Cluster Of Differentiation 200 (CD200), also known as OX-2 or MOX1/MOX2, is a membrane glycoprotein belonging to the immunoglobulin superfamily. This protein is widely expressed in various cell types, including neurons, endothelial cells, certain immune cells (such as B cells, dendritic cells, T cell subsets), and various tumor cells. CD200 is a multifaceted protein with important roles in immune regulation, neuronal function, and disease pathogenesis. Besides, Cluster Of Differentiation 8a (CD8a) has been identified as an interactor of CD200, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant rat CD200 and recombinant human CD8a. Briefly, CD200 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to CD8a-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-CD200 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 450/630 nm immediately. The binding activity of recombinant rat CD200 and recombinant human CD8a was shown in Figure 1, the EC50 for this effect is 0.09 ug/mL.