Generation of an In Vitro Cell Culture Model of Malaria-HIV Co-Infection

To set up the co-infection cultures, begin by seeding 1 x 10⁶ of freshly-isolated HIV-positive and HIV-negative PBMC in 100 microliters of RPMI-S+ per well into a 24-well plate. Bring the total volume of each well up to 500 microliters with 400 microliters of RPMI-S+.

Then, in triplicate and in a total volume of 500 microliters per well, add 3 x 10⁶ uninfected red blood cells or 3 x 10⁶ parasitized red blood cells to the HIV-infected and HIV-uninfected cells. Place the co-infection plate in a cell culture incubator.

Then, after the appropriate experimental time period, pellet the cells, and collect 700 microliters of culture supernatant from each well. Centrifuge the supernatant to remove any debris. Then, aliquot the samples as appropriate; label the tubes, and freeze them at minus 20 degrees Celsius or below for later analysis.