FRAP analysis of eosin-labeled band 3 in control and parasitized erythrocytes. Aliquots of cultures of wild-type (K⁺) and KAHRP knock-out (K^-) 3D7 strain malaria parasites in erythrocytes (∼5% parasitemia) were treated with eosin-maleimide to label band 3, then permeabilized with SLO. (A) The inset shows a differential interference contrast (DIC) image of an erythrocyte infected with a mature-stage parasite containing the dark hemozoin pigment with an uninfected erythrocyte below it. The fluorescence images of the same cells are shown before, immediately after, and 10 minutes after application of the bleach pulse at the center of each cell. Scale bar is 2 μm. The graph shows the typical kinetics of fluorescence recovery into the bleached region for an uninfected erythrocyte (○) and for erythrocytes infected with mature stage K⁺ (▵) or K^- (▿) parasites. Data points shown at negative times represent fluorescence intensities prior to the bleach event. (B) The f_r(300) values for uninfected erythrocytes, K⁺ ring stage- and mature stage-infected erythrocytes and K^- ring stage- and mature stage-infected erythrocytes. The top number reflects the number of separate experiments and the bottom number the total number of individual measurements. Inset shows immunofluorescence analysis of KAHRP in K⁺ and K^- strains carried out using the B89 monoclonal anti-KAHRP antibody. Shown are bright field and corresponding fluorescence images of erythrocytes infected with ring (non–hemozoin-containing) and mature (hemozoin-containing) parasites. Scale as in panel A. Error bars indicate standard error.