Mouse thymic dendritic cells (DC) have been isolated after collagenase digestion, selection of the low-density cell fraction, then depletion of T-lineage cells and other non-DC by treatment with specific monoclonal antibodies (mAb) and removal with anti-Ig-coated magnetic beads. The resulting DC preparation represented 0.1-0.2% of total thymic cells and contained 70-80% DC. Flow cytometry analysis of MHC class II (MHC II) expression by DC showed that 40% of DC expressed intermediate levels of MHC II, and 60% expressed high levels of this marker. Moreover, immunofluorescent 2-colour staining allowed the characterization of two clearly distinguishable DC subpopulations: MHC IIinter DC were CD45hi, CD44hi, HSAhi, whereas MHC IIhi DC were CD45lo, CD44lo, HSAlo. These results are discussed with regard to the functional significance of MHC IIinter and MHC IIhi DC subpopulations in the mouse thymus.