Efficient malaria treatment is a major healthcare challenge. Addressing this challenge requires in-depth understanding of malaria parasite maturation during the intraerythrocytic cycle. Exploring the structural and functional changes of the parasite through the intraerythrocytic stages and their impact on red blood cells (RBCs) is a cornerstone of antimalarial drug development. In order to precisely trace such changes, we performed a thorough imaging study of RBCs infected by Plasmodium falciparum, by using atomic force microscopy (AFM) and total internal reflection fluorescence microscopy (TIRF) supplemented with bright field microscopy for stage assignment. This multifaceted imaging approach allows to reveal structure–function relations via correlations of the parasite maturation with morphological and fluorescence properties of the stages. We established diagnostic patterns characteristic to the parasite stages based on the topographical profile of infected RBCs, which show close correlation with their fluorescence (TIRF) map. Furthermore, we found that hemozoin crystals exhibit a strong optical contrast, possibly due to the quenching of fluorescence. The topographical and optical features provide a tool for locating the hemozoin crystals within the RBCs and following their growth.