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Mouse Breast cancer type 1 susceptibility protein homolog (Brca1) ELISA Kit (MOEB0673)

SKU:
MOEB0673
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P48754
ELISA Type:
Sandwich
Reactivity:
Mouse
€649
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Description

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Mouse Breast cancer type 1 susceptibility protein homolog (Brca1) ELISA Kit

The Mouse Breast Cancer Type 1 Susceptibility Protein Homolog (BRCA1) ELISA Kit is specially designed for the accurate and efficient detection of BRCA1 levels in mouse serum, plasma, and tissue samples. This kit offers high sensitivity and specificity, ensuring precise and reproducible results for a variety of research applications.BRCA1 is a key protein involved in the repair of DNA damage and the maintenance of genomic stability. Mutations in the BRCA1 gene are associated with an increased risk of developing breast and ovarian cancer in humans.

Studying the expression levels of BRCA1 in mouse models can provide valuable insights into the mechanisms of tumorigenesis and potential therapeutic targets for cancer treatment.With this ELISA kit, researchers can easily quantify BRCA1 levels in mouse samples, allowing for a better understanding of its role in cancer development and progression. This kit is an essential tool for cancer research and drug development studies targeting BRCA1-related pathways.

Product Name:Mouse Breast cancer type 1 susceptibility protein homolog (Brca1) ELISA Kit
SKU:MOEB0673
Size:96T
Target:Mouse Breast cancer type 1 susceptibility protein homolog (Brca1)
Synonyms:RING-type E3 ubiquitin transferase BRCA1
Assay Type:Sandwich
Detection Method:ELISA
Reactivity:Mouse
Detection Range:0.312-20ng/mL
Sensitivity:0.072ng/mL
Intra CV:Provided with the Kit
Inter CV:Provided with the Kit
Linearity:Provided with the Kit
Recovery:Provided with the Kit
Function:E3 ubiquitin-protein ligase that specifically mediates the formation of 'Lys-6'-linked polyubiquitin chains and plays a central role in DNA repair by facilitating cellular responses to DNA damage. It is unclear whether it also mediates the formation of other types of polyubiquitin chains. The E3 ubiquitin-protein ligase activity is required for its tumor suppressor function. The BRCA1-BARD1 heterodimer coordinates a diverse range of cellular pathways such as DNA damage repair, ubiquitination and transcriptional regulation to maintain genomic stability. Regulates centrosomal microtubule nucleation. Required for normal cell cycle progression from G2 to mitosis. Required for appropriate cell cycle arrests after ionizing irradiation in both the S-phase and the G2 phase of the cell cycle. Involved in transcriptional regulation of P21 in response to DNA damage. Required for FANCD2 targeting to sites of DNA damage. May function as a transcriptional regulator. Contributes to homologous recombination repair (HRR) via its direct interaction with PALB2, fine-tunes recombinational repair partly through its modulatory role in the PALB2-dependent loading of BRCA2-RAD51 repair machinery at DNA breaks. Component of the BRCA1-RBBP8 complex which regulates CHEK1 activation and controls cell cycle G2/M checkpoints on DNA damage via BRCA1-mediated ubiquitination of RBBP8. Acts as a transcriptional activator (By similarity). Inhibits lipid synthesis by binding to inactive phosphorylated ACACA and preventing its dephosphorylation.
Uniprot:P48754
Sample Type:Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:Natural and recombinant mouse Breast cancer type 1 susceptibility protein homolog
Sub Unit:Heterodimer with BARD1. Part of the BRCA1-associated genome surveillance complex (BASC), which contains BRCA1, MSH2, MSH6, MLH1, ATM, BLM, PMS2 and the MRE11-RAD50-NBN protein (MRN) complex. This association could be a dynamic process changing throughout the cell cycle and within subnuclear domains. Component of the BRCA1-A complex, at least composed of BRCA1, BARD1, UIMC1/RAP80, FAM175A/Abraxas, BRCC3/BRCC36, BRE/BRCC45 and BABAM1/NBA1. Interacts (via the BRCT domains) with FAM175A (phosphorylated form); this is important for recruitment to sites of DNA damage. Can form a heterotetramer with two molecules of FAM175A (phosphorylated form). Component of the BRCA1-RBBP8 complex. Interacts (via the BRCT domains) with RBBP8 ('Ser-327' phosphorylated form); the interaction ubiquitinates RBBP8, regulates CHEK1 activation, and involves RBBP8 in BRCA1-dependent G2/M checkpoint control on DNA damage. Associates with RNA polymerase II holoenzyme. Interacts with SMC1A, COBRA1, DCLRE1C, CLSPN. CHEK1, CHEK2, BAP1, BRCC3, AURKA, UBXN1 and PCLAF. Interacts (via BRCT domains) with BRIP1 (phosphorylated form). Interacts with FANCD2 (ubiquitinated form). Interacts with H2AFX (phosphorylated on 'Ser-140'). Interacts (via the BRCT domains) with ACACA (phosphorylated form); the interaction prevents dephosphorylation of ACACA. Part of a BRCA complex containing BRCA1, BRCA2 and PALB2. Interacts directly with PALB2; the interaction is essential for its function in HRR. Interacts directly with BRCA2; the interaction occurs only in the presence of PALB2 which serves as the bridging protein. Interacts (via the BRCT domains) with LMO4; the interaction represses the transcriptional activity of BRCA1. Interacts (via the BRCT domains) with CCAR2 (via N-terminus); the interaction represses the transcriptional activator activity of BRCA1 (By similarity). Interacts with EXD2.
Research Area:Cancer
Subcellular Location:Nucleus Chromosome Cytoplasm Localizes at sites of DNA damage at double-strand breaks (DSBs); recruitment to DNA damage sites is mediated by the BRCA1-A complex. Translocated to the cytoplasm during UV-induced apoptosis.
Storage:Please see kit components below for exact storage details
Note:For research use only
UniProt Protein Function:BRCA1: the BRCA1-BARD1 heterodimer coordinates a diverse range of cellular pathways such as DNA damage repair, ubiquitination and transcriptional regulation to maintain genomic stability. Acts by mediating ubiquitin E3 ligase activity that is required for its tumor suppressor function. Plays a central role in DNA repair by facilitating cellular response to DNA repair. Required for appropriate cell cycle arrests after ionizing irradiation in both the S-phase and the G2 phase of the cell cycle. Involved in transcriptional regulation of P21 in response to DNA damage. Required for FANCD2 targeting to sites of DNA damage. May function as a transcriptional regulator. Inhibits lipid synthesis by binding to inactive phosphorylated ACC1 and preventing its dephosphorylation. Defects in BRCA1 are a cause of genetic susceptibility to breast cancer. Mutations in BRCA1 are thought to be responsible for more than 80% of inherited breast-ovarian cancer. Part of the BRCA1-associated genome surveillance complex (BASC), which contains BRCA1, MSH2, MSH6, MLH1, ATM, BLM, PMS2 and the RAD50-MRE11-NBN protein complex. This association may be a dynamic process changing throughout the cell cycle and within subnuclear domains. Interacts (via BRCT domains) with CTIP. Associates with RNA polymerase II holoenzyme. Interacts with SMC1 and COBRA1. Interacts (via BRCT domains) with BRIP1. Interacts with FANCD2 (ubiquitinated). Interacts with BAP1. Interacts with Artemis and claspin. Interacts with H2AFX (phosphorylated on S140). Interacts with CHK1. Interacts with BRCC3. Five human isoforms are produced by alternative splicing and alternative initiation. Isoform 1 and isoform 3 are widely expressed. Isoform 1 is largely nuclear, while isoforms 3 and 5 are cytoplasmic.Protein type: Ubiquitin ligase; Ubiquitin conjugating system; Transcription, coactivator/corepressor; Tumor suppressor; EC 6.3.2.-; Nuclear receptor co-regulatorCellular Component: ruffle; centrosome; protein complex; condensed nuclear chromosome; mitochondrial matrix; BRCA1-BARD1 complex; cytoplasm; plasma membrane; condensed chromosome; chromosome; nucleus; ribonucleoprotein complexMolecular Function: enzyme binding; DNA binding; zinc ion binding; coenzyme F420-2 alpha-glutamyl ligase activity; RNA binding; metal ion binding; UDP-N-acetylmuramoylalanyl-D-glutamyl-2,6-diaminopimelate-D-alanyl-D-alanine ligase activity; ubiquitin protein ligase binding; ubiquitin-protein ligase activity; ribosomal S6-glutamic acid ligase activity; damaged DNA binding; coenzyme F420-0 gamma-glutamyl ligase activity; ligase activityBiological Process: genetic imprinting; dosage compensation, by inactivation of X chromosome; centrosome cycle; protein ubiquitination; negative regulation of histone H3-K4 methylation; positive regulation of histone H3-K4 methylation; fatty acid metabolic process; chromosome segregation; negative regulation of histone acetylation; DNA recombination; double-strand break repair; DNA replication; fatty acid biosynthetic process; negative regulation of fatty acid biosynthetic process; positive regulation of histone H3-K9 methylation; negative regulation of histone H3-K9 methylation; protein autoubiquitination; positive regulation of DNA repair; chordate embryonic development; postreplication repair; cell cycle; DNA repair; double-strand break repair via homologous recombination; regulation of cell proliferation; positive regulation of angiogenesis; DNA damage response, signal transduction resulting in induction of apoptosis; positive regulation of protein ubiquitination; response to estrogen stimulus; positive regulation of histone acetylation; positive regulation of transcription from RNA polymerase II promoter; response to ionizing radiation; lipid metabolic process; G2/M transition DNA damage checkpoint; negative regulation of transcription, DNA-dependent; centrosome duplication; response to DNA damage stimulus
UniProt Protein Details:
NCBI Summary:
UniProt Code:P48754
NCBI GenInfo Identifier:161016835
NCBI Gene ID:12189
NCBI Accession:NP_033894.3
UniProt Secondary Accession:P48754,Q60957, Q60983, A2A4Q4
UniProt Related Accession:P48754
Molecular Weight:198,795 Da
NCBI Full Name:breast cancer type 1 susceptibility protein homolog
NCBI Synonym Full Names:breast cancer 1
NCBI Official Symbol:Brca1
NCBI Official Synonym Symbols:
NCBI Protein Information:breast cancer type 1 susceptibility protein homolog
UniProt Protein Name:Breast cancer type 1 susceptibility protein homolog
UniProt Synonym Protein Names:
Protein Family:BRCA1-A complex
UniProt Gene Name:Brca1
UniProt Entry Name:BRCA1_MOUSE
Component Quantity (96 Assays) Storage
ELISA Microplate (Dismountable) 8×12 strips -20°C
Lyophilized Standard 2 -20°C
Sample Diluent 20ml -20°C
Assay Diluent A 10mL -20°C
Assay Diluent B 10mL -20°C
Detection Reagent A 120µL -20°C
Detection Reagent B 120µL -20°C
Wash Buffer 30mL 4°C
Substrate 10mL 4°C
Stop Solution 10mL 4°C
Plate Sealer 5 -

Other materials and equipment required:

  • Microplate reader with 450 nm wavelength filter
  • Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
  • Incubator
  • Deionized or distilled water
  • Absorbent paper
  • Buffer resevoir

*Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

Allow all reagents to reach room temperature (Please do not dissolve the reagents at 37°C directly). All the reagents should be mixed thoroughly by gently swirling before pipetting. Avoid foaming. Keep appropriate numbers of strips for 1 experiment and remove extra strips from microtiter plate. Removed strips should be resealed and stored at -20°C until the kits expiry date. Prepare all reagents, working standards and samples as directed in the previous sections. Please predict the concentration before assaying. If values for these are not within the range of the standard curve, users must determine the optimal sample dilutions for their experiments. We recommend running all samples in duplicate.

Step
1. Add Sample: Add 100µL of Standard, Blank, or Sample per well. The blank well is added with Sample diluent. Solutions are added to the bottom of micro ELISA plate well, avoid inside wall touching and foaming as possible. Mix it gently. Cover the plate with sealer we provided. Incubate for 120 minutes at 37°C.
2. Remove the liquid from each well, don't wash. Add 100µL of Detection Reagent A working solution to each well. Cover with the Plate sealer. Gently tap the plate to ensure thorough mixing. Incubate for 1 hour at 37°C. Note: if Detection Reagent A appears cloudy warm to room temperature until solution is uniform.
3. Aspirate each well and wash, repeating the process three times. Wash by filling each well with Wash Buffer (approximately 400µL) (a squirt bottle, multi-channel pipette,manifold dispenser or automated washer are needed). Complete removal of liquid at each step is essential. After the last wash, completely remove remaining Wash Buffer by aspirating or decanting. Invert the plate and pat it against thick clean absorbent paper.
4. Add 100µL of Detection Reagent B working solution to each well. Cover with the Plate sealer. Incubate for 60 minutes at 37°C.
5. Repeat the wash process for five times as conducted in step 3.
6. Add 90µL of Substrate Solution to each well. Cover with a new Plate sealer and incubate for 10-20 minutes at 37°C. Protect the plate from light. The reaction time can be shortened or extended according to the actual color change, but this should not exceed more than 30 minutes. When apparent gradient appears in standard wells, user should terminatethe reaction.
7. Add 50µL of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
8. Determine the optical density (OD value) of each well at once, using a micro-plate reader set to 450 nm. User should open the micro-plate reader in advance, preheat the instrument, and set the testing parameters.
9. After experiment, store all reagents according to the specified storage temperature respectively until their expiry.

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample Type Protocol
Serum If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.
Plasma Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 × g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.
Urine & Cerebrospinal Fluid Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.
Cell culture supernatant Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.
Cell lysates Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Tissue homogenates The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.
Tissue lysates Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.
Breast Milk Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

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