Shelf-life study of Human Erythrocytes Cryopreserved for the Laboratory Culture of Plasmodium falciparum

Malaria is caused by multiple species of the parasite Plasmodium and disproportionately affects people living in the developing world who often do not have access or funds to allow for effective control of or protection from the parasite. This study is intended to support ongoing research at the Macha Research Trust (MRT) [also known as the Malaria Institute at Macha (MIAM)], which is located in Macha, a rural area in the Southern Province of Zambia where the virulent species, Plasmodium falciparum is prevalent. Our goal is to support the capacity of the laboratory at MRT to culture (propagate and preserve) locally isolated or laboratory strains of Plasmodium. Laboratory cultivation of P. falciparum requires fresh human blood. However, it is difficult to assure the steady supply of fresh, uninfected human blood needed to sustain culture experiments at MRT because blood from local residents cannot be used, and because many visiting scientists and physicians routinely take prophylactic anti-malarial drugs which can make their erythrocytes unable to support asexual propagation of, or gametocyte generation by P. falciparum in culture. We are investigating methods that should allow for the cryopreservation of erythrocytes obtained from uninfected individuals in the U.S. with subsequent shipment to Zambia. We have cryopreserved preparations of fresh, leukocyte-depleted erythrocyte suspensions using minimal aqueous volumes of solutions containing the macromolecular starchbased cryo-protectant hydroxy ethyl starch (HES) with or without polyvinyl alcohol (PVA). Such compounds act as ice recrystallization inhibitors (IRIs), preventing formation of ice crystals during the thawing process. We will describe ongoing efforts to determine if cryopreserved red blood cells (RBCs) will effectively support asexual propagation or gametocyte formation for the P. falciparum laboratory strain NF54 following long-term storage at -80°C. Further, the effect of a single short-term transfer to liquid nitrogen during the storage period (to stimulate transport) will be assessed