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Each lot of this antibody is quality control tested by Western blotting. Western blotting, suggested working dilution(s): Use 10 μl per 5 ml antibody dilution buffer for each mini-gel. It is recommended that the reagent be titrated for optimal performance for each application.
COA:
Application Notes:
This antibody reacts against Thr210-phosphorylated PLK-1 (Cross-reacts with unknown phosphorylated kinase in non-mitotic and mitotic cells. To increase specificity, immunoprecipitate with anti-PLK-1 antibody prior to blotting with phospho-specific PLK-1.)
Panel A. Extracts from untreated Hela cells (Lane 1) or overnight nocodazole-treated Hela cells (Lane 2) were immunoprecipitated with the pan-PLK monoclonal antibody (clone 3F8), and then western blotted with Poly6186 against phosphorylated PLK-1 (Thr210). Proteins were visualized using a donkey anti-rabbit secondary conjugated to HRP and a chemiluminescence detection system. Panel B. Extracts from untreated Hela cells (Lane 1) or overnight nocodazole-treated Hela cells (Lane 2) in Panel A were stripped and re-probed with a polyclonal antibody against pan-PLK-1 (Poly6185) to document equivalent loading of the PLK-1 protein.
Immunofluorescent microscope analysis of nocodazole treated Hela cells (10μM for overnight), using PLK-1 Phosphorylated (Thr210) polyclonal antibody (red). Nuclei were stain with DAPI (blue).
PLK-1 (polo-like kinase 1) is a member of te serine/threonine protein kinase family, cdc5/polo subfamily. Highly homologous to polo-like kinase (Drosophila), PLK-1 contains two polo box domains with a predicted molecular weight of 68 kD. This nuclear protein is highly expressed in placenta and colon and has been shown to regulate cdc2/cyclin B through phosphorylation and activation of cdc25c phosphatase. PLK-1 may also be required for cell division; depletion of PLK-1 results in apoptosis. PLK-1 is upregulated by growth stimulating agents and is regulated by cell cycle position (highest in G2/M phase, declining to nearly undetectable levels after mitosis and throughout G1). PLK-1 is modified by phosphorylation (Thr210 is the major phosphorylation site in activated PLK-1 from mitotic cells) and has been shown to interact with nuclear distribution gene C. The Poly6186 antibody recognizes human phosphorylated PLK-1 (Thr210) and has been shown to be useful for Western blotting. The Poly6186 antibody cross-reacts with an unknown kinase in non-mitotic cells. To increase specificity, it is recommended that the Poly6186 antibody be used for Western blotting after immunoprecipitation with a pan-specific PLK-1 antibody (clone 3F8).
Other Names:
Serine/Threonine protein kinase PLK, PLK, Serine-threonine protein kinase 13
Structure:
Serine/Threonine family of protein kinases, cdc5/polo subfamily. Highly homologous to polo-like kinase (Drosophila). Contains two polo box domains. Predicted molecular weight 68 kD
Distribution:
Nuclear protein, highly expressed in placenta and colon
Function:
Regulates cdc2/cyclin B through phosphorylation and activation of cdc25c phosphatase. May be required for cell division. Depletion of PLK-1 results in apoptosis
Regulation:
Upregulated by growth stimulating agents. Regulated by cell cycle position (highest in G2/M phase and declines to nearly undetectable levels after mitosis and throughout G1)
Modification:
Phosphorylation (Thr210 is major phosphorylation site in activated PLK-1 from mitotic cells)
Interaction:
Interacts with nuclear distribution gene C
Antigen References:
1. Hamanaka R, et al. 1994. Cell Growth Differ. 5:249. 2. Lake RJ, et al. 1993. Mol. Cell. Biol. 13:7793. 3. Holtrich U, et al. 1994. P. Natl. Acad. Sci. USA 91:1736.
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Purified anti-PLK-1 Phosphorylated (Thr210)
Panel A. Extracts from untreated Hela cells (Lane 1) or overnight nocodazole-treated Hela cells (Lane 2) were immunoprecipitated with the pan-PLK monoclonal antibody (clone 3F8), and then western blotted with Poly6186 against phosphorylated PLK-1 (Thr210). Proteins were visualized using a donkey anti-rabbit secondary conjugated to HRP and a chemiluminescence detection system. Panel B. Extracts from untreated Hela cells (Lane 1) or overnight nocodazole-treated Hela cells (Lane 2) in Panel A were stripped and re-probed with a polyclonal antibody against pan-PLK-1 (Poly6185) to document equivalent loading of the PLK-1 protein.
Immunofluorescent microscope analysis of nocodazole treated Hela cells (10μM for overnight), using PLK-1 Phosphorylated (Thr210) polyclonal antibody (red). Nuclei were stain with DAPI (blue).
