Mentions: Insulin is stored in the secretory granules of beta cells inpancreatic islets, where it is cocrystallized with zinc ions in a 6:2 ratio(insulin:Zn2+) (see Figure 1). Upon secretion from normal islets, insulin and zincions are released into the extracellular space in this 6:2 ratio. In order toimage real-time insulin and zinc secretions from islets, droplet microfluidicsis utilized to collect secretions with minimal dilution. Through a novelapproach, combining lock-in spatial filtering and droplet fluidics, highlysensitive measurements are made via confocal fluorescence microscopy. Using acompetitive zinc-binding assay between EDTA and the fluorescent zinc indicator,FluoZin-3, this approach results in sensitive and quantitative real-timeimaging of zinc as it is secreted from live pancreatic islets duringglucose-stimulated insulin secretion (GSIS). Furthermore, time traces ofdroplet-confined secretions can be stored in tubes, and can later be used toquantify insulin using radioimmunoassays (RIAs) or enzyme-linked immunosorbentassays (ELISAs). We are utilizing these methods to investigate insulin storagedefects that occur in diabetic mice, particularly those with mutations of the SLC30A8 gene which encodes for the zinc transporter ZnT-8 that is present exclusivelyin secretory vesicles of pancreatic beta cells. Deletion of this gene leads toimpaired glucose-tolerance in knockout mouse models.
Affiliation: P S Analytical, Arthur House, Crayfields Industrial Estate, Main Road, Orpington, Kent BR5 3Hp, UK.
No MeSH data available.