Simple and inexpensive fluorescence-based technique for high-throughput antimalarial drug screening.
Radioisotopic assays involve expense, multistep protocols, equipment, and radioactivity safety requirements which are problematic in high-throughput drug testing. This study reports an alternative, simple, robust, inexpensive, one-step fluorescence assay for use in antimalarial drug screening. Parasite growth is determined by using SYBR Green I, a dye with marked fluorescence enhancement upon contact with Plasmodium DNA. A side-by-side comparison of this fluorescence assay and a standard radioisotopic method was performed by testing known antimalarial agents against Plasmodium falciparum strain D6. Both assay methods were used to determine the effective concentration of drug that resulted in a 50% reduction in the observed counts (EC(50)) after 48 h of parasite growth in the presence of each drug. The EC(50)s of chloroquine, quinine, mefloquine, artemisinin, and 3,6-bis-epsilon-(N,N-diethylamino)-amyloxyxanthone were similar or identical by both techniques. The results obtained with this new fluorescence assay suggest that it may be an ideal method for high-throughput antimalarial drug screening.
Smilkstein
Martin
M
Medical Research Service, Department of Veterans Affairs Medical Center, Oregon Health and Science University, Portland, Oregon, USA. smilkste@earthlink.net
Sriwilaijaroen
Nongluk
N
Kelly
Jane Xu
JX
Wilairat
Prapon
P
Riscoe
Michael
M
D000818:Animals; D000962:Antimalarials / Q000494:pharmacology*; D016054:DNA, Protozoan / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004912:Erythrocytes / Q000469:parasitology; D019856:Ethanolamine / Q000378:metabolism; D005456:Fluorescent Dyes; D005615:Freezing; D006801:Humans; D066298:In Vitro Techniques; D008856:Microscopy, Fluorescence; D010963:Plasmodium falciparum / Q000187:drug effects / Q000378:metabolism; D016053:RNA, Protozoan / Q000737:chemistry
D000962:Antimalarials; D016054:DNA, Protozoan; D005456:Fluorescent Dyes; D016053:RNA, Protozoan; D019856:Ethanolamine