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|Sample type||Average %||Range|
|Cell culture media||120||115% - 129%|
Human p21 beta SimpleStep ELISA® kit (ab171579) has been re-developed with new capture and detector antibodies. This new kit has the same name but a different product number (ab214568). We have identified new recombinant monoclonal antibodies to use in the SimpleStep ELISA platform that provide higher sensitivity when quantification of p21 in human cell culture extracts and tissue lysates.
p21 in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of p21 protein in human cell and tissue extracts.
The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.
p21 plays a critical role in the cellular response to DNA damage, and its overexpression results in cell cycle arrest. Upregulation of p21 mRNA and protein following ionizing radiation is dependent on p53 (TP53), and p21 mediates cell cycle arrest in response to the p53 checkpoint pathway. p21 binds to and inhibits cyclin-dependent kinase activity, preventing phosphorylation of critical cyclin-dependent kinase substrates and blocking cell cycle progression. p21 functions in the nuclear localization and assembly of cyclin D-CDK4 complex and promotes its kinase activity towards RB1. At higher stoichiometric ratios, p21 inhibits the kinase activity of the cyclin D-CDK4 complex.
|Components||1 x 96 tests|
|10X p21 Capture Antibody||1 x 600µl|
|10X p21 Detector Antibody||1 x 600µl|
|10X Wash Buffer PT (ab206977)||1 x 20ml|
|50X Cell Extraction Enhancer Solution (ab193971)||1 x 1ml|
|5X Cell Extraction Buffer PTR (ab193970)||1 x 10ml|
|ab221825 - Antibody Diluent 5BI||1 x 6ml|
|p21 Human Lyophilized Recombinant Protein||2 vials|
|Plate Seals||1 unit|
|Sample Diluent NS||1 x 12ml|
|SimpleStep Pre-Coated 96-Well Microplate (ab206978)||1 x 96 tests|
|Stop Solution||1 x 12ml|
|TMB Development Solution||1 x 12ml|
Our Abpromise guarantee covers the use of ab214658 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Sandwich ELISA||Use at an assay dependent concentration.|
Background-subtracted data values (mean +/- SD) are graphed.
The concentrations of p21 were measured in duplicates in a two-fold dilution series and interpolated from the p21 standard curve and corrected for sample dilution. The interpolated dilution factor corrected values are plotted (mean +/- SD, n=2). The mean p21 concentration was determined to be 1,811 pg/mL in extract sample of MCF-7 cells treated for 18 hours with 1 mM camptothecin.
Interpolated concentrations of native p21 in human extract samples of HeLa cells, vehicle (DMSO)-treated MCF-7 cells and 18 hours 1 µM camptothecin-treated MCF-7 cells samples. The concentrations of p21 were measured in duplicates at 3 different dilutions, interpolated from the p21 standard curve. Note that Undiluted HeLa cell extract and Undiluted vehicle (DMSO)-treated MCF-7cell extract samples were at 32 g/mL. Note that Undiluted 18 hours 1 M camptothecin-treated MCF-7 cells samples were at 8 µg/mL. The interpolated, dilution factor-corrected values are graphed in pg of p21 per µg of total protein (mean +/- SD, n=2).
ab214658 has not yet been referenced specifically in any publications.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"