Microscopic analysis of H-RS cells from the lymph nodes of an HD patient and CD99-deficient B-cell lines. (A) Examination on the distribution of CD30 and CD99 molecules in lymph nodes of an HD patient by confocal microscopy. The frozen sections of lymph nodes were double-stained with FITC-conjugated anti-CD30 and biotinylated anti-CD99 antibodies. CD99 expression was identified by incubation with Texas Red–conjugated streptoavidin. CD99 molecule is absent in H-RS cells, whereas a considerable number of surrounding activated lymphocytes show strong reactivity (original magnification × 200). A CD30⁺ H-RS cell is shown at higher magnification (inset; original magnification × 1,000). (B) Localization of CD15 molecules in AS-TF IM9 cells with confocal microscopic analysis. AS-TF cells were stained with FITC-conjugated anti-CD15 antibody. Propidium iodide (PI) was included for nuclear staining. Most of large cells showed intense expression of CD15 in the Golgi and cytoplamic regions as well as on plasma membrane (original magnification × 630). Both (C) Vec-TF and (D) AS-TF IM9 cells were morphologically examined after Wright and Giemsa staining. AS-TF IM9 cells show typical H-RS morphology. (E and F) Restored cell morphology by exogenous expression of CD99 in Mut-IM9 cells. (E) Spontaneous CD99-deficient IM9 cells (Mut-IM9) and (F) CD99-TF Mut-IM9 cells were examined with Wright and Giemsa staining. All slides (C-F) were processed in parallel and photographed under identical magnification (40× objective).