Immunoprecipitation and immunoblot for VEGF receptors and PTK7. (A) Confluent MS1 cells were collected and lysed 2 hours after treatment with 20 ng/mL VEGF-A (control cells were not so treated). Immunoprecipitation was performed using 3 μg of antibodies specific for Flt-1, Flk-1/KDR, and Flt-4, and with 350-μg amounts of protein samples. Precipitates were collected and immunoblotting performed using antibodies against either PTK7 or VEGF receptors (**P < .01, compared with control). (B) Receptor complex formation between PTK7 and Flt-1 was assessed over time by immunoprecipitation and immunoblotting. MS1 cells, starved of serum and cytokines for 24 hours, were treated with 20 ng/mL VEGF-A and collected at various time points (5, 15, 30, 60, and 120 minutes). Immunoprecipitation and immunoblotting were performed as for panel A.