To study the protein function of Tilapia Lake Virus (TiLV) ORF10, the TiLV ORF1 0.coding region was amplified from the cDNA of spleen and kidney tissues of TiLV-infected tilapia.The eukaryotic expression vector pEGFP-ORF1 0.and recombinant expression vector pET3 2.a-ORF1 0.were then constructed.pEGFP-ORF1 0.was transfected into Epithelioma papulosum cyprini (EPC) cells for fluorescence microscopy observation.Green fluorescence signals were observed in the nuclei of EPC, indicating that TiLV ORF1 0.was localized in the nucleus.The recombinant vector pET3 2.a-ORF1 0.was transformed into E.coli DE 3.for protein expression analysis, and the results showed that His-TiLV ORF1 0.was successfully expressed in the supernatant form.Furthermore, the fusion protein His-TiLV ORF1 0.was purified by Ni-NTA affinity chromatography and was subsequently used to immunize Balb/C mice to prepare polyclonal antibodies.Western blot analysis showed that mice antibodies can specifically recognize His-TiLV ORF10.To further analyze the protein content of TiLV ORF1 0.in the muscle, spleen, liver, and kidney tissues of TiLV-infected tilapia, the polyclonal antibodies were used for western blot detection.The results revealed differential protein contents of TiLV ORF1 0.in the muscle, spleen, liver, and kidney tissues.The highest expression was observed in the liver and kidney tissues, followed by the spleen.The lowest expression was found in the muscle tissues.The subcellular localization, polyclonal antibody preparation, and tissue expression analysis of TiLV ORF1 0.in this article provide an important preliminary basis for the further study of the function of the ORF1 0.protein.