Abstract Some research groups have produced immunodeficient mice with human liver tissue as a model system for the analysis of drug metabolism and liver regeneration. Mouse models are important for research and development of drugs and vaccines for viral infections. Recent progress in developing humanized mouse models permits studies of adaptive immune responses, innate host responses, and therapeutic approaches for several liver diseases of viral etiology. In this study, we generated a humanized liver mouse model by transplant with fresh patient-derived hepatocytes (1 × 106 cells/mouse, intrasplenic injection) into preconditioned (50 mg/kg ganciclovir, intraperitoneal injection) mice (herpes simplex virus type 1 thymidine kinase [TK] transgene expressed within the liver of a highly immunodeficient mouse strain [NOG]). Successful reconstitution of human hepatocytes in TK-NOG mouse liver tissues was observed with a strong proliferation of human cells in a time-dependent manner, using cytokeratin 8/18 stain. Similarly, we detected significantly increased human albumin levels in TK-NOG mouse liver tissue and blood sera on immune staining and enzyme-linked immunosorbent assay. Therefore, this humanized liver mouse model provides a biomedical tool for studying human liver physiology, drug metabolism, and liver pathogenesis of viral etiology or liver regeneration.