Extracorporeal photopheresis (ECP) is a therapeutic procedure that is increasingly recognized for its efficacy in treating immune-mediated diseases, including transplant rejection. Its main mechanism is ex vivo apoptosis induction in leukocytes from patients by incubation with 8-methoxypsoralen and irradiation with ultraviolet A light. The process involves DNA cross-linking, which leads to a cascade of events within the cell and ultimately to apoptosis induction. Although ECP has been used for almost 40 y, there remain many questions about its immunological mechanisms and therapeutic potential. Here, we review current knowledge about mechanisms of apoptosis induction in subsets of peripheral blood mononuclear cells and interactions of apoptotic leukocytes with immune cells. We also highlight the challenges of reproducibly inducing cell death in a clinical manufacturing procedure and propose innovative ways to improve and quality-control ECP photopheresates.