Inhibin Beta A (INHbA) is a member of the transforming growth factor-beta (TGFb) superfamily that exerts a variety of biological functions, including immune response, sex determination, stem cell differentiation, and control of cellular migration and proliferation. Besides INHbA is aberrantly expressed in multiple tumor types, including breast, lung, esophageal, bladder, and colorectal cancers, and this overexpression is positively correlated with poor prognosis. Mature INHbA homodimerizes to form activin A, which has roles in the immune response, cell differentiation, and glucose metabolism, and heterodimerizes with INHbB to form inhibin, a complex that negatively regulates gonadal stromal cell proliferation and has tumor suppressor activity. Thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human INHbA and recombinant rat INHbB. Briefly, biotin-linked INHbA were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100ul were then transferred to INHbB-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 50 μl stop solution to the wells and read at 450 nm immediately. The binding activity of recombinant human INHbA and recombinant rat INHBB was shown in Figure 1, the EC50?for this effect is 2.2 ug/mL.