Western blot analysis of extracts from HeLa and HT-1080 cells, untreated or treated with UV, using Phospho-Mre11 (Ser676) Antibody (upper) or total Mre11 Antibody #4895 (lower).
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
Phospho-Mre11 (Ser676) Antibody detects endogenous levels of Mre11 only when phosphorylated at Ser676.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser676 of human Mre11. Antibodies are purified by protein A and peptide affinity chromatography.
Mre11, originally described in genetic screens from the yeast Saccharomyces cerevisiae in which mutants were defective in meiotic recombination (1), is a central part of a multisubunit nuclease composed of Mre11, Rad50 and Nbs1 (MRN) (2,3). The MRN complex plays a critical role in sensing, processing and repairing DNA double strand breaks. Defects lead to genomic instability, telomere shortening, aberrant meiosis and hypersensitivity to DNA damage (4). Hypomorphic mutations of Mre11 are found in ataxia-telangiectasia-like disease (ATLD), with phenotypes similar to mutations in ATM that cause ataxia-telangiectasia (A-T), including a predisposition to malignancy in humans (5). Cellular consequences of ATLD include chromosomal instability and defects in the intra-S phase and G2/M checkpoints in response to DNA damage. The MRN complex may directly activate the ATM checkpoint kinase at DNA breaks (6).
Phospho-Mre11 (Ser676) Antibody is directed to a site that was identified at Cell Signaling Technology (CST) using PhosphoScan®, CST's LC-MS/MS platform for modification site discovery. Phosphorylation at Ser676 was discovered using an ATM/ATR substrate antibody and was shown to be induced by UV treatment (7). Please visit PhosphoSitePlus®, CST's modification site knowledgebase, at www.phosphosite.org for more information.
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