A549, HepG2, HAP1, and 634T WT, ACLY and/or ACSS2-KO cell lines were lysed in mammalian protein extraction reagent (M-PER) buffer (Thermo Fisher Scientific, 78501) with 1× protease inhibitor (Sigma-Aldrich). Protein concentrations were determined using a Pierce BCA protein assay kit (Thermo Fisher Scientific, 23225). Fifteen micrograms of protein was loaded and separated using 4 to 15% Mini-PROTEAN TGX Precast SDS-PAGE Gels (Bio-Rad, #4561086). Samples were then transferred to nitrocellulose membranes for immunoblotting. Total acetylated-lysine (Cell Signaling Technology, 9441), β-actin (Sigma-Aldrich, A5441), ACLY (Cell Signaling Technology, 13390), ACSS2 (Cell Signaling Technology, 3658), and PEX5(D7V4D) (Cell Signaling Technology, 83020) antibodies were used to probe their respective targets. Anti-rabbit horseradish peroxidase–conjugated secondary antibodies (Millipore, AP132P) were used for imaging.
Acss2
Manufactured by Thermo Fisher Scientific
The ACSS2 is a laboratory instrument designed for conducting Accelerated Solvent Extraction (ASE) procedures. It is capable of performing automated extraction of organic compounds from solid and semi-solid samples using elevated temperatures and pressures. The ACSS2 allows for the efficient and consistent extraction of analytes from a wide range of sample matrices.
Automatically generated - may contain errors
Lab products found in correlation
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Acss2, by Cell Signaling Technology (1 mentions)
Protease inhibitor, by Merck Group (1 mentions)
Ap132p, by Merck Group (1 mentions)
M per buffer, by Thermo Fisher Scientific (1 mentions)
Mini protean tgx precast sds page gel, by Bio-Rad (1 mentions)
Total acetylated lysine, by Cell Signaling Technology (1 mentions)
β actin, by Merck Group (1 mentions)
Enhanced chemiluminescence reagent, by Bio-Rad (1 mentions)
Protease and phosphatase inhibitor cocktail, by GenDEPOT (1 mentions)
β actin, by Santa Cruz Biotechnology (1 mentions)
2 protocols using acss2
1
Acetylome Analysis of ACLY and ACSS2 KOs
2
Western Blot Analysis of ACSS2
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The cells were lysed with radioimmunoprecipitation assay (RIPA) buffer (P3200-010; Biosesang, Gyeonggi-do, Korea) containing protease and phosphatase inhibitor cocktails (GenDepot, Barker, TX, USA). The proteins were resolved by sodium dodecyl sulfate‒polyacrylamide gel electrophoresis (SDS‒PAGE) and transferred onto nitrocellulose membranes. The blots were immunostained with appropriate primary antibodies, followed by incubation with secondary antibodies conjugated to horseradish peroxidase. Antibody binding was detected with an enhanced chemiluminescence reagent (Bio-Rad. Hercules, CA, USA). Primary antibodies against the following molecules were used: ACSS2 (Thermo Fisher Scientific) and β-actin (Santa Cruz Biotechnology). The secondary antibodies were purchased from Santa Cruz Biotechnology.
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