Berok Growth Inhibition Assay by Thiol-reactive Probe Based Flow Cytometric Measurement.

The relapsing malaria species, , is the most widely distributed and difficult-to-treat cause of human malaria. The merozoites of preferentially invade ephemeral human CD71 reticulocytes (nascent reticulocytes), thereby limiting the development of a robust continuous culture . Fortunately, 's sister species, Berok, can be cultured continuously, providing the ability to screen novel therapeutics drug and vaccine candidates in a reliable and high-throughput manner. Based on well-established growth inhibition activity (GIA) assays against and , this protocol adopts the current flow cytometry assay methodology and investigates inhibitory antibodies using the Berok invasion model based on the thiol-reactivity and DNA abundance of viable parasites in macaque erythrocytes. Established GIA assays screen antibodies at either a single concentration or high/low dose concentrations to provide quick insights for prioritizing potential antibodies capable of specifically interrupting parasite ligand and host receptor binding with minimal concentrations. Hence, this protocol expands on the existing GIA assay by using serially diluted antibodies and generating a dose-response curve to better quantify the inhibitory efficacy amongst selected vaccine candidates.