The initial wave of thymus colonization can be anatomically separated into 2 distinct stromal compartments. Frozen section from E12 BALB/c thymic rudiment (A; original magnification × 400) showing the cytokeratin⁺ epithelial core compartment (green) surrounded by fibronectin⁺ perithymic mesenchyme (blue). Note that CD45⁺ lymphoid progenitors (red) are present in both the epithelial compartment and the outer layers of mesenchyme. Combined enzymatic and mechanical separation of isolated intact lobes (B, arrowhead; original magnification × 50) produces smooth epithelial cores (arrow) stripped of mesenchyme and separate mesenchyme fragments (asterisk) devoid of epithelium. Following complete disaggregation of separated mesenchyme and epithelial components, small but distinct populations of CD45⁺ lymphoid precursors can be detected by flow cytometry in each preparation (mesenchyme, C; epithelium, D) allowing the identification and isolation of CD45⁺ precursor subsets before and after contact with thymic epithelium. Vertical bars show levels of background staining (panels C and D). Total disaggregation of whole freshly isolated E12 thymic lobes and surface labeling for EpCAM1 and PDGFRα (E) identifies nonoverlapping populations of epithelial cells and mesenchymal cells, respectively. Note that, apart from a small cohort of CD45⁺ cells representing the earliest thymic migrants (F), the thymus at this stage consists predominantly of EpCAM1⁺PDGFRα^– epithelium and EpCAM1^–PDGFRα⁺ mesenchyme. Numbers indicate percentages of positive cells. Data are typical of 5 separate experiments.