Fig. 2. Expression of IGF-IR and IR and IGF-IR kinase activity. A, nonmalignant and malignant mammary cell lines growing in complete growth medium were lysed with RIPA buffer. Five hundred µg of lysate protein were immunoprecipitated (IP) with anti-IGF-IR, resolved by 10% SDS-PAGE, and immunoblotted (IB) with anti-IGF-IR. B, cells were grown to confluence, serum-starved for 16 h, and incubated for 10 min with or without IGF-I. Cell lysates were prepared with RIPA buffer, and 500 µg of each lysate were immunoprecipitated with anti-IGF-IR. Immunoprecipitates were subjected to in vitro kinase assay and analyzed as described in "Materials and Methods." C, 20 µg of cell lysate from A was resolved by SDS-PAGE and immunoblotted with anti-IR. The positions of the molecular weight markers are indicated. *, nonmalignant.