Advantages and disadvantages of silver protein staining in polyacrylamide gel

Morozov, V A; Il'inskii, P. O.; Ильин, К. В.

doi:10.1007/bf00854708

Cited by 5 publications

(4 citation statements)

References 6 publications

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“…As underlined by Jiang et al (Jiang et al, 2022), the binding capability of silver ion are affected by several factors including reactivity of protein towards silver ion. The staining of silver on the gel are selective, with certain proteins staining weakly and others not being detected at all (Morozov et al, 1986). The ndings in this study extend this analysis whereby the glycine-HCL extracted surface protein that were observed at several location on the coomassie stained gel could not be detected by silver staining Therefore, using both staining procedures can be advantageous to identify all surface proteins without any loss.…”

Section: Discussionsupporting

confidence: 60%

Application of Liquid-Phase Preparative Isoelectric Focusing for Comparative Analysis of Abundancy in Differentially Extracted Bacterial Protein of S.Typhi and Invasive Non-Typhoidal Salmonella

Khan

Kader

2023

Preprint

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Isolation, purification, and separation of complex mixtures is crucial in proteomic research. The conventional electrophoresis method for antigen characterization has limitations in separating low abundance components and selectively enriching important proteins with high degree of purity. Two-dimensional gel electrophoresis was introduced to overcome these limitations, but it also has inherent shortcomings in detecting hydrophobic proteins, low abundance proteins, or samples with proteins of various concentrations. Therefore, this study aims to develop an innovative approach for the enrichment and characterization of immunoreactive components found in differentially extracted whole cell bacterial protein derived from S.Typhi and S.spp. A modified liquid phase preparative isoelectric focusing (IEF) and SDS-PAGE method was used to purify and characterize the proteins. The modified liquid phase IEF efficiently fractionated the proteins into 20 fractions based on the pI value, providing a high-resolution power for protein separation, high throughput, and ease of performance. The fractionated proteins were then analysed by SDS-PAGE for their molecular weight, providing a simple and cost-efficient method for protein analysis. This innovative approach for the enrichment and characterization of immunoreactive components in differentially extracted whole cell bacterial protein derived from S.Typhi and S.spp has the potential to revolutionize the diagnosis and treatment of typhoid fever and other related diseases. By improving the sensitivity and accuracy of protein analysis, this study may lead to identification of exclusive disease biomarkers for early, accurate diagnosis of diseases, improved prognosis and treatment outcomes.

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Section: Discussionsupporting

confidence: 60%

Application of Liquid-Phase Preparative Isoelectric Focusing for Comparative Analysis of Abundancy in Differentially Extracted Bacterial Protein of S.Typhi and Invasive Non-Typhoidal Salmonella

Khan

Kader

2023

Preprint

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“…1C compared to Fig. 1F) may be that silver staining does not have a good linear relationship with protein concentration; or that silver staining is more selective, and the decisive factor influencing the effectiveness of silver staining is the amino-acid composition of the protein [22]. That is one reason for us to choose only one spot for each cell type, that we could with confidence judged to be specific from silver stained as well as from CBB-stained gels.…”

Section: Protein Spots Specific For Hpt Sgc and Gcmentioning

confidence: 99%

Hemocyte‐lineage marker proteins in a crustacean, the freshwater crayfish, Pacifastacus leniusculus

Söderhäll

Kim

et al. 2008

Proteomics

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To identify proteins associated with development of different hemocyte types in the freshwater crayfish Pacifastacus leniusculus, 2-DE followed by MS analysis was carried out with hematopoietic tissue (Hpt) cells, semigranular cells (SGC) and granular cells (GC). Within the hemocyte lineages one two-domain Kazal proteinase inhibitor (KPI) was found to be specific for SGC, while a superoxide dismutase (SOD) was specific for GC at protein as well as at mRNA level. The proliferation cell nuclear antigen (PCNA) was detected at the mRNA level in Hpt cells only. We also provide evidence that SGC and GC most likely differentiate to maturation as separate lineages. We found that after laminarin or lipopolysaccharide (LPS) injection into crayfish, the transcript levels of PCNA and SOD increased in the Hpt cells, whereas the KPI transcript never was present in Hpt regardless of any challenge. RNA interference of PCNA in the Hpt cells led to that most of the cells did not spread or attach to the tissue culture dish. These results suggest that PCNA, KPI and SOD can be used as markers for Hpt cells, SGC and GC, respectively, and in conjunction with these results, a model is proposed how the Hpt responds to a microbial challenge by proliferation and release of Hpt cells.

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“…Coomassie brilliant blue is more convenient, reproducible and an endpoint procedure compared with the multi-step silver stain procedures. Furthermore, variations in the efficacy silver staining for some proteins were reported by Morozov, et al in 1986 [ 31 ], so this variation together with protein load variations (300μg of protein for Coomassie brilliant blue R-250 versus 200μg of protein for silver staining) could account for other differences in the visual data occurring between the two stains.…”

Section: Discussionmentioning

confidence: 99%

Exploring Early and Late Toxoplasma gondii Strain RH Infection by Two-Dimensional Immunoblots of Chicken Immunoglobulin G and M Profiles

et al. 2015

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Toxoplasma gondii is an intracellular apicomplexan parasite infecting warm-blooded vertebrate hosts, with only early infection stage being contained with drugs. But diagnosis differencing early and late infection was not available. In the present investigation, 2-dimensional immunobloting was used to explore early and late infections in chickens. The protein expression of T. gondii was determined by image analysis of the tachyzoites proteome separated by standard-one and conventional two-dimentional gel polyacrylamide electrophoresis (2D- PAGE). Pooled gels were prepared from tachyzoites of T. gondii. A representative gel spanning a pH range of 3-10 of the tachyzoite proteome consisted of 1306 distinct polypeptide spots. Two-dimensional electrophoresis (2-DE) combined with 2-DE immunoblotting was used to resolve and compare immunoglobulins (Igs) M & G patterns against Toxoplasma gondii strain RH (mouse virulent strain). Total tachyzoite proteins of T. gondii were separated by two-dimensional gel electrophoresis and analyzed by Western blotting for their reactivity with the 7 and 56 days post-infection (dpi) SPF chicken antisera. Different antigenic determinant patterns were detected during analysis with M and G immunoglobulins. Of the total number of polypeptide spots analyzed (1306 differentially expressed protein spots), 6.97% were identified as having shared antigenic polypeptide spots on immunoblot profiles with IgG and IgM antibodies regardless the time after infection. Furthermore, some of the immunoreactive polypeptide spots seemed to be related to the stage of infection. Interestingly, we found natural antibodies to toxoplasmic antigens, in addition to the highly conserved antigenic determinants that reacted with non-specific secondary antibody; goat anti-chicken IgG antibodies conjugated with horseradish peroxidase. In conclusion, unique reactive polypeptide spots are promising candidates for designation of molecular markers to discriminate early and late chicken infection.

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Advantages and disadvantages of silver protein staining in polyacrylamide gel

Cited by 5 publications

References 6 publications

Application of Liquid-Phase Preparative Isoelectric Focusing for Comparative Analysis of Abundancy in Differentially Extracted Bacterial Protein of S.Typhi and Invasive Non-Typhoidal Salmonella

Application of Liquid-Phase Preparative Isoelectric Focusing for Comparative Analysis of Abundancy in Differentially Extracted Bacterial Protein of S.Typhi and Invasive Non-Typhoidal Salmonella

Hemocyte‐lineage marker proteins in a crustacean, the freshwater crayfish, Pacifastacus leniusculus

Exploring Early and Late Toxoplasma gondii Strain RH Infection by Two-Dimensional Immunoblots of Chicken Immunoglobulin G and M Profiles

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