NHEK were lysed in 2x SDS gel loading buffer (Sigma), heated for 5 min to 95 °C and spun at 16,000 rpm at room temperature for 15 min. Protein lysates were subjected to SDS-gel electrophoresis (Life Technologies) and SDS-gels were immunoblotted. p63 protein variants were detected with the monoclonal p63 antibody 4A4 (Ventana) following separation of proteins in non-gradient 8 % Tris-Glycine SDS-gels. Monoclonal antibodies were used for the detection of γH2A.X (Cell Signaling) and βActin (Sigma), and polyclonal antisera were used for detection of S*66/68 (corresponding to S160/162 in TAp63, Cell Signaling) and for LINE-1 ORF1p. Horseradish peroxidase signals were recorded with X-ray films (Kodak) and quantified with ImageJ (National Institutes of Health).
Tap63
Manufactured by Cell Signaling Technology
Sourced in United States
TAp63 is a protein that functions as a transcription factor, regulating the expression of various genes involved in cellular processes. It is a member of the p63 family of proteins, which play important roles in the development and maintenance of epithelial tissues. The core function of TAp63 is to act as a regulator of gene expression, though its specific roles and mechanisms of action may vary depending on the cellular context.
Automatically generated - may contain errors
Lab products found in correlation
γh2ax, by Cell Signaling Technology (1 mentions)
β actin, by Merck Group (1 mentions)
X ray film, by Kodak (1 mentions)
Hdac3, by Cell Signaling Technology (1 mentions)
Nupage tris hcl precast gels, by Thermo Fisher Scientific (1 mentions)
Chemidoc imaging system, by Bio-Rad (1 mentions)
Ifi44, by Thermo Fisher Scientific (1 mentions)
Hdac1, by Cell Signaling Technology (1 mentions)
Hdac2, by Cell Signaling Technology (1 mentions)
Hdac8, by Cell Signaling Technology (1 mentions)
Dnmt3a, by Cell Signaling Technology (1 mentions)
Dnmt1, by Cell Signaling Technology (1 mentions)
Dnmt3b, by Cell Signaling Technology (1 mentions)
Bcl xl, by Cell Signaling Technology (1 mentions)
T per tissue protein extraction reagent, by Thermo Fisher Scientific (1 mentions)
2 protocols using tap63
1
Immunoblotting of p63 Protein Variants
2
Western Blot Analysis of TNBC PDX Tumors
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About 50 mg of frozen TNBC PDX tumors were used to extract total protein with T-PER Tissue Protein Extraction Reagent (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s guidelines. The protein concentration was evaluated by Bradford Assay. The equal amount of denatured protein extracts was separated through electrophoresis in 4–15% NuPAGE Tris-HCl precast gels (Invitrogen, Waltham, MA, USA) and transferred onto nitrocellulose membranes. Membranes were then probed with primary antibodies including Cd74, Sat1, TAp63, Bcl-xL, Dnmt1, Dnmt3a, Dnmt3b, Tet2, Tet3, Hdac1, Hdac2, Hdac3, Hdac8 (Cell Signaling Technology, Danvers, MA, USA), Tet1, Ifi44, Fzd9, Wwc1 (Thermo Fisher Scientific, Waltham, MA, USA) and NF-κB and Lpl (Santa Cruz, Dallas, TX, USA). β-actin served as an internal control for each membrane. Immunoreactive bands were visualized using Clarity MaxTM Western ECL Blotting Substrates (Bio-Rad, Hercules, CA, USA) and images were captured using ChemiDocTM Imaging Systems (Bio-Rad, Hercules, CA, USA). The protein expression levels were quantified using Image J software (v1.53e).
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