A whole blood assay for antibody dependent phagocytosis of Plasmodium falciparum infected erythrocytes

Background: Antibodies are used to protect against Plasmodium falciparum malaria. One antibody target, the variant surface antigens, is expressed on infected erythrocytes (IEs). Antibodies to these antigens can either block IE sequestration in the tissues, facilitate natural killer cell-mediated killing, or opsonise IEs for phagocytic clearance by neutrophils and monocytes. 

Methods: We developed a high-throughput assay to measure antibody-dependent neutrophil phagocytosis (ADNP) and antibody-dependent cellular phagocytosis (ADCP, by blood monocytes) in the same sample of fresh whole blood. Results: Here we show that immune plasma mediates ADNP and ADCP in a concentration-dependent manner. Uptake is greater in the presence of complement proteins and is largely dependent on the expression of P. falciparum Erythrocyte Membrane Protein 1 located on the IE surface. Plasma from pregnant Papua New Guinean women with and without placental malaria shows that ADNP and ADCP are associated with protection from placental malaria. ADNP, but not ADCP, using IEs expressing IT4VAR19 (a PfEMP1 variant that binds to endothelial protein C receptor through a DC8 domain cassette) is higher at hospital presentation in children with uncomplicated malaria than in severe malaria. In pregnant women, ADNP and ADCP in whole blood are strongly correlated with one another (Spearman’s rho = 0.90), but not with ADNP or ADCP using purified neutrophils and monocytes in the absence of complement proteins.

Conclusions: The whole blood assay is a powerful new tool to assess functional antibodies that may protect against P. falciparum malaria. It allows simultaneous measurement of phagocytosis of opsonised IEs by monocytes and neutrophils.