The use of metrizamide for isopyknic separation and enrichment of Plasmodium falciparum schizonts from continuous culture.

Plasmodium falciparum-parasitized erythrocytes obtained from continuous in vitro cultures were fractionated over metrizamide density gradients. Late developmental stage schizonts were isolated from uninfected erythrocytes and other intracellular blood stage forms (rings and trophozoites) by centrifugation through 15% metrizamide. Schizonts comprised, on the average, 85% of the total number of cells recovered from the top fraction of the gradient with the remaining cells being predominantly uninfected erythrocytes. The mean percentage of schizonts recovered was 27% relative to the estimated number of mature forms present before fractionation. The concentrated schizonts were viable based upon their ability to complete schizogony and undergo a new cycle of reinvasion and schizogony in vitro. In addition, in these short term synchronous cultures the newly infected cells were able to incorporate radiolabeled hypoxanthine. This function was inhibited by the addition of chloroquine to these 45- to 46-hour cultures. Metrizamide-purified schizonts should be useful for further study and characterization of the unique metabolic, biochemical, and immunological properties of the malaria parasite.