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
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Thr50 of human CDCA2 protein.
Cell division cycle associated 2 (CDCA2, Repo-Man) is a cell-cycle protein that recruits protein phosphatase 1 (PP1) to mitotic chromatin at anaphase onset, which is essential for cell proliferation (1). Carboxy-terminal phosphorylation of CDCA2 at Ser893 by Aurora B inhibits the protein and leads to diffuse localization during prometaphase and metaphase. Dephosphorylation of CDCA2 by PP2A is necessary for CDCA2/PP1 complex reformation (2). The CDCA2/PP1 complex is required for chromatin binding and dephosphorylation of histone H3 at Thr3, Ser10, and Ser28 (2-4). The CDCA2/PP1 complex is also involved in nuclear envelope reformation during mitotic exit for proper progression through the M/G1 transition (4). The interaction of CDCA2 with importin beta and Nup153, which is required for nuclear envelope formation, is negatively regulated by CDK phosphorylation of the amino-terminal domain of CDCA2 (5). CDCA2 may play a role in DNA repair as the release of CDCA2 from chromatin at sites of DNA damage promotes the activation of DNA damage response (6). These results imply that the CDCA2/PP1 complex may play a part in cancer progression. Research studies indicate that CDCA2 may serve as a prognostic marker, as increased CDCA2 expression is seen in a number of cancers, including melanoma, neuroblastoma tumors, squamous cell carcinoma, and synovial sarcomas (7-9).
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