The invention relates to a
rapid detection method of TTK
enzyme activity and an application of the
rapid detection method. The detection method includes 1), taking a to-be-detected sample, blank control and
positive control to perform co-incubation with TTK
enzyme and MBP / ATP (
mannose-
binding protein /
adenosine triphosphate) mixture respectively to produce ADP (
adenosine diphosphate); 2), respectively adding ADP-Glo<TM> reaction reagents to three groups of
kinase reaction systems for co-incubation, and terminating the
kinase reaction and consuming the remaining TAP; 3), respectively adding
kinase detection reagents to the reaction systems for co-incubation to convert ADP to ATP, and reading the luminous value RLU of the newly synthesized ATP; 4), calculating the TTK
enzyme activity according to the formula: the enzyme activity=(RLU(Sample)-RLU(Blank)) / (RLU(Pos.Ctrl)-RLU(Blank))X100%. On the basis of ADP-Glo<TM> kinase experiment, the reaction process is quantified by direct direction ofATP consumed in the reaction; the reaction is carried out in 384-hole plates, which does not involve repeated suction and washing process, the operation error is small, the flux is increased, and themethod is suitable for a large number of microhole plate operation combined with high-flux
liquid transfer workstation equipment; the method is stable in
system, high in precision and can be used forscreening TTK target inhibitors and TTK target related drugs.