Blood (10 mL) was collected in ethylenediaminetetraacetic acid vacutainer tubes (Becton Dickinson and Company, Franklin Lakes, New Jersey) and centrifuged at 1250g for 10 minutes; then plasma was removed, aliquoted, and frozen at −80°C. Plasma interleukin-6 (IL-6) was assessed using high-sensitivity enzyme-linked immunosorbent assays (R&D Systems, Abingdon, United Kingdom). The limit of detection of the IL-6 assay was .039 pg/mL, with intra-assay and interassay coefficients of variation of 7.4% and 7.8%. Cytokine analysis was performed using repeated-measures ANOVA in SPSS 22 (IBM Corp., Armonk, New York).
High sensitivity enzyme linked immunosorbent assay
Manufactured by R&D Systems
Sourced in United Kingdom, Germany
The high-sensitivity enzyme-linked immunosorbent assay (HS-ELISA) is a quantitative analytical technique used to detect and measure the concentration of specific analytes in a sample. It utilizes the principle of antigen-antibody interaction, wherein an enzyme-labeled antibody is used to detect the presence and quantity of the target analyte. The HS-ELISA offers increased sensitivity compared to traditional ELISA methods, allowing for the detection of low-abundance analytes.
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Lab products found in correlation
Vacutainer tube, by BD (2 mentions)
Vidas system, by bioMérieux (2 mentions)
Spss 22, by IBM (1 mentions)
Spss statistics version 21, by IBM (1 mentions)
Automated particle enhanced immunoturbidimetric assay, by Roche (1 mentions)
Vascular injury 2 panel, by Mesoscale (1 mentions)
Multiplex bead suspension array immunoassays, by Merck Group (1 mentions)
12 protocols using high sensitivity enzyme linked immunosorbent assay
1
Plasma IL-6 Quantification Protocol
2
Plasma IL-6 Quantification Protocol
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Blood (20 mL) was drawn into BD Vacutainer tubes (Becton, Dickinson and Company, Franklin Lakes, New Jersey) containing ethylenediamine tetraacetic acid (EDTA) anticoagulant and centrifuged immediately at 1250g for 10 min. Plasma was removed, aliquoted, and frozen at −80°C before analysis. Plasma IL-6 was assessed using high-sensitivity enzyme-linked immunosorbent assays (R&D Systems, Abingdon, United Kingdom). The limit of detection of the IL-6 assay is .039 pg/mL, with intra-assay and interassay coefficients of variation of 7.4% and 7.8%, respectively. Cytokine analysis was performed using mixed measures analyses of variance and subsequent paired sample t tests in IBM SPSS Statistics version 21.0 (IBM Corp, Armonk, New York).
3
Quantifying Inflammatory Cytokines in Typhoid Vaccine Model
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We used a S. typhi vaccination model known to induce low-grade inflammation without body temperature change (34) (link). Blood (10 mL) was drawn into ethylenediamine tetraacetic acid BD Vacutainer tubes (Franklin Lakes, New Jersey) and centrifuged at 1250 × g for 10 min, and plasma was removed, aliquoted, and frozen at −80°C. Plasma IL-6, IL-1 receptor antagonist, and tumor necrosis factor alpha were assessed using high-sensitivity enzyme-linked immunosorbent assays (R&D Systems, Abingdon, United Kingdom). Limits of detection were .039 pg/mL, 6.26 pg/mL, and .038 pg/mL with associated intra-assay and interassay coefficients of variation of 7.4% and 7.8%, 5.3% and 8.6%, and 5.3% and 8.4%.
4
Quantifying Serum Inflammation Biomarkers
5
Comprehensive Metabolic Assessment in Diabetes
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Questionnaires for diabetes duration; current and past medication history; and past medical history for cardiovascular disease, alcohol history, and smoking status were completed. Anthropometric measures (height, weight, waist circumference), systolic blood pressure, diastolic blood pressure, heart rate, and clinical laboratory data were assessed at baseline and after 14 days of treatment. Venous blood and urine samples were obtained in the morning after a 12-hour overnight fast and 2 hours after intake of trial regimen. hemoglobin A1c level was measured using high performance liquid chromatography. Serum insulin level was measured using an immunoradiometric assay. Insulin resistance was estimated using homeostatic model assessment for insulin resistance (HOMA-IR), defined as [fasting plasma insulin (mU/L)×fasting plasma glucose (mmol/L)]/22.5 [15 (link)]. Betacell function was estimated using homeostatic model assessment for β cell function (HOMA-β), calculated as fasting plasma insulin (μU/mL)×20/fasting plasma glucose (mmol/L)–3.5 [15 (link)]. Chemistry values were determined using standard assays in each local laboratory. Glomerular filtration rate was estimated by the Modification of Diet in Renal Disease Study Group formula [16 (link)]. The levels of CRP and CD40L were measured using a high sensitivity enzyme-linked immunosorbent assay (R&D Systems, Minneapolis, MN, USA).
6
Inflammatory Cytokine IL-6 and Mood Assessment
7
Biomarkers in Frozen Samples
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All measurements were performed on samples stored at −80 °C. CRP was measured by automated particle-enhanced immunoturbidimetric assay (Roche, UK). This method has an inter- and intra-assay coefficient of variation of 3%. IL-6 was determined using a high-sensitivity enzyme-linked immunosorbent assay (R&D Systems, Abingdon, UK) with inter- and intra-assay coefficients of variation of <6% and sensitivity of 0.16 pg/mL. White blood cell count (WBC) was measured by a fully automated system Sysmex XE-2100 (TOA Medical Electronics, Kobe, Japan).
8
Plasma Biomarkers of Inflammation and Coagulation
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Using plasma collected at baseline and the month 8 visit, which was stored centrally at –70°C, 7 plasma biomarkers were measured that reflected systemic inflammation (IL-6, high-sensitivity C-reactive protein [hsCRP] and serum amyloid A [SAA]), vascular inflammation (soluble intercellular adhesion molecule-1 [sICAM] and vascular adhesion molecule-1 [sVCAM]), immune activation (interleukin-27 [IL-27]), and coagulation activation (D-dimer). IL-6 was measured using high-sensitivity enzyme-linked immunosorbent assay (R&D Systems), D-dimer using the VIDAS system (BioMerieux), and IL-27 using immunoassay (MesoScale). A multiplex platform was used to measure hsCRP, SAA, sICAM, and sVCAM (Vascular Injury II Panel, MesoScale). All samples were analyzed by researchers blinded to treatment group.
9
Quantification of Nitric Oxide Metabolites and Inflammatory Cytokines
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Greiss method was used to ascertain nitrite, the stable metabolite of NO. The formation of nitrite (NO2-) and nitrate (NO3-) was measured in cell culture supernatants. The results are presented as μmol NOx of NO3-/NO2- per liter of medium. According to the manufacturer's instructions, high-sensitivity enzyme-linked immunosorbent assay (R&D Systems, Wiesbaden, Germany) was used to measure the levels of VEGF, GM-CSF, TNF-α, and IL-6 in the plasma.
10
Plasma IL-6 Measurement by ELISA
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Plasma levels of IL-6 were measured by the high sensitivity enzyme-linked immunosorbent assay (R & D Systems, Wiesbaden, Germany) as previously described [15 (link)].
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