Abstract
For studying stem cell–derived islet organoids (SC-islets) in an organ-on-chip (OoC) platform, we have developed a reversed-phase liquid chromatography–tandem mass spectrometry (RPLC–MS/MS) method allowing for simultaneous determination of insulin, somatostatin-14, and glucagon, with improved matrix robustness compared to earlier methodology. Combining phenyl/hexyl-C18 separations using 2.1 mm inner diameter LC columns and triple quadrupole mass spectrometry, identification and quantification were secured with negligible variance in retention time and quantifier/qualifier ratios, negligible levels of carryover (<2%), and sufficient precision (±10% RSD) and accuracy (±15% relative error) with and without use of an internal standard. The obtained lower limits of quantification were 0.2 µg/L for human insulin, 0.1 µg/L for somatostatin-14, and 0.05 µg/L for glucagon. The here-developed RPLC–MS/MS method showed that the SC-islets have an insulin response dependent on glucose concentration, and the SC-islets produce and release somatostatin-14 and glucagon. The RPLC–MS/MS method for these peptide hormones was compatible with an unfiltered offline sample collection from SC-islets cultivated on a pumpless, recirculating OoC (rOoC) platform. The SC-islets background secretion of insulin was not significantly different on the rOoC device compared to a standard cell culture well-plate. Taken together, RPLC–MS/MS method is well suited for multi-hormone measurements of SC-islets on an OoC platform.
Electrophoresis. 2023