P falciparum merozoites bind to CD44. (A) Flow cytometry–based assay to quantify binding between free P falciparum merozoites and recombinant CD44-Fc or CD99-Fc proteins. Free merozoites were incubated with 3 μg recombinant proteins, and binding was detected using anti-CD44 BRIC 222 (IBGRL, 1:1000) or anti-CD99 (Invitrogen; 1:250) antibodies. (B) Flow cytometry–based assay to quantify binding between free P falciparum merozoites and recombinant CD44-His or GYPC-His proteins. Histograms depicting merozoite fluorescence upon incubation with recombinant His-tagged proteins and detection with anti–6x-His mouse monoclonal antibody (Invitrogen; 1:300). (C) Images of immunofluorescence assays of free P falciparum merozoites after binding to rCD44-His, and its colocalization with the merozoite protein RAP1. RAP1 was detected using anti-RAP1/2 mouse monoclonal antibody (from Anthony Holder; 1:500), and CD44 was detected with anti-CD44 rabbit polyclonal antibody (Invitrogen; 1:500), followed by goat anti-mouse AlexaFluor 488 and goat anti-rabbit AlexaFluor 555. Images were taken with a Keyence BZ-X700 all-in-1 fluorescence microscope at ×100 original magnification under oil immersion.