Ysi 2700 biochemistry analyzer

Biochemical analysis was performed as previously described (Culpepper et al. 2016 (link)). Blood was analyzed immediately after collection for pH, O₂, CO₂, and hemoglobin (ABL825; Radiometer America Inc.; Brea, CA, USA), as well as plasma concentrations of glucose and lactate (YSI 2700 biochemistry analyzer; Yellow Springs Instruments; Yellow Springs, OH, USA). Additional plasma was stored at −80°C for measurement of insulin (intra- and inter-assay CVs 5.6% and 4.7%, respectively), insulin like growth factor-1 (IGF-I; intra- and inter-assay CVs 3.1% and 5.6%, respectively), and cortisol (intra- and inter-assay CVs 5.6% and 4.7%, respectively) by ELISA (ALPCO Immunoassays; Salem, NH, USA); glucagon by radioimmunoassay (intra- and inter-assay CVs 4.8% and 11.7%, respectively; Millipore; Burlington, MA, USA) and norepinephrine by HPLC (intra- and inter-assay CVs 9.2% and 9.0%, respectively).

Arterial blood was collected in heparinized syringes and immediately analyzed for pH, blood gases and hematocrit (ABL520; Radiometer America Inc., Brea, CA). Additional arterial blood samples were collected in EDTA coated syringes and centrifuged at 4°C for 3 min. Plasma concentrations of glucose and lactate were determined using a YSI 2700 biochemistry analyzer (Yellow Springs Instruments, Yellow Springs, OH). Additional plasma aliquots were stored at −80°C before analyses. Plasma concentrations of insulin, insulin-like growth factor-1 (IGF-I) and cortisol were measured by ELISA (ALPCO Immunoassays, Salem, NH). Intra- and inter-assay coefficient of variation (CV) was 5.6 and 4.7%, respectively, for insulin (sensitivity, 0.14 ng/ml); 3.1 and5.6%, respectively, for IGF-I (sensitivity 0.09 ng/ml); and 3.1 and5.6%, respectively, for cortisol (sensitivity 0.1 ng/ml). Glucagon and norepinephrine were measured in plasma by radio-immunoassay and HPLC, respectively, as previously described.^{16 (link),19 (link)} Intra- and inter-assay CV for glucagon were 4.8 and11.7%, respectively (Millipore; sensitivity, 18.5 pg/ml) and 9.2 and9.0%, respectively, for norepinephrine (sensitivity 170 pg/ml).