Mouse Cyclin Dependent Kinase Inhibitor 2A (CDKN2A) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to CDKN2A. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to CDKN2A. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain CDKN2A, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of CDKN2A in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For Use with serum, plasma, and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

Capable of inducing cell cycle arrest in G1 and G2 phases (PubMed:8521522, PubMed:9393858). Acts as a tumor suppressor (PubMed:8521522, PubMed:9393858, PubMed:15601844, PubMed:17936562). Binds to MDM2 and blocks its nucleocytoplasmic shuttling by sequestering it in the nucleolus (PubMed:9529248, PubMed:10359817). This inhibits the oncogenic action of MDM2 by blocking MDM2-induced degradation of p53 and enhancing p53-dependent transactivation and apoptosis (PubMed:10359817). Also induces G2 arrest and apoptosis in a p53-independent manner by preventing the activation of cyclin B1/CDC2 complexes (PubMed:15361884). Binds to BCL6 and down-regulates BCL6-induced transcriptional repression (PubMed:15567177). Binds to E2F1 and MYC and blocks their transcriptional activator activity but has no effect on MYC transcriptional repression (By similarity). Binds to TOP1/TOPOI and stimulates its activity (By similarity). This complex binds to rRNA gene promoters and may play a role in rRNA transcription and/or maturation (By similarity). Interacts with NPM1/B23 and promotes its polyubiquitination and degradation, thus inhibiting rRNA processing (By similarity). Plays a role in inhibiting ribosome biogenesis, perhaps by binding to the nucleolar localization sequence of transcription termination factor TTF1, and thereby preventing nucleolar localization of TTF1 (PubMed:20513429). Interacts with COMMD1 and promotes its 'Lys63'-linked polyubiquitination (By similarity). Interacts with UBE2I/UBC9 and enhances sumoylation of a number of its binding partners including MDM2 and E2F1 (By similarity). Binds to HUWE1 and represses its ubiquitin ligase activity (By similarity). May play a role in controlling cell proliferation and apoptosis during mammary gland development (By similarity).

GENE ID	12578
SWISS PROT	Q64364
SYNONYMS	P16; ARF; CDK4I; CMM2; INK4; INK4a; MLM; MTS1; TP16; P14; P14ARF; P16INK4a; P19; Melanoma P16; Cyclin-dependent kinase 4 inhibitor A; Multiple tumor suppressor 1

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30 ℅ concentrate)	1 tube
Product data sheet	1 copy

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level CDKN2A were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level CDKN2A were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 0.108ng/mL.
ASSAY RANGE	0.312-20ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of CDKN2A. No significant cross-reactivity or interference between CDKN2A and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between CDKN2A and all analogs, therefore, cross reactivity may still exist.