Interferon Alpha (IFNa) is a pleiotropic cytokine belonging to the type I interferon family, primarily secreted by immune cells such as plasmacytoid dendritic cells in response to viral infections and other pathogens. It plays a central role in the innate immune response by inducing the expression of hundreds of interferon-stimulated genes (ISGs) that exert antiviral, antiproliferative, and immunomodulatory effects. IFNa signals through a cell-surface receptor complex composed of IFNAR1 and IFNAR2, initiating a cascade of intracellular events that are critical for host defense against viruses and for regulating immune homeostasis. Dysregulation of IFNa signaling is implicated in various autoimmune diseases, highlighting its importance in maintaining immune balance. IFNa binding to its receptor induces the activation of TYK2, which is constitutively associated with the IFNAR1 subunit, thereby initiating downstream JAK-STAT signaling.Thus a functional ELISA assay was conducted to detect the interaction of recombinant cattle IFNa and recombinant human TYK2. Briefly, IFNa was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to TYK2-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-IFNa pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µL stop solution to the wells and read at 450/630nm immediately. The binding activity of recombinant cattle IFNa and recombinant hunan TYK2 was shown in Figure 1, the EC50 for this effect is 1.14µg/mL.