The left panel shows the timeline of infection and/or drug exposure experiments. (A-D) Multicolor images of vDNA, vRNA, DAPI and/or GFP in monocyte derived macrophages (MDMs) from two different donors, infected with HIV1. Cells from donor 1 were infected with VSV-G-pseudotyped HIV-1ΔEnv Vpx carrying a GFP reporter (A) and cells from donor 2 were infected with a VSV-G-pseudotyped HIV-1ΔEnv without Vpx (B-D). (A,B) Infected cells were left untreated and fixed at 6 d p.i. (C) Infected cells were treated with 10 µM Nevirapine (NVP) throughout the experiment and fixed at 6 d p.i. (D) Infected cells were exposed to NVP for 3 d p.i., then NVP was washed out and cells were cultured for another 4 d and fixed at 7 d p.i. Arrows in (A-D) show the position of selected vRNA foci. (E) Boxplot shows the size of DNA foci in untreated infected cells from donor 2 imaged at 7 d p.i. (B), measured by the FWHM of n=24 foci. The red line defines the median, top and bottom limits define upper and lower quartiles, respectively. Whiskers show the full data range, except for outliers. Grey dots are individual data points. (F) Boxplot shows Pearson's r, measuring colocalization of DNA with RNA foci in the same cells (n=20). Center line defines the median, top and bottom limits define upper and lower quartiles, respectively. Whiskers show the full data range, except for outliers. Grey dots are individual data points. 