Human Tumor Protein p53 (p53)
|Product Family||Product Number||Product Description||Technical Manual|
|Human p53 Assay Kit||IB25001-48||Human p53 Assay Kit,
2×48 p53 assays in 96-well format
Human Tumor Protein p53 Assay Kit
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™ 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.
Bulk assay reagents can be custom manufactured to accommodate any scale of HTS. Please inquire.
Human Tumor Protein p53 Assay Services
The primary application of INDIGO’s cell-based receptor assays are to quantitatively assess the bioactivity of a test compound as an agonist (activator) or antagonist (inhibition of an agonist response) of a given receptor. Service assays include a positive control reference compound and ‘vehicle’ control for every experiment. A formal study report and all data files are provided to the client upon completion of the study. To receive a quote for your proposed study, complete & submit the online “Request a Quote” form or contact an INDIGO Customer Service Representative to discuss your desired study parameters.
Human Tumor Protein p53 Background
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.
<Synonyms: TP53, Tumor Protein P53, P53, LFS1, P%# Tumor Suppressor, Tumor Suppressor P53, Cellular Tumor Antigen P53, Phosphoprotein p53
Human Tumor Protein p53 Assay Data
Activation of p53 is demonstrated by treating reporter cells with Doxorubicin hydrochloride (provided), Quinacrine hydrochloride hydrate, (±)-Nutlin-3, Tenovin-1 (all from Cayman Chemical), and Cisplatin (Tocris). Reporter Cells were treated following the ‘~24 hour + ~24 hour’ dosing regimen described in the Technical Manual. Average relative light units (RLU) and corresponding standard deviation (SD) values were determined for each treatment concentration (n = 3). Fold-activation and Z’ values were calculated as described by Zhang, et al. (1999). All graphical manipulations were performed using GraphPad Prism software. The high Z' score confirms the robust performance of this assay.