Ab6533

Whole animal or human blood samples were separated by being allowed to sit for 30 min or using EDTA-coated tubes, followed by centrifugation at 4000 ×g for 15 min and storage at -80°C until use. Blood glucose, triglycerides, total cholesterol, glucose, and AST/ALT were measured using Fuji Dri-Chem Slides (Fujifilm). Leptin, ghrelin, GLP-1, adiponectin, and glucagon concentrations were determined in duplicate according to the manufacturer’s ELISA instructions. ELISA kits for mouse adiponectin and leptin were purchased from Abcam (ab108785 and ab100718; Cambridge, UK), and ELISA kits for mouse ghrelin, GLP-1, and glucagon were from Cusabio (CSB-E09817m, CSB-E08118m, and CSB-E15775m, respectively; Houston, TX). Enzyme immunoassay (EIA) kits for human/mouse apelin-13 were purchased from Phoenix Pharmaceutical (EK-057-19, Burlingame, CA). ELISA kits for glucose (Ab6533, Abcam), triglycerides (ETGA-200, Bioassay Systems, Hayward, CA, USA), and cholesterol (ab65390, Abcam) were used to measure levels in the liver tissues and sera saved after the experimental diets and from control and NAFLD patients.

Plasma total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), non-HDL cholesterol (n-HDL-C), and TG were determined by automated enzymatic kits (Sekisui Diagnostics, Lexington, MA, USA) on an ABX Pentra 400 autoanalyzer (Horiba Instruments Inc., Irvine CA, USA). Direct low-density lipoprotein cholesterol (d-LDL) was assessed with a commercial Elisa kit (KT-60293, Kamiya BioMedical Company, Seattle, WA, USA). Direct assessment of lipoprotein particle number and size was conducted by nuclear magnetic resonance spectroscopy (NMR) using automated signal acquisition followed by computational analysis and proprietary signal processing algorithms (Liposcience, Raleigh, NC) [57] (link). Serum insulin was analysed by ELISA (EZRMI-13K, Millipore, Billerica, MA) and glucose was measured by colorimetric analysis (ab6533, abcam, Cambridge, MA). Proprotein convertase subtilisin/kexin type 9 (PCSK9) was measured in serum samples using a commercial ELISA kit (CY-8078, MBL International, Woburn, MA).