Plasmodium falciparum: Development and validation of a measure of intraerythrocytic growth using SYBR Green I in a flow cytometer

• Published in: Experimental parasitology 2009-02, Vol.121 (2), p.144-150
• Main Authors: Izumiyama, Shinji, Omura, Mako, Takasaki, Tomohiko, Ohmae, Hiroshi, Asahi, Hiroko
• Format: Article
• Language: English
• Subjects: Animals; Erythrocytes - parasitology; Flow cytometry; Flow Cytometry - instrumentation; Flow Cytometry - methods; Fluorescent Dyes; L-Lactate Dehydrogenase - analysis; Merozoite test; Organic Chemicals; Parasite growth; Plasmodium falciparum; Plasmodium falciparum - enzymology; Plasmodium falciparum - growth & development; Protozoan Plasmodium falciparum; Sensitivity and Specificity; SYBR Green I
• ISSN: 0014-4894; 1090-2449
• DOI: 10.1016/j.exppara.2008.10.008

Reliable analytical techniques to test growth-promoting and antimalarial efficacy on plasmodia are very important. Flow cytometry (FCM) offers the possibility to study developmental stages of intraerythrocytic growth of malaria parasites using nucleic acid staining. To analyze the growth of Plasmodium falciparum SYBR Green I was introduced as an intercalating dye with FCM for the 488 nm line of an argon laser. Procedures employing FCM, including fixatives, dye concentrations, dilution buffer, and staining period, were optimized to simplify the method. FCM as described here allows parasitemia and parasites of different stages to be quantified according to the DNA content. The proportion of parasitized erythrocytes estimated by FCM and the Giemsa method agreed with determination by parasite lactate dehydrogenase. The protocol was extended to merozoite counting as a sensitive assay of growth inhibition of the parasite.