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Rat DNA mismatch repair protein Msh2 (Msh2) ELISA Kit (RTEB1583)

SKU:
RTEB1583
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P54275
ELISA Type:
Sandwich
Reactivity:
Rat
€649
Frequently bought together:

Description

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Rat DNA mismatch repair protein Msh2 (Msh2) ELISA Kit

The Rat DNA Mismatch Repair Protein MSH2 ELISA Kit is a powerful tool for quantitative analysis of MSH2 levels in rat samples. This kit is specifically designed for the accurate detection of MSH2 in serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, researchers can trust the reliability and reproducibility of their results, making it an invaluable resource for a variety of research applications.MSH2 is a key protein involved in the DNA mismatch repair pathway, playing a crucial role in maintaining genomic stability and preventing mutations.

Dysregulation of MSH2 has been implicated in various diseases, including cancer and hereditary nonpolyposis colorectal cancer (HNPCC). By measuring MSH2 levels, researchers can gain valuable insights into the mechanisms underlying these diseases and potentially identify new therapeutic targets.Overall, the Rat DNA Mismatch Repair Protein MSH2 ELISA Kit offers researchers a comprehensive solution for studying MSH2 expression in rat models, advancing our understanding of DNA repair mechanisms and their implications in disease development and progression.

Product Name:Rat DNA mismatch repair protein Msh2 (Msh2) ELISA Kit
SKU:RTEB1583
Size:96T
Target:Rat DNA mismatch repair protein Msh2 (Msh2)
Synonyms:MutS protein homolog 2
Assay Type:Sandwich
Detection Method:ELISA
Reactivity:Rat
Intra CV:3.9%
Inter CV:7.3%
Linearity:
Sample1:21:41:81:16
Serum(N=5)91-100%97-107%107-117%103-113%
EDTA Plasma(N=5)88-98%90-100%103-112%89-99%
Heparin Plasma(N=5)111-120%99-109%96-106%86-94%
Recovery:
Sample TypeAverage(%)Recovery Range(%)
Serum9690-102
Plasma9892-104
Function:Component of the post-replicative DNA mismatch repair system (MMR). Forms two different heterodimers: MutS alpha (MSH2-MSH6 heterodimer) and MutS beta (MSH2-MSH3 heterodimer) which binds to DNA mismatches thereby initiating DNA repair. When bound, heterodimers bend the DNA helix and shields approximately 20 base pairs. MutS alpha recognizes single base mismatches and dinucleotide insertion-deletion loops (IDL) in the DNA. MutS beta recognizes larger insertion-deletion loops up to 13 nucleotides long. After mismatch binding, MutS alpha or beta forms a ternary complex with the MutL alpha heterodimer, which is thought to be responsible for directing the downstream MMR events, including strand discrimination, excision, and resynthesis. ATP binding and hydrolysis play a pivotal role in mismatch repair functions. The ATPase activity associated with MutS alpha regulates binding similar to a molecular switch: mismatched DNA provokes ADP-->ATP exchange, resulting in a discernible conformational transition that converts MutS alpha into a sliding clamp capable of hydrolysis-independent diffusion along the DNA backbone. This transition is crucial for mismatch repair. MutS alpha may also play a role in DNA homologous recombination repair. In melanocytes may modulate both UV-B-induced cell cycle regulation and apoptosis.
Uniprot:P54275
Sample Type:Serum, plasma, tissue homogenates, cell culture supernates and other biological fluids
Specificity:Natural and recombinant rat DNA mismatch repair protein Msh2
Sub Unit:Heterodimer consisting of MSH2-MSH6 (MutS alpha) or MSH2-MSH3 (MutS beta). Both heterodimer form a ternary complex with MutL alpha (MLH1-PMS1). Interacts with EXO1. Part of the BRCA1-associated genome surveillance complex (BASC), which contains BRCA1, MSH2, MSH6, MLH1, ATM, BLM, PMS2 and the RAD50-MRE11-NBS1 protein complex. This association could be a dynamic process changing throughout the cell cycle and within subnuclear domains. Interacts with ATR. Interacts with SLX4/BTBD12; this interaction is direct and links MutS beta to SLX4, a subunit of different structure-specific endonucleases (By similarity). Interacts with SMARCAD1.
Research Area:Cancer
Subcellular Location:Nucleus
Storage:Please see kit components below for exact storage details
Note:For research use only
UniProt Protein Function:MSH2: Component of the post-replicative DNA mismatch repair system (MMR). Forms two different heterodimers: MutS alpha (MSH2- MSH6 heterodimer) and MutS beta (MSH2-MSH3 heterodimer) which binds to DNA mismatches thereby initiating DNA repair. When bound, heterodimers bend the DNA helix and shields approximately 20 base pairs. MutS alpha recognizes single base mismatches and dinucleotide insertion-deletion loops (IDL) in the DNA. MutS beta recognizes larger insertion-deletion loops up to 13 nucleotides long. After mismatch binding, MutS alpha or beta forms a ternary complex with the MutL alpha heterodimer, which is thought to be responsible for directing the downstream MMR events, including strand discrimination, excision, and resynthesis. ATP binding and hydrolysis play a pivotal role in mismatch repair functions. The ATPase activity associated with MutS alpha regulates binding similar to a molecular switch: mismatched DNA provokes ADP-->ATP exchange, resulting in a discernible conformational transition that converts MutS alpha into a sliding clamp capable of hydrolysis-independent diffusion along the DNA backbone. This transition is crucial for mismatch repair. MutS alpha may also play a role in DNA homologous recombination repair. In melanocytes may modulate both UV-B-induced cell cycle regulation and apoptosis. Heterodimer consisting of MSH2-MSH6 (MutS alpha) or MSH2- MSH3 (MutS beta). Both heterodimer form a ternary complex with MutL alpha (MLH1-PMS1). Interacts with EXO1. Part of the BRCA1- associated genome surveillance complex (BASC), which contains BRCA1, MSH2, MSH6, MLH1, ATM, BLM, PMS2 and the RAD50-MRE11-NBS1 protein complex. This association could be a dynamic process changing throughout the cell cycle and within subnuclear domains. Interacts with ATR. Interacts with SLX4/BTBD12; this interaction is direct and links MutS beta to SLX4, a subunit of different structure-specific endonucleases. Interacts with SMARCAD1. Ubiquitously expressed. Belongs to the DNA mismatch repair MutS family. Two isoforms of the human protein are produced by alternative splicing.Protein type: DNA-binding; Tumor suppressorChromosomal Location of Human Ortholog: 6q12Cellular Component: membrane; MutSalpha complex; MutSbeta complex; nuclear chromosome, telomeric region; nucleusMolecular Function: ADP binding; ATP binding; ATPase activity; centromeric DNA binding; damaged DNA binding; dinucleotide insertion or deletion binding; dinucleotide repeat insertion binding; DNA binding; DNA-dependent ATPase activity; double-strand/single-strand DNA junction binding; double-stranded DNA binding; enzyme binding; four-way junction DNA binding; guanine/thymine mispair binding; loop DNA binding; magnesium ion binding; mismatched DNA binding; MutLalpha complex binding; oxidized purine DNA binding; protein C-terminus binding; protein homodimerization activity; protein kinase binding; single base insertion or deletion binding; single guanine insertion binding; single thymine insertion binding; single-stranded DNA binding; Y-form DNA bindingBiological Process: B cell differentiation; B cell mediated immunity; cell cycle arrest; cellular response to DNA damage stimulus; determination of adult lifespan; DNA damage response, signal transduction resulting in induction of apoptosis; DNA repair; double-strand break repair; germ cell development; in utero embryonic development; intra-S DNA damage checkpoint; intrinsic apoptotic signaling pathway in response to DNA damage by p53 class mediator; isotype switching; maintenance of DNA repeat elements; male gonad development; meiotic gene conversion; mismatch repair; negative regulation of DNA recombination; negative regulation of neuron apoptosis; negative regulation of reciprocal meiotic recombination; oxidative phosphorylation; positive regulation of helicase activity; positive regulation of isotype switching to IgA isotypes; positive regulation of isotype switching to IgG isotypes; postreplication repair; response to amino acid; response to drug; response to organic cyclic compound; response to UV-B; response to X-ray; somatic hypermutation of immunoglobulin genes; somatic recombination of immunoglobulin gene segments; somatic recombination of immunoglobulin genes during immune response; spermatogenesis
UniProt Protein Details:
NCBI Summary:mismatch repair protein; may have an important role in spermatogenesis [RGD, Feb 2006]
UniProt Code:P54275
NCBI GenInfo Identifier:13591999
NCBI Gene ID:81709
NCBI Accession:NP_112320.1
UniProt Secondary Accession:P54275
UniProt Related Accession:P54275
Molecular Weight:104,028 Da
NCBI Full Name:DNA mismatch repair protein Msh2
NCBI Synonym Full Names:mutS homolog 2
NCBI Official Symbol:Msh2
NCBI Official Synonym Symbols:
NCBI Protein Information:DNA mismatch repair protein Msh2
UniProt Protein Name:DNA mismatch repair protein Msh2
UniProt Synonym Protein Names:MutS protein homolog 2
Protein Family:DNA mismatch repair protein
UniProt Gene Name:Msh2
UniProt Entry Name:
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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