T-lymphotropic feline leukemia virus (FeLV-T) induces immunodeficiency in cats. FeLV-T is fusion-defective and requires a cofactor, termed FeLIX, for infection. FeLIX is a truncated envelope glycoprotein of an endogenous FeLV and mediates infection by binding a phosphate transporter Pit-1. In this study, we established a feline sarcoma-positive leukemia-negative cell line expressing FeLIX, named QN/FeLIX cells. Upon infection, FeLV-T induced prominent foci with syncytia in QN/FeLIX cells and could be titrated by the focus assay. In addition, we established a FeLIX-expressing feline fibroblast cell line, named AH/FeLIX cells. FeLV-T productively infected AH/FeLIX cells and induced severe CPE with syncytia. QN/FeLIX and AH/FeLIX cells will be useful for the study of FeLIX-dependent mutants in FeLV-infected cats.