin-embedded cell blocks is an excellent alternative to immunofluorescent staining.
Osteogenic capability of autologous rabbit adipose-derived stromal cells in repairing calvarial defects.

PubMed

Cheng, Shao-Wen; Lin, Zhong-Qin; Wang, Wei; Zhang, Wei; Kou, Dong-Quan; Ying, Xiao-Zhou; Chen, Qing-Yu; Shen, Yue; Cheng, Xiao-Jie; Peng, Lei; Lv, Chuan-Zhu

2011-01-01

To evaluate the in vitro and in vivo osteogenic capability of adipose-derived stromal cells (ASCs). ASCs were isolated from New Zealand white rabbits and determined by alkaline phosphatase (ALP) staining, von Kossa staining and alizarin red staining. Some specific markers of osteogenic differentiation, including ALP, osteocalcin (OCN), osteopontin (OPN) were examined by reverse transcription-polymerase chain reaction (RT-PCR). In vivo, demineralized bone matrix (DBM)-ASCs composites were implanted into the rabbit calvarial defects created at each side of the longitudinal midline. After 6 weeks, histologic properties of the transplants were analyzed. ASCs were successfully induced into osteogenesis. ALP staining, von Kossa staining and alizarin red staining showed positive results. The expressions of ALP, OCN and OPN were detected in ASCs after cultivation in osteogenic medium. Extensive new bone was observed in the defects transplanted with DBM-ASCs composites. ASCs have the potential to differentiate into osteogenic lineage and DBM-ASCs constructs are a promising method for regeneration in bone defects.
Automated robust registration of grossly misregistered whole-slide images with varying stains

NASA Astrophysics Data System (ADS)

Litjens, G.; Safferling, K.; Grabe, N.

2016-03-01

Cancer diagnosis and pharmaceutical research increasingly depend on the accurate quantification of cancer biomarkers. Identification of biomarkers is usually performed through immunohistochemical staining of cancer sections on glass slides. However, combination of multiple biomarkers from a wide variety of immunohistochemically stained slides is a tedious process in traditional histopathology due to the switching of glass slides and re-identification of regions of interest by pathologists. Digital pathology now allows us to apply image registration algorithms to digitized whole-slides to align the differing immunohistochemical stains automatically. However, registration algorithms need to be robust to changes in color due to differing stains and severe changes in tissue content between slides. In this work we developed a robust registration methodology to allow for fast coarse alignment of multiple immunohistochemical stains to the base hematyoxylin and eosin stained image. We applied HSD color model conversion to obtain a less stain color dependent representation of the whole-slide images. Subsequently, optical density thresholding and connected component analysis were used to identify the relevant regions for registration. Template matching using normalized mutual information was applied to provide initial translation and rotation parameters, after which a cost function-driven affine registration was performed. The algorithm was validated using 40 slides from 10 prostate cancer patients, with landmark registration error as a metric. Median landmark registration error was around 180 microns, which indicates performance is adequate for practical application. None of the registrations failed, indicating the robustness of the algorithm.
Gram Stains: A Resource for Retrospective Analysis of Bacterial Pathogens in Clinical Studies

PubMed Central

Srinivasan, Usha; Ponnaluri, Sreelatha; Villareal, Lisa; Gillespie, Brenda; Wen, Ai; Miles, Arianna; Bucholz, Brigette; Marrs, Carl F.; Iyer, Ram K.; Misra, Dawn; Foxman, Betsy

2012-01-01

We demonstrate the feasibility of using qPCR on DNA extracted from vaginal Gram stain slides to estimate the presence and relative abundance of specific bacterial pathogens. We first tested Gram stained slides spiked with a mix of 108 cfu/ml of Escherichia coli and 105 cfu/ml of Lactobacillus acidophilus. Primers were designed for amplification of total and species-specific bacterial DNA based on 16S ribosomal gene regions. Sample DNA was pre-amplified with nearly full length 16S rDNA ribosomal gene fragment, followed by quantitative PCR with genera and species-specific 16S rDNA primers. Pre-amplification PCR increased the bacterial amounts; relative proportions of Escherichia coli and Lactobacillus recovered from spiked slides remained unchanged. We applied this method to forty two archived Gram stained slides available from a clinical trial of cerclage in pregnant women at high risk of preterm birth. We found a high correlation between Nugent scores based on bacterial morphology of Lactobacillus, Gardenerella and Mobiluncus and amounts of quantitative PCR estimated genus specific DNA (rrn copies) from Gram stained slides. Testing of a convenience sample of eight paired vaginal swabs and Gram stains freshly collected from healthy women found similar qPCR generated estimates of Lactobacillus proportions from Gram stained slides and vaginal swabs. Archived Gram stained slides collected from large scale epidemiologic and clinical studies represent a valuable, untapped resource for research on the composition of bacterial communities that colonize human mucosal surfaces. PMID:23071487
Gram and acridine orange staining for diagnosis of septic arthritis in different patient populations.

PubMed

Cunningham, Gregory; Seghrouchni, Khalid; Ruffieux, Etienne; Vaudaux, Pierre; Gayet-Ageron, Angèle; Cherkaoui, Abdessalam; Godinho, Eduardo; Lew, Daniel; Hoffmeyer, Pierre; Uçkay, Ilker

2014-06-01

The sensitivity of Gram staining is known to be suboptimal for the diagnosis of native joint septic arthritis. We lack information about the accuracy of Gram compared to other microscopic staining techniques for predicting infection in different patient populations. This was a cohort study with cost evaluations at the Orthopaedic Service of Geneva University Hospitals (January 1996-October 2012). Among 500 episodes of arthritis (196 with immunosuppression, 227 with underlying arthroplasties and 69 with gout or other crystals in synovial fluid), Gram staining revealed pathogens in 146 episodes (146/500, 29 %) or in 146 of the 400 culture-positive episodes (37 %). Correlation between the Gram and acridine staining of the same sample was good (Spearman 0.85). Overall, the sensitivity, specificity, positive predictive value and negative predictive value of Gram stain for rapid diagnosis of septic arthritis was 0.37, 0.99, 0.99 and 0.28, respectively, compared to microbiological cultures. Quite similar values were recorded across the different patient subpopulations, in particular for sensitivity values that were 0.33 for patients with prosthetic joint infections, 0.40 for immunosuppressed patients, 0.36 for patients under antibiotic administration and 0.52 for patients with concomitant crystalline disease. The sensitivity of Gram or acridine orange staining for a rapid diagnosis of episodes of septic arthritis is suboptimal compared to microbiological culture, regardless of underlying conditions, immunosuppression or antibiotic therapy. The sensitivity in the presence of synovial fluid crystals is moderate. Acridine orange and Gram stains are equivalent.
A Flow-Cytometric Gram-Staining Technique for Milk-Associated Bacteria

PubMed Central

Holm, Claus; Jespersen, Lene

2003-01-01

A Gram-staining technique combining staining with two fluorescent stains, Oregon Green-conjugated wheat germ agglutinin (WGA) and hexidium iodide (HI) followed by flow-cytometric detection is described. WGA stains gram-positive bacteria while HI binds to the DNA of all bacteria after permeabilization by EDTA and incubation at 50°C for 15 min. For WGA to bind to gram-positive bacteria, a 3 M potassium chloride solution was found to give the highest fluorescence intensity. A total of 12 strains representing some of the predominant bacterial species in bulk tank milk and mixtures of these were stained and analyzed by flow cytometry. Overall, the staining method showed a clear differentiation between gram-positive and gram-negative bacterial populations. For stationary-stage cultures of seven gram-positive bacteria and five gram-negative bacteria, an average of 99% of the cells were correctly interpreted. The method was only slightly influenced by the growth phase of the bacteria or conditions such as freezing at −18°C for 24 h. For any of these conditions, an average of at least 95% of the cells were correctly interpreted. When stationary-stage cultures were stored at 5°C for 14 days, an average of 86% of the cells were correctly interpreted. The Gram-staining technique was applied to the flow cytometry analysis of bulk tank milk inoculated with Staphylococcus aureus and Escherichia coli. These results demonstrate that the technique is suitable for analyzing milk samples without precultivation. PMID:12732558
Gram stains: a resource for retrospective analysis of bacterial pathogens in clinical studies.

PubMed

Srinivasan, Usha; Ponnaluri, Sreelatha; Villareal, Lisa; Gillespie, Brenda; Wen, Ai; Miles, Arianna; Bucholz, Brigette; Marrs, Carl F; Iyer, Ram K; Misra, Dawn; Foxman, Betsy

2012-01-01

We demonstrate the feasibility of using qPCR on DNA extracted from vaginal Gram stain slides to estimate the presence and relative abundance of specific bacterial pathogens. We first tested Gram stained slides spiked with a mix of 10(8) cfu/ml of Escherichia coli and 10(5) cfu/ml of Lactobacillus acidophilus. Primers were designed for amplification of total and species-specific bacterial DNA based on 16S ribosomal gene regions. Sample DNA was pre-amplified with nearly full length 16S rDNA ribosomal gene fragment, followed by quantitative PCR with genera and species-specific 16S rDNA primers. Pre-amplification PCR increased the bacterial amounts; relative proportions of Escherichia coli and Lactobacillus recovered from spiked slides remained unchanged. We applied this method to forty two archived Gram stained slides available from a clinical trial of cerclage in pregnant women at high risk of preterm birth. We found a high correlation between Nugent scores based on bacterial morphology of Lactobacillus, Gardenerella and Mobiluncus and amounts of quantitative PCR estimated genus specific DNA (rrn copies) from Gram stained slides. Testing of a convenience sample of eight paired vaginal swabs and Gram stains freshly collected from healthy women found similar qPCR generated estimates of Lactobacillus proportions from Gram stained slides and vaginal swabs. Archived Gram stained slides collected from large scale epidemiologic and clinical studies represent a valuable, untapped resource for research on the composition of bacterial communities that colonize human mucosal surfaces.
Effect of Photo-Fenton Bleaching on Tetracycline-stained Dentin in vitro.

PubMed

Bennett, Zackary Yale; Walsh, Laurence James

2015-02-01

Tetracycline-stained tooth structure is difficult to bleach using nightguard tray methods. The possible benefits of in-office light-accelerated bleaching systems based on the photo-Fenton reaction are of interest as possible adjunctive treatments. This study was a proof of concept for possible benefits of this approach, using dentine slabs from human tooth roots stained in a reproducible manner with the tetracycline antibiotic demeclocycline hydrochloride. Color changes overtime in tetra-cycline stained roots from single rooted teeth treated using gel (Zoom! WhiteSpeed(®)) alone, blue LED light alone, or gel plus light in combination were tracked using standardized digital photography. Controls received no treatment. Changes in color channel data were tracked overtime, for each treatment group (N = 20 per group). Dentin was lighter after bleaching, with significant improvements in the dentin color for the blue channel (yellow shade) followed by the green channel and luminosity. The greatest changes occurred with gel activated by light (p < 0.0001), which was superior to effects seen with gel alone. Use of the light alone did not significantly alter shade. This proof of concept study demonstrates that bleaching using the photo-Fenton chemistry is capable of lightening tetracycline-stained dentine. Further investigation of the use of this method for treating tetracycline-stained teeth in clinical settings appears warranted. Because tetracycline staining may respond to bleaching treatments based on the photo-Fenton reaction, systems, such as Zoom! WhiteSpeed, may have benefits as adjuncts to home bleaching for patients with tetracycline-staining.
A flow-cytometric gram-staining technique for milk-associated bacteria.

PubMed

Holm, Claus; Jespersen, Lene

2003-05-01

A Gram-staining technique combining staining with two fluorescent stains, Oregon Green-conjugated wheat germ agglutinin (WGA) and hexidium iodide (HI) followed by flow-cytometric detection is described. WGA stains gram-positive bacteria while HI binds to the DNA of all bacteria after permeabilization by EDTA and incubation at 50 degrees C for 15 min. For WGA to bind to gram-positive bacteria, a 3 M potassium chloride solution was found to give the highest fluorescence intensity. A total of 12 strains representing some of the predominant bacterial species in bulk tank milk and mixtures of these were stained and analyzed by flow cytometry. Overall, the staining method showed a clear differentiation between gram-positive and gram-negative bacterial populations. For stationary-stage cultures of seven gram-positive bacteria and five gram-negative bacteria, an average of 99% of the cells were correctly interpreted. The method was only slightly influenced by the growth phase of the bacteria or conditions such as freezing at -18 degrees C for 24 h. For any of these conditions, an average of at least 95% of the cells were correctly interpreted. When stationary-stage cultures were stored at 5 degrees C for 14 days, an average of 86% of the cells were correctly interpreted. The Gram-staining technique was applied to the flow cytometry analysis of bulk tank milk inoculated with Staphylococcus aureus and Escherichia coli. These results demonstrate that the technique is suitable for analyzing milk samples without precultivation.
Detection of alkaline phosphatase in canine cells previously stained with Wright-Giemsa and its utility in differentiating osteosarcoma from other mesenchymal tumors.

PubMed

Ryseff, Julia K; Bohn, Andrea A

2012-09-01

Osteosarcoma (OSA) is a common primary bone tumor in dogs. Demonstration of alkaline phosphatase (ALP) reactivity by tumor cells on unstained slides is useful in differentiating osteosarcoma from other types of sarcoma. However, unstained slides are not always available. The objectives of this study were to evaluate the diagnostic utility of detecting ALP expression in differentiating osteosarcoma from other sarcomas in dogs using cytologic material previously stained with Wright-Giemsa stain and to assess the sensitivity and specificity of ALP expression for diagnosing osteosarcoma using a specific protocol. Archived aspirates of histologically confirmed sarcomas in dogs that had been previously stained with Wright-Giemsa stain were treated with 5-bromo, 4-chloro, 3-indolyl phosphate/nitroblue tetrazolium (BCIP/NBT) as a substrate for ALP. Cells were evaluated for expression of ALP after incubation with BCIP/NBT for 1 hour. Sensitivity and specificity of ALP expression for diagnosis of OSA were calculated. In samples from 83 dogs, cells from 15/17 OSAs and from 4/66 tumors other than OSA (amelanotic melanoma, gastrointestinal stromal tumor, collision tumor, and anaplastic sarcoma) expressed ALP. Sensitivity and specificity of ALP expression detected using BCIP/NBT substrate applied to cells previously stained with Wright-Giemsa stain for OSA were 88 and 94%, respectively. ALP expression detected using BCIP/NBT substrate applied to previously stained cells is useful in differentiating canine OSA from other mesenchymal neoplasms. © 2012 American Society for Veterinary Clinical Pathology.
Phenol Oxidase Mediated Protein Cross-Linking

DTIC Science & Technology

1989-06-26

proteins and Fasciola eggshell precursor proteins have their tyrosine residues oxidized to DOPA prior to secretion. We have been trying to determine...reproducibly with "Arnow’s Stain" although Fasciola DOPA protein kindly supplied by Herb Waite stains reproducibly in our hands. NOVEL SILVER STAINING OF
Gestation-related gene expression and protein localization in endometrial tissue of Suffolk and Cheviot ewes at gestation Day 19, after transfer of Suffolk or Cheviot embryos.

PubMed

Sequeira, M; Pain, S J; de Brun, V; Meikle, A; Kenyon, P R; Blair, H T

2016-10-01

The objective of this study was to investigate the gene expression of progesterone and estrogen receptor α (PR, ERα), insulin-like growth factor (IGF) 1, IGF-2, their receptor (IGFR1), IGF-binding proteins (BP) 1 to 6, insulin receptor, adiponectin receptors (AdipoR1/2), cyclooxygenase 2 (PTGS2), mucin 1 and to localize PR, ERα, IGF-1, IGFR1, PTGS2, and proliferating cellular nuclear antigen (PCNA) in the endometrium of pregnant (Day 19) Suffolk and Cheviot ewes carrying Suffolk and Cheviot embryos transferred within and reciprocally between breeds. Gene expression was determined by real-time quantitative polymerase chain reaction (RT-qPCR), and antigen determination was measured by immunohistochemistry in the luminal epithelium (LE), superficial and deep glands (SG, DG, respectively) and superficial and deep stroma. Gene expression of PR, IGF-1, IGFBP2, and IGFBP5 was higher in Suffolk than that in Cheviot ewes (P < 0.05). Greater abundance of IGF-2 and IGBP3 expression was found in Cheviot ewes carrying Cheviot embryos than Cheviot ewes carrying Suffolk embryos (P < 0.05). No staining for PR and ERα was observed in the LE, very scarce staining in SG and DG, whereas positive staining was observed in both superficial and deep stroma. No differences were found for PR staining, but Cheviot ewes had higher ERα staining intensity than Suffolk ewes (P < 0.05). Positive staining for IGF-1 was observed in all cell types except DG, and staining of IGFR1 was observed in all cell types. No differences among groups in staining were found for IGF-1 or IGFR1 in any cell type. Positive staining of PTGS2 was observed in LE and SG in all groups. An interaction between ewe and embryo breed affected PTGS2 staining (P < 0.05), whereby Cheviot ewes carrying Suffolk embryos had a lower PTGS2 staining than Suffolk ewes carrying Suffolk embryos. Positive staining of PCNA was found in LE and SG. Suffolk ewes carrying Suffolk embryos showed lower PCNA immunostaining than Cheviot ewes carrying Suffolk embryos (P < 0.05), whereas no differences were observed in ewes carrying Cheviot embryos. This study showed that gestation-related protein expression in the endometrium of Suffolk and Cheviot ewes is affected by both ewe and embryo breed at Day 19 of pregnancy. Copyright © 2016 Elsevier Inc. All rights reserved.
[Histological structure of the trabecular meshwork in the eyeball: challenging the traditional concept and preliminary findings in rabbits, rats and mice].

PubMed

Shi, Yun; Zhou, Fan-Qi; Luo, Zhou-Cai; Chen, Ying-Hua; Chen, Yu; Dong, Wei-Ren

2017-10-20

To verify that the trabecular meshwork (TM) in the wall of the eyeball consists of smooth muscle fibers instead of collagen fibers or endothelial cells. Eighteen fresh eyeballs from 3 rabbits, 3 SD rats and 3 mice were sectioned along the sagittal plane and sliced after paraffin embedding for HE staining, VG staining, Masson staining, α-SMA immunohistochemistry or CD31 immunohistochemistry. These slices were observed under microscope and the structure of the TM was compared with those of scleral collagen fibers, ciliary muscles and endothelial cells. HE staining of the eyeball slices from the 3 animal species resulted in purplish red staining of the TM, which was highly consistent with ciliary muscle fibers. The cell?like structures on the surface of the TM were not clearly outlined, with flat nuclei showing a dark purple staining; these structures did not show obvious boundaries from the TM. Ciliary muscle fibers, which were smooth muscle cells in nature, were aligned in bundles in various directions. The longitudinally sectioned cells were flat and contained purplish cytoplasm and highly flattened nuclei. Scleral collagen fibers were stained dark red with a few fibroblasts sandwiched among them. The long axis of the fibroblasts was in parallel with that of the collagen fibers. The outline of the fibroblast was not clear and the nucleus was flat in dark blue. The vascular endothelial cells presented with different morphologies and contained light purplish cytoplasm and dark nuclei, protruding into the vascular cavity. VG staining of the TM revealed a pale red filamentous structure, and the collagen fibers were stained bright red. Masson staining of the TM showed a reticular structure consisting mainly of dark red fibers intermingled with thin green fibers. Scleral collagen fibers presented with a cord?like green wavy structure. The endothelial cells were green and flat, while the ciliary smooth muscle fibers were purple. In immunohistochemistry for α?SMA, the TM and the ciliary smooth muscle fibers showed a strong positivity in the cytoplasm, while the scleral collagen fibers and vascular endothelial cells showed negative staining; immunohistochemistry for CD31 showed no obvious positive staining in the TM, collagen fibers or ciliary smooth muscle cells from all the animals in spite of slight differences among them. The TM consists mainly of smooth muscle fibers with a thin layer of peripheral endomysium without endothelial cells.
Immuno-Oncology Biomarkers for Gastric and Gastroesophageal Junction Adenocarcinoma: Why PD-L1 Testing May Not Be Enough.

PubMed

Weinberg, Benjamin A; Xiu, Joanne; Hwang, Jimmy J; Shields, Anthony F; Salem, Mohamed E; Marshall, John L

2018-04-27

The treatment of patients with advanced gastric and gastroesophageal junction (G/GEJ) adenocarcinomas has been transformed by the U.S. Food and Drug Administration approval of pembrolizumab. Tumor and adjacent tissue must stain positively for the programmed cell death ligand 1 (PD-L1) protein by companion diagnostic testing. However, some patients with PD-L1-negative tumors also benefit from pembrolizumab. High microsatellite instability (MSI) and tumor mutational load (TML) are positive predictive biomarkers for immune checkpoint inhibition (ICI) in other tumors. We sought to identify more patients who could benefit from ICI using alternative PD-L1 thresholds, MSI, and TML. Tumor specimens underwent next-generation sequencing (NGS) and PD-L1 testing using immunohistochemistry. NGS was used to determine TML and MSI. We profiled 581 G/GEJ adenocarcinoma specimens. PD-L1 staining was scored for intensity (0, none; 1+, weak; 2+, moderate; 3+, strong). Using 2+ staining at a 5% threshold, 9.3% of tumors were PD-L1 positive, and using 1+ staining at 1%, 16.2% were PD-L1 positive. 6.9% of tumors had high MSI. High TML (≥17 mutations per megabase) was seen in 6.9%, and medium TML (≥7) was seen in 56.5% of tumors. Thirty (5.2%) PD-L1-negative tumors at the 1+, 1% threshold had high TML or high MSI. Primary tumors had higher rates of high TML (8.8% vs. 3.9%; p = .0377) and high MSI (8.5% vs. 3.9%; p = .0471) than metastases. PD-L1 testing alone fails to detect patients who may benefit from ICI. Lower PD-L1 thresholds and TML testing should be considered in future clinical trials. Pembrolizumab is approved by the U.S. Food and Drug Administration for patients with refractory gastric and gastroesophageal cancers if the tumor and adjacent tissue stain positively for the programmed cell death ligand 1 (PD-L1) protein by companion diagnostic testing. Tumor mutational load, microsatellite instability (MSI), and alternative PD-L1 testing thresholds may serve as predictive biomarkers for response to immune checkpoint inhibition, and standard PD-L1 testing will not identify all patients who may benefit from this therapy. © AlphaMed Press 2018.
7 CFR 51.2547 - Definitions.

Code of Federal Regulations, 2013 CFR

2013-01-01

... shells, shells not split on suture, adhering hull material, light stained, or dark stained. (1) Damage by... shell surface or, on dyed nuts, when readily noticeable. (v) Dark stained on raw or roasted nuts, means an aggregate amount of dark brown, dark gray or black discoloration affects more than one-eighth of...
7 CFR 51.2547 - Definitions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... hull material, light stained, or dark stained. (1) Damage by external (shell) defects means any... noticeable. (v) Dark stained on raw or roasted nuts, means an aggregate amount of dark brown, dark gray or... than three-fourths, but not less than one-half the shell cavity. (ii) Kernel spotting refers to dark...
7 CFR 51.2547 - Definitions.

Code of Federal Regulations, 2012 CFR

2012-01-01

... hull material, light stained, or dark stained. (1) Damage by external (shell) defects means any... noticeable. (v) Dark stained on raw or roasted nuts, means an aggregate amount of dark brown, dark gray or... than three-fourths, but not less than one-half the shell cavity. (ii) Kernel spotting refers to dark...

7 CFR 51.2547 - Definitions.

Code of Federal Regulations, 2014 CFR

2014-01-01

... shells, shells not split on suture, adhering hull material, light stained, or dark stained. (1) Damage by... shell surface or, on dyed nuts, when readily noticeable. (v) Dark stained on raw or roasted nuts, means an aggregate amount of dark brown, dark gray or black discoloration affects more than one-eighth of...
7 CFR 51.2547 - Definitions.

Code of Federal Regulations, 2011 CFR

2011-01-01

... hull material, light stained, or dark stained. (1) Damage by external (shell) defects means any... noticeable. (v) Dark stained on raw or roasted nuts, means an aggregate amount of dark brown, dark gray or... than three-fourths, but not less than one-half the shell cavity. (ii) Kernel spotting refers to dark...
7 CFR 3201.104 - Metal cleaners and corrosion removers.

Code of Federal Regulations, 2014 CFR

2014-01-01

...) Definition. (1) Products that are designed to clean and remove grease, oil, dirt, stains, soils, and rust..., dirt, stains, and soils from stainless steel surfaces. (iii) Other metal cleaners. Products that are designed to clean and remove grease, oil, dirt, stains, and soils from metal surfaces other than stainless...
Trypan blue staining of the anterior capsule under an air bubble with a modified cannula.

PubMed

Toprak, Ahmet Baris; Erkin, Esin Fatma; Guler, Cenap

2003-01-01

To attain good visibility of the anterior capsule in the advanced or white cataract, trypan blue 0.1% is used to stain the anterior capsule. The dye is usually injected under an air bubble. However, it is difficult to inject the dye properly due to capillary forces. An ordinary anterior chamber cannula was modified and its coverage area increased to facilitate the staining of the anterior capsule under an air bubble. The anterior capsule was stained properly by using the modified cannula in all of the cases.
Improved silver staining of nucleolar organiser regions in paraffin wax sections using an inverted incubation technique.

PubMed Central

Coghill, G; Grant, A; Orrell, J M; Jankowski, J; Evans, A T

1990-01-01

A new simple modification to the silver staining of nucleolar organiser regions (AgNORs) was devised which, by performing the incubation with the slide inverted, results in minimal undesirable background staining, a persistent problem. Inverted incubation is facilitated by the use of a commercially available plastic coverplate. This technique has several additional advantages over other published staining protocols. In particular, the method is straightforward, fast, and maintains a high degree of contrast between the background and the AgNORs. Images PMID:1702451
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
Stem Cell Therapy for Healing Wounded Skin and Soft Tissues

DTIC Science & Technology

2014-03-01

histological analyses were performed. Immunofluorescence staining with chicken anti-GFP antibody showed that transplanted ASCs were evenly distributed in the...stained v.-ith chicken anti-GFP and mouse anti-a-SMA antibodies. Nuclei were stained v.·ith DAPl Merged image o f a -SM.b.. (red) and G FP (p ·Hn...Expressio n ofCD31 (PEC.-\\.M 4 1) in tr an$pbnted ASCs. Rabbit ear wounds at POD𔄁 were stained with chicken anti-GFP (A) and mouse anti-CD31 (B
Acetylcholinesterase and Nissl staining in the same histological section.

PubMed

Shipley, M T; Ennis, M; Behbehani, M M

1989-12-18

Acetylcholinesterase (AChE) enzyme histochemistry and Nissl staining are commonly utilized in neural architectonic studies. However, the opaque reaction deposit produced by the most commonly used AChE histochemical methods is not compatible with satisfactory Nissl staining. As a result, precise correlation of AChE and Nissl staining necessitates time-consuming comparisons of adjacent sections which may have differential shrinkage. Here, we have modified the Koelle-Friedenwald histochemical reaction for AChE by omitting the final intensification steps. The modified reaction yields a non-opaque reaction product that is selectively visualized by darkfield illumination. This non-intensified darkfield AChE (NIDA) reaction allows clear visualization of Nissl staining in the same histological section. This combined AChE-Nissl method greatly facilitates detailed correlation of enzyme and cytoarchitectonic organization.
Influence of orthodontic adhesives and clean-up procedures on the stain susceptibility of enamel after debonding.

PubMed

Joo, Hyun-Jin; Lee, Yong-Keun; Lee, Dong-Yul; Kim, Yae-Jin; Lim, Yong-Kyu

2011-03-01

To determine the influence of the type of orthodontic adhesive system, such as conventional acid-etching (CE) and self-etching primers (SEPs), on the stain susceptibility of enamel surface after debonding. Effects of clean-up procedures on the enamel surface were also determined. Two types and four brands of adhesive systems were investigated using 135 human premolars. Unbonded teeth were used as controls. Three-dimensional scanning of the enamel surface was performed before bracket bonding, after debonding, and after clean-up procedures. The color of each tooth was measured before bracket bonding and again after debonding and clean-up procedures. This was followed by methylene blue staining. The stain susceptibility of the enamel surface was measured after finishing only (F-condition) and after finishing/polishing (FP-condition). After debonding, the amount of residual adhesive resins in CE materials was greater than that in SEP materials. For the F-condition, staining color change in SEP materials was significantly higher than that in CE materials. For the FP-condition, staining color change in both CE and SEP materials was not different from those of the control. The SEP system would show less stain susceptibility if the thin residual adhesive resin layer after debonding is removed by polishing.
The distribution of lectin receptor sites in human breast lesions.

PubMed

Skutelsky, E; Hoenig, S; Griffel, B; Alroy, J

1988-08-01

Conflicting data regarding the status of A, B, H and T antigens in epithelium of normal, mastopathies, fibroadenomas and carcinomas of the breast stimulated us to re-examine the carbohydrate residues in these condition. Currently, we extended the number of carbohydrate residues studied by using ten different biotinylated lectins as probes and avidin-biotin-peroxidase complex (ABC) as a visualant. In addition, the pattern of lectin staining of cancerous cells in primary and metastatic sites was compared. In primary and metastatic breast carcinomas, lectin receptor sites were stained more intensely with Concanavalia ensiformi agglutinin (*Con A), Ricinus communis agglutinin-I (RCA-I) and wheat germ agglutinin (WGA), than in normal breast, in mastopathies or in fibroadenomas. Cryptic receptor sites for peanut agglutinin (PNA) were stained in all cases of breast carcinomas, while free PNA sites stained only in a few cases of well-differentiated carcinomas. Receptors sites for Ulex europaeus agglutinin-I (UEA-I) stained non-malignant epithelium of patients with blood group H but did not stain malignant cells. The results show significant differences in lectin-binding patterns and staining intensities between normal and non-malignant, and malignant epithelial breast cells. Furthermore, these results indicate that in malignant cells, there is an increased content of sialic acid-rich carbohydrates but not of asialylated glycoconjugates.
Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets

PubMed Central

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

2015-01-01

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

PubMed

Tissot, F; Prod'hom, G; Manuel, O; Greub, G

2015-09-01

The impact of round-the-clock cerebrospinal fluid (CSF) Gram stain on overnight empirical therapy for suspected central nervous system (CNS) infections was investigated. All consecutive overnight CSF Gram stains between 2006 and 2011 were included. The impact of a positive or a negative test on empirical therapy was evaluated and compared to other clinical and biological indications based on institutional guidelines. Bacterial CNS infection was documented in 51/241 suspected cases. Overnight CSF Gram stain was positive in 24/51. Upon validation, there were two false-positive and one false-negative results. The sensitivity and specificity were 41 and 99 %, respectively. All patients but one had other indications for empirical therapy than Gram stain alone. Upon obtaining the Gram result, empirical therapy was modified in 7/24, including the addition of an appropriate agent (1), addition of unnecessary agents (3) and simplification of unnecessary combination therapy (3/11). Among 74 cases with a negative CSF Gram stain and without formal indication for empirical therapy, antibiotics were withheld in only 29. Round-the-clock CSF Gram stain had a low impact on overnight empirical therapy for suspected CNS infections and was associated with several misinterpretation errors. Clinicians showed little confidence in CSF direct examination for simplifying or withholding therapy before definite microbiological results.
Differential Lectin Binding Patterns Identify Distinct Heart Regions in Giant Danio (Devario aequipinnatus) and Zebrafish (Danio rerio) Hearts

PubMed Central

Manalo, Trina; May, Adam; Quinn, Joshua; Lafontant, Dominique S.; Shifatu, Olubusola; He, Wei; Gonzalez-Rosa, Juan M.; Burns, Geoffrey C.; Burns, Caroline E.; Burns, Alan R.; Lafontant, Pascal J.

2016-01-01

Lectins are carbohydrate-binding proteins commonly used as biochemical and histochemical tools to study glycoconjugate (glycoproteins, glycolipids) expression patterns in cells, tissues, including mammalian hearts. However, lectins have received little attention in zebrafish (Danio rerio) and giant danio (Devario aequipinnatus) heart studies. Here, we sought to determine the binding patterns of six commonly used lectins—wheat germ agglutinin (WGA), Ulex europaeus agglutinin, Bandeiraea simplicifolia lectin (BS lectin), concanavalin A (Con A), Ricinus communis agglutinin I (RCA I), and Lycopersicon esculentum agglutinin (tomato lectin)—in these hearts. Con A showed broad staining in the myocardium. WGA stained cardiac myocyte borders, with binding markedly stronger in the compact heart and bulbus. BS lectin, which stained giant danio coronaries, was used to measure vascular reconstruction during regeneration. However, BS lectin reacted poorly in zebrafish. RCA I stained the compact heart of both fish. Tomato lectin stained the giant danio, and while low reactivity was seen in the zebrafish ventricle, staining was observed in their transitional cardiac myocytes. In addition, we observed unique staining patterns in the developing zebrafish heart. Lectins’ ability to reveal differential glycoconjugate expression in giant danio and zebrafish hearts suggests they can serve as simple but important tools in studies of developing, adult, and regenerating fish hearts. PMID:27680670
Staining human lymphocytes and onion root cell nuclei with madder root.

PubMed

Cücer, N; Guler, N; Demirtas, H; Imamoğlu, N

2005-01-01

We performed staining experiments on cells using natural dyes and different mordants using techniques that are used for wool and silk dyeing. The natural dye sources were madder root, daisy, corn cockle and yellow weed. Ferrous sulfate, copper sulfate, potassium tartrate, urea, potassium aluminum sulfate and potassium dichromate were used as mordants. Distilled water, distilled water plus ethanol, heptane, and distilled water plus methanol were used as solvents. All dye-mordant-solvent combinations were studied at pH 2.4, 3.2 and 4.2. The generic staining procedure was to boil 5-10 onion roots or stimulated human lymphocyte (SHL) preparations in a dye bath on a hot plate. Cells were examined at every half hour. For multicolor staining, madder-dyed lymphocytes were decolorized, then stained with Giemsa. The AgNOR technique was performed following the decolorization of Giemsa stained lymphocytes. Good results were obtained for both onion root cells and lymphocytes that were boiled for 3 h in a dye bath that included 4 g madder root, 4 g ferrous sulfate as mordant in 50 ml of 1:1 (v/v) methanol:distilled water. The pH was adjusted to 4.2 with 6 ml acetic acid. We conclude that madder root has potential as an alternative dye for staining biological materials.
Dirt detection on brown eggs by means of color computer vision.

PubMed

Mertens, K; De Ketelaere, B; Kamers, B; Bamelis, F R; Kemps, B J; Verhoelst, E M; De Baerdemaeker, J G; Decuypere, E M

2005-10-01

In the last 20 yr, different methods for detecting defects in eggs were developed. Until now, no satisfying technique existed to sort and quantify dirt on eggshells. The work presented here focuses on the design of an off-line computer vision system to differentiate and quantify the presence of different dirt stains on brown eggs: dark (feces), white (uric acid), blood, and yolk stains. A system that provides uniform light exposure around the egg was designed. In this uniform light, pictures of dirty and clean eggs were taken, stored, and analyzed. The classification was based on a few standard logical operators, allowing for a quick implementation in an online set-up. In an experiment, 100 clean and 100 dirty eggs were used to validate the classification algorithm. The designed vision system showed an accuracy of 99% for the detection of dirt stains. Two percent of the clean eggs had a light-colored eggshell and were subsequently mistaken for showing large white stains. The accuracy of differentiation of the different kinds of dirt stains was 91%. Of the eggs with dark stains, 10.81% were mistaken for having bloodstains, and 33.33% of eggs with bloodstains were mistaken for having dark stains. The developed system is possibly a first step toward an on line dirt evaluation technique for brown eggs.
Visualizing the Effects of a Positive Early Experience, Tactile Stimulation, on Dendritic Morphology and Synaptic Connectivity with Golgi-Cox Staining

PubMed Central

Mychasiuk, Richelle; Gibb, Robbin; Kolb, Bryan

2013-01-01

To generate longer-term changes in behavior, experiences must be producing stable changes in neuronal morphology and synaptic connectivity. Tactile stimulation is a positive early experience that mimics maternal licking and grooming in the rat. Exposing rat pups to this positive experience can be completed easily and cost-effectively by using highly accessible materials such as a household duster. Using a cross-litter design, pups are either stroked or left undisturbed, for 15 min, three times per day throughout the perinatal period. To measure the neuroplastic changes related to this positive early experience, Golgi-Cox staining of brain tissue is utilized. Owing to the fact that Golgi-Cox impregnation stains a discrete number of neurons rather than all of the cells, staining of the rodent brain with Golgi-Cox solution permits the visualization of entire neuronal elements, including the cell body, dendrites, axons, and dendritic spines. The staining procedure is carried out over several days and requires that the researcher pay close attention to detail. However, once staining is completed, the entire brain has been impregnated and can be preserved indefinitely for ongoing analysis. Therefore, Golgi-Cox staining is a valuable resource for studying experience-dependent plasticity. PMID:24121525
Toluidine Blue 0.05% Vital Staining for the Diagnosis of Ocular Surface Squamous Neoplasia in Kenya.

PubMed

Gichuhi, Stephen; Macharia, Ephantus; Kabiru, Joy; Zindamoyen, Alain M'bongo; Rono, Hilary; Ollando, Ernest; Wanyonyi, Leonard; Wachira, Joseph; Munene, Rhoda; Onyuma, Timothy; Jaoko, Walter G; Sagoo, Mandeep S; Weiss, Helen A; Burton, Matthew J

2015-11-01

Clinical features are unreliable for distinguishing ocular surface squamous neoplasia (OSSN) from benign conjunctival lesions. To evaluate the adverse effects, accuracy, and interobserver variation of toluidine blue 0.05% vital staining in distinguishing OSSN, confirmed by histopathology, from other conjunctival lesions. Cross-sectional study in Kenya from July 2012 through July 2014 of 419 adults with suspicious conjunctival lesions. Pregnant and breastfeeding women were excluded. Comprehensive ophthalmic slitlamp examination was conducted. Vital staining with toluidine blue 0.05% aqueous solution was performed before surgery. Initial safety testing was conducted on large tumors scheduled for exenteration looking for corneal toxicity on histology before testing smaller tumors. We asked about pain or discomfort after staining and evaluated the cornea at the slitlamp for epithelial defects. Lesions were photographed before and after staining. Diagnosis was confirmed by histopathology. Six examiners assessed photographs from a subset of 100 consecutive participants for staining and made a diagnosis of OSSN vs non-OSSN. Staining was compared with histopathology to estimate sensitivity, specificity, and predictive values. Adverse effects were enumerated. Interobserver agreement was estimated using the κ statistic. A total of 143 of 419 participants (34%) had OSSN by histopathology. The median age of all participants was 37 years (interquartile range, 32-45 years) and 278 (66%) were female. A total of 322 of the 419 participants had positive staining while 2 of 419 were equivocal. There was no histological evidence of corneal toxicity. Mild discomfort was reported by 88 (21%) and mild superficial punctate keratopathy seen in 7 (1.7%). For detecting OSSN, toluidine blue had a sensitivity of 92% (95% CI, 87%-96%), specificity of 31% (95% CI, 25%-36%), positive predictive value of 41% (95% CI, 35%-46%), and negative predictive value of 88% (95% CI, 80%-94%). Interobserver agreement was substantial for staining (κ = 0.76) and moderate for diagnosis (κ = 0.40). With the high sensitivity and low specificity for OSSN compared with histopathology among patients with conjunctival lesions, toluidine blue 0.05% vital staining is a good screening tool. However, it is not a good diagnostic tool owing to a high frequency of false-positives. The high negative predictive value suggests that a negative staining result indicates that OSSN is relatively unlikely.
Cationic Organochalcogen with Monomer/Excimer Emissions for Dual-Color Live Cell Imaging and Cell Damage Diagnosis.

PubMed

Chao, Xi-Juan; Wang, Kang-Nan; Sun, Li-Li; Cao, Qian; Ke, Zhuo-Feng; Cao, Du-Xia; Mao, Zong-Wan

2018-04-25

Studies on the development of fluorescent organic molecules with different emission colors for imaging of organelles and their biomedical application are gaining lots of focus recently. Here, we report two cationic organochalcogens 1 and 2, both of which exhibit very weak green emission (Φ 1 = 0.12%; Φ 2 = 0.09%) in dilute solution as monomers, but remarkably enhanced green emission upon interaction with nucleic acids and large red-shifted emission in aggregate state by the formation of excimers at high concentration. More interestingly, the monomer emission and excimer-like emission can be used for dual color imaging of different organelles. Upon passively diffusing into cells, both probes selectively stain nucleoli with strong green emission upon 488 nm excitation, whereas upon 405 nm excitation, a completely different stain pattern by staining lysosomes (for 1) or mitochondria (for 2) with distinct red emission is observed because of the highly concentrated accumulation in these organelles. Studies on the mechanism of the accumulation in lysosomes (for 1) or mitochondria (for 2) found that the accumulations of the probes are dependent on the membrane permeabilization, which make the probes have great potential in diagnosing cell damage by sensing lysosomal or mitochondrial membrane permeabilization. The study is demonstrative, for the first time, of two cationic molecules for dual-color imaging nucleoli and lysosomes (1)/mitochondria (2) simultaneously in live cell based on monomer and excimer-like emission, respectively, and more importantly, for diagnosing cell damage.
In vitro engineering of fibrocartilage using CDMP1 induced dermal fibroblasts and polyglycolide.

PubMed

Zhao, Guiqing; Yin, Shuo; Liu, Guangpeng; Cen, Lian; Sun, Jian; Zhou, Heng; Liu, Wei; Cui, Lei; Cao, Yilin

2009-07-01

This study was designed to explore the feasibility of using cartilage-derived morphogenetic protein-1 (CDMP1) induced dermal fibroblasts (DFs) as seed cells and polyglycolide (PGA) as scaffold for fibrocartilage engineering. DFs isolated from canine were expanded and seeded on PGA scaffold to fabricate cell/scaffold constructs which were cultured with or without CDMP1. Proliferation and differentiation of DFs in different constructs were determined by DNA assay and glycosaminoglycan (GAG) production. Histological and immunohistochemical staining of the constructs after being in vitro cultured for 4 and 6 weeks were carried out to observe the fibrocartilage formation condition. The fibrocartilage-specific gene expression by cells in the constructs was analyzed by real-time PCR. It was shown that in the presence of CDMP1 the proliferation and GAG synthesis of DFs were significantly enhanced compared to those without CDMP1. Fibrocartilage-like tissue was formed in the CDMP1 induced construct after being cultured for 4 weeks, and it became more matured at 6 weeks as stronger staining for GAG and higher gene expression of collagen type II was observed. Since only weak staining for GAG and collagen type II was observed for the construct engineered without CDMP1, the induction effect on the fibrocartilage engineering can be ascertained when using DFs as seed cells. Furthermore, the potential of using DFs as seed cells to engineer fibrocartilage is substantiated and further study on using the engineered tissue to repair fibrocartilage defects is currently ongoing in our group.
A study of peripheral blood in hedgehogs in Turkey.

PubMed

Ozparlak, Haluk; Celik, Ilhami; Sur, Emrah; Ozaydin, Tuğba; Arslan, Atilla

2011-09-01

The aim of this study was to determine diameters of blood cells, differential counts of peripheral blood leukocytes, alpha-naphthyl acetate esterase (ANAE), acid phosphatase (ACP-ase) activity of some leukocyte types, and enzymatic positivity percentages of peripheral blood lymphocytes in two hedgehogs species, Hemiechinus auritus, the long-eared hedgehog, and Erinaceus concolor, the southern white-breasted hedgehog. Air-dried peripheral blood smears were stained with May-Grünwald-Giemsa stain. ANAE and ACP-ase were stained in glutaraldehyde-acetone-fixed smears. ANAE-positive lymphocytes displayed a dot-like positivity pattern characterized with 1-5 reddish brown cytoplasmic granules, whereas ACP-ase positive lymphocytes displayed a dot-like positivity pattern characterized with 1-3 pinkish cytoplasmic granules. Monocytes gave a diffuse and strong reaction while neutrophils displayed a weak positive reaction for ANAE and ACP-ase. No difference was observed in mean diameters of peripheral blood cells of these species. It was found that lymphocytes made up the majority (64.3% and 65.5%) of leukocytes, followed by neutrophils (23.9% and 23.3%), eosinophils (9.0% and 7.6%), monocytes (1.8% and 2.3%), and basophils (1.0% and 1.3%) in H. auritus and E. concolor, respectively. Mean ANAE positivity oflymphocytes was 36.6% and 51.3% and ACP-ase positivity was 32.1% and 37.5% for H. auritus and E. concolor, respectively. The ANAE positivity of lymphocytes in E. concolor was significantly (P < 0.05) higher than that of H. auritus.

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