Application of 3D-ExM in visualiz ing virus architecture. (A) Representative images of iD98HR1 cells that have been induced to enter EBV’s lytic phase and stained for the EBV envelope protein GP350 (red) and DNA (white) before and after 12× 3D-ExM. Images on the right are the zoom-in images shown in the orange box on the left images. (B) Left: representative images of uninfected and HIV-infected HeLa cells stained with DNA (magenta), HIV Gag (green) and HIV gRNA (red). Right: schematic diagram of how HIV genome integration sites were visualized using HIV genome sequence-specific biotinylated RNA probes. (C) Representative 12× 3D-ExM images of HIV gRNA and Gag proteins. HIV gRNA was labeled using specific FISH probes. Arrows show integration sites of HIV-1 genome in a nucleus. (D) Larger FOV images of Fig. 6 B. The enlarged images within the square boxes are shown in Fig. 6 B. DNA was stained with DAPI (pre-ExM) or DRAQ5 (post-ExM).