A unique sandwich enzyme-linked immunosorbent assay (ELISA) for the determination of ABH antigens in bloodstains has been developed. Human anti-A and anti-B sera and Ulex europaeus anti-H lectin were coated on the inner surface of microplate wells. The sample antigens from bloodstains, solubilized with n-octyl-ß-D-glucopyranoside which was then removed by passing through a Sephadex G-25 (G-25) mini-column, were placed in the wells. After washing the wells repeatedly, peroxidase-conjugated Ulex europaeus lectin I was added and incubated. Antigen activities were determined by the development of colors using o-phenylenediamine/H2O2. This technique permitted clear detection of all ABH antigens corresponding to the antisera and lectin with high sensitivities. The A and B antigens were solubilized as aggregates with H antigen from the erythrocyte membrane. Excess detergent remaining in the sample reduced the sensitivity and accuracy of this ELISA, probably due to the removal of antibody from the wells by the effect of the surfactant. The treatment of solubilized antigens with G-25, an indispensable step, eliminated the adverse effect of the detergent on the ELISA. The ELISA method reported here is described by the authors as easy, economical, sensitive, and potentially useful for clinical tests as well as for forensic purposes. Figures, references