Lymphoid potential of Wnt3a^−/− hematopoietic progenitors. (A) Flow cytometric analysis of common lymphocyte progenitors (CLPs). Lineage-negative IL-7R⁺ cells were gated and analyzed for c-Kit and Sca1 expression. Numbers indicate the percentage of cells in each gate. (B) Percentage of CLPs in total FLs from Wt and Wnt3a^−/− embryos. The averages are indicated by a dash. Data from 11 Wt and 6 Wnt3a^−/− FLs, belonging to 6 different litters identified by different colors. (C) To evaluate B-cell potential Wt and Wnt3a^−/− FL cells were cultured for 7 days on confluent layers of OP9 BM derived stromal cells. Cells were harvested and presence of cells from different lineages was analyzed by flow cytometry. Data are representative of 4 independent experiments. (D) Reduced cellularity in Wnt3a^−/− thymic anlage revealed by immunohistochemistry analysis. Tissue sections of E12.5 embryos were stained with HE to visualize different tissues (left panels). Thymic epithelial cells stained with the thymic stroma antibody ER-TR4 (right panels) to confirm identification of thymic anlage in Wnt3a^−/− embryos. Staining with an isotype control for ER-TR4 was completely blank. Thymic anlage is indicated by ▼. Stainings shown are representative of 4 Wt and 4 Wnt3a^−/− embryos examined. (E) Thymic lobes from Wt and Wnt3a^−/− E12.5 embryos were cultured in FTOC to allow T-cell development to proceed, and harvested at the indicated time points. Thymocytes subsets were analyzed by flow cytometry. Data are from 5 Wt and 3 Wnt3a^−/− embryos analyzed.