High sensitivity plasma CRP was assayed using the N Latex CRP mono immunoassay on the Behring Nephelometer II Analyzer (Dade Behring, Milton Keynes, UK) [47 ]. Exclusion criteria for blood sampling included: clotting or bleeding disorders, history of fits or convulsions, or being on anticoagulant medication [48 ]. For the purpose of this analysis, CRP concentration was expressed in mg/L.
Behring nephelometer 2 analyzer
Manufactured by Siemens
Sourced in Germany, United Kingdom
The Behring Nephelometer II Analyzer is a laboratory instrument used to measure the concentration of specific proteins, such as antibodies or antigens, in a sample. It utilizes the principle of nephelometry, which involves the measurement of the intensity of light scattered by particles in a solution, to quantify the analytes of interest. The device provides accurate and reliable results for clinical diagnostic applications.
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Lab products found in correlation
N latex cystatin c, by Siemens (2 mentions)
Hlv 723 g8, by Tosoh (2 mentions)
Hem 907, by Omron (1 mentions)
Modular 700, by Roche (1 mentions)
Bio plex 200 array reader, by Bio-Rad (1 mentions)
Vitros 4600 system, by Ortho Clinical Diagnostics (1 mentions)
Uf 5000, by Sysmex (1 mentions)
Cobas 8000, by Roche (1 mentions)
Xe 5000 hematology analyzer, by Sysmex (1 mentions)
Architect c16000, by Abbott (1 mentions)
Cytometric bead array, by BD (1 mentions)
Alinity c, by Abbott (1 mentions)
Xe 5000, by Sysmex (1 mentions)
Cobas c701, by Roche (1 mentions)
16 protocols using behring nephelometer 2 analyzer
1
Plasma CRP Quantification Protocol
2
Kidney Function Assessment Protocols
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Data regarding markers of kidney function were collected at the baseline. The serum creatinine level was measured using a colorimetric technique on the Johnson & Johnson VITROS 950 Chemistry Analyzer using the enzymatic method. The baseline serum creatinine measurements were not calibrated. The cystatin C level was measured at the HABC core laboratory, at the University of Vermont, Burlington, Vermont, using the Behring nephelometer II analyzer (Dade Behring Inc), which used a particle-enhanced immunonepholometric assay (N Latex Cystatin C).8 (link)
The eGFRs were calculated using cystatin C-based (eGFRCys) and creatinine-based (eGFRCr) Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations.9 (link) The cystatin C-based CKD-EPI equation includes the baseline cystatin C level, sex, and age. The creatinine-based CKD-EPI equation includes the baseline creatinine level, sex, age, and race. The variable of interest, eGFRDiff, was calculated as eGFRCys − eGFRCr. We performed sensitivity analyses using the newly published 2021 CKD-EPI eGFRCr equations without the race factor.10 (link)
The eGFRs were calculated using cystatin C-based (eGFRCys) and creatinine-based (eGFRCr) Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations.9 (link) The cystatin C-based CKD-EPI equation includes the baseline cystatin C level, sex, and age. The creatinine-based CKD-EPI equation includes the baseline creatinine level, sex, age, and race. The variable of interest, eGFRDiff, was calculated as eGFRCys − eGFRCr. We performed sensitivity analyses using the newly published 2021 CKD-EPI eGFRCr equations without the race factor.10 (link)
3
Cardiovascular Risk Factors and Chronic Illness
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During study waves 2 (2004/5) and 4 (2008/9), a nurse measured systolic (SBP) and diastolic blood pressure (DBP) (Omron HEM 907) on three occasions at 1-minute intervals with the subjects seated; SBP and DBP are derived as the mean of the first two readings. Hypertension was considered as SBP ≥ 140 or DBP ≥ 90 or taking anti-hypertensive drugs, whereas normotension was taken as SBP < 120 and DBP < 80. A blood sample was also drawn from consenting respondents in order to measure levels of fibrinogen, total cholesterol, triglycerides and C-reactive protein. High total cholesterol was defined for values ≥ 5.2 mmol/l and high triglycerides for values > 1.7 mmol/l.
High-sensitivity C-reactive protein (CRP) was analysed using the N Latex CRP mono Immunoassay on the Behring Nephelometer II Analyzer (Dade Behring, Milton Keynes, UK). In all analyses, CRP data, originally skewed, were normalized using log-transformation. Fibrinogen levels were ascertained using a modification of the Clauss thrombin clotting method on the Organon Teknika MDA 180 analyser. High CRP and high fibrinogen were represented by the highest tertiles of their distribution.
Prevalent chronic illness (coronary heart disease, stroke and/or cancer) at baseline, as well as age and sex, were considered as confounding factors in all analyses.
High-sensitivity C-reactive protein (CRP) was analysed using the N Latex CRP mono Immunoassay on the Behring Nephelometer II Analyzer (Dade Behring, Milton Keynes, UK). In all analyses, CRP data, originally skewed, were normalized using log-transformation. Fibrinogen levels were ascertained using a modification of the Clauss thrombin clotting method on the Organon Teknika MDA 180 analyser. High CRP and high fibrinogen were represented by the highest tertiles of their distribution.
Prevalent chronic illness (coronary heart disease, stroke and/or cancer) at baseline, as well as age and sex, were considered as confounding factors in all analyses.
4
Chronic Inflammation's Role in Health
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At each nurse visit, a blood sample was drawn. High sensitivity serum CRP (mg/L) was analyzed using the N Latex CRP mono Immunoassay on the Behring Nephelometer II Analyzer (Dade Behring, Milton Keynes, UK) (11 ). For the purposes of the present analyses, CRP was treated as a continuous variable. We excluded study members with values of CRP > 10 mg/L, reasoning that they may be cases of acute inflammation and immune activation due to current infection rather than chronic inflammation.
We used covariates collected at baseline: age, sex, educational level [low (no qualification), moderate (up to high school diploma), and high (university degree or higher)], current smoking status (yes/no), and total physical activity (four levels from low to high). Use of anti-inflammatory and antihypertensive medication was self-reported. These constitute the set of common covariates to all models. We used additional covariates measured at wave 1, specific to each outcome, described hereafter.
We used covariates collected at baseline: age, sex, educational level [low (no qualification), moderate (up to high school diploma), and high (university degree or higher)], current smoking status (yes/no), and total physical activity (four levels from low to high). Use of anti-inflammatory and antihypertensive medication was self-reported. These constitute the set of common covariates to all models. We used additional covariates measured at wave 1, specific to each outcome, described hereafter.
5
Biochemical Markers of Metabolic Health
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Heparinized plasma glucose, triglycerides, total and HDL-cholesterol concentrations in samples were determined using enzymatic methods and spectrophotometry (Modular 700, Roche Diagnostics, Meylan France). Low-density lipoprotein (LDL)-cholesterol was calculated by Fried-wald’s formula [Total cholesterol–(HDL cholesterol + triglycerides/5)]. Serum insulin was measured by immunoradiometric sandwich assay (Bis-Insulin IRMA®, CisBio international, Gif-Sur-Yvette, France). The homeostasis model assessment resistance (HOMA-RI) index was calculated by the following equation: insulin (μIU/mL) x glucose (mmol/L)/22.5. Serum high-sensitive C-reactive protein (hsCRP) level was measured using automated immunonephelometry (Behring Nephelometer II Analyzer®, Dade Behring, Germany). Leptin and TNFα were measured by commercially available multiplex beads immunoassays (Fluorokine MAP Multiplex Human Cytokine Panel and Obesity Panel, R&D Systems, Minneapolis, USA) and read by the Bioplex 200 array reader (Bio-Rad Laboratories, Hercules, CA, U.S.A.) which uses Luminex xMAPTM Technology (Luminex Corporation, Austin, TX, U.S.A.).
6
Kidney Function Decline Predictors
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The outcome for the cross-sectional analyses was the presence of CKD, defined as an eGFR ≤60 ml/min per 1.73 m2 at baseline (year 2). The primary outcome for change analyses was 30% decline in eGFR between year 2 and year 9 adjusted for baseline eGFR. The outcomes for sensitivity analyses were (1) rapid decline in kidney function defined as an annualized eGFR decline >3 ml/min per 1.73 m2 per year between year 2 and year 9 adjusted for baseline eGFR and (2) change in eGFR between year 2 and year 9 adjusted for baseline eGFR.
Laboratory measurements for serum creatinine were conducted at year 2 and year 9 at the CHS Core Laboratory using standardized assays. A particle-enhanced immunonephelometric assay (N Latex Cystatin C, Dade Behring) with a nephelometer (Behring Nephelometer II Analyzer, Dade Behring) was used to measure and calibrate Cystatin C.15 (link) eGFR was calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) creatinine-cystatin C equation.16 (link)
Laboratory measurements for serum creatinine were conducted at year 2 and year 9 at the CHS Core Laboratory using standardized assays. A particle-enhanced immunonephelometric assay (N Latex Cystatin C, Dade Behring) with a nephelometer (Behring Nephelometer II Analyzer, Dade Behring) was used to measure and calibrate Cystatin C.15 (link) eGFR was calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) creatinine-cystatin C equation.16 (link)
7
Inflammatory Markers Measurement Protocol
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Inflammatory markers at wave 2 C-reactive protein (CRP) was analysed from serum using the N latex CRP mono assay on the Behring Nephelometer II Analyzer (Dade Behring, Milton Keynes, UK). Intra and inter assay coefficients of variation were less than 2%. Systemic inflammation is defined as CRP>3 mg/L levels. As per previous research, participants with CRP levels higher than 10 mg/L (5.6% of the analytic sample) (likely due to infection) were excluded. CRP was log transformed and used as a continuous variable.
Fibrinogen was analysed from citrate plasma samples using a modification of the Clauss thrombin clotting method on the IL-ACS-TOPS analyser. Intra and inter-assay coefficients of variation were less than 7%. Fibrinogen was used as a continuous variable as there are no established clinical cutpoints and was also log transformed for analyses, given its skewed distribution.
Fibrinogen was analysed from citrate plasma samples using a modification of the Clauss thrombin clotting method on the IL-ACS-TOPS analyser. Intra and inter-assay coefficients of variation were less than 7%. Fibrinogen was used as a continuous variable as there are no established clinical cutpoints and was also log transformed for analyses, given its skewed distribution.
8
Measuring Inflammatory and Lipid Markers
9
Measuring Inflammatory Biomarkers in Plasma
10
Metabolic Biomarker Profiling Protocol
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A sterile antecubital vein puncture was conducted using Vacutainer® tubes without anticoagulants. The sample was then centrifuged at 3500 rpm for 15 min at room temperature and aliquoted in micro vials. Blood samples came from 12 h fasted patients in the morning. All serum samples were stored at −20 °C for ulterior determination in the reference laboratory.
The rheumatoid factor (RF) was determined through nephelometry (Behring Nephelometer™ II Analyzer, BNII, Siemens, Marburg, Germany). Analyses included Total cholesterol (TC), high-density lipoprotein (HDL), and triglycerides (TG). The Friedwald formula [LDL = TC – HDL − TG/5] was used to determine low-density lipoprotein (LDL) [17 (link)].
Glucose was determined with the glucose oxidase technique (VITROS® 4600 System, Ortho-Clinical Diagnostics, Inc, Rochester, NY, USA) [18 (link)]. Type 2 DM diagnosis was considered if patients had a previous diagnosis, were under antidiabetic treatment, or had a fasting glucose value of ≥126 mg/dL [19 (link)].
The rheumatoid factor (RF) was determined through nephelometry (Behring Nephelometer™ II Analyzer, BNII, Siemens, Marburg, Germany). Analyses included Total cholesterol (TC), high-density lipoprotein (HDL), and triglycerides (TG). The Friedwald formula [LDL = TC – HDL − TG/5] was used to determine low-density lipoprotein (LDL) [17 (link)].
Glucose was determined with the glucose oxidase technique (VITROS® 4600 System, Ortho-Clinical Diagnostics, Inc, Rochester, NY, USA) [18 (link)]. Type 2 DM diagnosis was considered if patients had a previous diagnosis, were under antidiabetic treatment, or had a fasting glucose value of ≥126 mg/dL [19 (link)].
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