Transmembrane Protease, Serine 2 (TMPRSS2) is a type II transmembrane serine protease widely expressed in epithelial tissues, especially in the respiratory and gastrointestinal tracts. It plays critical roles in physiological processes like proprotein activation and tissue remodeling. Importantly, TMPRSS2 mediates viral entry by cleaving viral spike proteins, making it a key host factor in viral infections. TMPRSS2 binds to ACE2, facilitating the activation and entry of certain viruses into host cells.Thus a functional ELISA assay was conducted to detect the interaction of recombinant human TMPRSS2 and recombinant human ACE2. Briefly, TMPRSS2 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to ACE2-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-TMPRSS2 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 human TMPRSS2 and recombinant human ACE2 was shown in Figure 1, the EC50?for this effect is 2.14076μg/mL.