Avian erythroblastosis (AE; erythroid leukosis) was detected in a 78-day-old Japanese native chicken. At necropsy, the liver was enlarged and diffusely dark red in color. Moderate splenomegaly was observed. Histologically, round to polygonal tumor cells were observed only in the blood vessels of the liver and other organs. Immunohistochemistry (IHC) was performed on formalin-fixed, paraffin-embedded (FFPE) sections to characterize tumor cells. Tumor cells, as well as normal erythrocytes as positive controls, were consistently positive for IHC by using the commercially available anti-human hemoglobin antibody. Exogenous avian leukosis virus (ALV) subgroup B and endogenous ALV-E genes were detected by PCR, although ultrastructural observation revealed no viral particles associated with tumor cells. Results suggest that the commercial anti-human hemoglobin antibody used in the study cross-react to chicken erythrocytes on FFPE sections by IHC and can also be used for definitive diagnosis of the spontaneous case of AE in chickens.