Expression profile of RSP-2 and PfEMP1 at the erythrocyte surface during the blood-stage cycle. RSP-2^CSA was stained with mAb B4 (green) and PfEMP1^CSA with mAb 1B4/D4 (red) by L-IFA on IE. Parasite nuclei are stained by DAPI (blue). (A) Positive anti–RSP-2 staining on synchronized rIEs (< 16 hours after invasion) and early trophozoites (16 to 20 hours after invasion). No surface staining was detectable on mature forms (> 20 hours after invasion). (B) Absence of PfEMP1 staining on rings (< 16 hours) but strong IFA signal with early trophozoite and mature stages. (C) Superposition of anti–RSP-2 and anti-PfEMP1 staining shows colocalization in early trophozoite stages. Scale bar measures 10 μm.