Purpose: Cholangiocarcinoma (CC) is characterized by a still unfavorable prognosis due to a late diagnosis and high recurrence rate. In this retrospective study prognostic factors for long-term survival and an immunohistochemical panel of markers (IHC) for distinction of CC and other primary liver malignancies were analyzed. Materials and Methods: In 208 patients with CC clinical data, tumor characteristics and the primary mode of treatment were analyzed using univariate and multivariate statistics. In 145 cases the immunohistochemical profile of the tumors was established using markers such as CK7, CK20, CA19-9, HepPar-1, AFP, CD34 and CDX2 routinely in comparison to 60 cases of HCC (control group). Significance of marker expression in relation to histological subtype were estimated using SPSS 10.0. Results: Median overall survival (OS) was 18.8±22.5 months. Multivariate analysis identifies subtype of CC / tumor localisation (dCC vs. iCC or pCC), tumor size (P=0.001), grading (P=0.002) and tumor resection (P<0.0001) as independent prognostic factors for survival. CK7, CK20 and CA19-9 were the most commonly overexpressed markers in CC (86.2%, 55.2% and 50%), whereas expression of these markers in HCC was significantly less frequent (24.3%, 0% and 8.3%, respectively; P<0.001 and P<0.0001). Conclusions: Tumor localization and stage, surgical resection and tumor biology as expressed by tumor cell differentiation are the most important factors for OS in CC. Best predictive markers for differentiation between CC and HCC were CK7, CK20 and HepPar-1. Using this panel a fast and accurate differentiation was possible in more than 95% of all analyzed liver tumors.