T-lymphocyte reconstitution in ZAP-70–transduced mice.

Splenocytes were collected from euthanized animals at 18 weeks following BMT and analyzed by flow cytometry. (A) Cells were stained with PE-conjugated αCD8 and Cy-conjugated αCD4 mAbs. The percentages of CD4⁺ and CD8⁺ T cells in each dot blot are indicated. (B) The activation status of the CD4⁺ T-cell population was determined using PE-conjugated αCD25 and αCD69 mAbs, and the relative percentages of naive and memory CD4⁺ T cells were monitored with a mAb recognizing CD62L (CD62L⁺ and CD62L⁻, respectively). The percentages of stained cells are indicated in each histogram. These parameters could not be analyzed in ZAP-70^−/− mice undergoing transplantation with ZAP-70^−/− progenitor cells harboring the EGFP vector. Data are representative of results obtained from 3 to 4 mice in each group. (C) Expression of ZAP-70 in EGFP-transduced splenocytes, ZAP-70–transduced splenocytes, and ZAP-70^−/− mice undergoing transplantation with WT BM as well as WT mice and the human T-cell line, Jurkat, was assessed in Western blots using an αZAP-70 mAb that reacts against both human (hZAP-70) and murine ZAP-70 (mZAP-70). Protein loading was monitored by immunoblotting with an αErk2 mAb. Data are representative of results obtained using splenocytes isolated from 2 mice in each group.