Introduction: The development of an effective vaccine against Hepatitis C virus (HCV) is still a target of intensive vaccine research. The HCV envelope proteins E1 and E2 which can induce broadly neutralizing antibodies are the major candidate for this purpose. Different types of expression systems have been used to express these glycoproteins. In this study, an expression system using Pichia pastoris was used to express E1 and E2 in full length. Methods: E1 and E2 regions containing the restriction sites from HCV 1b were separately amplified and cloned into a pPICZAa vector. The km71h strain of P. pastoris was transfected with the confirmed vectors separately using electroporation. The recombinant E1 and E2 proteins were evaluated for their antigenicity in an ELISA test and the induction of humoral immunity in mice. Results: The expression of full length HCV glycoproteins E1 and E2 in P. pastoris strain km71h was successfully achieved and their specific antibody was detected in serum samples from HCV infected patients. Furthermore, the recombinant glycoproteins could elicit a significant humoral immunity in mice as a vaccine candidate. Conclusion: P. pastoris is one of the best eukaryotic expression systems for the production of HCV glycoproteins in full length and the expressed proteins could be used in diagnostic tests such as ELISA. The induction of humoral immune responses in mice should lead to further studies on these glycoproteins for designing an effective vaccine.