Glucagon (GCG) is a critical peptide hormone produced by alpha cells in the pancreatic islets. It serves as the primary counter-regulatory hormone to insulin, playing a central role in maintaining glucose homeostasis. During fasting states or low blood sugar (hypoglycemia), glucagon is secreted and binds to its receptor on liver cells. This binding triggers a signaling cascade that stimulates glycogenolysis (the breakdown of glycogen into glucose) and gluconeogenesis (the production of new glucose from non-carbohydrate sources). The resulting release of glucose into the bloodstream provides vital energy to the brain and other organs. Thus, glucagon is essential for preventing dangerously low blood sugar levels.Besides,Taste Receptor Type 1 Member 3 (TAS1R3) has been identified as an interactor of GCG, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human GCG and recombinant human TAS1R3. Briefly, biotin-linked TAS1R3 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100µl were then transferred to GCG-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 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. Measured by its binding ability in a functional ELISA. When Recombinant GCG is lmmobilized at 2 µg/mL(100 uLwell), the concentration of TAS1R3 that produces 50% optimal bindingresponse is found to be approximately 1.38 µg/mL.