AMPK alpha 1, also known as AMP-activated protein kinase alpha 1 subunit, is a crucial enzyme that plays a significant role in energy metabolism. It is is involved in regulating several metabolic processes, including glucose and fatty acid uptake, oxidation, and storage. It promotes energy-producing pathways, such as glycolysis, fatty acid oxidation, and mitochondrial biogenesis, while inhibiting energy-consuming processes, like protein synthesis and lipogenesis. Besides, PRKAb1 has been identified as an interactor of AMPK Alpha 1, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human AMPK Alpha 1 and recombinant human PRKAb1. Briefly, AMPK Alpha 1 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to PRKAb1-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-AMPK Alpha 1 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 450/630 nm immediately. The binding activity of recombinant human AMPK Alpha 1 and recombinant human PRKAb1 was shown in Figure 1, the EC50?for this effect is 0.94 ug/mL.