Figure 1.

Identification of Thy-1^hiCD2⁻Lin⁻ T cell progenitors in the adult bone marrow. C57BL/6 Ly5.2 bone marrow cells were enriched for Thy-1⁺ cells with immunomagnetic beads, and cells were subsequently stained for Thy-1 with one fluorochrome; TCRαβ, B220, Mac-1, and NK1.1 with a second fluorochrome; and CD2 with a third fluorochrome. Analysis of Thy-1 vs. B220, Mac-1, TCRαβ, and NK1.1 (A); the box encloses Thy-1^hiTCRαβ⁻B220⁻Mac-1⁻NK1.1⁻ (Thy-1^hiLin⁻) cells. The latter cells were gated and reanalyzed for Thy-1 vs. CD2 (B); boxes 1 and 2 enclose Thy-1^hiCD2⁻Lin⁻ and Thy-1^hiCD2⁺Lin⁻ cells, respectively. Progenitor activity of sorted Thy-1^hiCD2⁻Lin⁻ Ly5.2 marrow cells was tested by incubating 5 × 10⁴ cells with 4 × 10⁶ Ly5.1 T cell-depleted marrow cells for 48 hr. The cultured cells were analyzed for CD4 and CD8 markers (C) or TCRαβ vs. Ly5.2 markers (D). Boxes enclose Ly5.2⁺ T cells. In a repeat experiment, the sorted Thy-1^hiCD2⁻Lin⁻ cells were cocultured with T cell-depleted Ly5.1 marrow cells, and cells were harvested at either 24 hr (E) or 48 hr (F). Gated Ly5.2⁺ cells harvested from the cultures were stained for TCRαβ vs. CD2, and boxes enclose CD2⁺TCRαβ⁻ (E) or CD2⁺TCRαβ⁺ (F) cells. The sorted Thy-1^hiCD2⁻Lin⁻ cells were also cocultured with whole instead of T cell-depleted Ly5.1 marrow cells, and cultures were stained for Ly5.2 vs. CD2 after 24 and 48 hr in G and H, respectively. Gated Ly5.2 cells from the latter cultures were stained for TCRαβ vs. CD2 at 24 and 48 hr (I and J, respectively), and boxes enclose CD2⁺TCRαβ⁺ cells. Expression of CD44 on sorted Thy-1^hiCD2⁻Lin⁻ cells before culture (K) is compared with that after coculture with T cell-depleted Ly5.1 marrow cells for 24 (L) or 48 hr (M). Analyses of CD44 vs. CD2 on gated Ly5.2 cells are shown, and boxes enclose CD44^hiCD2⁺ cells. Each pair of profiles is from a separate representative experiment of at least four replicate experiments.