p56Lck tyrosine phosphorylation and activity in different Jurkat cell clones. Cells were stimulated with 100 nmol/L thrombin for the times indicated. (A) Cell lysates were immunoprecipitated with 4G10 antibody preadsorbed on protein A-Sepharose. Immunoprecipitated proteins were subjected to SDS-PAGE and transferred to Immobilon membranes for Western blotting with anti-p56Lck antibody. Development was performed as described above. (B) Cell lysates were immunoprecipitated with anti-p56Lck antibody preadsorbed to protein A-Sepharose. After extensive washing, p56Lck activity was determined with enolase as exogenous substrate. (C) Equal amounts of p56Lck were immunoprecipitated in JE6.1 and J45.01 cells. Note the lack of p56Lck and p56Lck activity in JCaM1.