The only generally accepted serological marker currently used for the diagnosis of autoimmune pancreatitis (AIP) is IgG4. Our aim was mainly to determine whether hybrid kappa\lambda antibody can help to diagnose AIP and to differentiate it from pancreatic cancer. We established an enzyme-linked immunosorbent assay (ELISA) system to measure the levels of hybrid kappa\lambda antibodies in human sera. Sera were obtained from 338 patients, including 61 with AIP, 74 with pancreatic cancer, 50 with acute pancreatitis, 40 with ordinary chronic pancreatitis, 15 with miscellaneous pancreatic diseases, and 98 with normal pancreas. Our study showed levels of hybrid kappa\lambda antibodies in the AIP group were significantly higher than in the non-AIP group (P < 0.001). The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) for the diagnosis of AIP were 80.3%, 91%, 66.2% and 95.5% respectively. Furthermore, the combined measurement of serum hybrid kappa\lambda antibody and IgG4 tended to increase the sensitivity although the difference was not statistically significant (90.2% vs. 78.7%, P = 0.08), compared to measurement of IgG4 alone. Our findings suggest that hybrid kappa\lambda antibody could be a new serological marker to diagnose AIP and differentiate it from pancreatic cancer.