Improvement of SYBR Green I−based fluorescence assay reading procedure for anti−babesial drugs screening in vitro

抄録

Drug susceptibility of the Babesia parasite in vitro has been used to assess the anti-babesial activities of new candidates. Recently, the in vitro drug susceptibility assay has been performed by SYBR Green I (SG I) -based fluorescence assay. Herein, we investigated the validity of a fluorescence assay for the shipping process under various culture conditions. The highest fluorescence signal was observed in freeze-thaw of Babesia and Theileria parasites cultures incubated with lysis buffer containing SG I for a half hour. This result indicates the ability of this additional step to lysis of infected RBCs completely with releasing of parasite nucleic acid. Such that, the nucleic acid can interact with SG I stain. All tested drugs showed potential anti-babesial activity and slightly high selective index against the in vitro growth of the Babesia and Theileria parasites. The IC₅₀ values generated using the fluorescence assay with additional freeze-thaw step is comparable with those of IC₅₀ values obtained from the unmodified fluorescence assay and microscopy method.