Serine phosphorylation of STAT3 in murine calvaria cells treated with IL-11 and/or heparin.

Phosphate-starved calvaria cells were labeled with ³²P for 2 hours and then either left untreated (lane 1) or treated for 10 minutes with heparin alone (25 μg/mL; lane 2), IL-11 alone (20 ng/mL; lane 3), or heparin plus IL-11 (25 μg/mL and 20 ng/mL, respectively; lane 4). The cells were lysed and the lysate was immunoprecipitated with a polyclonal rabbit antimouse STAT3 antibody prior to immunoblotting the precipitates with a polyclonal anti-STAT3 antibody (A) or running them on a second gel and immunoblotting with a STAT-phosphoserine727 Ab (B). Alternatively, total³²P labeling of STAT3 was visualized by autoradiography (C).