Nonfasting blood samples were taken by venipuncture in EDTA-containing vacutainers at the baseline clinic visit and processed at the Department of Clinical Biochemistry, University of Cambridge (UK), where all lipids assays were performed.41 (link) Plasma was isolated by centrifugation for 15 minutes, 3000 rpm, 4 °C and stored at -80 °C for future analysis. Circulating levels of total cholesterol, HDL-C and TG were measured on fresh samples with the RA 1000 (Bayer Diagnostics, Basingstoke, United Kingdom) and LDL-C levels were calculated with the Friedewald formula.42 (link) Non-HDL-cholesterol was calculated by subtracting HDL-C from total cholesterol. Apo B was measured by using rate immunonephelometry (Behring Nephelometer BNII, Marburg, Germany).43 (link) HbA1c was measured using Diamat ion exchange HPLC (Bio-Rad Laboratories, Hemel Hempstead, United Kingdom).39 (link)In the case-control subset additional parameters were tested. Lipoprotein particle size and particle number was analyzed with an automated nuclear magnetic resonance (NMR) spectroscopy by Liposcience Co as described.44 (link) Plasma LPL mass and Apo A-V were measured as described.16 (link) Samples were analyzed in random order to avoid systemic bias. Researchers and laboratory personnel had no access to identifiable information and could identify samples by number only.
Ra 1000
Manufactured by Bayer
Sourced in United States, United Kingdom, China
The RA-1000 is a laboratory analysis instrument that performs automated radiochemical analysis. It is designed to process and analyze radioactive samples with precision and efficiency. The core function of the RA-1000 is to quantify and characterize radioactive substances in various sample types.
Automatically generated - may contain errors
18 protocols using ra 1000
1
Blood Lipids and Lipoprotein Analysis Protocol
2
Serum Biochemical Profiling Assay
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The concentrations of total protein, glucose, albumin and globulin in the serum samples were analyzed with an Automatic Biochemical Analyzer (RA-1000, Bayer Corp., Tarrytown, NY, USA) using colorimetric methods. Serum total cholesterol and triglycerides (glycerol-3-oxidase-peroxidase-enzymatic-colorimetric) levels were analyzed by an automated Vitros 350 analyzer (Otho-Clinical Diagnostic, Johnson & Johnson, Rochester, NY, USA). The levels of enzyme activities including aspartate aminotransferase and alanine aminotransferase were analyzed using an Automated Clinical Chemistry Analyzer (Roche, Cobus-Mira-Plus, Roche Diagnostic System, Inc., NY, USA).
3
Urine and Blood Analysis in Mice
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Mice in each group were placed in independent metabolic cages, with free access to drink. The mice were fasted overnight before euthanasia, and 24 h urine samples were collected. All urine specimens are preserved with toluene. After 24 h, the urine volume was assessed. After centrifugation to remove the sediments, the samples were separately placed into 1.5 mL Eppendorf tubes, which were then stored in a freezer at -20 °C for subsequent use. The 24 h urine albumin excretion rate (UAER) was determined. Upon the end of the experiment, blood was collected by the inner canthal blood sampling method. After standing for 1 h, the blood samples were centrifuged at 3000 rpm for 10 min, and the supernatant was extracted and stored in an Eppendorf tube at − 20 °C. Finally, the levels of blood urea nitrogen (BUN) and serum creatinine (Scr) were determined using an automatic biochemical analyzer (RA-1000, Bayer, Leverkusen, Germany) with its supporting kits.
4
Comprehensive Serum Biomarker Profiling
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Specific ELISA kits (Leadman Group Co., Ltd., Beijing, China) were used to assess the contents of serum immunoglobulins (IgA, IgM, and IgG), following the manufacturer’s instructions. Biochemical indices such as glucose (GLU), total cholesterol (TC) and triacylglycerol (TG), total protein (TP), albumin (ALB), urea (UREA), and creatinine (CREA) in the serum were measured by an automatic biochemical analyzer (RA-1000, Bayer Corp., Tarrytown, NY, USA).
5
Comprehensive Metabolic Health Assessment
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We examined changes in BMI, waist circumference, the ratio of total to HDL cholesterol, glycated hemoglobin (HbA1c), and blood pressure (systolic and diastolic) as primary outcomes. Changes in weight, waist-to-hip ratio, total cholesterol, HDL cholesterol, LDL cholesterol, triglycerides, and a metabolic-risk z score were examined as secondary outcomes. Weight, height, waist circumference, and hip circumference were measured with a standardized protocol. BMI was defined as weight divided by height squared. Blood pressure was measured twice (using the average) with an Accutorr sphygmomanometer (Datascope). Total cholesterol, HDL cholesterol, triglycerides (RA 1000; Bayer Diagnostics) and HbA1c (Diamat ion exchange HPLC; Bio-Rad Laboratories) were measured in nonfasting blood. LDL cholesterol was calculated using the Friedewald formula (20 (link)). The metabolic risk z score was calculated as previously described (21 (link)) by averaging z scores of blood pressure (systolic and diastolic), HbA1c, waist circumference, HDL cholesterol multiplied by −1, and log-transformed triglycerides.
6
Comprehensive Serum Biomarker Analysis
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The serum immunoglobulins (IgA, IgG and IgM ) and cytokines (TNF-α and IL-1β ) concentrations were measured using the commercially available ELISA kits (Shanghai Junshi Biosciences Co. Ltd., Shanghai, China), and absorbance was measured at 450 nm. The concentrations of total cholesterol (TC ), total triglyceride (TG ), low density lipoprotein (LDL) cholesterol, high density lipoprotein (HDL ) cholesterol and urea nitrogen (urea-N ) in serum samples were analyzed by an automatic biochemical analyzer (RA-1000, Bayer Corp., Tarrytown, NY) following the instructions of the commercial kits (Zhongsheng Biochemical Co., Ltd., Beijing, China). The concentration of growth hormone (GH ) in serum samples was determined by using a radioimmunoassay (Sn-96513, Shanghai, China).
7
Antioxidant and Inflammatory Markers in Poultry
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The concentrations of total antioxidant capacity (T-AOC ), superoxide dismutase, glutathione peroxidase (GSH-Px ), catalase (CAT ), and malondialdehyde (MDA ) in serum (n = 8) were determined by an automatic biochemical analyzer (RA-1000, Bayer Corp., Tarrytown, NY) using colorimetric methods following instructions of the manufacturer of the corresponding reagent kit (Zhongsheng Biochemical Co., Ltd., Beijing, China). After slaughter, the liver samples (n = 8) on days 21 and 42 were used to determine the contents of interleukin-1β (IL-1β ), interleukin-6 (IL-6 ), and tumor necrosis factor-alpha (TNF-α ) by ELISA test (Shanghai Lengton Biosicences Co. Ltd., Shanghai, China) at a wavelength of 450 nm.
8
Serum Lipid and sPLA2 Measurement
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Blood samples were stored at -80°C at the Department of Clinical Biochemistry, University of Cambridge. Serum levels of total cholesterol, high-density lipoprotein (HDL) cholesterol, and triglycerides were measured on fresh samples with the RA 1000 (Bayer Diagnostics, Basingstoke, UK), and LDL cholesterol levels were calculated with the Friedewald formula. Serum sPLA2 activity was measured by a selective fluorometric assay 5 by using fluorescent substrate 1-hexadecanoyl-2-(1-pyrenedecanoyl)-sn-glycero-3 phosphomethanol, sodium salt (Interchim, Montluçon, France), as previously described. 6 (link) One hundred percent hydrolysis of the fluorescent substrate was measured using 0.1 U sPLA2 from bee venom (Sigma Chemical Co). The hydrolysis of substrate in the absence of plasma was used as negative control and deduced from sPLA2 activity. All samples were tested in duplicate, and plasma activity was expressed as nmol/min.mL. The minimum detectable activity was 0.10 nmol/min/mL. Plasma concentrations of CRP were measured with a sandwich-type enzyme-linked immunosorbent assay as previously described. 19 (link) Results were related to a standard consisting of commercially available CRP (Behringwerke AG, Marburg, Germany). The lower detection limit was 0.1 mg/L.
9
Serum Cytokine and Lipid Profile Analysis
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The serum IgA, IgG, IgM, IL-6, IL-1 β, and tumor necrotic factor-α (TNF-α) were measured using ELISA Kits (Shanghai Junshi Biosciences Co. Ltd., Shanghai, China), and value was measured at 450 nm. The concentrations of total cholesterol (TC), triglyceride, high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol in serum sample were measured using commercial kits (Zhongsheng Biochemical Co. Ltd., Beijing, China) via an automatic biochemical analyzer (RA-1000; Bayer Corp., Tarrytown, NY).
10
Blood Analysis of Avian Subjects
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Occurring at week 4 and at the end of the experiment, 5 mL blood samples were collected from the branchial vein of 20 birds/treatment using a sterilized syringe and stored K3EDTA (Becton Dickinson Vacutainer Systems, Franklin Lakes, NJ, USA) The collected blood samples were taken to the laboratory and stored at 4 °C. Regarding serum analysis, approximately 3 mL blood samples were centrifuged at 4000× g for 15 min at 4 °C, to separate the serum. The white blood cell (WBC), red blood cells (RBC), and lymphocyte counts of the whole blood samples were determined using an automatic blood analyzer (ADVIA 120; Bayer). Protein, albumin, globulin, creatinine, uric acid, triglyceride, and phosphorus in the serum samples were analyzed with an automatic biochemical analyzer (RA-1000, Bayer Corp., Tarrytown, NY, USA) using colorimetric methods.
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