Small-scale culture of Plasmodium falciparum using μ-Slide Angiogenesis followed by automatic infection rate counting to assess drug effects

Abstract

It is very important to reduce the costs involved in malarial drug development by small-scale culture of Plasmodium falciparum, and automation of the assay system for drug efficacy against the parasites for high-throughput screening. In this study, we report that P. falciparum-infected erythrocytes can be stably cultured on μ-Slide Angiogenesis, which is used to investigate angiogenesis in tube formation assays, followed by automatic counting of the infection rate (parasitaemia). After 10 μL of parasite-infected erythrocytes were added to the inner well of μ-Slide Angiogenesis to prevent a multilayer of erythrocytes, 30 μL of silicon oil was overlaid on the culture medium to avoid evaporation of the medium, leading to stable small-scale parasite cultivation. The parasites were stained with a cell-permeant fluorescent nucleic acid stain (SYTO21) followed by cultivation. After taking bright field and fluorescent images using an inverted microscope, the infection rate could be calculated automatically by counting the number of erythrocytes and parasites using MetaMorph Offline software. The effect of anti-malarial drugs on parasite growth could be investigated on μ-Slide Angiogenesis, in which the parasite culture was added to the inner wells containing the drugs followed by their cultivation. Taken together, this method may be useful for image-based screening for anti-malarial drug candidates with automatic counting of parasite infection rates.

Keywords: Drug screening; Image-based analysis; Plasmodium falciparum; Small-scale culture; μ-Slide Angiogenesis.