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Alliance"],"published-print":{"date-parts":[[2021,7]]},"abstract":"<jats:p>Progress towards a protective vaccine against malaria remains slow. To date, only limited protection has been routinely achieved following immunisation with either whole-parasite (sporozoite) or subunit-based vaccines. One major roadblock to vaccine progress, and to pre-erythrocytic parasite biology in general, is the continued reliance on manual salivary gland dissection for sporozoite isolation from infected mosquitoes. Here, we report development of a multi-step method, based on batch processing of homogenised whole mosquitoes, slurry, and density-gradient filtration, which combined with free-flow electrophoresis rapidly produces a pure, infective sporozoite inoculum. Human-infective<jats:italic>Plasmodium falciparum<\/jats:italic>and rodent-infective<jats:italic>Plasmodium berghei<\/jats:italic>sporozoites produced in this way are two- to threefold more infective than salivary gland dissection sporozoites in in vitro hepatocyte infection assays. In an in vivo rodent malaria model, the same<jats:italic>P. berghei<\/jats:italic>sporozoites confer sterile protection from mosquito-bite challenge when immunisation is delivered intravenously or 60\u201370% protection when delivered intramuscularly. 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