Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC) is antigenic and an important epitope of oxLDL. This study validates the assay for autoantibodies against oxPAPC (anti-oxPAPC-Ab) and investigates the possible association between anti-oxPAPC-Ab and cardiovascular disease. A synthetic PAPC was oxidatively modified as an antigen for the anti-oxPAPC-Ab assay. The concentrations of the antibody in serum were measured by EIA. The analytical parameters of the anti-oxPAPC-Ab assay were validated. Levels of anti-oxPAPC-Ab were prevalent in patients with hypertension, myocardial infarction (MI) and healthy subjects. Anti-oxPAPC-Ab specifically reacts with oxPAPC, but not with 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC). The characteristics of the assay included precision (inter-assay coefficients of variation were 7.9% for IgG and 13.2% for IgM), cross-reactivity, clinical sensitivity for hypertension (43% and 47%) and MI (37% and 41%), clinical specificity (95.2%) and normal values (less than 13 Unit/mL for IgG and less than 7 Unit/mL for IgM). Elevated levels of anti-oxPAPC-Ab were found in smoking populations, in patients with hypertension and MI. Anti-oxPAPC-Ab are significantly elevated in patients with hypertension and MI. A synthetic PAPC, after oxidation, was used to detect anti-oxPAPC-Ab, which may greatly enhance the reliability of the assay. The determination of anti-oxPAPC-Ab could serve as an autoimmune marker in the associating diagnosis of cardiovascular disease.