IDO expression in human brain tissue with and without HIVE. Double immunostaining demonstrates accumulation of CD8⁺ T cells (purple) around IDO-positive cells (brown) in the microglial nodule in HIVE (A). CD68⁺ macrophages/microglia (brown) in the microglial nodule correspond to IDO-positive cells on serial section (B). IDO (brown) is mainly localized in HIVp24⁺ cells (purple) in microglia nodule (C) and perivascular cells (D). Occasional IDO-expressing astrocytes were seen in HIVE cases (arrows). Note the absence of purple stain in the cytoplasm of astrocytes. Fewer CD8⁺ T cells (E) and CD68⁺ cells (F) and lack of cells staining for IDO (E) are shown in HIV-negative control. Human brain sections were double-immunostained with antibodies to CD8⁺ and HIVp24 (purple, VIP substrate) and IDO (brown, DAB substrate;A,C-E) or CD68 (brown, DAB substrate; B,F). Original magnifications: panels A-D: ×200; inset, panels C and D: ×600. Western blot analysis for IDO and CD68 expression in brain homogenates from control (HIV-), HIV+, and HIVE cases. Representative immunoblots of CD68, IDO, and internal standard α actin, and ratios of IDO/α actin and CD68/α actin are shown (G). IDO expression detected by Western blot in uninfected or HIV-1-infected MDMs in vitro with or without IFN-γ stimulation (H).