GDF9 (Growth/differentiation factor 9) is an oocyte derived growth factora which belongs to the transforming growth factor-beta (TGFβ) superfamily. GDF9 is required for ovarian folliculogenesis and promotes primordial follicle development. S100A8 has been identified as an interactor of GDF9 through two-hybrid assay, thus a binding ELISA assay was conducted to detect the interaction of recombinant human GDF9 and recombinant human S100A8. Briefly, GDF9 were diluted serially in PBS, with 0.01%BSA (pH 7.4). Duplicate samples of 100uL were then transferred to S100A8-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-GDF9 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 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 of GDF9 and S100A8 was shown in Figure 1, and this effect was in a dose dependent manner.