Circulating autoantibodies to phospholipids (aPLs), such as cardiolipin (CL), are found in patients with antiphospholipid antibody syndrome (APS). We recently demonstrated that many aPLs bound to CL only after it had been oxidized (OxCL), but not to a reduced CL analogue that could not undergo oxidation, We now show that the neoepitopes recognized by some aPLs consist of adducts formed between breakdown products of oxidized phospholipid and associated proteins, such as beta(2) glycoprotein 1 (beta(2)GP1). Addition of human beta(2)GP1, polylysine, native low-density lipoprotein, or apolipoprotein AI to OxCL-coated wells increased the anticardiolipin antibody (aCL) binding from APS sera that first had been diluted so that no aCL binding to OxCL could be detected, No increase in aCL binding was observed when these proteins were added to wells coated with reduced CL. The ability of beta(2)GP1, polylysine, or low-density lipoprotein to be a ''cofactor'' for aCL binding to OxCL was greatly reduced when the proteins were methylated, Incubation of beta(2)GP1 with oxidized 1-palmitoyl-2-linoleyl-[1-C-14]-phosphatidylcholine (PC), but not with dipalmitoyl-[1-C-14]-PC, led to formation of covalent adducts with beta(2)GP1 recognized by APS sera, These data suggest that the reactive groups of OxCL, such as aldehydes generated during the decomposition of oxidized polyunsaturated fatty acids, form covalent adducts with beta(2)GP1 (and other proteins) and that these are epitopes for aCLs. Knowledge that the epitopes recognized by many aPLs are adducts of oxidized phospholipid and associated proteins, including beta(2)GP1, may give new insights into the pathogenic events underlying the clinical manifestations of APS.