Transfusion-related acute lung injury (TRALI) occasionally causes serious symptoms that may be fatal to recipients. Polymorphonuclear neutrophils (PMNs) and alloantibodies specific to PMN cell surface antigens are suspected to cause TRALI. The aim of this study is to establish a sensitive and stable procedure of detecting alloantibodies not only in donor blood, but also in recipient's plasma. We have introduced a new method of detecting alloantibodies based on double-determinant enzyme-linked immunosorbent assay (DD-ELISA) and a monoclonal antibody-immobilized granulocyte antigen (MAIGA) test (arbitrarily designated as modified DD-ELISA). We verified the specificity of alloantibodies against PMN cell surface antigens in plasma samples from three normal healthy donors of blood that induced respiratory distress in recipients after a blood transfusion. Anti-CD32 (Fc γ RIII) alloantibodies were detected in all the plasma samples using two different clones of the monoclonal anti-CD32 antibody. The specificities of these plasma samples could not be identified by the granulocyte immunofluorescence test (GIFT) using typed test cells. Except for the anti-CD32 alloantibodies, one plasma sample was proved to have the anti-HNA-1a alloantibodies. In another plasma sample, the anti-HNA-2a alloantibodies were detected. By modified DD-ELISA, we could clearly specify the presence of alloantibodies in the three plasma samples. Our results also suggest that the anti-CD32 alloantibodies can be generated in vivo and may play some roles in the development of TRALI.