The Mcl-1 transgene enhances the survival of mature lymphoid (B and T) as well as myeloid cells but does not override endogenous determinants of viability. (A) Spleen cells from transgenic and nontransgenic mice were assayed by two-color flow cytometry for the presence of B and T cell cell markers, either immediately on explantation (day 0) or after 1 or 3 days of of incubation, as in Fig2B. The dot plots shown were obtained by gating on the viable cell population and the percentages of B220⁺ and CD3⁺ cells, and cell-surface IgM⁺ and CD5⁺ cells, represent the mean ± SD of two to three animals. Results from parallel assays are shown in Table 1. The B/T cell ratio indicated was calculated from both sets of markers. The number of cells recovered from the spleen in these experiments averaged 2.9 ± 0.6 (SD) × 10⁸ for nontransgenic and 4.6 ± 1.3 × 10⁸ for transgenic mice. The percentage of viable cells on day 0 was greater than 95% and the average percentage of viable cells on days 1, 2, and 3, respectively, was 51% ± 6% (SD), 19% ± 4%, and 2.4% ± 0.7% for nontransgenic mice and 75% ± 8%, 56% ± 5%, and 40% ± 8% for transgenic mice. (B) The fractions of T lymphoid, B lymphoid, and myeloid cells surviving on days 1 and 3 (compared with the initial numbers present on day 0) were calculated from the experiment in (A). The values for the T-cell markers CD3⁺ and CD5⁺ cells were averaged, as were those for the B-cell markers B220⁺ and IgM⁺; CD11b was assayed in duplicate as a myeloid marker. Bars represent the SE of two to three animals. On day 0, the total number of viable nontransgenic T, B, and myeloid cells averaged 9.7 ± 0.1 × 10⁷, 1.7 ± 0.2 × 10⁸, and 1.5 ± 0.4 × 10⁷ (SE of three animals), and the total number of viable transgenic T, B, and myeloid cells averaged 1.4 ± 0.1 × 10⁸, 2.6 ± 0.3 × 10⁸, and 2.8 ± 0.5 × 10⁷. (C) Spleen cells from Mcl-1 transgenic and nontransgenic mice were incubated in vitro as in (A), with IL-2 or IL-3 (100 pmol/L) being added on day 1 as indicated. On day 9, the viable cell population was assayed by flow cytometry for CD11b and F4/80, CD11b and CD5, or B220. At this time, the percentage of viable cells in transgenic cultures was 26% to 28%, as determined by propidium iodide staining. Essentially no viable cells remained in nontransgenic cultures to which no factor had been added on day 1, and only small numbers of viable cells (≤1%) could be detected in nontransgenic cultures to which growth factors had been added. B, B220⁺ cells; T, CD5⁺ cells; M, CD11b⁺ and/or F4/80⁺ cells. The CD11b⁺ cells shown were CD5⁻, and the CD5⁺ cells shown were CD11b⁻, with ≤3% of cells being CD11b⁺CD5⁺.