Lipid extraction kit

Insulin levels were measured using a mouse insulin ELISA kit (Crystal Chem, 90080). GLP-1 was measured with the total GLP-1 kit (MesoScale Discovery, K150JVC-1). For GLP-1 measurement, mice from the separate cohort used for macronutrient test, were fasted for 5 hours prior to an oral administration of liquid mixed meal (200uL of Ensure Plus, Abbott Laboratories). Plasma mouse Reg3g was measured using Proteome Profiler Mouse XL Cytokine Array (R&D Systems, ARY028). Human REG3A was determined by using R-PLEX Human Reg-3-alpha Antibody Set (MesoScale Diagnostics, F21U8-3). All assays were performed according to the manufacturer’s instructions. For hepatic triglyceride measurement, liver lipids were extracted with Lipid Extraction Kit (Abcam, ab211044). Triglycerides (Biovision, K622) were measured using the extracted liver lipids. Malondialdehyde (MDA) concentrations of intestines were determined using an Oxiselect TBARS assay kit (Cell Biolabs, STA-330). Cellular ROS production was determined using DCFDA/H2DCFDA assay kit (Abcam, ab113851).

Total unsaturated fatty acid levels in neurons were prepared by lipid extraction Kit (Abcam) and measured by Lipid Assay Kit for unsaturated fatty acids (Abcam) following the manufacturer’s instructions. 1 × 10⁶ neurons were collected in 1.5 mL micro centrifuge tube and centrifuged at 1000 × g for 5 minutes. Cell pellets were washed and resuspended in 25 μL PBS, and 500 μL extraction buffer was added. After agitating the mixture for 15–20 minutes at room temperature, sample tubes were centrifuged at 10,000 × g for 5 minutes. The supernatant was carefully collected and dried in a 37°C incubator overnight. Dried lipid extract was resuspended in 50 μL suspension buffer and sonicated for 15–20 minutes at 37°C before quantification. Absorbance was measured at OD 540 nm.