V-Erb B2 Erythroblastic Leukemia Viral Oncogene Homolog 3 (ErbB3) is a member of the ErbB family of receptor tyrosine kinases. It is also known as human epidermal growth factor receptor 3 (HER3). ErbB3 plays a crucial role in various cellular processes including proliferation, differentiation, and survival. Neuregulin 1 (NRG1) is one of ligands of ErbB3, when NRG1 binds to ErbB3, it triggers dimerization and self-phosphorylation of the receptor, thereby activating downstream signaling pathways such as the PI3K/Akt and MAPK pathways. These signaling pathways are essential for cell growth and survival. Thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human ErbB3 and recombinant human NRG1. Briefly, NRG1 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to ErbB3-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-NRG1 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. When recombinant human ErbB3 is immobilized at 2 ug/mL (100 uL/well), the concentration of NRG1 that produces 50% optimal binding response is found to be approximately 0.14 ug/mL.