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. 2005 May 25;1:2005.0008. doi: 10.1038/msb4100012

Figure 5.

Figure 5

Co-immunoprecipitation of STAT5 and EGFR. (A) In co-immunoprecipitation experiments, EGFR was detected by Western blot (WB) in immunoprecipitations (IP) with STAT5, and STAT5 was detected in immunoprecipitations of EGFR. Co-immunoprecipitation was reduced by addition of phosphorylated peptides pY978 and pY998 of EGFR. There was no effect when these peptides were added in their nonphosphorylated form as control (Y978 and Y998). In addition, equal amounts of EGFR and STAT5 were immunoprecipitated, as shown by the IP with STAT5, WB with STAT5 and the IP with EGFR, WB with EGFR, respectively. The experiments were performed on HeLa cells growing in 10% FBS. (B) Phospho-EGFR, STAT5 and Crk after immunoprecipitation of EGFR from normal growing HeLa cells (basal), serum-starved cells for 14 h (starved) and EGF-stimulated cells for 2 and 10 min. The antibody was used for IP and WB of EGFR against codons 998-1022, while the antibody against phospho-EGFR was directed against pYl 197 of EGFR. STAT5 antibody is directed against the C-terminal domain of Stat5b, but recognized isoforms STAT5a/b, and Crk antibody is directed against the N-terminus of Crk.