TF-Detect™ Human P53 Activity Assay Kit enables fast and sensitive detection and quantification of p53 in a 96-well format. Double-stranded oligonucleotides containing a p53 consensus binding site are immobilized in a 96-well plate. The p53 proteins present in nuclear extracts are captured by the immobilized oligonucleotides specifically and then detected by a p53 antibody and a HRP-conjugated secondary antibody. The colorimetric signal generated by HRP substrate TMB can be easily quantified by spectrophotometry.
A purified recombinant human p53 protein is also provided in the kit for use as a protein standard to provide quantitative data for comparing p53 activities of different sample types and/or time points.
- Detects as little as 0.8 ng of human p53 protein
- Purified recombinant human p53 protein included for use as a protein standard to provide quantitative comparison of p53 activities of different sample types and/or time points
- Optimized 96-well format for high-throughput analysis on 96-well plate readers
- Single strip (8-well) assay can also be performed
- 3 and 1/2 hours from preparation to detection
Figure 1. Performance comparison between GeneCopoeia's TF-Detect Human p53 Acitivity Assay kit and a similar competitor product. A human recombinant p53 protein was detected and quantified using both kits.
Figure 2. The activity of p53 proteins from the nuclear extracts of MCF-7 (Left) and HEK293 (Right) cells were detected using the TF-Detect P53 activity assay kit. Both cell types were treated with 0.2mM H2O2 for 3 hours before harvesting. The cell nuclear extracts were prepared following the Preparation of Nuclear Extract protocol in the manual.
Background of tumor suppressor p53
When responding to diverse cellular stresses, tumor suppressor p53 regulates target genes that induce cell cycle arrest, apoptosis, senescence, DNA repair, or changes in metabolism. p53 protein binds to a consensus p53-binding site and activates expression of downstream genes that inhibit growth and/or invasion. Mutations in p53 frequently occur in numerous types of human cancers. p53 mutants lose the tumor suppressor activity by failing to bind to the consensus DNA binding site.
- Wu, J. et al. (2012) MicroRNA-34a inhibits migration and invasion of colon cancer cells via targeting to Fra-1. Carcinogenesis (2012) 33 (3): 519-528. doi: 10.1093/carcin/bgr304. [TF-Detect™ Human p53 activity assay kit].