Fig. 5

Immuno-electron microscopy (EM) of monocyte-derived macrophages (MDMs) infected with HIV-1 R3A PTAP⁻YP⁻. Seven-day-old MDMs were infected with HIV-1 R3A PTAP⁻YP⁻ for a further 7 days, fixed and processed for cryosection immunolabelling with antibodies against p24/55, and protein A-gold as in Fig. 4. A rabbit-anti-mouse Alexa-488 bridging antibody was used to locate infected cells by on-grid correlative light and EM. a A representative cell from Donor 429 located by on-grid immunolabelling. The marked area from the intracellular plasma membrane-connected compartment (IPMC) is shown at higher magnification in (b). c View of the area marked in (b) to show the immature phenotype of the budding-arrested virus particles. d Segmentation of the cell surface (grey) and IPMC membranes (black). Virus buds were counted and the lengths of membrane in the section calculated from the segmentations. e–g A partial cell profile from another infected cell, Donor 432. e Low power overview of the cell profile as seen emerging from under the grid bar. f Higher magnification detail of the marked area showing the IPMC. The arrow indicates an opening from the IPMC to the cell surface and arrowheads in (e) and (f) mark the five immature virus particles at the cell surface. g Segmentation of this cell as in (d). The length of the grid bar indicated by the dashed line was not included in the analysis. Scale bars: a, d and g, 5 μm; b and e, 2 μm; c, 200 nm; and f, 1 μm