IP-Kinase Assay

Abstract

Immunoprecipitation (IP)- Kinase assays are an invaluable tool to assess the activation status of intracellular signaling cascades within a specific cellular state and also to confirm the enzymatic activity of a specific kinase towards a putative substrate of interest. Intracellular signal transduction cascades play an important role in modulating the localization of transcription factors and thus impact the cellular transcriptome. This in turn regulates key cell fate decisions including cell survival, apoptosis, proliferation, and differentiation. Here we describe an in vitro non-radioactive method to assess kinase activity towards a specific substrate. In this protocol we outline the method for Akt, however the basic protocol may be applied to any kinase and putative substrate of interest.