Objective To establish a red-green dual-color fluorescence tracing orthotopic transplantation model of HCC.Methods Stable high red fluorescent protein (RFP)-expressing cells HepG2-RFP and Hepa1-6-RFP were injected into the right liver lobe of green fluorescent protein (GFP)-expressing nude mice.The growth and metastasis of tumor were visualized by whole-body in vivo fluorescence imaging system in real time.HCC tissues were removed from tumor-bearing mice, and 5-μm-thick serial frozen slices were made.Later, the transplanted tumors was observed under the microscope.