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XRCC1 Colorimetric Cell-Based ELISA Kit (CBCAB00909)

SKU:
CBCAB00909
Product Type:
ELISA Kit
ELISA Type:
Cell Based
Research Area:
Epigenetics and Nuclear Signaling
Reactivity:
Human
Mouse
Rat
Detection Method:
Colorimetric
€769

Description

system_update_altDatasheetMSDS

XRCC1 Colorimetric Cell-Based ELISA Kit

The XRCC1 Colorimetric Cell-Based ELISA Kit is a powerful tool for the detection and quantification of XRCC1 protein levels in cell lysates. This kit offers high sensitivity and specificity, providing reliable and reproducible results for a variety of research applications.XRCC1 is a key protein involved in DNA repair pathways, playing a critical role in maintaining genomic stability and preventing mutations. Dysregulation of XRCC1 has been linked to various diseases, including cancer and neurodegenerative disorders, making it a valuable biomarker for studying these conditions and exploring potential therapeutic interventions.

With the XRCC1 Colorimetric Cell-Based ELISA Kit, researchers can accurately measure XRCC1 levels in cell samples, allowing for in-depth analysis of DNA repair processes and their impact on disease development. Trust in this kit to deliver precise and meaningful data for your research needs.

Product Name:XRCC1 Colorimetric Cell-Based ELISA
Product Code:CBCAB00909
ELISA Type:Cell-Based
Target:XRCC1
Reactivity:Human, Mouse, Rat
Dynamic Range:> 5000 Cells
Detection Method:Colorimetric 450 nmStorage/Stability:4°C/6 Months
Format:96-Well Microplate

The XRCC1 Colorimetric Cell-Based ELISA Kit is a convenient, lysate-free, high throughput and sensitive assay kit that can detect XRCC1 protein expression profile in cells. The kit can be used for measuring the relative amounts of XRCC1 in cultured cells as well as screening for the effects that various treatments, inhibitors (ie siRNA or chemicals), or activators have on XRCC1.

Qualitative determination of XRCC1 concentration is achieved by an indirect ELISA format. In essence, XRCC1 is captured by XRCC1-specific primary antibodies while the HRP-conjugated secondary antibodies bind the Fc region of the primary antibody. Through this binding, the HRP enzyme conjugated to the secondary antibody can catalyze a colorimetric reaction upon substrate addition. Due to the qualitative nature of the Cell-Based ELISA, multiple normalization methods are needed:

1. A monoclonal antibody specific for human GAPDH is included to serve as an internal positive control in normalizing the target absorbance values.
2. Following the colorimetric measurement of HRP activity via substrate addition, the Crystal Violet whole-cell staining method may be used to determine cell density. After staining, the results can be analysed by normalizing the absorbance values to cell amounts, by which the plating difference can be adjusted.
Database Information:Gene ID: 7515, UniProt ID: P18887, OMIM: 194360, Unigene: Hs.98493
Gene Symbol:XRCC1
Sub Type:None
UniProt Protein Function:XRCC1: Corrects defective DNA strand-break repair and sister chromatid exchange following treatment with ionizing radiation and alkylating agents. Homodimer. Interacts with polynucleotide kinase (PNK), DNA polymerase-beta (POLB) and DNA ligase III (LIG3). Interacts with APTX and APLF. Interacts with APEX1; the interaction is induced by SIRT1 and increases with the acetylated form of APEX1.
UniProt Protein Details:

Protein type:DNA repair, damage

Chromosomal Location of Human Ortholog: 19q13.2

Cellular Component: nucleoplasm; nucleus

Molecular Function:protein binding; enzyme binding; damaged DNA binding

Biological Process: single strand break repair; base-excision repair; DNA repair

NCBI Summary:The protein encoded by this gene is involved in the efficient repair of DNA single-strand breaks formed by exposure to ionizing radiation and alkylating agents. This protein interacts with DNA ligase III, polymerase beta and poly (ADP-ribose) polymerase to participate in the base excision repair pathway. It may play a role in DNA processing during meiogenesis and recombination in germ cells. A rare microsatellite polymorphism in this gene is associated with cancer in patients of varying radiosensitivity. [provided by RefSeq, Jul 2008]
UniProt Code:P18887
NCBI GenInfo Identifier:317373290
NCBI Gene ID:7515
NCBI Accession:P18887.2
UniProt Secondary Accession:P18887,Q6IBS4, Q9HCB1,
UniProt Related Accession:P18887
Molecular Weight:69,477 Da
NCBI Full Name:DNA repair protein XRCC1
NCBI Synonym Full Names:X-ray repair complementing defective repair in Chinese hamster cells 1
NCBI Official Symbol:XRCC1  
NCBI Official Synonym Symbols:RCC  
NCBI Protein Information:DNA repair protein XRCC1; X-ray repair cross-complementing protein 1
UniProt Protein Name:DNA repair protein XRCC1
UniProt Synonym Protein Names:X-ray repair cross-complementing protein 1
Protein Family:DNA repair protein
UniProt Gene Name:XRCC1  
UniProt Entry Name:XRCC1_HUMAN
ComponentQuantity
96-Well Cell Culture Clear-Bottom Microplate2 plates
10X TBS24 mL
Quenching Buffer24 mL
Blocking Buffer50 mL
15X Wash Buffer50 mL
Primary Antibody Diluent12 mL
100x Anti-Phospho Target Antibody 60 µL
100x Anti-Target Antibody60 µL
Anti-GAPDH Antibody60 µL
HRP-Conjugated Anti-Rabbit IgG Antibody12 mL
HRP-Conjugated Anti-Mouse IgG Antibody12 mL
SDS Solution12 mL
Stop Solution24 mL
Ready-to-Use Substrate12 mL
Crystal Violet Solution12 mL
Adhesive Plate Seals2 seals

The following materials and/or equipment are NOT provided in this kit but are necessary to successfully conduct the experiment:

  • Microplate reader able to measure absorbance at 450 nm and/or 595 nm for Crystal Violet Cell Staining (Optional)
  • Micropipettes with capability of measuring volumes ranging from 1 µL to 1 ml
  • 37% formaldehyde (Sigma Cat# F-8775) or formaldehyde from other sources
  • Squirt bottle, manifold dispenser, multichannel pipette reservoir or automated microplate washer
  • Graph paper or computer software capable of generating or displaying logarithmic functions
  • Absorbent papers or vacuum aspirator
  • Test tubes or microfuge tubes capable of storing ≥1 ml
  • Poly-L-Lysine (Sigma Cat# P4832 for suspension cells)
  • Orbital shaker (optional)
  • Deionized or sterile water

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

StepProcedure
1.Seed 200 µL of 20,000 adherent cells in culture medium in each well of a 96-well plate. The plates included in the kit are sterile and treated for cell culture. For suspension cells and loosely attached cells, coat the plates with 100 µL of 10 µg/ml Poly-L-Lysine (not included) to each well of a 96-well plate for 30 minutes at 37°C prior to adding cells.
2.Incubate the cells for overnight at 37°C, 5% CO2.
3.Treat the cells as desired.
4.Remove the cell culture medium and rinse with 200 µL of 1x TBS, twice.
5.Fix the cells by incubating with 100 µL of Fixing Solution for 20 minutes at room temperature. The 4% formaldehyde is used for adherent cells and 8% formaldehyde is used for suspension cells and loosely attached cells.
6.Remove the Fixing Solution and wash the plate 3 times with 200 µL 1x Wash Buffer for five minutes each time with gentle shaking on the orbital shaker. The plate can be stored at 4°C for a week.
7.Add 100 µL of Quenching Buffer and incubate for 20 minutes at room temperature.
8.Wash the plate 3 times with 1x Wash Buffer for 5 minutes each time.
9.Add 200 µL of Blocking Buffer and incubate for 1 hour at room temperature.
10. Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
11.Add 50 µL of 1x primary antibodies (Anti-XRCC1 Antibody and/or Anti-GAPDH Antibody) to the corresponding wells, cover with Parafilm and incubate for 16 hours (overnight) at 4°C. If the target expression is known to be high, incubate for 2 hours at room temperature.
12.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
13.Add 50 µL of 1x secondary antibodies (HRP-Conjugated AntiRabbit IgG Antibody or HRP-Conjugated Anti-Mouse IgG Antibody) to corresponding wells and incubate for 1.5 hours at room temperature.
14.Wash 3 times with 200 µL of 1x Wash Buffer for 5 minutes each time.
15.Add 50 µL of Ready-to-Use Substrate to each well and incubate for 30 minutes at room temperature in the dark.
16.Add 50 µL of Stop Solution to each well and read OD at 450 nm immediately using the microplate reader.

(Additional Crystal Violet staining may be performed if desired – details of this may be found in the kit technical manual.)

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