p53 Assay Kit [Human]

This Human Tumor Protein p53 (p53) assay kit is an all-inclusive luciferase reporter assay system that includes, In addition to p53 Reporter Cells, this kit provides two optimized media for use during cell culture and in diluting test samples, the reference p53 activator Doxorubicin (hydrochloride), Luciferase Detection Reagent, two cell culture-ready assay plates, and a detailed protocol. p53 Reporter Cells are prepared using INDIGO's proprietary CryoMite(TM) process. This cryo-preservation method yields high cell viability post-thaw, and provides the convenience of immediately dispensing healthy, division-competent reporter cells into assay plates. There is no need for intermediate spin-and-wash steps, viability determinations, or cell titer adjustments prior to assay setup. INDIGO's p53 assay kits feature a luciferase detection reagent specially formulated to provide stable light emission between 5 and 90+ minutes after initiating the luciferase reaction. Incorporating a 5-minute reaction-rest period ensures that light emission profiles attain maximal stability, thereby allowing assay plates to be processed in batch. By doing so, the signal output from all sample wells, from one plate to the next, may be directly compared within an experimental set. Tumor protein P53, also known as p53, acquired its reputation as "guardian of the genome" through its ability to sense and respond to cellular stress, preventing accumulation of DNA damage and subsequent formation of malignancies. p53 is a transcription factor and tumor suppressor that responds to cellular stress by regulating genes involved in a diverse array of cellular responses, including but not limited to, cell cycle arrest, DNA repair, apoptosis, senescence, and autophagy, thus minimizing the negative consequences of genetic mutation. p53 is expressed at low levels in cells in the absence of stress, regulated by various factors including MDM2 (also known as HDM2), which acts as a negative regulator through its E3 ubiquitin ligase activity, and WIP1/PPM1D, which also acts as a negative regulator of p53 through its ability to dephosphorylate specific residues on both p53 and MDM2, leading to the destabilization of p53 and the stabilization of HDM2, respectively. p53 in turn directly influences the expression of these two negative regulators, as both MDM2 and WIP1/PPM1D are target genes of p53. p53 is the most commonly mutated gene in human cancer formation, and is therefore of interest in cancer research and therapeutic development. Potential therapeutics may target the p53 pathway in several ways. These include restoration of mutated p53 to its wild-type conformation and targeting regulators of p53 activity, for example, by inhibiting MDM2. The small molecule PRIMA-1, as well as derivates of the thiosemicarbazone family, have been shown to restore wild-type p53 activity, and several classes of MDM2 inhibitors, including nutlins, have been developed and studied. Since the causes, results, and regulation of p53 activation are diverse and context-dependent, therapeutics targeting the p53 pathway are necessarily diverse as well. INDIGO's p53 Reporter Cells include the luciferase reporter gene functionally linked to a p53-responsive promoter. Thus, quantifying changes in luciferase expression in the treated reporter cells provides a sensitive surrogate measure of the changes in human p53 activity. Therefore, the principal application of this reporter assay is in the screening of test compounds to quantify any functional activity that they may exert against human p53. Protein function: Acts as a tumor suppressor in many tumor types, induces growth arrest or apoptosis depending on the physiological circumstances and cell type. Involved in cell cycle regulation as a trans-activator that acts to negatively regulate cell division by controlling a set of genes required for this process. One of the activated genes is an inhibitor of cyclin-dependent kinases. Apoptosis induction seems to be mediated either by stimulation of BAX and FAS antigen expression, or by repression of Bcl-2 expression. Its pro-apoptotic activity is activated via its interaction with PPP1R13B/ASPP1 or TP53BP2/ASPP2 (PubMed:12524540). However, this activity is inhibited when the interaction with PPP1R13B/ASPP1 or TP53BP2/ASPP2 is displaced by PPP1R13L/iASPP (PubMed:12524540). In cooperation with mitochondrial PPIF is involved in activating oxidative stress-induced necrosis, the function is largely independent of transcription. Induces the transcription of long intergenic non-coding RNA p21 (lincRNA-p21) and lincRNA-Mkln1. LincRNA-p21 participates in TP53-dependent transcriptional repression leading to apoptosis and seems to have an effect on cell-cycle regulation. Implicated in Notch signaling cross- over. Prevents CDK7 kinase activity when associated to CAK complex in response to DNA damage, thus stopping cell cycle progression. Isoform 2 enhances the transactivation activity of isoform 1 from some but not all TP53-inducible promoters. Isoform 4 suppresses transactivation activity and impairs growth suppression mediated by isoform 1. Isoform 7 inhibits isoform 1-mediated apoptosis. Regulates the circadian clock by repressing CLOCK-ARNTL/BMAL1-mediated transcriptional activation of PER2 (PubMed:24051492). [The UniProt Consortium]