Analysis of Protein-Protein Interaction by Co-IP in Human Cells While there are various approaches available to analyze protein-protein interactions, coimmunoprecipitation (co-IP) remains one of the most classic and commonly used methods to discover novel protein interactions or to determine the physical association of proteins The assay begins with the prepara …
Co-immunoprecipitation (co-IP) Troubleshooting Guide Immunoprecipitation methods (IP, co-IP, ChIP and others) typically require a great deal of optimization and troubleshooting While in most cases the critical factor is the antibody, several other factors can significantly influence the outcome of the IP These guidelines will help you to obtain the best possible results from your IP
RANK ligand converts the NCoR HDAC3 co-repressor to a PGC1β . . . co-activator 1 beta (PGC1b), analogous to the prior finding that NCoR HDAC3 complexes deacetylate PGC1a as a prerequisite for co-activation of estrogen-related receptor a (ERRa) and in-duction of thermogenesis in brown adipose tissue 10 We extend this concept to demonstrate that RANK signaling promotes the
Protein-Protein Interactions: Co-Immunoprecipitation Growing evidence shows that protein-protein interactions are crucial in many biological processes in living cells Genetic (such as yeast two-hybrid, Y2H) and biochemical (such as co-immunoprecipitation, co-IP) methods are the methods commonly used at the beginning of a study to identify the interacting proteins
The Ski protein can inhibit ligand induced RARα and HDAC3 . . . The immunocomplexes and 10% input were detected by an anti-Flag antibody (B) COS-1 cells transfected with Flag-HDAC3 were treated without or with RA for 18 h in the absence or presence of 5 μM MG132 (C) COS-1 cells were transfected with Flag-HDAC3 with or without T7-Ski, and CHX was added to the cells after 24 h transfection Flag-HDAC3
Analysis of Protein–Protein Interaction by Co-IP in Human Cells It is important to confirm the co-IP results using other methods such as immunofluorescence colocalization analysis, proximity ligation assay , and bimolecular fluorescence complementation assay Here we describe an updated protocol for Co-IP using calcium phosphate-transfected HEK293T cells, the most commonly used transfection method and cell