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BRCA2 ELISA 试剂盒

BRCA2 适用: 人 Colorimetric Sandwich ELISA 0.156-10 ng/mL Tissue Homogenate
产品编号 ABIN578767
发货至: 中国
  • 抗原 See all BRCA2 ELISA试剂盒
    BRCA2 (Breast Cancer 2, Early Onset (BRCA2))
    适用
    • 3
    • 2
    • 2
    • 1
    • 1
    • 1
    • 1
    • 1
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    检测方法
    Colorimetric
    实验类型
    Sandwich ELISA
    检测范围
    0.156-10 ng/mL
    最低检测浓度
    0.156 ng/mL
    应用范围
    ELISA
    原理
    This immunoassay kit allows for the in vitro quantitative determination of human breast cancer susceptibility protein 1, BRCA-2 concentrations in tissue homogenates and other biological fluids.
    样品类型
    Tissue Homogenate
    Analytical Method
    Quantitative
    特异性
    This assay recognizes recombinant and natural human BRCA-2.
    交叉反应 (详细)
    No significant cross-reactivity or interference was observed.
    灵敏度
    The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest detectable concentration that could be differentiated from zero.
    产品特性
    Homo sapiens,Human,Breast cancer type 2 susceptibility protein,Fanconi anemia group D1 protein,BRCA2,FACD,FANCD1
    组件
    Reagent (Quantity ): Assay plate (1), Standard (2), Sample Diluent (1 × 20ml), Assay Diluent A (1x10ml), Assay Diluent B (1x10ml), Detection Reagent A (1 × 120μl), Detection Reagent B (1 × 120μl), Wash Buffer (25 x concentrate) (1 × 30ml), Substrate (1x10ml), Stop Solution (1x10ml), Plate sealer for 96 wells (5), Instruction (1)
    试剂未包括
    Luminometer. Pipettes and pipette tips. EP tube Deionized or distilled water.
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  • 样本量
    100 μL
    板类型
    Pre-coated
    实验流程
    The microtiter plate provided in this kit has been pre-coated with an antibody specific to BRCA-2. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated polyclonal antibody preparation specific for BRCA-2 and Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. Then a TMB substrate solution is added to each well. Only those wells that contain BRCA-2, biotin-conjugated antibody and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of BRCA-2 in the samples is then determined by comparing the O.D. of the samples to the standard curve.
    试剂准备

    Bring all reagents to room temperature before use. Wash Buffer - If crystals have formed in the concentrate, warm to room temperature and mix gently until the crystals have completely dissolved. Dilute 30 mL of Wash Buffer Concentrate into deionized or distilled water to prepare 750 mL of Wash Buffer. Standard - Reconstitute the Standard with 0.5 mL of Sample Diluent. This reconstitution produces a stock solution of 10 ng/ml. Allow the standard to sit for a minimum of 15 minutes with gentle agitation prior to making serial dilutions (Making serial dilution in the wells directly is not permitted). The undiluted standard serves as the high standard (10 ng/ml). The Sample Diluent serves as the zero standard (0 ng/ml). ng/mL 10 5 2.5 1.25 0.625 0.312 0.156 0 Detection Reagent A and B - Dilute to the working concentration using Assay Diluent A and B (1:100), respectively.

    样品收集
    Tissue homogenates - The preparation of tissue homogenates will vary depending upon tissue type. For this assay, tissue was rinsed with 1X PBS to remove excess blood, homogenized in 5~10 mL of 1X PBS and stored overnight at ≤ -20 °C. After two freeze-thaw cycles were performed to break the cell membranes, the homogenates were centrifuged for 5 minutes at 5000 x g. Remove the supernate and assay immediately or aliquot and store at ≤ -20 °C. Other biological fluids - Remove particulates by centrifugation and assay immediately or aliquot and store samples at -20 C or -80 C . Avoid repeated freeze-thaw cycles. Note: Tissue homogenates to be used within 7 days may be stored at 2-8 C, otherwise samples must stored at -20 C ( ≤ 1 months) or -80 C ( ≤ 2 months) to avoid loss of 3 bioactivity and contamination. Avoid freeze-thaw cycles. When performing the assay slowly bring samples to room temperature.
    实验流程

    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 4 C until the kits expiry date. Prepare all reagents, working standards and samples as 4 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 particular experiments.
    1. Add 100 μ l of Standard, Blank, or Sample per well. Cover with the Plate sealer. Incubate for 2 hours at 37 C .
    2. Remove the liquid of each well, don ’ t wash.
    3. Add 100 μ l of Detection Reagent A working solution to each well. Cover with the Plate sealer. Incubate for 1 hour at 37 °C. Detection Reagent A working solution may appear cloudy. Warm to room temperature and mix gently until solution appears uniform.
    4. Aspirate each well and wash, repeating the process three times for a total of three washes. Wash by filling each well with Wash Buffer (approximately 400 μ l) using a squirt bottle, multi-channel pipette, manifold dispenser or autowasher. Complete removal of liquid at each step is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating or decanting. Invert the plate and blot it against clean paper towels.
    5. Add 100 μ l of Detection Reagent B working solution to each well. Cover with a new Plate sealer. Incubate for 1 hours at 37 C .
    6. Repeat the aspiration/wash five times as in step
    4. 7. Add 90 μ l of Substrate Solution to each well. Cover with a new Plate sealer. Incubate within 30 minutes at 37 °C. Protect from light.
    8. Add 50 μ l of Stop Solution to each well. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
    9. Determine the optical density of each well at once, using a microplate reader set to 450 nm.
    Important Note:1. Absorbance is a function of the incubation time. Therefore, prior to starting the assay it is recommended that all reagents should be freshly prepared prior to use and all required strip-wells are secured in the microtiter frame. This will ensure equal elapsed time for each pipetting step, without interruption.
    2. Please carefully reconstitute Standards or working Detection Reagent A and B according to the instruction, and avoid foaming and mix gently until the crystals have completely dissolved. The reconstituted Standards can be used only once. This assay requires pipetting of small volumes. To minimize imprecision caused by pipetting, ensure that pipettors are calibrated. It is recommended to suck more than 10 μ l for once pipetting.
    3. To ensure accurate results, proper adhesion of plate sealers during incubation steps is necessary. Do not allow wells to sit uncovered for extended periods between incubation steps. Once reagents have been added to the well strips, DO NOT let the strips DRY at any time during the assay.
    4. For each step in the procedure, total dispensing time for addition of reagents to the assay plate should not exceed 10 minutes. 5
    5. To avoid cross-contamination, change pipette tips between additions of each standard level, between sample additions, and between reagent additions. Also, use separate reservoirs for each reagent.
    6. The wash procedure is critical. Insufficient washing will result in poor precision and falsely elevated absorbance readings.
    7. Duplication of all standards and specimens, although not required, is recommended.
    8. Substrate Solution is easily contaminated. Please protect it from light.

    结果分析

    Average the duplicate readings for each standard, control, and sample and subtract the average zero standard optical density. Create a standard curve by reducing the data using computer software capable of generating a four parameter logistic (4-PL) curve-fit. As an alternative, construct a standard curve by plotting the mean absorbance for each standard on the x-axis against the concentration on the y-axis and draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the BRCA-2 concentrations versus the log of the O.D. and the best fit line can be determined by regression analysis. It is recommended to use some related software to do this calculation, such as curve expert 13.0. This procedure will produce an adequate but less precise fit of the data. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.

    限制
    仅限研究用
  • 注意事项
    1. The kit should not be used beyond the expiration date on the kit label.
    2. Do not mix or substitute reagents with those from other lots or sources.
    3. If samples generate values higher than the highest standard, further dilute the samples and repeat the assay. Any variation in standard diluent, operator, pipetting technique, washing technique,incubation time or temperature, and kit age can cause variation in binding.
    4. This assay is designed to eliminate interference by soluble receptors, ligands, binding proteins, and other factors present in biological samples. Until all factors have been tested in the Immunoassay, the possibility of interference cannot be excluded.
    储存条件
    4 °C/-20 °C
    储存方法
    The Standard, Detection Reagent A, Detection Reagent B and the 96-well strip plate should be stored at -20 °C upon being received. The other reagents can be stored at 4 °C.
  • 抗原 See all BRCA2 ELISA试剂盒
    BRCA2 (Breast Cancer 2, Early Onset (BRCA2))
    别名
    BRCA2 (BRCA2 产品)
    别名
    BRCC2 ELISA Kit, BROVCA2 ELISA Kit, FACD ELISA Kit, FAD ELISA Kit, FAD1 ELISA Kit, FANCB ELISA Kit, FANCD ELISA Kit, FANCD1 ELISA Kit, GLM3 ELISA Kit, PNCA2 ELISA Kit, 30169 ELISA Kit, BcDNA:SD25109 ELISA Kit, CG13583 ELISA Kit, CG13584 ELISA Kit, CG30169 ELISA Kit, Dmel\\CG30169 ELISA Kit, brca2 ELISA Kit, dmbrca2 ELISA Kit, fancd1 ELISA Kit, BRCA2 ELISA Kit, brcc2 ELISA Kit, brovca2 ELISA Kit, facd ELISA Kit, fad ELISA Kit, fad1 ELISA Kit, fancb ELISA Kit, AI256696 ELISA Kit, AW045498 ELISA Kit, Fancd1 ELISA Kit, RAB163 ELISA Kit, BRCA2, DNA repair associated ELISA Kit, Breast cancer 2, early onset homolog ELISA Kit, breast cancer 2, early onset ELISA Kit, BRCA2 ELISA Kit, Brca2 ELISA Kit, brca2 ELISA Kit, LOC584780 ELISA Kit
    背景
    BRCA2 (Breast Cancer Type 2 susceptibility protein) is a human gene that is involved in the repair of chromosomal damage and belongs to a class of genes known as tumor suppressor genes. Tumor suppressor genes regulate the cycle of cell division by keeping cells from growing and dividing too rapidly or in an uncontrolled way. Although the structures of the BRCA1 and BRCA2 genes are very different, their functions appear to be similar. The proteins made by both genes are essential for repairing damaged DNA. The BRCA2 protein binds to and regulates the protein produced by the RAD51 gene to fix breaks in DNA. These breaks can be caused by natural and medical radiation or other environmental exposures, but also occur when chromosomes exchange genetic material during a special type of cell division that creates sperm and eggs (meiosis). The BRCA1 protein also interacts with the RAD51 protein. By repairing DNA, these three proteins play a role in maintaining the stability of the human genome. Like BRCA1, BRCA2 probably regulates the activity of other genes and plays a critical role in embryo development.The BRCA2 gene is located on the long (q) arm of chromosome 13 at position 12.3, from base pair 31,787,616 to base pair 31,871,804. Certain variations of the BRCA2 gene cause an increased risk for breast cancer. Researchers have identified about 450 mutations in the BRCA2 gene, many of which cause an increased risk of cancer. BRCA2 mutations are usually insertions or deletions of a small number of DNA base pairs (the building material of chromosomes) in the gene. As a result of these mutations, the protein product of the BRCA2 gene is abnormally short and does not function properly. Researchers believe that the defective BRCA2 protein is unable to help fix mutations that occur in other genes. As a result, mutations build up and can cause cells to divide in an uncontrolled way and form a tumor. 2
    基因ID
    3026
    途径
    DNA Damage Repair, M Phase, Maintenance of Protein Location
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