Tumor Necrosis Factor Related Apoptosis Inducing Ligand (TRAIL), also designated TNFSF10, is a type II transmembrane protein belonging to the TNF superfamily. It functions as a key cytokine that selectively triggers extrinsic apoptosis in cancer cells while sparing most normal cells, making it a promising candidate for anticancer therapy. TRAIL exists as a homotrimer and binds to death receptors DR4 and DR5 to activate caspase?mediated apoptotic signaling. It is widely expressed in immune cells and various tissues, participating in immune surveillance and tumor suppression. Unlike many TNF family members, TRAIL exhibits minimal systemic toxicity due to decoy receptors in normal tissues that block excessive signaling.TRAIL does not directly bind FAS; they share parallel apoptotic pathways via distinct death receptors.To detect the activity of recombinant TRAIL , a functional ELISA assay was performed to evaluate the interaction between recombinant dog TRAIL and recombinant human FAS. Briefly, TRAIL was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to FAS-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-TRAIL 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 dog TRAIL and recombinant human FAS was shown in Figure 1, the EC50?for this effect is 0.012 μg/mL.