Western blot analysis of extracts from 293T and Colo205 cells (high expression), and A549 and SNB19 cells (low expression), using XPA (D9U5U) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 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.
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: 263
XPA (D9U5U) Rabbit mAb recognizes endogenous levels of total XPA protein.
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Arg158 of human XPA protein.
Nucleotide excision repair (NER) is a process by which cells identify and repair DNA lesions that result from chemical and radiation exposure (1). The DNA binding protein XPA is an essential part of a pre-incision complex that forms at sites of damage, and is necessary for the initiation of nucleotide excision repair (2). XPA is one of eight NER proteins (XPA-G, XPV) encoded by genes that are defective in cases of xeroderma pigmentosum, a disorder characterized by sensitivity to sunlight, predisposition to exposed tissue cancers, and neurological defects in some patients (3). Activation of XPA follows phosphorylation at Ser196 and results in increased NER activity. Phosphorylation of XPA at Ser196 is induced by UV exposure in an ATR-dependant fashion (4) and promotes nuclear accumulation of XPA (5). Research studies suggest that XPA may be a direct substrate of the serine/threonine kinase ATR (4) and that NER activity may be negatively regulated through dephosphorylation of Ser196 by the phosphatase WIP1 (6).
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