Plasma glucose during the euglycemic–hyperinsulinemic clamp was determined in duplicate by the glucose oxidase method (Analox GM7 or GM9, Analox Instruments, London, UK). Serum insulin concentration, determined every 30 min during the clamp, was measured by a double antibody RIA (Phadeseph Insulin RIA kit, Pharmacia & Upjohn, Uppsala, Sweden) or automatized electro-chemiluminescence immunoassay (Cobas 8000, Roche Diagnostics GmbH), and serum FFA concentration, determined every 60 min during the clamp was measured using an enzymatic assay (ACS-ACOD, Wako Chemicals GmbH, Neuss, Germany).
Acs acod
Manufactured by Fujifilm
Sourced in Germany
The ACS-ACOD is a laboratory equipment product manufactured by Fujifilm. It is designed for automated chemical synthesis and analysis. The core function of the ACS-ACOD is to facilitate controlled and precise chemical reactions and measurements.
Automatically generated - may contain errors
Lab products found in correlation
Analox gm7, by Analox Instruments (2 mentions)
Cobas 8000, by Roche (2 mentions)
God pap, by Roche (2 mentions)
Metrolab 2300 auto analyzer, by Wiener Lab (1 mentions)
Chod pod, by Spinreact (1 mentions)
Autodelfia, by PerkinElmer (1 mentions)
Quantitative sandwich elisa, by MyBioSource (1 mentions)
7 protocols using acs acod
1
Euglycemic-Hyperinsulinemic Clamp Glucose Measurements
2
Metabolic Biomarker Measurement Methods
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The main analyses applied in the study are described in the following sections (details about the remaining analyses can be found in Supplementary methods). Plasma glucose was measured bedside using the glucose oxidase method (YSI model 2900D, Yellow Springs, OH, USA). For serum insulin and C-peptide concentrations, a two-sided chemical luminescence immunoassay (Siemens Atellica IM analyser, Erlangen, Germany) was used. Plasma glucagon was measured using a C-terminally directed antiserum (code no 4305) measuring glucagon of pancreatic origin as described earlier (32 (link)). For analysis of free fatty acids, an enzymatic colorimetric method was used (manual ELISA (NEFA-HR (2 (link))), ACS-ACOD, Wako Chemicals, Neuss, Germany).
3
Bovine Plasma Metabolite Analysis
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Blood samples were obtained by venipuncture coccygeal with heparinized tubes from each cow after the afternoon milking on three occasions (weeks 4, 8, and 11 of the trial). Samples were centrifuged at 800× g for 10 min on the day of sampling, and the obtained plasma was aliquoted and frozen at −20 °C in 1.5 mL microtubes for later analysis. At the end of the trial, β-hydroxybutyrate (β-hydroxybutyrate Rambut, Randox®, Crumlin, UK), non-esterified fatty acids (NEFA, ACS-ACOD, Wako®, Neuss, Deutschland), albumin (BCG, Human®, Wiesbaden, Germany), and urea (GLDH, Human®) were determined in plasma in a Metrolab 2300 autoanalyzer (WienerLab®, Rosario, Argentina).
4
Biochemical Markers of Metabolism
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Glucose ((GOD-PAP); Roche Diagnostics, Barcelona, Spain), triacylglycerides ((LPL- GPO); Roche Diagnostics), cholesterol ((CHOD-POD); Spinreact, Girona, Spain), glycerol (GPO-Trinder, Sigma Diagnostic, Madrid, Spain) and NEFA ((ACS-ACOD); Wako Chemicals, Neuss, Germany) levels were determined by enzymatic colorimetric tests. Plasma insulin measurement (Mercodia, Uppsala, Sweden) was determined using immunoassay kits. The HOMA-IR insulin resistance index and QUICKI insulin sensitivity index were calculated as previously described [33 (link)].
5
Quantification of Metabolic Markers
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Plasma glucose was determined in duplicate by the glucose oxidase method (Analox GM7 or GM9, Analox Instruments, London, UK). Serum insulin concentration was measured by a double antibody radioimmunoassay (Phadeseph Insulin RIA kit, Pharmacia & Upjohn, Uppsala, Sweden), fluoroimmunometric assay (AutoDELFIA, PerkinElmer Inc, Turku, Finland) or automatized electro-chemiluminescence immunoassay (Cobas 8000, Roche Diagnostics GmbH, Mannheim, Germany). Alanine aminotransferase and aspartate aminotransferase were measured using automatized enzymatic method (Cobas 8000, Roche Diagnostics GmbH, Mannheim, Germany) and serum FFA concentration using an enzymatic assay (ACS-ACOD, Wako Chemicals GmbH, Neuss, Germany).
6
Blood Biomarker Analysis Protocol
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Capillary blood was analyzed immediately for glucose and lactate using a Biosen analyser (Biosen C-line, EKF-diagnostic GmbH, Magdeburg, Germany). Venous blood was drawn in vacutainer EDTA tubes and serum separation tubes. EDTA tubes were centrifuged immediately at 4°C and plasma stored at -80°C until analysis. Blood in serum separation tubes was allowed to coagulate at room temperature, centrifuged and serum stored at -80°C until analysis. To determine Cortisol and Insulin concentrations in serum, Quantitative Sandwich Elisa was used (MyBioSource, San Diego, CA 92195-3308, USA, Cat. No. MBS043519, Human Hydrocortisone Elisa Kit). The Insulin concentrations were determined using Human Insulin Kit from Invitrogen Corporation (7335 Executive Way, Frederick, MD 21704, USA Cat. No. KAQ1251). Free fatty acid analysis of plasma was performed by the accredited Laboratory for Clinical Chemistry, Sahlgrenska University Hospital (SWEDAC ISO 15189) using an enzymatic colorimetric method (ACS-ACOD by Wako Chemicals, Germany).
7
Plasma Biochemical Analyses
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Commercial enzymatic tests were used to determine in plasma samples: glucose (GOD-PAP, Roche Diagnostics, Spain), triacylglycerides (LPL/GPO-Trinder Roche Diagnostics, Spain), glycerol (GPO-Trinder, Sigma Diagnostic, Spain), cholesterol (Menarini, Italy), non-esterified fatty acids (NEFA) (ACS-ACOD, Wako Chemicals GmbH, Germany) and ketone bodies (Wako Chemicals GmbH, Germany). Plasma insulin was determined with a specific EIA kit for rats (Mercodia, Denmark). Corticosterone, leptin and adiponectin were determined in plasma samples by X-Map Technology (Bioplex 100X, Spain) using the "rat stress hormone" RSH69K and "rat adipocyte" RADPCYT-82 panels (Millipore, Spain).
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