The mutation of tumor suppressor p53 gene probably produce p53 antibodies. p53 antibodies are of importance to tumor's diagnosis, prognosis and relapse monitor. Current method to detect p53 antibodies, i.e. enzyme-linked immunosorbent assay (ELISA), requires long time with multi-step, and the assay is only semi-quantitative. Immunomagnetic electrochemiluminescence (IM-ECL) assay was devoloped for quantitative detection of p53 antibodies in human serum. Results indicate that the detection limit of this assay is 10 ng/L. A stable calibration curve with a wide linear range from 0.01 to 1 000 μg/L was established, thus, made quantitation possible. Serum samples from lung cancer patients were tested using the IM-ECL assay. The assay uses only 50 μl of sample per test and requires a 30 min incubation period in addition to a 50 s acquisition time. The positive rate of p53 antibodies was 28.6% in lung cancer sera. Concentrations of p53 antibody in the positive carcinoma sera were quantified from the calibration curve. A trend was found that higher the p53 antibody concentration in the lung cancer serum likely linked to a higher diagonostic stage of the cancer. IM-ECL assay is significantly better in terms of detection limit, linear range, and assay time, compared to the similar paramenters of common ELISA. The results suggests that IM-ECL is a feasible method for rapid, sensitive and quantitative detection of p53 antibodies in human serum.