Human BRCA2 (Breast Cancer Susceptibility Protein 2) CLIA Kit (HUES00402)
- Product type:
- ELISA Kit
- ELISA Type:
|Detection range:||62.50-4000 pg/mL|
|Sample type:||Serum, plasma and other biological fluids|
|Repeatability:||CV < 15%|
|Specificity:||This kit recognizes Human BRCA2 in samples. No significant cross-reactivity or interference between Human BRCA2 and analogues was observed.|
This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Human BRCA2. Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human BRCA2 and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human BRCA2, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Human BRCA2. The concentration of Human BRCA2 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
|UniProt Protein Function:||BRCA2: involved in double-strand break repair and/or homologous recombination. May participate in S phase checkpoint activation. Interacts with RAD51 and DSS1. Interacts with ubiquitinated FANCD2. Interacts with PALB2, enables the recombinational repair and checkpoint functions. Interacts with WDR16. Defects in BRCA2 are a cause of genetic susceptibility to breast cancer and may underlie susceptibility to uveal melanoma.|
|UniProt Protein Details:|
Protein type:Tumor suppressor; Nuclear receptor co-regulator; DNA repair, damage
Chromosomal Location of Human Ortholog: 13q12. 3
Cellular Component: nucleoplasm; centrosome; protein complex; cytoplasm; BRCA2-MAGE-D1 complex; nucleus; secretory granule
Molecular Function:gamma-tubulin binding; protein binding; H4 histone acetyltransferase activity; histone acetyltransferase activity; protease binding; H3 histone acetyltransferase activity; single-stranded DNA binding
Biological Process: positive regulation of transcription, DNA-dependent; cytokinesis; cell aging; positive regulation of mitotic cell cycle; DNA repair; regulation of cytokinesis; oocyte maturation; inner cell mass cell proliferation; negative regulation of mammary gland epithelial cell proliferation; response to UV-C; double-strand break repair via homologous recombination; DNA damage response, signal transduction by p53 class mediator resulting in transcription of p21 class mediator; male meiosis I; nucleotide-excision repair; double-strand break repair; response to gamma radiation; hemopoiesis; spermatogenesis; replication fork protection; brain development; DNA damage response, signal transduction by p53 class mediator resulting in induction of apoptosis; centrosome duplication; female gonad development; response to X-ray
Disease: Pancreatic Cancer, Susceptibility To, 2; Breast-ovarian Cancer, Familial, Susceptibility To, 2; Prostate Cancer; Breast Cancer; Medulloblastoma; Glioma Susceptibility 3; Tracheoesophageal Fistula With Or Without Esophageal Atresia; Fanconi Anemia, Complementation Group D1; Wilms Tumor 1
|NCBI Summary:||Inherited mutations in BRCA1 and this gene, BRCA2, confer increased lifetime risk of developing breast or ovarian cancer. Both BRCA1 and BRCA2 are involved in maintenance of genome stability, specifically the homologous recombination pathway for double-strand DNA repair. The BRCA2 protein contains several copies of a 70 aa motif called the BRC motif, and these motifs mediate binding to the RAD51 recombinase which functions in DNA repair. BRCA2 is considered a tumor suppressor gene, as tumors with BRCA2 mutations generally exhibit loss of heterozygosity (LOH) of the wild-type allele. [provided by RefSeq, Dec 2008]|
|NCBI GenInfo Identifier:||14424438|
|NCBI Gene ID:||675|
|NCBI Accession:||P51587. 2|
|UniProt Secondary Accession:||P51587,O00183, O15008, Q13879, Q5TBJ7,|
|UniProt Related Accession:||P51587|
|NCBI Full Name:||Breast cancer type 2 susceptibility protein|
|NCBI Synonym Full Names:||breast cancer 2, early onset|
|NCBI Official Symbol:||BRCA2|
|NCBI Official Synonym Symbols:||FAD; FACD; FAD1; GLM3; BRCC2; FANCD; PNCA2; FANCD1; XRCC11; BROVCA2|
|NCBI Protein Information:||breast cancer type 2 susceptibility protein; Fanconi anemia group D1 protein; breast cancer 2 tumor suppressor; BRCA1/BRCA2-containing complex, subunit 2; breast and ovarian cancer susceptibility gene, early onset|
|UniProt Protein Name:||Breast cancer type 2 susceptibility protein|
|UniProt Synonym Protein Names:||Fanconi anemia group D1 protein|
|Protein Family:||BRCA2 and CDKN1A-interacting protein|
|UniProt Gene Name:||BRCA2|
|UniProt Entry Name:||BRCA2_HUMAN|
As the RLU values of the standard curve may vary according to the conditions of the actual assay performance (e. g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human BRCA2 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human BRCA2 were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||12.73||9.08||8.15||9.84||9.54||6.58|
The recovery of Human BRCA2 spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||88-99||94|
|Cell culture media (n=5)||90-101||95|
Samples were spiked with high concentrations of Human BRCA2 and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
An unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
|Micro CLIA Plate(Dismountable)||8 wells ×12 strips||-20°C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100×)||1 vial, 120 µL|
|Concentrated HRP Conjugate (100×)||1 vial, 120 µL||-20°C(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4°C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25×)||1 vial, 30 mL|
|Substrate Reagent A||1 vial, 5 mL||4°C (shading light)|
|Substrate Reagent B||1 vial, 5 mL||4°C (shading light)|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
- Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- Aliquot 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids. ) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37°C.
- Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
- Determine the RLU value of each well immediately.