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Figure 1 | Retrovirology

Figure 1

From: Partial rescue of V1V2 mutant infectivity by HIV-1 cell-cell transmission supports the domain’s exceptional capacity for sequence variation

Figure 1

Assessing free virus infection and cell-cell transmission with the inGLuc reporter system. (A) Evaluation of cell-cell transmission between 293-T donor cells and A301-CCR5 target cells. The optimal input of 293-T donor cells in co-culture setups with A3.01-CCR5 target cells was determined by titration of 293-T donor cells transfected with the indicated JR-FL env variants or mock control (no env). A3.01-CCR5 target cells were kept constant at 50.000 cells. 100 μl donor cell input corresponds to approximately 15.000 293-T cells. The grey box highlights the donor cell input chosen for subsequent assays (100 μl) since this input best covered a wide range of envelope infectivities while still being in the dynamic range of the assay. Mean and SD of two independent experiments with two replicates are shown. Inset: an input of 100 μl 293-T cells transfected with JR-FL wt reporter virus was treated with the RT inhibitor Tenofovir or the protease inhibitor Lopinavir prior to starting the cell-cell co-culture. Both inhibitors completely abolished target cell infection. (B) Absolute GLuc reporter signals obtained in cell-cell transmission and different free virus infection setups were compared. The relative light units (RLUs) are in all cases normalized to 100 μl infectious input derived from the same donor cells, either transfected 293-T cells (cell-cell assay) or free virus stock (free virus assays). As target cells either A3.01-CCR5 cells (50.000 cells for both cell-cell and free virus assays) or TZM-bl cells (15.000 cells) were used. Mean and SD of two independent experiments with 2 replicates are shown.

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