The disease-causing blood-stage of the Plasmodium falciparum lifecycle begins with invasion of human erythrocytes by merozoites. P. falciparum merozoites have two types of secretory organelles (micronemes and rhoptries) whose sequential release is essential for invasion and intracellular development. During invasion, large bulb-like rhoptry organelles inject an array of invasion and virulence factors into the cytoplasm of the host red blood cell, but the molecular mechanism mediating rhoptry exocytosis is unknown. Here we identify a set of parasite specific proteins, termed P. falciparum Cytosolically Exposed Rhoptry Leaflet Interacting protein (PfCERLI) 1 and 2 that cap the extremity of the rhoptry. Reduction in CERLI expression interferes with proteolytic processing blocking secretion of key rhoptry antigens that coordinate merozoite invasion and therefore proliferation in P. falciparum. Lipid binding domains identified within CERLIs are likely contributors to assembling the machinery that docks the rhoptry to the plasma membrane prior to fusion. While further studies are needed to determine the fine detail of how PfCERLI knockdown causes these changes in rhoptry function, identification of PfCERLI1’s direct association with release of rhoptry antigens is a key step in understanding the complex molecular events that control rhoptry secretion during invasion. This study makes extensive use of quantitative super-resolution microscopy, image analysis and computer vision to provide important mechanistic insight into a parasite specific exocytic pathway, essential for the establishment of infection.