Nondenaturing

For this purpose, either Flag epitope tagged IKK-α, - β or - ε was co-transfected with Myc tagged IKKγ-WT or IKKγΔ and subjected to nondenaturing coimmunoprecipitation. ❋ Ping Liu Et Al. (2009)

IKKε was transfected in the absence or presence of TANK. 48 h later, WCEs were collected and subjected to nondenaturing co-immunoprecipitation using anti-Myc Ab. IKKα, IKKβ or IKKε association was detected by anti-Flag antibody (top panel). ❋ Ping Liu Et Al. (2009)

IKKγ −/− MEFs were cotransfected with eukaryotic expression vectors encoding Myc-IKKγ-WT and FLAG-MAVS in the absence or presence of Myc-IKKγΔ. 36 h later, nondenaturing coimmunoprecipation experiments were conducted using anti-FLAG. ❋ Ping Liu Et Al. (2009)

A nondenaturing co-immunoprecipitation assay experiment was then performed using anti-Myc Ab.as the primary immunoprecipitating Ab. followed by Western immunoblot of the immunoprecipitates using anti-V5 Ab. V5-TANK was only observed in immunoprecipitates in cells expressing IKKγ-WT, but not IKKγΔ ( ❋ Ping Liu Et Al. (2009)