Paired primary and recrudescent Plasmodium falciparum isolates were collected from treatment failures identified during the course of antimalarial drug studies on the Thai-Cambodian border. Ten paired samples were subjected to PCR-restriction fragment length polymorphism (RFLP) and PCR-single-strand conformational polymorphism (PCR-SSCP) analysis of the MSP-2 gene. PCR-SSCP analysis of paired samples demonstrated that each recrudescent isolate was identical to, or a subpopulation of, its matched primary isolate and was distinct from all unrelated isolates. This method represents a field applicable method to distinguish re-infections from treatment failures when antimalarial drug studies are performed in malaria-endemic areas.