B. Preparation of compound solution: The test compound can be dissolved in the organic solvent at a concentration of 5mg/ml. It is recommended to use DMSO as solvent has no significant effect on parasite growth.

C. Setup of the assay: Parasite culture synchronized at ring stage by D-sorbitol treatment brought to the 1% parasitemia with 3% hematocrit. In a total volume of 100µl, 50µl parasite culture is mixed with the various concentration of test compound (0, 1.5, 3.0, 6.25, 12.5, 25, 50µg/ml) in 25µl and remaining complete media. Chloroquinine can be added as “positive control” and sovent as “negative control”. Incubate the compounds for 48hrs. Monitor the appearance of hemolysis or any such effect. If appeared, stop the assay and screen the compounds using other assay.  

D. Monitoring the growth of parasite: After 48hrs, After exposure, smears were made. Parasitemia has been determined after JSB staining (Fields’ stain) using oil immersion objective.

Figure 30.5: Over-view of the microscopy based antimalarial assay