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PubMed1993

Nuclear immunostaining of hepatitis C infected hepatocytes with monoclonal antibodies to C100-3 nonstructural protein. Comparison of immunogold silver staining with other immunohistochemical methods.

Immunohistochemical methods have been used to localize an HCV antigen on paraffin embedded liver tissue sections by means of monoclonal antibodies to C100-3 nonstructural protein. Peroxidase-antiperoxidase, alkaline phosphatase-antialkaline phosphatase, biotin-streptavidin-peroxidase and immunogold silver staining methods showed a nuclear staining of the hepatocytes in cases of chronic hepatitis with positive HCV serology, alcoholic liver disease and hepatocarcinoma. No cross reactions were observed with viral hepatitis B and delta antigens. The strongest reaction without background staining was obtained with immunogold silver staining. Nuclear localization was compared to the cytoplasmic staining described in the literature.

Antibodies, Monoclonal↗
PubMed1989

The silver-stained proteins of interphasic nucleolar organizer regions as a parameter of cell duplication rate.

The relationship between interphasic silver-stained proteins of the nucleolar organizer regions (Ag-NOR proteins) and cell replication rate has been studied in 13 established neuroblastoma cell lines. The quantity of Ag-NOR proteins was measured in silver-stained cells by means of an automated image analyzer. The results indicated that the amount of Ag-NOR proteins is strictly proportional to the proliferative activity of the cells. Cell lines with a difference of only 4 h in doubling time were characterized by a statistically significant difference in Ag-NOR protein amount. The Ag-NOR protein quantity can therefore be used as a parameter for evaluating the cell proliferation rate.

Cell Division↗
PubMed1989

Thiosemicarbazide used after periodic acid makes methenamine silver staining of renal glomerular basement membranes faster and cleaner.

The periodic acid-methenamine silver staining technique, which is frequently used for demonstrating the renal glomerular basement membrane, requires a high degree of skill, and in some cases it may be difficult to obtain a good result. To overcome such difficulty and inconsistency, we have improved the method by performing methenamine silver staining after oxidation with periodic acid and subsequent application of thiosemicarbazide. In this procedure, this semicarbazide enhanced the reaction of methenamine silver with the glomerular basement membrane and the reaction was completed within a shorter time in comparison with the conventional method. This modification also eliminated any nonspecific reaction with the surface of the glass slide and the solution container and yielded excellent and reproducible results irrespective of the fixation method and material employed. It was also found to stain the renal glomerular basement membrane of rabbits, which is demonstrable only with difficulty by the conventional method.

Animals↗
PubMed1999

On the mechanism of background silver staining during sodium dodecyl sulphate-polyacrylamide gel electrophoresis.

A novel experimental procedure to study the factors that contribute to background silver staining in sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) is described. A mechanism is proposed for background silver staining, which was found to result primarily from the redox initiator system using ammonium persulphate (APS) and an amine such as N,N,N',N'-tetramethylethylenediamine (TEMED).

Ammonium Sulfate↗
PubMed2004

[Comparison of several silver staining methods for DNA polyacrylamide gel electrophoresis].

OBJECTIVE: To compare 4 silver staining methods for DNA detection in polyacrylamide gel electrophoresis. METHODS: After the electrophoresis was completed, the gels were stained separately by four different methods, namely using ammonia-silver-citric acid, low-concentration silver nitrate, 0.1% and 0.2% silver nitrate. RESULTS: DNA was detected only by staining with 0.1% and 0.2% silver nitrate. CONCLUSION: 0.2% silver nitrate used along with sodium hydroxide shows the highest sensitivity in DNA detection, while its use with sodium carbonate produces the best quality of the image.

DNA↗
PubMed2002

Silver stain removal using H2O2 for enhanced peptide mass mapping by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Hydrogen peroxide solutions are reported for the removal of silver stain from proteins isolated in polyacrylamide gels. Removal of silver stain prior to in-gel digestion is shown to enhance sensitivity and sequence coverage of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) peptide mass maps. The rate of silver removal using H2O2 is influenced by H2O2 concentration and increases with increasing pH. The presence of complexation reagents such as ammonia from mass spectrometry compatible ammonium bicarbonate solutions enhances the efficiency and speed of H2O2-mediated silver removal. H2O2-mediated silver removal using the described procedure does not appear to have any detrimental effects on proteins but is observed to produce a slightly elevated level of methionine oxidization over that usually observed in in-gel tryptic digestion.

Bicarbonates↗
PubMed2002

[A study of forensic individual identification used STR locus with silver staining and multiplex PCR methods].

Amplification of short tandem repeat(STR) loci has become a useful tool for human identification applications. To improve throughput and efficiency for the forensic materials and gain foure and six STR locis multiplex methods with silver staining, CSF1PO,TPOX,THO1 and vWA(referred to as multiplex A), D18S51, D7S820, D13S317, D5S818, D3S1358 and Amelogenin(referred to as multiplex B) have been evaluated for use in a rape case. The products of multiplex amplication were separated in a denaturing polyacrylamide gel and analyzed with silver staining. Two multiplex amplications used in this case could provide a power of discrimination of approximately 2.43 x 10(-19). Silver staining was shown to be a validation methods for analysing the products of four and six multiplex amplications.

English Abstract↗
PubMed1992

Silver-stained nucleolar organizer proteins in urothelial bladder lesions. A morphometric study.

The diagnostic value of image analysis of silver-stained nucleolar organizer regions (AgNORs) has been investigated in 170 urothelial bladder lesions obtained by transurethral biopsies in the same number of patients. According to the WHO classification 25 specimens showed a flat non-invasive growth pattern with simple hyperplasia or mild dysplasia (H/D1 = 10), as well as moderate (D2 = 10) to severe atypia (Cis = 5). 135 samples were of papillary and/or infiltrating type including polypoid cystitis (pC = 5), true papillomas (G0 = 7), and carcinomas of different malignancy grade (G1 = 30, G2 = 55, G3 = 38). 10 biopsies served as normal controls. Benign non-neoplastic urothelium (controls, H/D1, pC) exhibited few but large AgNORs (mean number [MNN] = 3.3 +/- 0.5, mean area [MNA] = 0.29 +/- 0.08 micron 2), whereas carcinomatous lesions (Cis, G1-G3) showed numerous small silver-stained dots within their nuclei (MNN = 6.9 +/- 1.1; MNA = 0.12 +/- 0.04 micron 2). Both parameters, MNN and MNA, were inversely correlated (r = -0.69, p less than 0.001); their quotient (NQ = MNN/MNA) revealed a clear cut difference in the AgNOR content of benign (control, H/D1:NQ = 12.8 +/- 2.5) and moderate to severe atypical flat urothelial lesions (D2, Cis: NQ = 44.2 +/- 12.8, p less than 0.001). This parameter also discriminated between G1 (NQ = 40.7 +/- 17.3), G2 (NQ = 57.5 +/- 18.8), and G3 carcinomas (NQ = 79.0 +/- 21.8, p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Histocytochemistry↗
PubMedDate not supplied

The isoelectric focusing of keratins in hair followed by silver staining.

An isoelectric focusing method followed by silver staining has been developed for the study of keratins which is as effective as two-dimensional electrophoresis and fluorography for hair species identification. Hair from dogs, rabbits, horses, cows, guinea-pigs, donkeys, sheep and cats were successfully identified. Narrow pH ranges were used to observe heterogeneity in human hair. Although this heterogeneity may be affected by environmental conditions, it may be of use in criminalistics.

Animals↗
PubMedDate not supplied

An improved silver staining procedure for schizodeme analysis in polyacrylamide gradient gels.

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.

Animals↗
PubMed1993

Comparison of silver stainings and immunohistology for the detection of neurofibrillary tangles and extracellular cerebral amyloid in paraffin sections.

The sensitivities of six silver-staining methods and immunohistology for beta and tau protein were compared for their ability to demonstrate neurofibrillary tangles (NFT) and senile plaques (SP) in paraffin sections. Serial sections of the hippocampal area of 35 brains showing these neuropathological findings were cut and stained by the methods of Cross, Campbell, Bielschowsky, Gallyas, Yamaguchi, our variant (method of Reusche) and immunohistology. In the detection of NFT, the techniques of Gallyas, Bielschowsky, our method and tau protein immunostaining were the most sensitive methods. The procedure of Campbell and again our method were proven to be superior to the other stainings in demonstrating SP as well as diffuse and subpial amyloid. Moreover, our method reliably stained vascular and perivascular amyloid which can be identified in brains with congophilic angiopathy. Due to a lack of control in certain steps of the procedures most of the silver-staining methods are complicated and to not present reliable results. Our variant is easy to perform and, thus, may be used as a sensitive, simple and reliable alternative for the impregnation of the main lesions (NFT and SP) occurring in senile dementia of Alzheimer type and brains with normal aging for screening, retrospective and quantitative studies and for routine purposes.

Aging↗
PubMed1981

A silver staining technique for investigating wear of restorative dental composites.

A silver staining technique was developed to demonstrate microdefects in dental restorative composites. Fine silver particles were preferentially introduced into the damaged region to provide optical contrast between the damaged and the undamaged regions. The amount of silver deposition determined with an electron probe microanalyzer, provided an indication of the extent of damage within the dental composites. Examples to demonstrate this technique were given with one clinically worn dental composite restoration and one in vitro worn composite sample.

Dental Materials↗
PubMed2006

Formation of epsilon-formyllysine on silver-stained proteins: implications for assignment of isobaric dimethylation sites by tandem mass spectrometry.

Considerable effort is focused presently on the detection and comprehensive assignment of post-translational modifications of proteins. Obviously attention must be paid to the possibility of chemical modifications that may occur to protein samples during sample handling and manipulation prior to analysis by tandem mass spectrometry. This is of particular concern when a modification is isobaric with the mass differential in common with a known post-translational analog. Here we provide evidence that silver staining protocols that use formaldehyde can result in epsilon-formylation of lysine residues. This modification is in fact isobaric with the important product of methyltransferases, epsilon,epsilon-dimethyllysine. Without exercising proper caution the analysis of silver-stained protein samples by mass spectrometry looking for dimethylation of lysine will yield a significant number of misassigned sites of modification. High accuracy measurements of the mass of the precursor ions and their fragments are required to eliminate this uncertainty. The occurrence of dimethylation of the epsilon-amino function of lysine residues has been reported often in histones. For histone samples excised from silver-stained gels, we found that most sites initially assigned to be dimethylated by automatic search engines under standard search parameters (100 ppm error tolerance) are actually in fact formylated. Caution must be exercised when data obtained from instruments unable to perform high accuracy mass measurements (better than 5 ppm) are to be interpreted.

Amino Acid Sequence↗
PubMed1980

Light microscopic observations on the behavior of silver-stained trivalents in pachytene cells of Sigmodon fulviventer (Rodentia, Muridae) heterozygous for centric fusion.

Silver-stained pachytene cells of male cotton rats, Sigmodon fulviventer, heterozygous for a naturally occurring Robertsonian fusion, were examined by light microscopy. Silver staining clearly revealed synaptonemal complexes of all autosomal bivalents and of the translocation trivalent, as well as X-Y conformations within the sex vesicle. The synaptonemal complex of the trivalent conforms closely in morphology to that observed by electron microscopy in hybrids of other species. In all cases, the proximal telomeric knobs of the two acrocentric elements were paired and projected in the same direction from the metacentric element. The geometry of trivalent pairing in pachytene may predispose to normal disjunction and, therefore, account for the high fertility observed in S. fulviventer heterozygotes.

Animal Population Groups↗
PubMed1989

Silver staining of Pneumocystis carinii in the rat's lung.

The backscattered electron imaging mode of the scanning electron microscope was used to study the ontogenetic acquisition of argyrophilia in Pneumocystis carinii in rats. Silver staining continually increased from the late trophozoite to the mature cyst stage. The silver uptake began with a fine outline at the surface of the bodies of the late trophozoites; their cellular extensions, however, did not stain. The oblate precyst forms acquired the silver in heterogeneous patches. On spherical cysts the silver staining became more uniform and intense with at least one dense spot. The spherical and collapsed cysts also had short silver staining projections that may represent microvilli. Collapsed forms were paler than spherical ones and appear to be cysts that have undergone partial or complete release of sporozoites. These cell surface observations confirm and amplify previous transmission electron microscopical and histochemical studies indicating that silver staining correlates with the acquisition of the cell pellicle.

Animals↗
PubMed1990

Silver staining of interphase nucleolar organizer regions in cytologic smears previously stained by the Papanicolaou and May-Grünwald-Giemsa techniques.

Malignant cells are known to display a distinctive argyrophilic nucleolar organizer region (Ag-NOR) protein distribution compared with distribution in their benign counterparts. In this study, Ag-NOR staining was applied to smears from reactive and malignant serous effusions. The smears were either unstained or had been previously stained with the Papanicolaou or May-Grünwald-Giemsa (MGG) technique. The purpose of this study was to assess the feasibility and usefulness of Ag-NOR staining in diagnostic cytopathology and to set up a procedure that could be used on prestained smears in retrospective studies. The one-step silver staining method was applied to prestained smears after a weak destaining step in trichloracetic acid and to unstained smears after a postfixation step in Carnoy's fluid. Positive results were observed on both destained and unstained preparations, MGG-stained smears showing a better visualization of the silver deposits than Papanicolaou-stained smears. Major differences in size, shape, and distribution patterns of the Ag-NOR-positive granules were observed between neoplastic and reactive cells. Furthermore, the mean number of silver-stained dots per cell was significantly higher in malignant cells (33.32 +/- 2.23) than in reactive mesothelial cells (9.71 +/- 0.66). These data indicate that this Ag-NOR technique can be profitably applied to prestained cytologic smears to assist in the diagnosis of malignancy and that the technique has the advantage that cellular morphology and silver staining can be evaluated on the same slide.

Ascitic Fluid↗
PubMed1990

Modification of the silver staining technique to detect lipopolysaccharide in polyacrylamide gels.

A silver staining method used routinely for detecting bacterial lipopolysaccharide (LPS) in sodium dodecyl sulfate-polyacrylamide gels (C. Tsai and E. Frasch, Anal. Biochem. 119:115-119, 1982) appeared to be inappropriate for visualizing certain LPS preparations. It did not stain S-form fractions of polyagglutinable Pseudomonas aeruginosa LPS or several partly deacylated (alkali-treated) S-form LPS after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. However, these LPS preparations could be detected by anti-LPS sera after electroblotting onto nitrocellulose, thereby confirming their integrity and presence in the polyacrylamide gel. This is because LPS fractions containing a low number of fatty acids are washed out of the gel during the initial fixing step (40% ethanol-4% acetic acid, overnight). By omitting this fixing step, which was originally developed for detecting proteins, and by increasing the LPS oxidation time (from 5 to 20 min), we restored the ability to detect LPS fractions that otherwise would not be stained. These modifications did not affect the detection of other S- and R-form LPSs. Thus, differences in the number of fatty acids present in polyagglutinable P. aeruginosa LPS may result in a selective loss of fatty acid-deficient S-form LPS in these apparent R-form LPS preparations. This modified procedure provides a fast, simple, and sensitive way to analyze LPS in polyacrylamide gels despite the number of acyl groups present.

Electrophoresis, Polyacrylamide Gel↗
PubMed1991

The immunogold-silver staining procedure in the study of freshly suspended Langerhans cells at the transmission electron microscopic level.

The potential of an immunogold-silver staining for the study of human suspended Langerhans cells at the transmission electron microscopic level was evaluated. Cells were labeled, by using a preembedding technique, with 5-nm colloidal gold particles followed by silver enhancement. The use of small colloidal gold particles permits a detection of small quantities of antigen; the metallic silver deposition around gold granules gives rise to a large electron-dense marker which can be easily detected even at low magnification. Ultrastructural details were well preserved, and the background was not significant. The major advantage of the present immunogold-silver staining is that it enables to detect labeled cells easily, even when limited amounts of antigenic moieties are present on a low percentage of cells. Therefore, a rapid and simultaneous evaluation of both immunophenotype and ultrastructural details of investigated cells is allowed.

Antigens, CD↗