Interferon Alpha (IFNa) is a cytokine belonging to the type I interferon family, primarily produced by plasmacytoid dendritic cells and other immune cells in response to viral infections. It functions as a critical mediator of innate immunity by binding to the interferon-alpha receptor (IFNAR) complex, activating the JAK-STAT signaling pathway. This induction leads to the expression of interferon-stimulated genes (ISGs), which establish an antiviral state in cells, inhibit viral replication, and modulate immune responses. IFNa also exhibits antitumor and immunoregulatory properties and is used therapeutically against viral infections and certain cancers.Besides,Interferon Alpha 10 (IFNa10) has been identified as an interactor of IFNa, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant dog IFNa and recombinant human IFNa10 .Briefly, biotin-linked IFNa were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100µl were then transferred to IFNa10-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50µl stop solution to the wells and read at 450nm immediately. The binding activity of IFNa and IFNa10 was shown in Figure 1, the EC50 for this effect is 0.38 µg/mL.