Adult humans have roughly one million pancreatic islets, which are clusters of cells measuring 100 to 250 μm in diameter that secrete endocrine hormones to control blood glucose levels. Glucagon is released by the islets, particularly during fasting or hypoglycemia. Precise, real-time measurement of glucagon secretion from individual islets is crucial for understanding islet function and metabolic diseases, such as diabetes. We recently designed a fluorescence anisotropy immunoassay (FAIA) to measure mouse and human insulin secretion from living islets in a continuous-flow microfluidic device (InsC-chip). FAIAs are commonly employed in high-throughput screening assays in both industrial and academic settings. They quantify target protein concentration based on competitive binding to a monoclonal antibody with a fluorophore-labelled analogue (i.e., tracer), resulting in a lower steady-state fluorescence anisotropy. Based on this platform, we are now developing an FAIA to measure glucagon secretion (Glu-chip). We are working with two monoclonal antibodies that have distinct epitopes on the N and C termini of glucagon. Our preliminary data show that neither antibody can be presently used on-chip due to low-dynamic range (i.e., minimal change in anisotropy upon binding) and slow kinetics (i.e., equilibrium is not reached on-chip). To overcome these limitations, we are truncating the tracers to the minimal epitope and designing point mutations to accelerate the binding kinetics. Finally, our goal will be to validate the FAIA by measuring glucagon secretion from individual mouse and human islets. This project will develop a novel assay to measure glucagon secretion with high temporal resolution and outline a general strategy to optimize FAIAs for continuous flow detection (i.e., organ-on-chip).
Chee et al. (2026) studied this question.