We want to perform a K-LISA (ELISA for kinase activity) for mTOr, but we do not have many cells. So we would like to do it without the immunoprecipitation step, is it possible? Also, how many ug of protein are needed?
The IP step should concentrate the sample. If you only have a small amount of starting material why do you want to skip the IP step?
As an alternative I would suggest measuring phosphorylation levels of mTOR downstream targets, such as S6K by Western blot. In this assay relatively small number of cells is needed.
Is there a reason you dont want to use WB? There you can easily distinguish between mTORC1/2 and the phospho-ABs for S6K1/ S6 or Akt work really well!!
Alternatively you could do intracellular flow cytometry http://www.ncbi.nlm.nih.gov/pubmed/23449254.
- Badges
- Science method
- Similar topics
- Medicine
- Immunology
- Immunology Techniques
- Immunoassay
- ELISA