The protein levels in renal tissues were detected by Western blotting analysis. Briefly, the renal tissues from different groups were lyzed by Radio-Immunoprecipitation Assay lysis buffer (Beyotime Bio Inc, Shanghai, China). The protein concentrations were determined by BCA protein estimation kit (Beyotime Bio Inc). Then, equal amounts of protein samples were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Beyotime Bio Inc), and transferred to polyvinylidene difluoride membranes (Beyotime Bio Inc). After blocking with 5% non-fat dry milk, the membranes were incubated with primary antibodies at 4°C overnight. The primary antibodies used in this study were as follows: GRα (BOSTER Bio Inc, Wuhan, China), GRβ (Bioss Bio Inc, Beijing, China), TGF-β1 (BOSTER Bio Inc), activator protein (AP)-1 (Bioss Bio Inc), and GAPDH (Santa Cruz Bio Inc, Santa Cruz, USA). Then the corresponding secondary antibodies conjugated with horseradish peroxidase were added. ECL detection reagent (7 Sea Biotech, Shanghai, China) was used to visualize the blots. The results were quantified with Gel-Pro-Analyzer (Media Cybernetics, Maryland, USA).
Ecl detection reagent
Manufactured by 7Sea Biotech
Sourced in China
The ECL detection reagent is a laboratory reagent used for chemiluminescent detection in various biochemical and molecular biology applications. The reagent generates a luminescent signal upon reaction with the target analyte, which can be detected and quantified using specialized imaging equipment.
Automatically generated - may contain errors
Lab products found in correlation
Radioimmunoprecipitation assay lysis buffer, by Beyotime (3 mentions)
Gel pro analyzer, by Media Cybernetics (3 mentions)
Sodium dodecyl sulfate polyacrylamide gel electrophoresis, by Beyotime (2 mentions)
Bca protein estimation kit, by Beyotime (2 mentions)
Polyvinylidene difluoride membrane, by Beyotime (1 mentions)
Tgf β1, by Boster Bio (1 mentions)
Polyvinylidene difluoride, by Beyotime (1 mentions)
Bca protein evaluation kit, by Beyotime (1 mentions)
Sodium dodecyl sulphate polyacrylamide gel electrophoresis, by Beyotime (1 mentions)
3 protocols using ecl detection reagent
1
Western Blot Analysis of Renal Proteins
2
Spinal Cord Tissue Protein Quantification Using Western Blotting
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Western blotting was used for determining the protein concentration in the tissues of the spinal cord. In brief, we lyse tissues from various groups with Radio-Immunoprecipitation assay lysis buffer (Beyotime Bio Inc, Shanghai, China) and measure protein quantities with the BCA protein evaluation kit (Beyotime Bio Inc). Later, using a sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Beyotime Bio Inc), protein specimens of identical amounts were separated and moved to membranes fabricated from polyvinylidene difluoride (Beyotime Bio Inc). Following blocking with 5% non-fat dry milk, the membranes were treated at 4° C during the night hours with primary antibodies. GAPDH (Santa Cruz Bio Inc, USA), caspase-8 (Santa Cruz Bio Inc, USA) and STAT3 (Santa Cruz Bio Inc, USA) were the set of primary antibodies employed in the work. Then, to bind with the horseradish peroxidase, the appropriate secondary antibodies were added. The blotting findings were seen using an ECL detection reagent provided by 7 Sea Biotech, Shanghai, China. A Gel-Pro-Analyzer (Media Cybernetics, USA) was implemented to quantify the data.
3
Quantification of Rat Macrophage Proteins
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The protein levels in rat monocyte-derived macrophages were detected by Western blotting analysis. Brie y, the cells from different groups were lysed by Radio-Immunoprecipitation Assay lysis buffer (Beyotime Bio Inc, Shanghai, China). The protein concentrations were determined by BCA protein estimation kit (Beyotime Bio Inc). Then, equal amounts of protein samples were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (Beyotime Bio Inc), and transferred to polyvinylidene di uoride membranes (Beyotime Bio Inc). After blocking with 5% non-fat dry milk, the membranes were incubated with primary antibodies at 4°C overnight. Then the corresponding secondary antibodies conjugated with horseradish peroxidase were added. ECL detection reagent (7 Sea Biotech, Shanghai, China) was used to visualize the blots. The results were quanti ed with Gel-Pro-Analyzer (Media Cybernetics, USA).
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