What kind of assay method would you use to detect kinase activation?

[beachbum]

say if you used a cytokine or a gas to test its effect on the kinase.

[Yggdrasil]

You could try measuring the activity of the enzyme if you know a target of the kinase. Just add the substrate and radiolabeled ATP and check for the presence of radiolabeled substrate after the reaction.

 

Alternatively, if you know the downstream effects of the kinase (such as activation of a specific gene), then you can test the effects of the effector in vivo (although you would need to design some control experiements as well).

[Bluenoise]

It would be sweat if you have a flourogenic substrate so you could map the activity real time with some sort of spectrophotometer.

 

We may be able to help more if you can tell us the exact kinase and substrate that it acts upon.

[beachbum]

its actually more of a kinase signalling system. its called mTOR (mammalian homolog: Tor2). its supposed to activate/modulate autophagosome formation in autophagy. I wanna see the effects of some stuff on this "pathway" or i dunno if its even a gene. you can check some background info on it (paper: Klionsky D. Emr J. Autophagy as a Regulated Pathway of Cellular Degradation Science 2000 Volume???) Say its a gene, can you test the effect of gene activation? Thankyou.

[KrusherX]

Identify the target you're interested in, find a phospho-specific antibody (specific to the phosphorylated form of the target) and you have your level of activity for this kinase