Flow cytometric readout based on Mitotracker Red CMXRos staining of live asexual blood stage malarial parasites reliably assesses antibody dependent cellular inhibition

摘要: Background: Functional in vitro assays could provide insights into the efficacy of malaria vaccine candidates. For estimating anti-parasite effect induced by a candidate, an accurate determination live parasite count is essential component most bioassays. Although traditionally parasites are counted microscopically, faster, more and less subjective method for counting desirable. In this study mitochondrial dye (Mitotracker Red CMXRos) was used obtaining reliable counts through flow cytometry. Methods: Both asynchronous tightly synchronized asexual blood stage cultures Plasmodium falciparum were stained with CMXRos subjected to detection cytometry fluorescence microscopy. The obtained compared standard microscopic examination Giemsa-stained thin smears. A comparison ability stain compromised (induced either medium starvation or anti-malarial drug treatment) carried out. Finally, staining determine specific growth inhibition index (SGI) antibody-dependent cellular (ADCI) assay. Results: Mitotracker can reliably detect intra-erythrocytic stages P. falciparum. Comparison between shows that predominantly stains functional mitochondria. Parasite highly reproducible IgG from hyper-immune individuals inhibit presence monocytes ADCI Further, dose-dependent inhibitory be detected both total purified sera affinity IgGs against N-terminal non-repeat region GLURP coupled Conclusions: based on populations has been optimized. This rapid sensitive high inter-assay reproducibility which mediated antibodies such as