Phosphorylation of Tyr319 in ectopically expressed ZAP-70 is significantly less efficient in BJAB B cells than in Jurkat T cells. Immunoblotting analysis of immunoprecipitated ZAP-70 protein from transiently transfected BJAB B cells and Jurkat T-cells was done with the phospho-ZAP-70^Tyr319/Syk^Tyr352 antibody. The pcDNA3-ZAP-70-Myc construct was introduced into both cell lines by nucleofection. In the top panels both Myc-tagged and endogenous ZAP-70 proteins were immunoprecipitated with an anti–ZAP-70 antibody. The 2 proteins differ in size by 1 kDa because of the Myc tag. The bottom panels show analysis only of Myc-tagged ZAP-70 that was immunoprecipitated with an anti-Myc antibody.