Effect of the intermolecular CSA distance on cytoadhesion of P falciparum–infected erythrocytes. (A) Cytoadhesion behavior of erythrocytes infected with FCR3^CSA or FCR3^Δvar2csa (trophozoite stage) or of uninfected erythrocytes (RBC) on functionalized membranes with different CSA spacing intervals (<d>). In panels A-D, 4 × 10⁷ cells mL⁻¹ were allowed to settle on the membranes for 60 minutes under controlled atmospheric conditions before unattached cells were washed out at a wall shear stress of 0.8 Pa. The means ± standard deviation (SD) of at least 5 biological replicates are shown. (B) Cytoadhesion behavior of erythrocytes infected with FCR3^CSA as a function of the intermolecular CSA distance and the incubation time. The following intermolecular distances between CSA molecules were investigated: 4, 6, 7, 8, 9, 11, 13, 16, 17, 34, and 54 nm, with an SD of 0.6 nm for all CSA distances. The data points were fitted using hyperbolic decay functions with Hill coefficient. (C) Cytoadhesion specificity of erythrocytes infected with FCR3^CSA versus erythrocytes infected with FCR3^Δvar2csa on supported membranes functionalized with pullulan or with the glycosaminoglycans CSA, hyaluronic acid (HA), or chondroitin sulfate C (CSC). (D) Area of tight contact between the functionalized membrane and the cells indicated as a function of the intermolecular CSA distance. Effect of CSA distance on cell deformation is shown in the reconstructed 3-dimensional height profiles of the bottom planes of a single erythrocyte infected with FCR3^CSA or an uninfected erythrocyte (RBC) resting on functionalized membranes, with a CSA spacing of 6 nm (E) and 11 nm (F). Representative height profiles of at least 10 biological replicates are shown.