Direct quantification of polo-like kinase 1 activity in cells and tissues using a highly sensitive and specific ELISA assay.
Polo-like kinase 1 (Plk1) plays a pivotal role in the regulation of cellular proliferation. Plk1 is overexpressed in approximately 80% of human tumors of diverse origins, and overexpression of Plk1 promotes neoplastic transformation of human cells. A growing body of evidence suggests that deregulation of Plk1 closely correlates with prognosis of various cancers in humans. Thus, accurate assessment of Plk1 deregulation would provide clear clinical advantages. However, because of the limited amount of cancer tissues available, quantification of the Plk1 activity has not been feasible. Here, we report the development of a rapid, highly sensitive, and specific ELISA-based Plk1 assay that can quantify the level of Plk1 activity with a small amount (2-20 microg) of total cellular proteins. Unlike the conventional immunocomplex kinase assay, this assay directly utilizes total cellular lysates and does not require a Plk1 enrichment step such as immunoprecipitation or affinity purification. Using this assay, we demonstrated that Plk1 activity is elevated in tumors but not in the surrounding normal tissues and that the level of Plk1 activity significantly diminishes after an antiproliferative chemotherapy. The method described here may provide an innovative tool for assessing the predisposition for cancer development, monitoring early tumor response after therapy, and estimating the prognosis of patients with cancers from multiple organ sites.
Park
Jung-Eun
JE
Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Li
Luowei
L
Park
Joobae
J
Knecht
Rainald
R
Strebhardt
Klaus
K
Yuspa
Stuart H
SH
Lee
Kyung S
KS
D000951:Antigens, Neoplasm / Q000032:analysis; D000970:Antineoplastic Agents / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000032:analysis*; D002477:Cells / Q000201:enzymology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D006801:Humans; D009369:Neoplasms / Q000175:diagnosis / Q000201:enzymology / Q000473:pathology*; D017346:Protein Serine-Threonine Kinases / Q000032:analysis*; D011518:Proto-Oncogene Proteins / Q000032:analysis*; D000097869:Polo-Like Kinase 1
D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D018797:Cell Cycle Proteins; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D000097869:Polo-Like Kinase 1