Accubind elisa kit

Two groups of six male C57BL/6 mice (10–12 weeks, Charles Rivers) were used. Both groups were allowed ad libitum access to food containing PTU (Teklad + 0.15% propylthiouracil) for 44 days to induce hypothyroidism. Six mice were injected (IP) with a supra-physiological dose of 5 μg T3 in 20 μl saline at day 42, 43, and 44 (corresponding to 0.21–0.24 μg T3/g BW); the remaining six hypothyroid mice were injected with 20 μl saline. Animals were sacrificed at day 45. The heart was excised and left and right ventricle (LV, RV) and septum were dissected, weighed, immediately frozen in liquid nitrogen, and stored at −80°C. Tibia length was determined for normalization purposes. Plasma T3 levels were determined by AccuBind ELISA kits (Monobind Inc., Lake Forest, CA) according to the manufacturer’s instructions. Animals were housed individually and all experiments complied with the Guide for Care and Use of Laboratory Animals of the National Institutes of Health (NIH Publication no. 86-23, revised 1996) and were approved by the Institutional Animal Care and use Committees of VU University Medical Center Amsterdam.

The free T₃ (fT3), free T₄ (fT4), TSH and insulin concentrations were determined using AccuBind ELISA kits, as recommended by the manufacturer (MonoBind Inc., Lake Forest, CA, USA). Glucose was determined using the glucose oxidase method (ParsAzmoon, Tehran, Iran). The HbA1c level was measured using a latex agglutination immunoassay kit (Man Co., Ltd., Tehran, Iran) and an automatic analyzer (BT4500, Biotechnica Instruments, Roma, Italy). Serum TPOAb was measured with Rapid ELISA (Monobind, Lake Forest, CA, USA). The intra-assay and inter-assay CVs were 6 and 4%, respectively. An anti-TPO concentration of more than 9 IU/mL was considered as positive [27 (link)].