Exendin 4

Peptide agonists (exendin‐4, exendin‐phe1, exendin‐asp3) were purchased from by Wuxi Apptec at more than 90% purity.

Exendin-4, exendin-F1 and P5 were all custom synthesised by Wuxi Apptec (Wuhan, China) with > 90% purity confirmed by HPLC. The synthesis of Luxendin645 and Exendin-4-Cy5 have been described previously [18] (link), [19] (link). Unless otherwise specified, all chemicals were obtained from Sigma Aldrich (Gillingham, UK).

Insulin secretion assays were performed in triplicate on size-matched islets, isolated from male mice (8–10 weeks of age) and incubated for 1 h in modified Krebs-Ringer bicarbonate buffer containing 3 mmol/l glucose. Subsequently, islets were either perifused (~50 islets/chamber) or batch incubated (10 islets/well) for 30 min in Krebs-Ringer solution with either 3 mmol/l or 17 mmol/l glucose, 10 mmol/l glucose supplemented with 100 nmol/l exendin-4 (Wuxi Apptec, Shanghai, China), or the glucose-dependent insulinotropic peptide (GIP; Wuxi Apptec). Secreted and total insulin content were quantified using a homogeneous time-resolved fluorescence (HTRF) insulin kit (Cisbio, Codolet, France) in a PHERAstar reader (BMG Labtech, Aylesbury, UK), following the manufacturer’s guidelines. Data are presented as secreted insulin/insulin content.