Plasmodium falciparum: development of a transgenic line for screening antimalarials using firefly luciferase as the reporter.
High-throughput screening (HTS) of small-molecule libraries against pharmacological targets is a key strategy of contemporary drug discovery. This study reports a simple, robust, and cell-based luminescent method for assaying antimalarial drugs. Using transfection technology, we generated a stable Plasmodium falciparum line with high levels of firefly luciferase expression. A luciferase assay based on this parasite line was optimized in a 96-well plate format and used to compare with the standard [(3)H] hypoxanthine radioisotope method. The 50% inhibitory concentrations (IC(50)s) of chloroquine, artesunate, artemether, dihydroartemisinin and curcumin obtained by these two methods were not significantly different (P>0.05, ANOVA). In addition, this assay could be performed conveniently with a luminescence plate reader using unsynchronized stages within as early as 12h. Furthermore, the luciferase assay is robust with a Z' score of 0.77-0.92, which suggests the feasibility for further miniaturization and automation.
Cui
Long
L
Department of Entomology, The Pennsylvania State University, 501 ASI Building, University Park, PA 16802, USA.
Miao
Jun
J
Wang
Jiaxu
J
Li
Qianjun
Q
Cui
Liwang
L
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics* / Q000378:metabolism; D000962:Antimalarials / Q000494:pharmacology*; D003001:Cloning, Molecular; D004912:Erythrocytes / Q000469:parasitology; D017930:Genes, Reporter; D006801:Humans; D020128:Inhibitory Concentration 50; D016014:Linear Models; D049409:Luciferases, Firefly / Q000096:biosynthesis / Q000235:genetics*; D049408:Luminescent Agents; D010963:Plasmodium falciparum / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics*; D015203:Reproducibility of Results; D014162:Transfection
D000962:Antimalarials; D049408:Luminescent Agents; D049409:Luciferases, Firefly