Figure 1

The enhancement assay. (A) Schematic diagram of the assay. (B) Flow cytometry dot plots from a typical enhancement assay. The gate is set against uninfected control SupT1/R5 cells. The upper panel shows the percentage of SupT1/R5 cells infected by virus pre-incubated with C' and NHS control serum, the lower panel with C' and an example autologous patient serum. (C) Light microscopy images from a typical enhancement assay. Images were taken of the cells analysed in 1B, prior to intracellular p24 staining and flow cytometry. The upper panel shows cells infected by virus pre-incubated with C' and NHS control serum, the lower panel with C' and autologous patient serum. Arrows on the upper panel indicate the presence of syncytia, which are large and numerous on the lower panel. (D) Calculation of fold enhancement. All experiments are performed alongside an HIV-antibody-negative (NHS) control culture, in the presence (C') and absence (HIC') of active complement. Enhancement is observed in the presence of C', therefore fold enhancement is calculated as the ratio of infected cells in the presence of C' and test (patient) serum to infected cells in the presence of C' and NHS. Figures 1B, C and D are all derived from experiments using MM32.10 virus and MM32 day 15 autologous serum (see Figure 2). The NHS + C' control experiments yielded 0.4% infected cells and the patient serum + C' cultures 8.8% infected cells, equating to a 22-fold enhancement of infection.