Alpha-L-fucosidase (AFU) has been reported to be a predictor of survival in patients with several cancers, but it is unclear whether AFU is associated with prognosis in patients with intrahepatic cholangiocarcinoma (iCCA). In this study, we used receiver operating characteristic (ROC) analysis to generate the cutoff point of AFU for overall survival (OS). The prognostic influence of the AFU level in serum on OS was studied using Kaplan-Meier curves. Moreover, invasion assays and Western blotting were performed to explore the effects of AFU on iCCA invasion in vitro. We found that higher AFU levels (≥20.85 U/L) were significantly associated with favorable median OS (44.3 months versus 20.1 months; P = 0.022) in iCCA patients. Cox regression models' analyses showed that the AFU level was an independent predictor for OS (P = 0.006). Moreover, our results revealed that the AFU could impair the invasion capability of the iCCA cells, HuH28, and also downregulated the expression of matrix metalloproteinase 2 and matrix metalloproteinase 9. In conclusion, our results indicate that AFU is a significantly favorable prognostic factor in iCCA patients.