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Fig. 2 | Malaria Journal

Fig. 2

From: Progress and challenges in the use of fluorescence‐based flow cytometric assays for anti‐malarial drug susceptibility tests

Fig. 2

Proposed key features of fluorochrome-based flow cytometry for high-throughput antimalarial drug susceptibility testing. In a conventional drug susceptibility test, various doses of antimalarial drugs need to be evaluated; thus, the assay requires a protocol suitable for a multiwell format. Taking advantage of DNA-binding fluorochromes, key characteristics of flow cytometer-based drug susceptibility assays are proposed. Briefly, Plasmodium-infected erythrocytes are incubated with cell-permeant fluorochromes without cell fixation or harvesting. Given the fluorescence enhancement (increase in emitted fluorescence intensity or a Stokes shift upon binding to nucleic acids), the stained cells are directly exposed to laser without removal of unbound fluorochrome, eliminating a cell washing step. Highly selective DNA binding of fluorochromes minimizes the effects of confounding factors derived from RNA binding in reticulocytes. Since several wells are analysed, the fluorochrome must resist photobleaching. In addition, the high resolution of fluorescence intensity further allows discrimination of ring-form or mature trophozoites or schizonts, characterizing stage-specific drug resistance

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