Bs 180

Blood samples were collected from the external jugular vein of all calves at 10, 35, and 70 days of age for hematological (mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), erythrocytes, hematocrit, hemoglobin, leukocytes, thrombocytes, cell distribution width (RDW), and mean corpuscular volume (MPV)) and biochemical analyses (alanine transaminase (ALT) and aspartate transaminase (AST)) and for determination of selenium concentration. Hematological analyses were carried out using a HORIBA SCIL VET ABC instrument (Horiba Medical, USA) immediately after the blood samples were collected. For the assessment of biochemical parameters (ALT, AST, and selenium level), blood was collected into heparinized tubes and later centrifuged at 5000× g rpm for 10 min at room temperature. The obtained plasma was divided into Eppendorf tubes for individual analyses and frozen at −20 °C. Biochemical analyses of plasma parameters were performed on a Mindray BS-180 (China) apparatus using commercial CORMAY kits (Lublin, Poland). The selenium level was tested by IDEXX laboratory (Kornwesthei, Germany) using an ICP-MS analyzer with an Se-78 isotope.

The blood lipid-related indexes: total cholesterol (TC, mmol/L), triglyceride (TG, mmol/L), high-density lipoprotein cholesterol (HDL-C, mmol/L) and low-density lipoprotein cholesterol (LDL-C, mmol/L); uric acid (UA, mmol/L), aspartate aminotransferase, alanine aminotransferase, blood calcium and serum phosphate (mmol/L) using test kit and biochemical analyser (BS-180, Mindray company). Fasting plasma glucose (FPG, mmol/L) and 2 h blood glucose after OGTT load (2 h PG, mmol/L) were determined by glucose oxidase method using test kit and biochemical analyzer (BS-180, Mindray Company). Glycosylated hemoglobin (HbA1c, %) using BioRad reagent, measured by VARIANT II (BioRad Company). Thyroid-stimulating hormone (TSH, mIU/L), Free thyroxine (FT4, pmol/L) anti-thyroid peroxidase antibody (TPOAb, 0–34IU/L) and anti-thyroid globulin antibody (TgAb, IU/L) using ICMA (Roche Company). Urinary iodine (UIC, g/L) was determined by inductively coupled plasma mass spectrometer (7700×, Agilent, USA).

The serum was separated by centrifuging the blood samples for 10 minutes at 1500g, and was stored at −20°C until analyses of creatinine (Cr) and blood urea nitrogen (BUN) levels with an automatic biochemical analyzer (BS‐180; Shenzhen Mindray Bio‐Medical Electronics Co, Ltd, Shenzhen, China).

Serum creatinine (Scr) and blood urea nitrogen (BUN) were measured using BS-180 automatic biochemistry analyzer (Mindray, Shenzhen, China) following the manufacturer's instructions.