Characterization of NTAL phosphorylation after FcϵRI aggregation in HuMCs. (A) Immunodepletion of pp30 from HuMCs with anti-NTAL pAb. HuMCs were sensitized and triggered for 60 seconds, and aliquots of the cell lysates were immunoprecipitated using an anti-NTAL pAb as described in “Materials and methods.” The immunoprecipitates and aliquots of the lysates before and after immunoprecipitation were then probed with 4G10 antiphosphotyrosine mAb or an anti-NTAL mAb. *Band for NTAL (B) Comparative kinetics of NTAL and pp30 phosphorylation. In these and other blots, the observed decrease in antibody recognition of NTAL (eg, at 60 seconds) represents epitope masking due to phosphorylation. (C) Comparative concentration dependency of NTAL and pp30 phosphorylation following FcϵRI aggregation. The cell lysates were all prepared under nondenaturing conditions as for Figure 2E-F. Conc indicates concentration; l/c, IgG light chain; h/c, IgG heavy chain; pY, phosphotyrosine; and asterisk, phosphorylated NTAL. The blots are representative of 2 to 3 similar experiments.