Rat elisa kits

Serum concentration of TSH (NIDDK reagents, Bethesda, MD; sensitivity range = 2.5–80 ng/ml) and corticosterone (reagents from Merck-Millipore, Perkin Elmer and Sigma; limits of detection, 20–2,000 ng/ml) were quantified by radioimmunoassay. Total T3 (limits of detection, 0.75–10 ng/ml) and T4 (limits of detection, 1–30 μg/dl) were quantified by ELISA [kits from Diagnóstica Internacional (Zapopan, JAL., México)] following the manufacturer's instructions, except for one variation: standard curve was prepared by adding 25 μl of rat hypothyroid serum to the calibrators provided in the kit, as recommended (48 (link)). Leptin (assay range, 0.2–12.8 ng/ml) and insulin (sensitivity range, 0.1–12.8 ng/ml) were measured using rat ELISA kits from Crystal Chem Inc. (Downers Grove, IL), ACTH using an ELISA kit from USCNK Life Science (Wuhan P.R., China; detection range, 12.35–1,000 pg/ml), and 17β-estradiol using an ELISA kit from Arbor assays (Michigan, USA; limits of detection, 3.75–120 pg/ml). Experiments 1 and 2 were analyzed independently; for each experiment, samples from male and female rats were measured in duplicate within a single assay, and inter-assay and intra-assay coefficients of variation were below 10%.

For the rat samples, plasma levels of T₃ and T₄ were measured using rat ELISA kits (Crystal Chem Inc; Downers Grove, IL, USA) (Lopez et al. 2010 (link), Gonzalez et al. 2012 (link), Varela et al. 2012 (link)). For the human samples, serum glucose concentrations were measured in duplicate by the glucose oxidase method using a Beckman Glucose Analyser II (Beckman Instruments; Brea, CA, USA). Roche Hitachi Cobas c711 instrument (Roche) was used to perform HDL cholesterol and total serum triglycerides determinations. HDL cholesterol was quantified by a homogeneous enzymatic colorimetric assay through the cholesterol esterase/cholesterol oxidase/peroxidase reaction (Cobas HDLC3; Roche). Serum fasting triglycerides were measured by an enzymatic, colorimetric method with glycerol phosphate oxidase and peroxidase (Cobas TRIGL; Roche). LDL cholesterol was calculated using the Friedewald formula. Serum free T₄ was measured by electrochemiluminescence (Roche Diagnostics) with intra- and inter-assay coefficients of variation less than 5%. Methods have been previously reported (Ortega et al. 2015 (link), Gavalda-Navarro et al. 2016 (link)).