Interleukin 27A (IL27A, also known as p28) is a key subunit of the IL-27 cytokine, belonging to the IL-6/IL-12 family. It has a typical four-helix bundle structure, which is essential for its biological functions. Though it carries the core activity to regulate immune cell differentiation and mediate pro-inflammatory or anti-inflammatory effects, IL-27A is unstable and unable to be secreted or exert functions independently. IL-27A directly binds to EBI3 to form the functional IL-27 heterodimer.To detect the activity of recombinant IL27A, a functional ELISA assay was performed to evaluate the interaction between recombinant mouse IL27A and recombinant human EBI3.Briefly, biotin-linked IL27A were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100µl were then transferred to EBI3-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 IL27A and EBI3 was shown in Figure 1, the EC50 for this effect is 0.55µg/mL.