The six test proteins (BSA, avidin, myoglobin, recombinant protein G, jacalin, and apotransferrin) were characterized and visualized by silver stained SDS-PAGE (Thermo Fisher Scientific) according to the manufacturer’s protocol. Briefly, protein samples were mixed with an equivalent volume of 2x Laemmli buffer, containing 65.8 mM Tris-HCl (pH 6.8, Carl Roth), 26.3% glycerol (v/v, Carl Roth), 2.1% SDS (Carl Roth), 0.02% bromophenol blue (Sigma-Aldrich) and 5.0% 2-mercaptoethanol (Sigma-Aldrich), and heated at 95 ∘C for 5 min. The protein samples (50 ng in 10 μL) and the color-prestained standard marker (10 ng, broad range, 11–245 kDa, New-England Biolabs, Frankfurt, Germany) were loaded onto the gels (Protean Precast, 4–20%, Bio-Rad, Munich, Germany). The manufacturer’s silver stain kit protocol (Thermo Fisher Scientific) was applied. A ChemiDoc system (Bio-Rad) with Image Lab software 5.2.1 (Bio-Rad) was used for image acquisition. The SDS-PAGE gel image can be found in Figure S6 (Electronic Supplementary Material).
Silver stained sds page
Manufactured by Thermo Fisher Scientific
Silver stained SDS-PAGE is a laboratory technique used to separate and visualize proteins based on their molecular weight. It involves the use of polyacrylamide gel electrophoresis (SDS-PAGE) to separate proteins, followed by silver staining to detect and visualize the protein bands.
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Lab products found in correlation
Image lab software 5, by Bio-Rad (2 mentions)
Tris hcl, by Carl Roth (2 mentions)
Glycerol, by Carl Roth (2 mentions)
Bromophenol blue, by Merck Group (2 mentions)
2 mercaptoethanol, by Merck Group (2 mentions)
Chemidoc system, by Bio-Rad (2 mentions)
Sodium dodecyl sulfate (sds), by Carl Roth (2 mentions)
Silver stain kit, by Thermo Fisher Scientific (1 mentions)
Protean precast gels, by Bio-Rad (1 mentions)
2 protocols using silver stained sds page
1
SDS-PAGE Characterization of Test Proteins
2
Visualizing Protein Oligomerization by SDS-PAGE
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Protein oligomerization was visualized and quantified by silver stained SDS-PAGE (Thermo Fisher Scientific). Protein samples dissolved in PBS were mixed with an equivalent volume of 2x Laemmli buffer, containing 65.8 mM Tris-HCl (pH 6.8, Carl Roth), 26.3% glycerol (v/v, Carl Roth), 2.1% SDS (Carl Roth), 0.02% bromophenol blue (Sigma Aldrich) and 5.0% 2-mercaptoethanol (Sigma Aldrich), and heated at 95 °C for 5 min. The samples (50 ng in 10 μL) were loaded onto PROTEAN Precast gels (4–20%, Bio-Rad, Munich, Germany) together with 10 ng Color Prestained Protein Standard, Broad Range (11–245 kDa, New-England Biolabs, Frankfurt, Germany) and stained with a silver stain kit (Thermo Fisher Scientific) following manufacturer's instructions. For image acquisition and quantification of protein monomers, dimers, and higher oligomers, a ChemiDoc system (Bio-Rad) with Image Lab software 5.2.1 (Bio-Rad) was used. Protein dimers occurred in the modified samples of all proteins (Fig. S1 , Table S2 ), and signals of higher oligomers were observed but exceeded the analytical detection limit (3%) only for modified HSP60 (23 ± 13%).
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