Mucin 5AC (MUC5AC) is a glycoprotein found in different epithelial cancers, including biliary tract cancer (BTC). The aims of this study were to investigate the role of MUC5AC as serum marker for BTC and its prognostic value after operation with curative intent.From January 2007 to July 2012, a quantitative assessment of serum MUC5AC was performed with enzyme-linked immunoassay in a total of 88 subjects. Clinical and biochemical data (including CEA and Ca 19-9) of 49 patients with BTC were compared with a control population that included 23 patients with benign biliary disease (BBD) and 16 healthy control subjects (HCS).Serum MUC5AC was greater in BTC patients (mean 17.93 ± 10.39 ng/mL) compared with BBD (mean 5.95 ± 5.39 ng/mL; P < .01) and HCS (mean 2.74 ± 1.35 ng/mL) (P < .01). Multivariate analysis showed that MUC5AC was related with the presence of BTC compared with Ca 19-9 and CEA: P < .01, P = .080, and P = .463, respectively. In the BTC group, serum MUC5AC ≥ 14 ng/mL was associated with lymph-node metastasis (P = .050) and American Joint Committee on Cancer and International Union for Cancer Control stage IVb disease (P = .047). Moreover, in patients who underwent operation with curative intent, serum MUC5AC ≥ 14 ng/mL was related to a worse prognosis compared with patients with lesser levels, with 3-year survival rates of 21.5% and 59.3%, respectively (P = .039).MUC5AC could be proposed as new serum marker for BTC. Moreover, the quantitative assessment of serum MUC5AC could be related to tumor stage and long-term survival in patients with BTC undergoing operation with curative intent.