反应详情

Cells were starved (24 hours for MCF-7, 4 hours for 32D derived cell lines) before addition of the indicated amounts of growth factors. Total cell lysates were prepared by addition of SDS PAGE sample buffer (1% SDS, 0.15M Tris pH 8.6, 5% BME and 1 mM Sodium Ortho Vanadate) directly to cells. Cell lysates and were run on 8–16% Tris-Glycine gradient gels (Novex). Proteins were transferred onto Hybond ECL nitrocellulose membranes (Amersham) and were immunoblotted with anti-phosphotyrosine MAb.

1. additionbefore addition of the indicated amounts of growth factors