Effect of conditioning therapy on recipient dermal DC and macrophages. (A) Representative flow cytometry analysis of dermal cell suspensions derived from clinical biopsies by collagenase digestion before and after conditioning. Selective depletion of SSC^loAF^loCD14⁻CD1a⁺ DC is evident. (B) Paired analysis of dermal cells obtained before and after conditioning from a total of 27 patients showing a decline in CD1a⁺ DC but preservation of CD14⁺ DC and macrophages. Enumeration of cells per unit area was achieved using a standard 2-mm punch of shaved dermis freshly digested into single cell suspension with collagenase. The total number of cells present in each fraction was quantified with the aid of Trucount fluorescent beads. (C) Subgroup analysis of 17 patients treated with reduced intensity and 10 patients with full intensity conditioning showing similar depletion of dermal DC in both groups. P <0.001 for before and after counts (Wilcoxon rank sum test). There is also a slight but nonsignificant increase in CD14⁺ DC with full intensity conditioning. Error bars indicate SD. (D) Immunofluorescence images of skin before and after conditioning showing maintenance of CD163⁺ perivascular macrophages. AF in the green channel can be seen in some cells (arrows). Bar, 50 µm. Two representative examples of six patients are shown. (E) Expression of CD52 antigen by dermal APC (red) compared with isotype controls (blue). Numbers indicate median fluorescence intensity ratios. Dermal T cells in the same preparation are shown for comparision. Normal skin was prepared stained and gated as described in Fig. 1.