Edta treated tubes

Mice were euthanized by CO₂ inhalation and blood was collected by cardiac puncture. Serum was collected by centrifugation (15 min, 2000 g, 4°C) after allowing blood to clot on ice. Plasma was collected by centrifugation (15 min, 2000 g, 4°C) in EDTA-treated tubes (Sarstedt, Numbrecht, Germany). All serum and plasma samples were snap frozen and stored at −80°C. Kits were used to measure total cholesterol (Infinity Total Cholesterol, Fisher Scientific), free cholesterol (Free Cholesterol E, Wako Diagnostics, Richmond, VA), HDL cholesterol (HDL-C E, Wako Diagnostics), LDL cholesterol (LDL-C Reagent L-type, Wako Diagnostics), NEFAs [HR Series NEFA-HR(2), Wako Diagnostics], and TGs (Infinity Triglycerides, Fisher Scientific) according to the manufacturer's directions. For HFD and LFD male mice, n=5-8; for HFD and LFD female mice, n=8-9; and for CD male mice, n=6-8.

Treatment consisted of a single i.p. injection of 500 ng of diphtheria toxin in phosphate buffered saline (PBS). Blood glucose was measured using an Accu-Chek Aviva system (Roche, Bromma, Sweden). For the assessment of insulin levels, plasma was collected in EDTA-treated tubes (Sarstedt, Nümbrecht, Germany) and measured with alphaLISA (Perkin Elmer, Upplands Väsby, Sweden). Glucose tolerance tests were performed after 12 hours overnight fasting, by giving an i.p. injection of 2 g glucose per kg bodyweight. To avoid prolonged hyperglycemia, DT-treated RIP-DTR mice used for subsequent OPT imaging were transplanted with ~100–150 RIP-DTR negative islets into the ACE, normalising their blood glucose levels as shown previously for streptozotocin-treated mice^{43 (link)}.