Secreted Frizzled Related Protein 2 (SFRP2) is a key modulator of the Wnt signaling pathway. It harbors a frizzled-like cysteine-rich domain (CRD) that shares homology with the Wnt-binding region of Frizzled receptors, and a C-terminal netrin domain. As a core member of the SFRP family, SFRP2 localizes in the extracellular space and exerts dual regulatory effects on Wnt signaling: it acts as an antagonist of canonical Wnt signaling while activating the noncanonical Wnt/Ca²⁺ pathway, participating in embryonic development, angiogenesis, tissue regeneration and tumorigenesis. It is widely expressed in the alimentary system, brain, embryo mesenchyme and other tissues, and its abnormal expression is associated with cardiovascular diseases and various cancers. SFRP2 binds directly to WNT2 via its CRD domain, blocking WNT2's interaction with Frizzled receptors to inhibit canonical Wnt signaling.Thus a functional ELISA assay was conducted to detect the interaction of recombinant rat SFRP2 and recombinant human WNT2. Briefly, SFRP2 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to WNT2-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-SFRP2 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 rat SFRP2 and recombinant human WNT2 was shown in Figure 1, the EC50 for this effect is 0.181 µg/ml.