Stat1 response analysis of pre-B cells stimulated with bone marrow stromal cell supernatants. (A) Cultured pre-B cells (pool of either WT [B^+/+] or KO [B^−/−]) were stimulated 5 minutes with the supernatant of P7 TC-PTP^+/+ or TC-PTP^−/− bone marrow stromal cells cultures (with or without the presence of anti–IFN-γ antibody [α-IFN-γ]; 5 WT, 5 KO stroma). Intracellular expression of Stat1 and phosphorylated Stat1 (pStat1) was then analyzed by flow cytometry. The relative cell number (RCN) is plotted against pStat1 and Stat1 (inset) for unstimulated (time 0, dotted line) and stimulated (5 minutes, bold line) pre-B cells. (B) Normalization of the MFI of pStat1 versus Stat1 obtained by flow cytometry was obtained for each B cell/stromal cell supernatant stimulation condition. Data are provided as pStat1/Stat1 ratio ± SEM. *P < .02. (C) Leukemic pre-B–cell lines ABE 8.1/2 and 70Z/3 were cultured 24 hours in the presence of either TC-PTP^+/+ or TC-PTP^−/− bone marrow stromal cell culture supernatant (stroma +/+ or stroma −/−). Cells were stained with annexin V and propidium iodide (PI) for flow cytometry analysis. The combined percentage of preapoptotic (annexin V⁺, PI⁻) and apoptotic (annexin V⁺, PI⁺) cells is provided. Representative data are shown for each subset. (D) Quantitation of annexin V⁺ in leukemic cells; □, leukemic cells incubated with TC-PTP^+/+ stroma supernatant (n = 4); ▪, leukemic cells incubated with TC-PTP^−/− stroma supernatant (n = 4). Fold increase in the number of apoptotic annexin V⁺ cells ± SEM is provided relative to time 0 leukemic culture. **P < .005. All experiments were repeated at least 3 times.