Tissue and cultured cardiomyocytes were lysed in 1X lysis buffer (Cell Signaling Technology), protein concentration was measured by bicinchoninic acid assay (Pierce), and extracts were subjected to SDS-PAGE using Novex Tris-Glycine Gels (4–20% gradient gel, Life Technologies) or Mini-protean TGX Gels (4–20% gradient gel, Bio-Rad), transferred to nitrocellulose membranes, and probed with various primary antibodies from Cell Signaling Technology: CHIP (#2080S), Hsc70 (#8444S), Myc (#2276S), GAPDH (#2118S), and α-tubulin (#3878S), AbMart Inc.: p-CHIP S20 (Custom antibody, Shanghai, China; Project: 25011–1), Sigma: ubiquitin antibody (#SAB4503053), and Li-Cor: fluorescence-labeled secondary antibodies (#926-32211, 926-32210, 926-68023, or 926-68022). Gels were imaged (Odyssey, Li-Cor) and band intensity quantified (Odyssey Software 3.1).
α tubulin
Manufactured by Abmart
Sourced in China, United States
α-tubulin is a structural protein that is a major component of microtubules, which are cytoskeletal elements essential for various cellular processes. It plays a core role in the formation and organization of the cytoskeleton.
Automatically generated - may contain errors
Lab products found in correlation
Novex tris glycine gel, by Thermo Fisher Scientific (1 mentions)
Lysis buffer, by Cell Signaling Technology (1 mentions)
Mini protean tgx gel, by Bio-Rad (1 mentions)
Gapdh, by Abmart (1 mentions)
Pgc 1α, by Merck Group (1 mentions)
P akt, by Cell Signaling Technology (1 mentions)
Fluorochrome conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
Flag tag (flag), by Merck Group (1 mentions)
Srebp1, by Santa Cruz Biotechnology (1 mentions)
P ampk, by Cell Signaling Technology (1 mentions)
Srebp2, by BD (1 mentions)
β actin, by Abmart (1 mentions)
Sirt1, by Merck Group (1 mentions)
Leupeptin, by Roche (1 mentions)
Pepstatin, by Roche (1 mentions)
Aprotinin, by Roche (1 mentions)
Alphaeasefc software, by Bio-Techne (1 mentions)
3 protocols using α tubulin
1
Cardiomyocyte Protein Extraction and Western Blot
2
Immunoblotting and Immunoprecipitation Assays
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Immunoblotting and immunoprecipitation assays were performed as previously described78 (link), with antibodies recognizing CREB, Ser133-P-CREB, AMPK, P-AMPK, AKT, P-AKT (Cell Signaling), PGC-1α (Merck), CRTC2 (Calbiochem), PCK1 (Santa Cruz), SREBP1 (Santa Cruz), SREBP2 (BD), HA (Convance), LXRα (Santa Cruz), FLAG (Sigma), GST (Covance), HIS (Abmart), GAPDH (AOGMA), α-TUBULIN (Abmart), β-ACTIN (Abmart), and other antibodies (Supplementary Table 3 ). For immunostaining assays, primary cultured hepatocytes seeded on coverslips were fixed with 4% paraformaldehyde for 20 min, permeabilized with 0.3% Triton X-100 in PBS, and then blocked by 5% normal goat serum and 0.3% Triton X-100 in PBS, followed by incubation with anti-CREB (1:1000), anti-P-CREB (1:1000), or anti-HA (1:3000) antibodies for 1 h at 4 °C. Slides were then washed, and after washing, slides were incubated with fluorochrome-conjugated secondary antibodies (Invitrogen). Slides were then washed and mounted with Vectashield mounting media containing 4, 6-diamidino-2-phenylindole (DAPI).
3
Neonatal Mouse Cardiomyocyte Protein Extraction
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Neonatal mouse cardiomyocytes were lysed in 100 ml of lysis buffer (50 mmol/L Tris, pH 7.4, 1 mmol/L ethylene diamine tetraacetic acid, 150 mmol/L NaCl, 0.25% sodium deoxycholate, and 1% NP40) containing protease inhibitors (1 mmol/L phenylmethylsulfonyl fluoride, 1 mg/ml aprotinin, 1 mg/ml leupeptin, and 1 mg/ml pepstatin; Roche). Samples were incubated at °C for 1 h and then centrifuged at 12,000 ×g for 30 min, and the supernatant was collected for analysis. Lysates were resolved by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then transferred to poly (vinylidene fluoride) membranes. The membranes were blocked and incubated with specific antibodies against Sirt1 (Millipore, USA), superoxide dismutase (Sod2; Abclonal, USA) and α-tubulin (Abmart, China). Peroxidase-conjugated anti-mouse or anti-rabbit immunoglobulin secondary antibody was used. Proteins were visualized by enhanced chemiluminescence. Densitometry analysis of Western blots was performed using AlphaEaseFC software (Alpha Innotech, USA).
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