Synthetic peptides reproducing 4 DRADGQPAG (D4) and a sequential array of DRADGQPAG and DRAAGQPAG repeats (DDAAD) of the Plasmodium vivax circumsporozoite (CS) protein were investigated for their potential use in the detection of P. vivax sporozoite antibodies in human sera. These peptides specifically inhibited the binding of monoclonal antibodies to the P. vivax CS protein in Western blots. However, when D4 and DDAAD peptides were used in an enzyme-linked immunosorbent assay (ELISA) for the detection of human antibodies, more sera bound to the DDAAD (61%) than to the D4 peptide (22%). This binding was specific, and suggested that the DDAAD peptide contained epitopes constituted by the sequential array of DRADGQPAG and DRAAGQPAG repeat variants and absent in the D4 peptide. The ELISA using the DDAAD peptide was applied to the detection of P. vivax CS protein antibodies in a large number of sera from Kataragama, an endemic area in Sri Lanka. The prevalence of these antibodies increased with age, reaching 40% in adults > 50 years old. The ELISA employing the DDAAD peptide represents a simple and useful tool for the analysis of the antibody response to P. vivax sporozoites in naturally exposed individuals.