(A through E) Frozen sections from a case of classic (nodular sclerosis) HD tested by two-color staining with CD40L and BCL-6 MoAbs (A, C, and D) and CD40L and syn-1 MoAbs (B and E) (see Materials and Methods). Only scattered CD40L⁺(cytoplasmic, blue) lymphoid cells are detectable in the proximity of BCL-6⁺ (nuclear, red) RS cells (A), whereas expression of CD40L (cytoplasmic, blue) can be observed on several reactive lymphocytes that surround syn-1⁺ (cytoplasmic and membranous, red) RS cells (B). (A, inset) A higher magnification of BCL-6⁺ RS cells with two CD40L⁺ lymphoid cells (arrows). (C through E) RS cells of classic HD displaying the BCL-6⁺ (C), BCL-6⁻ (D), and syn-1⁺ (E) phenotype, respectively. The BCL-6⁺ RS cell does not display close association with CD40L⁺ T cells (C). Conversely, both BCL-6⁻(D) and syn-1⁺ (E) RS cells are accompanied by several CD40L⁺ T cells. Two-color staining, no counterstain. Original magnification × 180 (A), × 250 (inset A), × 400 (B), × 630 (C, D, and E).