Plasma cytokines, lipocalin 2, and serum amyloid A1 were assayed in duplicate with a multiplex immunoassay (Millipore, Molsheim, France) and Luminex xMap technology (Bio‐Rad Laboratories, Hercules, CA) or enzyme‐linked immunosorbent assay (ELISA) (Lipocalin‐2/NGAL Human ELISA Kit EHLCN2 and SAA Human ELISA Kit KHA0011; Invitrogen, Carlsbad, CA) following the manufacturer's instructions.
Multiplex immunoassay
Manufactured by Merck Group
Sourced in United States, Germany, Belgium
The Multiplex immunoassay is a laboratory equipment used for the simultaneous detection and quantification of multiple analytes or biomarkers in a single sample. It is a powerful tool that allows for the efficient and high-throughput analysis of complex biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Luminex xmap technology, by Bio-Rad (3 mentions)
Beckman glucose analyzer, by Beckman Coulter (2 mentions)
Synchron cx5ce, by Beckman Coulter (2 mentions)
Cx9pro, by Beckman Coulter (2 mentions)
Sta r analyzer, by Diagnostica Stago (1 mentions)
Elecsys 2010, by Roche (1 mentions)
Clinical chemistry system 950irc, by Johnson & Johnson (1 mentions)
Adiponectin, by ALPCO (1 mentions)
Trap5b, by Quidel (1 mentions)
Ifn λ2 3, by R&D Systems (1 mentions)
Ifn γ, by BD (1 mentions)
Liquichip luminex 200 system, by Qiagen (1 mentions)
Cytometric bead array cba, by BD (1 mentions)
Milliplex analyst 5, by Merck Group (1 mentions)
P800 blood collection system, by BD (1 mentions)
Microvue ykl 40, by Quidel (1 mentions)
Elecsys immunoassay, by Roche (1 mentions)
21 protocols using multiplex immunoassay
1
Multiplex Assay for Plasma Biomarkers
2
Metabolic Biomarkers in Bariatric Surgery
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Blood samples from overnight fasted BLC participants were analyzed by a commercial laboratory (West Coast Clinical Laboratories, Van Nuys, CA). The chemistry panel was measured on a Beckman Synchron CX5CE or CX9PRO. Insulin was determined by radioimmunoassay, and leptin and adiponectin concentrations were measured using a commercially available kit (Millipore, St. Charles, MO). Analysis of overnight fasted blood samples from RYGB subjects was performed at Vanderbilt University Medical Center. Glucose was measured at the bedside using the glucose oxidase method (Beckman Glucose Analyzer, Fullerton, CA). Insulin and leptin were determined by radioimmunoassay and adiponectin by multiplex immunoassay (Millipore, St. Charles, MO). Triglycerides and total, HDL, and LDL cholesterol were assayed with ACE reagents and instrumentation (Alfa Wassermann, Caldwell, NJ). Insulin resistance was calculated using the homeostasis model assessment of insulin resistance (HOMA-IR) using fasting measurements of glucose and insulin (18 (link)). In both BLC and RYGB samples, thyroid panel (T3, T4, TSH) was measured by immunoassay with chemiluminescent detection (Millipore Corporation, Billerica, MA). Samples from only 9 of the 13 pairs were available for analysis due to a lack of sufficient sample for 4 RGYB participants.
3
Metabolic Biomarkers in Bariatric Surgery
4
Biomarkers of Ischemic Stroke Risk
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Laboratory methods were described in detail [24 (link)]. Briefly, fasting baseline blood samples were drawn in the morning using standardized methods, centrifuged to separate plasma and serum, and shipped overnight on ice to the University of Vermont, where they were processed further and stored at −80 °C. HGF was measured in retrieved plasma of the cases of ischemic stroke and the cohort random sample using a Multiplex Immunoassay (Millipore/Luminex); the interassay coefficient of variation (CV) was 2.98% to 9.54%. N-terminal pro-B-type natriuretic peptide (NT pro-BNP) was measured using an electrochemiluminescence immunoassay (Roche Elecsys 2010; Roche Diagnostics; CV < 5%). D-dimer was measured using an immunoturbidometric assay on the STAR analyzer (Diagnostica Stago); the CV was 5% to 17%. C-reactive protein (CRP) was measured using a high-sensitivity particle-enhanced immunonephelometric assay on the BNII nephelometer (N High Sensitivity CRP, Dade Behring Inc); the CV was 2% to 6%. Total cholesterol, high-density lipoprotein (HDL) cholesterol, and triglycerides were measured by colorimetric reflectance spectrophotometry using the Ortho Vitros Clinical Chemistry System 950IRC instrument (Johnson & Johnson Clinical Diagnostics). Interleukin (IL)-6 was measured by ultrasensitive ELISA (Quantikine HS Human IL-6 Immunoassay) with a CV of 6.3%.
5
Biomarkers of Bone Turnover and Metabolic Factors
6
Lung Cytokine Profiling in Mice
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A single lung lobe from each mouse was excised and snap frozen before being homogenised in buffers recommended in the manufacturer's instructions. Levels of IL-5, IL-13, IFNγ (BD Biosciences; PharMingen), IFNα, IFNβ (Verikine; PBL Assay Science), CCL7/MCP3 and IFNλ2/3 (R&D Systems) were determined in clarified lung lysates by ELISA. CCL2/MCP1, CCL3/MIP1α, CCL4/MIP1β, CXCL9/MIG and CXCL10/IP10 were measured by employing a Multiplex Immunoassay (Millipore). All concentrations were normalised to lung weight.
7
Metabolic Biomarkers in Fasted Subjects
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All subjects were fasted for at least 12 h (maximum 14 h) before phlebotomy. Glycaemia, triglycerides, total cholesterol and high-density cholesterol levels were measured by an enzymatic colorimetric dry chemistry method (Johnsons & Johnsons, Raritan, USA, VITROS 5600). Low-density cholesterol was determined by Friedewald’s formula. Insulin levels were measured by chemiluminescence immunoassay (Linco Research Incorporated, St. Charles, USA, Lincoplex).
High-sensitivity C-reactive protein (hs-CRP) levels were determined through particle-enhanced immunonephelometry. Plasma IL-1β, IL-6, IL-8, IL-10, IL-12, TNF-α and adiponectin levels were measured using a Multiplex immunoassay (Milliplex, Merck Millipore, MA, USA) according to manufacturer’s instructions.
Insulin resistance was estimated by homeostasis model assessment (HOMA), which was calculated as [glycemia (mMol) x insulin (uU/mL) ÷ 22,5]. The insulin resistance was diagnosed if any of the following conditions were met: BMI > 28.9 kg/m2, homeostasis model assessment of insulin resistance (HOMA-IR) > 4.65, or BMI > 27.5 kg/m2 and HOMA-IR > 3.60 [21 (link)].
High-sensitivity C-reactive protein (hs-CRP) levels were determined through particle-enhanced immunonephelometry. Plasma IL-1β, IL-6, IL-8, IL-10, IL-12, TNF-α and adiponectin levels were measured using a Multiplex immunoassay (Milliplex, Merck Millipore, MA, USA) according to manufacturer’s instructions.
Insulin resistance was estimated by homeostasis model assessment (HOMA), which was calculated as [glycemia (mMol) x insulin (uU/mL) ÷ 22,5]. The insulin resistance was diagnosed if any of the following conditions were met: BMI > 28.9 kg/m2, homeostasis model assessment of insulin resistance (HOMA-IR) > 4.65, or BMI > 27.5 kg/m2 and HOMA-IR > 3.60 [21 (link)].
8
Multiplex Immunoassay for Adipocytokine Profiling
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Blood samples from all subjects were collected in monovettes prepared with the anticoagulant ethylene-diamine-tetra-acetic acid (EDTA). Plasma samples were obtained by centrifugation and stored at − 80 °C until analyzed.
Measurements of concentrations of the adipocytokines resistin, adiponectin, leptin, interleukin (IL)-6, IL-1β, and TNF-α in plasma samples were performed using a multiplex immunoassay (Merck Millipore, Darmstadt, Germany) following the manufacturer’s protocol. In brief, standards and samples were diluted in an equal volume of assay puffer and incubated with antibody-immobilized beads at 4 °C for 16 h. After washing, a biotinylated detection antibody was added for 1 h followed by incubation with streptavidin-phycoerythrin for 30 min at room temperature. Adipocytokine levels were determined using the LiquiChip luminex 200 system (Qiagen, Hilden, Germany).
Measurements of concentrations of the adipocytokines resistin, adiponectin, leptin, interleukin (IL)-6, IL-1β, and TNF-α in plasma samples were performed using a multiplex immunoassay (Merck Millipore, Darmstadt, Germany) following the manufacturer’s protocol. In brief, standards and samples were diluted in an equal volume of assay puffer and incubated with antibody-immobilized beads at 4 °C for 16 h. After washing, a biotinylated detection antibody was added for 1 h followed by incubation with streptavidin-phycoerythrin for 30 min at room temperature. Adipocytokine levels were determined using the LiquiChip luminex 200 system (Qiagen, Hilden, Germany).
9
Inflammatory and Cardiovascular Biomarker Profiling
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Plasma samples were used for the assessment of inflammatory markers by Cytometric Bead Array (CBA, BD Bioscience, San Diego, CA, USA), using the following kits: Cytokines Th1/Th2 [IL-2, IL-4, IL-10, tumor necrosis factor (TNF), and interferon-gamma (IFN-γ)], Chemokines [CCL5/regulated on activation, normal T cell expressed and secreted (RANTES), CXCL9/monokine induced by interferon-gamma (MIG), CCL2/monocyte chemoattractant protein (MCP)-1, CXCL10/interferon-gamma-induced protein (IP)-10], and human transforming growth factor-beta (TGF-β)-1 Single Plex Flex Set. In addition, multiplex immunoassay (Merck Millipore, Billerica, MA, USA) was used to assess the levels of cardiovascular disease-related proteins, using the Human Cardiovascular Disease-Panel II kit [a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), growth differentiation factor (GDF)-15, myoglobin, soluble intercellular adhesion molecule (sICAM)-1, p-selectin, neutrophil gelatinase-associated lipocalin (lipocalin-2/NGAL), soluble vascular cell adhesion protein (sVCAM)-1, and serum amyloid A (SAA)].
10
Plasma Cytokine Multiplex Assay
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Before the assay, blood samples were centrifuged at 500 g for 10 min at 4 °C and stored at − 80 °C. According to manufacturers’ instructions, the plasma cytokine (IFN-γ, IL-4, IL-17a, and IL-10) levels were assayed using the human cytokine magnetic beads multiplex immunoassay (Merck, Germany). Briefly, samples, standards, and quality control reagents were added to a 96-well plate pre-wetted with wash buffer. Next, premixed beads were added to each well and incubated overnight at 4 °C. After washing, a detection antibody solution was added and incubated for 1 h. Then, a streptavidin-phycoerythrin solution was added and incubated for 30 min. After washing and resuspending beads, the plate was processed and analyzed the Median Fluorescent Intensity (MFI) data using Luminex 2000 software. Cytokine concentrations were calculated from a calibration curve by 5-parametric curve fitting using MILLIPLEX analyst 5.1 software (Merck, Germany).
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