Figure 2.

Tyrosine phosphorylation of (A) Vav, Syk, and Lyn; (B) PLC-γ₂; (C) SHIP and CD79a; and (D) SHP1 after BCR crosslinking in B cells from CD22-deficient mice and wild-type littermates. Purified splenic B cells (10⁷/lane) were incubated with anti-IgM antibodies and processed as in Fig. 1. Proteins of interest were immunoprecipitated from cell lysates with specific antibodies either alone or in combination, as indicated. Control immunoprecipitations of lysates from unstimulated wild-type B cells with normal rabbit serum are shown (CTL). In some cases (A and C), MAPK was also simultaneously immunoprecipitated from the lysates. Immunoprecipitated proteins were fractionated by SDS/PAGE and transferred onto a membrane for subsequent anti-phosphotyrosine immunoblotting (Upper). All blots subsequently were stripped of anti-phosphotyrosine antibody and reprobed with precipitating antibodies to quantify the amount of proteins within the immunoprecipitates (Lower). Results are representative of those obtained with at least three littermate pairs of mice for each molecule shown. The same results were obtained when Vav, Syk, Lyn, SHIP, or CD79a were immunoprecipitated individually from lysates (data not shown).