A simple, ex vivo phagocytosis assay of P. vivax merozoites by flow cytometry

07 Oct 2019
Elizangela Farias, Fhabiane Bezerra, Djane Clarys Baia-da-Silva, Yury Oliveira Chaves, Tatiana Bacry Cardoza, Maria Edilene Martins de Almeida, Lucas Barbosa Oliveira, Pritesh Lalwani, Patrícia Puccinelli Orlandi, Marcus Vinicius Guimaraes Lacerda, Stefanie Costa Pinto Lopes, and Paulo Afonso Nogueira

As phagocytosis is the first line of defense against malaria, we developed a phagocytosis assay with Plasmodium vivax (P. vivax) merozoites that can be applied to evaluate vaccine candidates. Briefly, after leukocyte removal with loosely packed cellulose powder in a syringe, P. vivax trophozoites matured to the merozoite-rich schizont stages in the presence of the E64 protease inhibitor. The Percoll gradient-enriched schizonts were chemically disrupted to release merozoites that were submitted to merozoite opsonin-dependent phagocytosis in two phagocytic lines with human and mouse antibodies against the N- and C-terminus of P. vivax Merozoite Surface Protein-1 (Nterm-PvMSP1 and MSP119). The resulting assay is simple and efficient for use as a routine phagocytic assay for the evaluation of merozoite stage vaccine candidates.