Atp-dependent bioluminescence method and kit for detecting...

C - Chemistry – Metallurgy – 12 – Q

Patent

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C12Q 1/48 (2006.01) C12Q 1/66 (2006.01)

Patent

CA 2431274

The invention provides a method for measuring protein kinase activity, said method comprising: (a) providing a first solution comprising ATP and a protein kinase to be tested, and a second solution comprising ATP in the absence of said kinase to be tested; (b) adding a substrate capable of being phosphorylated by the protein kinase to be tested to the first and second solutions of step (a); (c) measuring the concentration of ATP and/or ADP, or the rate of change thereof with respect to time, in each of the reaction mixtures formed in step (b) using a bioluminescence reaction; and (d) using the information about the concentration of ATP and/or ADP to determine the activity of the protein kinase to be tested. A particular advantage of the methods and kits of the invention is that they provide a generic protein kinase activity test, that is, a test which is not specific for a single protein kinase or its substrate.

La présente invention concerne une méthode permettant de mesurer l'activité de protéine kinases, ladite méthode consistant à : (a) se munir d'une première solution comprenant de l'ATP et une protéine kinase à tester et une seconde solution comprenant de l'ATP en absence de ladite kinase à tester ; (b) ajouter un substrat pouvant être phosphorylé par la protéine kinase à tester à la première et à la seconde solution de l'étape (a) ; (c) mesurer la concentration en ATP et/ou en ADP, ou le taux de changement de ladite concentration par rapport au temps, dans chacun des mélanges de réaction formés dans l'étape (b) à l'aide d'une réaction de bioluminescence ; et (d) utiliser les informations concernant la concentration en ATP et/ou en ADP de manière à déterminer l'activité de la protéine kinase à tester. Les méthodes et les trousses selon l'invention offrent un avantage particulier résidant dans le fait qu'elles permettent d'effectuer un test générique d'activité de protéine kinases, c'est-à-dire un test qui n'est pas spécifique à une protéine kinase unique ou à son substrat.

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