FIG. 4.

Flow cytometric analysis and Western blot of phosphoproteins of thymocytes in adult DOX-treated DTg mice. (A) FACS analysis of thymocyte populations of male DTg mice from the F176042 and F176043 transactivator founder lines treated or not with 0.5 mg/ml DOX in drinking water for 1 week. The percentages of cells and standard deviations are indicated in each quadrant. Total cell numbers for each thymus are indicated in parentheses. (B) FACS analysis of thymocyte cell populations of male DTg mice from F148571 and female DTg mice from the F176042 and F176043 transactivator founder lines treated or not with 2 mg/ml DOX in drinking water for 6 weeks. The percentages of cells and standard deviations are indicated in each quadrant. (C) Western blot analysis of pY proteins detected with anti-pY MAb (4G10) in thymocyte lysates from DOX-treated and untreated DTg male mice from the F148571 founder line after stimulation with anti-CD3 antibody. The membrane was washed and reblotted with antiactin and anti-Nef antibodies to control loading and to detect Tg expression, respectively. Quantitation was performed as described in Materials and Methods. Data are expressed relative to uninduced non-Tg cells (assigned value of 1). Statistical analysis was performed with Student's t test, and P values are shown in panel B; *** in panel A, P < 0.001.