Anticardiolipin antibodies (aCL) detected by standard ELISA are found in association with autoimmune and infectious diseases. It is now recognized that beta 2 glycoprotein I (beta 2GPI) is a cofactor for aCL binding to cardiolipin (CL). To examine differences in cofactor requirements, aCL positive sera from patients with the antiphospholipid syndrome (APS) and syphilis were studied. Using an ELISA with human purified beta 2GPI adsorbed onto irradiated plates, we detected high binding activity in 29 out of 35 samples from APS patients and low in only 1 out of 37 aCL positive syphilis sera. Moreover, a good correlation (r = 0.79) was also observed in the former group between aCL and anti beta 2GPI. Whole IgG and affinity purified IgG aCL from APS patients did not bind to CL in the absence of beta 2GPI, but recognized beta 2GPI on irradiated plates in the absence of phospholipids. In contrast, IgG purified from syphilis patients only bound to CL alone. Taken together, these data indicate that performing both ELISA (aCL and anti beta 2GPI) it could be possible to distinguish aCL from autoimmune or infectious diseases.