Meso scale discovery two plex mouse metabolic immunoassay kit

At around 13–14 weeks of age, tissues and blood were collected for metabolic and biochemical profiling from a separate cohort of mice. Blood was collected by cardiac puncture under terminal anaesthesia into Microtube 1.1 mL Z-Gel (Starstedt AG & Co), centrifuged at 10,000 rpm for 10 min at 4 °C and stored at − 80 °C until use. Plasma Serum was submitted and analysed in the Core Biochemical Assay Laboratory (CBAL). Mouse insulin and leptin were measured using a Meso Scale Discovery two-plex mouse metabolic immunoassay kit according to the manufacturer’s instructions and using calibrators provided by Meso Scale Diagnostics. Blood glucose was measured using approximately 2 μl blood drops using a glucometer (AlphaTrak2; Abbot Laboratories) and glucose strips (AlphaTrak2 test 2 strips, Abbot Laboratories, Zoetis).

For mouse studies 1 and 2 (MS1 and MS2), human recombinant GDF15 (Cat# 4570, BioVision Inc.) was prepared in saline and administered via subcutaneous (s.c.) injection as a single dose in the morning (9 AM). For mouse studies, 12 to 14 human recombinant GDF15 (Cat# Qk017, Qkine) was administrated via s.c. injection as a single dose in the morning (9 AM). At 1 h and/or 4 h posthuman recombinant GDF15 treatment, mice were killed by CO₂ inhalation and blood was collected via cardiac puncture. Blood was collected in EDTA tubes (Becton Dickinson and Company) and centrifuged to collect plasma, which was stored at −80 °C until analysis. For MS1 and MS2, plasma human GDF15 was measured using the human GDF15 Quantikine ELISA (R&D Systems) and plasma mouse corticosterone was measured using the mouse corticosterone ELISA (Cat# 55-CORMS-E01, ALPCO) as per manufacturer instructions. For MS12 to MS14, GDF15 was measured using the human GDF15 ELISA (Cat#DY957, R&D Systems, BioTechne) and plasma mouse corticosterone was measured using the mouse corticosterone EIA (Cat# AC14F1, IDS). Plasma leptin was measured using a Meso Scale Discovery two-plex mouse metabolic immunoassay kit (Cat# K15124C, Meso Scale Diagnostics).