Interleukin 33 (IL33) is a dual-function protein that acts both as a traditional cytokine and an intracellular nuclear factor. It is primarily released by endothelial and epithelial cells upon cellular damage or stress, serving as an "alarmin" to alert the immune system. IL33 signals through its receptor ST2 to drive type 2 immune responses, which are crucial in allergic inflammation, host defense against parasites, and tissue repair.Besides,Thymic Stromal Lymphopoietin (TSLP) has been identified as an interactor of IL33, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant mouse IL33 and recombinant human TSLP. Briefly, IL33 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to TSLP-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-IL33 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 mouse IL33 and recombinant human TSLP was shown in Figure 1, the EC50?for this effect is 0.23 μg/mL..