L type wako glu2

Serum GH levels were determined by Electro Chemiluminescence Immunoassay using Cobas 8000 analyzer series module e801 (Roche Diagnostics, Mannheim, Germany; %CV of intra-assay variability <15%). The serum glucose level was determined by hexokinase enzymatic method (L-type Wako Glu2; Wako Pure Chemical Industries, Osaka, Japan) using LABOSPECT 008 α (HITACHI, Tokyo, Japan) and JCA-BM8060 (JEOL, Tokyo, Japan). The lactate level was determined by enzymatic method (Deteminer LA; Minaris Medical, Tokyo, Japan) using JCA-BM9130 (JEOL, Tokyo, Japan). Serum insulin levels were determined by Chemiluminescent Immunoassay using Architect i2000 SR (Abbott, Osaka, Japan). Plasma amino acids fractionation was determined by High-Pressure Liquid Chromatography using a Modification of the 6-Aminoquinolyl-N-Hydroxysuccinimidyl Carbamate (AQC) Method on a Waters Acquity UPLC system. GH, glucose, insulin, amino acids, and lactate levels were analyzed by the LSI Medience Corporation (Kyoto, Japan). Acylated ghrelin levels were measured using an Active Ghrelin ELISA Kit (# 97751, LSI Medience, Tokyo, Japan).

Body weight, blood glucose, and glycated hemoglobin (HbA1c) were measured once monthly. Blood samples were collected from the tail vein of nonfasting rats to measure the blood glucose and HbA1c. Blood glucose was measured by the hexokinase-G-6-PDH method (L type Wako Glu2, Wako Pure Chemical Industries, Ltd., Osaka, Japan). HbA1c was measured using an automated glycohemoglobin analyzer (HLC-723GHb V, Tosoh Corporation, Tokyo, Japan) [21 ].

Body weight, blood glucose, and glycated hemoglobin (HbA1c) were measured once monthly. Blood samples to measure the blood glucose and HbA1c were collected from the tail vein of nonfasting rats. Blood glucose was measured by the hexokinase/glucose-6-phosphate dehydrogenase method (L type Wako Glu2, Wako Pure Chemical Industries, Ltd., Osaka, Japan). HbA1c was measured using an automated glycohemoglobin analyzer (HLC-723GHb V, Tosoh Corporation, Tokyo, Japan).