Cell lysates was harvested from the cell lines using radio immunoprecipitation assay (RIPA) buffer (Thermo Fisher Scientific) with protease inhibitors, phosphatase inhibitors and EDTA (Thermo Fisher Scientific). Aliquots of protein were electrophoresed on SDS/PAGE Tris·HCl gels (Bio-Rad Laboratories). The proteins were separated by electrophoresis prior to transfer to PVDF membranes (Bio-Rad Laboratories) and incubated with primary antibodies overnight at 4 °C, followed by incubation with HRP-linked anti-rabbit or anti-mouse IgG (GE Healthcare Biosciences) at a dilution of 1:100,000 for 1 h at room temperature. The antigen–antibody complex was detected with the ECL Prime Western Blotting Detection Kit (GE Healthcare Biosciences). The intensity of the blots was quantified by densitometry analysis using Image lab software version 5.0 (Bio-Rad Laboratories).
Sds page tris hcl gels
Manufactured by Bio-Rad
Sourced in United States
SDS/PAGE Tris·HCl gels are laboratory equipment used for polyacrylamide gel electrophoresis (PAGE) to separate proteins based on their molecular weight. The gels are composed of Tris-HCl buffer and provide a stable matrix for the separation of proteins in the presence of the denaturing agent sodium dodecyl sulfate (SDS).
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Lab products found in correlation
Ecl prime western blotting detection kit, by GE Healthcare (2 mentions)
Ripa buffer, by Thermo Fisher Scientific (2 mentions)
Protease inhibitor, by Thermo Fisher Scientific (1 mentions)
Phosphatase inhibitor, by Thermo Fisher Scientific (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions)
Image lab software version 5, by Bio-Rad (1 mentions)
Pvdf membrane, by Bio-Rad (1 mentions)
Wet transfer system, by Bio-Rad (1 mentions)
Hrp linked anti rabbit igg, by GE Healthcare (1 mentions)
Radioimmunoprecipitation assay buffer, by Thermo Fisher Scientific (1 mentions)
Actb antibody, by Merck Group (1 mentions)
Immun blot pvdf membrane, by Bio-Rad (1 mentions)
2 protocols using sds page tris hcl gels
1
Western Blot Quantitative Analysis
2
Western Blot Analysis of SIRT4 Expression
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Total protein was extracted from the cell lines in radioimmunoprecipitation assay buffer (Thermo Fisher Scientific, Waltham, MA, USA). Aliquots of protein were electrophoresed on SDS–PAGE Tris-HCl gels (Bio-Rad, Hercules, CA, USA). The separated proteins were transferred to Immun-Blot PVDF membrane (Bio-Rad) using a wet transfer system (Bio-Rad) and incubated with SIRT4 antibody (ab105039; Abcam, Cambridge, MA, USA; 1 : 1000 dilution) or ACTB antibody (A2066; Sigma Aldrich; 1 : 2000 dilution) at 4 °C overnight, followed by incubation with HRP-linked anti-rabbit IgG (GE Healthcare Biosciences, Piscataway, NJ, USA) in a dilution of 1 : 100 000 for 1 h at room temperature. The antigen–antibody complex was detected with the ECL Prime Western Blotting Detection Kit (GE Healthcare Biosciences).
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