Malaria is associated with significant microcirculation disorders, especially when the infection reaches its severe stage. This can lead to a range of fatal conditions, from cerebral malaria to multiple organ failure, of not fully understood pathogenesis. It has recently been proposed that a breakdown of the glycocalyx, the carbohydrate-rich layer lining the vascular endothelium, plays a key role in severe malaria, but direct evidence supporting this hypothesis is still lacking. Here, the interactions betweenPlasmodium falciparuminfected red blood cells (PfRBCs) and endothelial glycocalyx are investigated by developing anin vitro, physiologically relevant model of human microcirculation based on microfluidics. Impairment of the glycocalyx is obtained by enzymatic removal of sialic acid residues, which, due to their terminal location and net negative charge, are implicated in the initial interactions with contacting cells. We show a more than twofold increase ofPfRBC adhesion to endothelial cells upon enzymatic treatment, relative to untreated endothelial cells. As a control, no effect of enzymatic treatment on healthy red blood cell adhesion is found. The increased adhesion ofPfRBCs is also associated with cell flipping and reduced velocity as compared to the untreated endothelium. Altogether, these results provide a compelling evidence of the increased cytoadherence ofPfRBCs to glycocalyx-impaired vascular endothelium, thus supporting the advocated role of glycocalyx disruption in the pathogenesis of this disease.