Detection of Endogenous SOCS1 and SOCS3 Proteins by Immunoprecipitation and Western Blot Analysis

The suppressors of cytokine signalling (SOCS) protein family consist of eight members (SOCS 1–7, and CIS). SOCS1 and SOCS3 are the best-studied family members and have been shown to act as negative feedback inhibitors of the JAK/STAT signalling pathway. To study the physiological roles of the SOCS proteins, it is necessary to establish methods for detecting endogenous proteins often expressed at low levels in cells after cytokine induction. To facilitate the detection of endogenous SOCS1 and SOCS3 proteins, we have generated in-house antibodies specific to these proteins, which we have used together with commercially available antibodies. Here, we describe the methods for immunoprecipitating SOCS1 and SOCS3 proteins from mouse tissue extracts and their subsequent detection by Western blot analysis. These methods can also be applied to the detection of SOCS1 and SOCS3 in cell lines.