Deficient MHC class II antigen presentation in a subset of HRS cells. Fresh lymph node tissue from a patient with nodular sclerosis Hodgkin's disease was disaggregated into a single-cell suspension and the viable cell fraction incubated for 90 minutes at 37°C with 5 μg/mL Hoechst 33258 (A and D) and stained with either an MHC class II αβ dimer-specific antibody (BU27) (B) or an antibody that recognizes CLIP bound to MHC class II molecules (CerCLIP.1) (E). Cell suspensions with surface-bound MoAbs or a mixture of synthetic beads with (G, H, I, arrows) or without (G, arrowheads) bound mouse IgG2a molecules (4.2 × 105 per bead) were fluorescently labeled with a PE-conjugated goat anti-mouse secondary antibody. Samples were processed for fluorescence microscopy and visualized by illumination with either ultraviolet (A, D), green (B, E, H), or white light (G). Enlarged images of labeled HRS cells (C, F, arrows) and synthetic beads (I, arrow) were used for densitometric analysis. Here, examples are shown in false colors to visualize the relative membrane densities of MHC class II molecules. A long wavelength (light blue) represents a high membrane density. Densities in the upper left-hand corner (C, F, I, stars) were used for background subtraction. Notice that a proportion of bystander lymphocytes express MHC class II αβ dimers (B) but not MHC class II αβ–CLIP complexes (E). Bar in (B) is 20 μm. Magnifications in (A, B, D, E, G, H) are the same.
