BACKGROUND: Vinculin is a cytoplasmic protein that binds to actin and is involved in cell adhesion. The presence of anti-vinculin autoantibodies in irritable bowel syndrome (IBS) is consistent with prior studies which have implicated vinculin as an important regulator of the enteric nervous system. OBJECTIVE: Therefore, in this study, we developed a testing protocol and optimized it before evaluating the value of the Anti-Vinculin quantitative test in the diagnosis of IBD and IBS. METHODS: To establish the procedure for quantification of anti-vinculin antibody (ATV) in serum by indirect ELISA and to investigate value of anti-vinculin testing in diagnosing irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD). RESULTS: After optimizing the procedure for ATV quantitative ELISA, we conducted the tests to determine the concentration of ATV in serum specimens. In this research, we use ATV results from 215 patients with IBS, 71 patients with IBD and 31 healthy subjects and assessed utility of ATV as a diagnostic test for IBS and to differentiate between IBS and IBD. On testing for ATV in patients with IBS and IBD, we found that it has potential for diagnosing patients with IBS, with an area under the curve (AUC) of 0.76, p <0.001With a cut-off of ATV concentration at 510.59 ng/mL, sensitivity of test was 48.8%, and specificity was 96.8%. CONCLUSION: The model of using ATVs to differentiate between IBD and IBS had not given optimal results. Further research is needed with larger sample sizes and uniformity among groups to come up with an effective model.