Cell lysates were incubated with anti-TRIM69 (Ab111943; Abcam), anti-p53 (Ab26; Abcam), or IgG at 4 °C for 2 h, then incubated with protein G agarose beads (Roche, Indianapolis, IN, USA) at 4 °C for 1 h. Subsequently, immunoprecipitates were washed three times with Radio Immunoprecipitation Assay (RIPA) lysis buffer, collected by centrifugation, resuspended in SDS-PAGE sample buffer, and analyzed with western blot assays.
Ab111943
Manufactured by Abcam
Sourced in United States
Ab111943 is a laboratory antibody product. It is a primary antibody that can be used for research purposes. The core function of this product is to detect and bind to a specific target in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Protein g agarose beads, by Roche (1 mentions)
Bicinchoninic acid assay kit, by Merck Group (1 mentions)
Ubiquitin, by Abcam (1 mentions)
Ab53287, by Abcam (1 mentions)
Ab32503, by Abcam (1 mentions)
Ab7780, by Abcam (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Bcl 2, by Abcam (1 mentions)
Bcl2 associated x (bax), by Abcam (1 mentions)
Ab32124, by Abcam (1 mentions)
Foxo3a, by Abcam (1 mentions)
3 protocols using ab111943
1
Immunoprecipitation and Western Blot Analysis
2
Protein Expression Analysis of TRIM69 and Apoptosis Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The specimens and HLECs were lysed with RIPA lysis buffer at 4 °C. A bicinchoninic acid assay kit (Sigma Chemical Co., St. Louis, MO) was used for protein quantification. Proteins of each sample (25 μg per sample) were subjected to 10% or 15% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Western blot assays were performed with primary antibodies to the following: TRIM69 (Ab111943; Abcam, Cambridge, MA, USA); p53 (Ab26; Abcam); Bax (Ab32503; Abcam); Bcl-2 (Ab32124; Abcam); Foxo3a (Ab53287; Abcam); ubiquitin (Ab7780; Abcam); and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (#5174; Cell Signaling Technology, Danvers, MA, USA). GAPDH protein was used as the loading control.
3
Protein Isolation and Western Blot Analysis
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For preparation of cell lysates, cell pellets were resuspended in SDS sample buffer (12.5% glycerol, 175 mM Tris-HCl [pH 8.5], 2.5% SDS, 70 mM 2-mercaptoethanol, 0.5% bromophenol blue). Proteins were subsequently separated on NuPage 4% to 12% Bis-Tris polyacrylamide gels and transferred onto nitrocellulose membranes. Blots were probed with either anti-actin (JLA20 hybridoma; courtesy of the Developmental Studies Hybridoma Bank, University of Iowa), one of two anti-TRIM69 antibodies (catalog no. ab111943 [Abcam] or PA5-12215 [Thermo Fisher]), anti-DENV-2 NS3 (PA5-32199; Thermo Fisher), anti-phospho-STAT1 (Tyr701) (58D6), or anti-c-myc (9E10 hybridoma; Developmental Studies Hybridoma Bank, University of Iowa). Thereafter, membranes were probed with DyLight-labeled goat secondary antibodies (Thermo) and scanned using a LiCor Odyssey scanner.
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