Abstract Background In the diagnosis of IgG4-related sclerosing cholangitis (IgG4-SC), differentiation from extrahepatic cholangiocarcinoma (ECC) is extremely important but is still a clinical challenge. This study aimed to elucidate the usefulness of peroral cholangioscopy (POCS) for the differential diagnosis between IgG4-SC and ECC. Methods POCS findings for bile duct stricture were retrospectively evaluated in 17 patients with IgG4-SC diagnosed at the Hiroshima University Hospital and 53 patients with surgically resected infiltrating ECC. Mucosal surface, dilated vessels (tortuosity, caliber alteration, and disruption), and easily bleeding were compared between the groups. Results The stricture sites of IgG4-SC evaluated by POCS were 10 extrapancreatic bile ducts and 9 intrapancreatic bile ducts. In patients with IgG4-SC, smooth mucosal surface was observed in 89% (17/19), dilated vessels in 58% (11/19) [tortuosity 82% (9/11), caliber alteration 18% (2/11), and disruption 9% (1/11)], and easily bleeding in 0%. Irregular mucosal surface and easily bleeding were observed significantly more frequently in ECC (both P < 0.001). The frequency of caliber alteration and disruption of dilated vessels was significantly less in IgG4-SC (P < 0.001 and 0.005, respectively). The sensitivity and specificity of POCS in the diagnosis of ECC were 96 and 89%, respectively. Dilated vessels in IgG4-SC were observed significantly more frequently in the extrapancreatic bile duct, especially the hilar bile duct (P = 0.006). Concerning image evaluation, the interobserver agreement was κ = 0.719, and the intraobserver agreement was κ = 0.768 and 0.754. Conclusions Characteristic POCS findings of the stricture sites in IgG4-SC were smooth mucosal surface, dilated vessels without caliber alteration and disruption, and lack of easily bleeding. These POCS findings are extremely useful for distinguishing between IgG4-SC and ECC.