Method for determining activity of cell cycle regulatory factor and method for diagnosing cancer using the same

摘要

A method for determining the activity of a cell cycle regulatory factor comprising the steps of: preparing a sample for measuring a cyclin-dependent kinase/cyclin complex from living cells; reacting adenosine 5'-O-(3-thiotriphosphate) (ATP-gammaS) with a substrate for the cyclin-dependent kinase in presence of the sample in order to introduce a monothiophosphate group into a serine or threonine residue of the substrate; labeling the substrate by coupling a labeling fluorophore or a labeling enzyme with a sulfur atom of the introduced monothiophosphate group; measuring the amount of fluorescence from the labeling fluorophore labeling the substrate, or reacting the labeling enzyme labeling the substrate with a substance which generates an optically detectable product by reaction with the labeling enzyme and optically measuring the amount of the generated product; and calculating the activity of the cyclin-dependent kinase from the measured amount of fluorescence or the measured amount of the generated product with reference to a pre-produced reference curve.