Cells were lysed in buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 0.1% SDS, and 1% NP-40) supplemented with protease inhibitors (Roche, Cat. No. 04 693 132 001). Cell extracts were separated by SDS-PAGE and transferred onto a nitrocellulose membrane (Amersham, Cat. No. 10600004). Membranes were then incubated with antibodies against p16 (Abcam, Cat. No. ab51243); p53 (Cat. No. sc-126), E2F1 (Cat. No. sc-251), PA28γ (Cat. No. sc-136025) and ubiquitin (Cat. No. sc-9133) (Santa Cruz Biotechnology); HA (Cell Signaling, Cat. No. 9301S); phosphorylated Rb (Cell signaling, Cat. No. 9301S); Rb (Oncogene, Cat. No. OP77-100UG); HCV core protein (Thermo Fisher Scientific, Cat. No. MA1-080); and γ-tubulin (Sigma, Cat. No. T6557), and subsequently with an appropriate horseradish peroxidase-conjugated secondary antibody: anti-mouse or anti-rabbit IgG (H + L)-HRP (Bio-Rad, Cat. No. 1706516 and 170–6515, respectively). The ECL kit (Advansta, Cat. No. K-12045-D50) was used to visualize the protein bands with the ChemiDoc XRS imaging system (Bio-Rad).
Anti mouse or anti rabbit igg h l hrp
Manufactured by Bio-Rad
Anti-mouse or anti-rabbit IgG (H + L)-HRP is a secondary antibody conjugated with horseradish peroxidase (HRP). It is used to detect and quantify mouse or rabbit primary antibodies in various immunoassays.
Automatically generated - may contain errors
Lab products found in correlation
Chemidoc xrs imaging system, by Bio-Rad (2 mentions)
Nitrocellulose membrane, by Cytiva (1 mentions)
Ecl kit, by Advansta (1 mentions)
Phosphorylated rb, by Cell Signaling Technology (1 mentions)
γ tubulin, by Merck Group (1 mentions)
Protease inhibitor, by Roche (1 mentions)
Nitrocellulose blotting membrane, by GE Healthcare (1 mentions)
Protein assay dye reagent, by Bio-Rad (1 mentions)
Westernbright ecl, by Advansta (1 mentions)
2 protocols using anti mouse or anti rabbit igg h l hrp
1
Western Blot Analysis of Cell Signaling
2
Western Blotting Analysis of Protein Expression
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Cells were lysed in buffer supplemented with protease inhibitors (50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.1% SDS, and 1% NP-40). Protein concentrations in cell lysates were quantified utilizing Protein Assay Dye Reagent (Bio-Rad, Hercules, CA, USA). Cell lysates were subjected to separation via SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred onto a nitrocellulose blotting membrane (GE Healthcare Life Science, Marlborough, MA, USA). Membranes were reacted with primary antibodies targeting HCV Core, DNA methyltransferase 1 (DNMT1) (Abcam, Cambridge, UK), Bax, DNMT3a, DNMT3b, HA, p53 upregulated modulator of apoptosis (PUMA), γ-tubulin, p21, p53 (Santa Cruz Biotechnology), and E6AP (Thermo Fisher Scientific). Subsequently, membranes were incubated with a horseradish peroxidase-conjugated secondary antibody, either anti-mouse or anti-rabbit IgG (H + L)-HRP (Bio-Rad). The protein bands were visualized using WesternBright ECL (Advansta, San Jose, CA, USA) and the ChemiDoc XRS imaging system (Bio-Rad).
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