PhosphoTracer p53 pSer15 + Total ELISA Kit (ab119666)
- Product namePhosphoTracer p53 pSer15 + Total ELISA Kit
- Tests1 x 96 well plate
- Sample typeCell culture extracts
- Assay typeSandwich
- Sensitivity20 µg protein/ml
- Range20 µg protein/ml - 2000 µg protein/ml
- Assay time1h 15m
- Species reactivityReacts with: Human
- Product overview
PhosphoTracer assays use a traditional immuno-sandwich format, but with a major difference both the analyte and the assay reagents are added to the PhosphoTracer assay microplate at the same time. After a short incubation period, unbound assay reagents and analytes are washed away, and immuno-complexes containing both antibodies are detected. The process can take as little as 60 minutes to complete.
PhosphoTracer kits also allow a higher degree of assay flexibility. In contrast to other ELISA formats, no antibodies are present on the assay microplate itself, so assays for several different targets can be performed in different wells on the same microplate. Simply mix the lysate with your target reagents of choice, using the microplate configuration of your choice.
A whole new way of performing cellular assays, PhosphoTracer takes the hard work out of running a standard ELISA, while still giving the high quality results expected from a sandwich immunoassay. Fully self-contained kits are supplied in convenient 96-well packs. Simple to use and highly sensitive PhosphoTracer kits are designed to get results, fast.
Abcam's PhosphoTracer p53 assay kits detect endogenous levels of p53 (GenBank Accession NP_000537) in cellular lysates. The phospho-p53 assay detects p53 only when phosphorylated at Ser15. The total p53 assay detects p53 irrespective of phosphorylation status.
The substrate used with the HRP conjugated detection antibody is a combination of 10-Acetyl-3,7-dihydroxyphenoxazine (ADHP) (wavelength exc/em = 530-540nm / 590-600nm), a highly sensitive and stable substrate for HRP) and ADHP Dilution Buffer (a stabilized H2O2 solution). Learn more about the fluorogenic substrate, ADHP.
- Tested applicationsSandwich ELISA more details
- Storage instructionsStore at +4°C. Please refer to protocols.
Components 1 x 96 tests 96-well PhosphoTracer assay plate (stripwell) 1 unit Adherent plate seal 2 units ADHP (100X) 1 x 120µl ADHP Dilution Buffer 1 x 15ml Assay Control Lysate (lyophilized) 1 x 0.25ml Enhancer Solution 1 x 1ml Lysis Buffer (5X) 1 x 15ml Mouse monoclonal p53 (HRP) (24 assay points) 1 x 0.75ml Mouse monoclonal Phospho-p53 (HRP) (72 assay points) 3 x 0.75ml Rabbit polyclonal p53 (24 assay points) 1 x 0.75ml Rabbit polyclonal Phospho-p53 (Ser15) (72 assay points) 3 x 0.75ml Stop Solution 1 x 2ml Wash Buffer (10X) 1 x 15ml
Defects in TP53 are a cause of esophageal cancer (ESCR) [MIM:133239].
Defects in TP53 are a cause of Li-Fraumeni syndrome (LFS) [MIM:151623]. LFS is an autosomal dominant familial cancer syndrome that in its classic form is defined by the existence of a proband affected by a sarcoma before 45 years with a first degree relative affected by any tumor before 45 years and another first degree relative with any tumor before 45 years or a sarcoma at any age. Other clinical definitions for LFS have been proposed (PubMed:8118819 and PubMed:8718514) and called Li-Fraumeni like syndrome (LFL). In these families affected relatives develop a diverse set of malignancies at unusually early ages. Four types of cancers account for 80% of tumors occurring in TP53 germline mutation carriers: breast cancers, soft tissue and bone sarcomas, brain tumors (astrocytomas) and adrenocortical carcinomas. Less frequent tumors include choroid plexus carcinoma or papilloma before the age of 15, rhabdomyosarcoma before the age of 5, leukemia, Wilms tumor, malignant phyllodes tumor, colorectal and gastric cancers.
Defects in TP53 are involved in head and neck squamous cell carcinomas (HNSCC) [MIM:275355]; also known as squamous cell carcinoma of the head and neck.
Defects in TP53 are a cause of lung cancer (LNCR) [MIM:211980].
Defects in TP53 are a cause of choroid plexus papilloma (CPLPA) [MIM:260500]. Choroid plexus papilloma is a slow-growing benign tumor of the choroid plexus that often invades the leptomeninges. In children it is usually in a lateral ventricle but in adults it is more often in the fourth ventricle. Hydrocephalus is common, either from obstruction or from tumor secretion of cerebrospinal fluid. If it undergoes malignant transformation it is called a choroid plexus carcinoma. Primary choroid plexus tumors are rare and usually occur in early childhood.
Defects in TP53 are a cause of adrenocortical carcinoma (ADCC) [MIM:202300]. ADCC is a rare childhood tumor of the adrenal cortex. It occurs with increased frequency in patients with the Beckwith-Wiedemann syndrome and is a component tumor in Li-Fraumeni syndrome.
modificationsAcetylated. Acetylation of Lys-382 by CREBBP enhances transcriptional activity. Deacetylation of Lys-382 by SIRT1 impairs its ability to induce proapoptotic program and modulate cell senescence.
Phosphorylation on Ser residues mediates transcriptional activation. Phosphorylated by HIPK1 (By similarity). Phosphorylation at Ser-9 by HIPK4 increases repression activity on BIRC5 promoter. Phosphorylated on Thr-18 by VRK1. Phosphorylated on Ser-20 by CHEK2 in response to DNA damage, which prevents ubiquitination by MDM2. Phosphorylated on Thr-55 by TAF1, which promotes MDM2-mediated degradation. Phosphorylated on Ser-46 by HIPK2 upon UV irradiation. Phosphorylation on Ser-46 is required for acetylation by CREBBP. Phosphorylated on Ser-392 following UV but not gamma irradiation. Phosphorylated upon DNA damage, probably by ATM or ATR. Phosphorylated on Ser-15 upon ultraviolet irradiation; which is enhanced by interaction with BANP.
Dephosphorylated by PP2A-PPP2R5C holoenzyme at Thr-55. SV40 small T antigen inhibits the dephosphorylation by the AC form of PP2A.
May be O-glycosylated in the C-terminal basic region. Studied in EB-1 cell line.
Ubiquitinated by MDM2 and SYVN1, which leads to proteasomal degradation. Ubiquitinated by RFWD3, which works in cooperation with MDM2 and may catalyze the formation of short polyubiquitin chains on p53/TP53 that are not targeted to the proteasome. Ubiquitinated by MKRN1 at Lys-291 and Lys-292, which leads to proteasomal degradation. Deubiquitinated by USP10, leading to its stabilization. Ubiquitinated by TRIM24, which leads to proteasomal degradation. Ubiquitination by TOPORS induces degradation. Deubiquitination by USP7, leading to stabilization. Isoform 4 is monoubiquitinated in an MDM2-independent manner.
Monomethylated at Lys-372 by SETD7, leading to stabilization and increased transcriptional activation. Monomethylated at Lys-370 by SMYD2, leading to decreased DNA-binding activity and subsequent transcriptional regulation activity. Lys-372 monomethylation prevents interaction with SMYD2 and subsequent monomethylation at Lys-370. Dimethylated at Lys-373 by EHMT1 and EHMT2. Monomethylated at Lys-382 by SETD8, promoting interaction with L3MBTL1 and leading to repress transcriptional activity. Demethylation of dimethylated Lys-370 by KDM1A prevents interaction with TP53BP1 and represses TP53-mediated transcriptional activation.
Sumoylated by SUMO1.
- Antigen NY-CO-13Cellular tumor antigen p53LFS1
- p53 tumor suppressorP53_HUMANPhosphoprotein p53Tp53Transformation related protein 53TRP53Tumor protein p53Tumor suppressor p53
Our Abpromise guarantee covers the use of ab119666 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Sandwich ELISA||sELISA. Tested in U2OS, A431, HEK293.|
PhosphoTracer p53 pSer15 + Total ELISA Kit images
Using p53 assay kits, a significant increase in both p53 phosphorylation at Ser15 and accumulation of total p53, is detected in U-2 OS cells treated with doxorubicin for 18 hours, compared with untreated cells.
Using the PhosphoTracer phospho-p53 assay, the length of assay incubation time that was required to detect recombinant phospho-p53 was examined. After 15 mins, phospho-p53 concentrations of approximately 2 ng/ml were readily detected, while after 2 hour incubation, the limit of detection was approximately 0.2 ng/ml. Longer incubation times had little effect on overall limit of detection that was observed.
U2OS Cells were seeded at 25K cells/well in a 96 well tissue culture plate. The following day, the media was aspirated and replaced with fresh media containing doxorubicin at various concentrations. The plate was incubated at 37°C for a further 18 hours. The media was removed from the wells, and the cells were lysed with 150 µL/well freshly prepared PhosphoTracer Lysis Mix, with shaking for 10 min at room temp. Lysates (50 µL) were transferred to replicate wells of a PhosphoTracer assay plate and analyzed for either phospho-p53, or Total p53 using the standard PhosphoTracer protocol. Briefly, Antibody Mix specific for either phospho-p53, or Total p53 was added to the lysates, and the plate was incubated for 1 hr at room temp, with shaking. The plate was washed and Substrate Mix was added. The plates were covered in foil and incubated for 10 min with shaking. Signal in the wells was determined using a plate reader.
References for PhosphoTracer p53 pSer15 + Total ELISA Kit (ab119666)
ab119666 has not yet been referenced specifically in any publications.