Measurement of PAI‐1 and C‐reactive protein (CRP) was performed at the Research Institute of Internal Medicine at Oslo University Hospital, Rikshospitalet. Plasma samples were thawed in a water bath at 37°C for 5 min, followed by centrifugation for 2 min at 13,500 g to obtain platelet‐free plasma. Plasma PAI‐1 antigen levels were measured in duplicate by enzyme‐immunoassay (EIA) with matched antibodies from R&D Systems designed to detect PAI‐1 in its active and latent forms but not in complex with tissue plasminogen activator. EIA was performed in a 384‐format using a combination of a SELMA (Jena) pipetting robot and a BioTek dispenser/washer. Absorption was read at 450 nm with wavelength correction set to 540 nm using an ELISA plate reader (Bio‐Rad). PAI‐1 intra‐ and inter‐assay coefficients of variation were <10%. CRP was measured by high‐sensitivity CRP (hsCRP) using EIA, as previously described.28
Matched antibodies
Manufactured by R&D Systems
Sourced in United States
Matched antibodies are a set of two or more antibodies that are specifically designed to work together in an assay or application. They are produced and validated to recognize different epitopes on the same target protein, providing a consistent and reliable tool for researchers. The core function of matched antibodies is to enable the detection and measurement of target proteins in various biological samples.
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Lab products found in correlation
6 protocols using matched antibodies
1
Quantifying PAI-1 and CRP Biomarkers
2
ELISA Quantification of IL-8 Levels
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IL-8 levels were measured in cell supernatants using a specific ELISA with matched antibodies from R&D Systems (Minneapolis, USA) according to the manufacturer’s descriptions.
3
Cytokine Measurements in Lung Cells
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Human MCP-1 and CXCL8 (IL-8) were measured using matched antibodies from R&D Systems. Soluble IL-6R was measured using Europium-Streptavidin as previously described (1 (link)). Serum samples were collected in clot activating tubes (Vacutainer, BD, UK). IL-6 and MCP-1 in serum was measured using the V-plex human assay kit (Meso Scale Discovery, MD, Maryland, US) with a lower limit of detection of 0.06 pg/ml and 0.13 pg/ml respectively.
Lung endothelial and epithelial cells. Human pulmonary artery endothelial cells (HPAECs) were from Lonza (Slough, UK) and stimulated TNF-α (5 ng/ml) or sIL-6R ± IL-6 under growth-factor-free conditions for 24 h in EBM-2 media (Lonza, Switzerland). Serum stimulations were performed in EBM-2 media with 65% serum from A/A or C/C genotyped donors. Human bronchial epithelial cells (HBECs) were originally derived in the laboratory of Professor Jerry Shay, and with his permission were gifted to us by Professor Gisli Jenkins (Nottingham Respiratory Research Unit, Nottingham City Hospital). The HBECs were incubated under growth-factor free conditions for 4-24 h with TNF-α (5 ng/ml) or sIL-6R ± IL-6 prior to collection of supernatants for measurement. For the serum stimulation experiments 65% serum (v/v) was added to each well to obtain a final sIL-6R concentration as close to 50 ng/ml in the C/C donors.
Lung endothelial and epithelial cells. Human pulmonary artery endothelial cells (HPAECs) were from Lonza (Slough, UK) and stimulated TNF-α (5 ng/ml) or sIL-6R ± IL-6 under growth-factor-free conditions for 24 h in EBM-2 media (Lonza, Switzerland). Serum stimulations were performed in EBM-2 media with 65% serum from A/A or C/C genotyped donors. Human bronchial epithelial cells (HBECs) were originally derived in the laboratory of Professor Jerry Shay, and with his permission were gifted to us by Professor Gisli Jenkins (Nottingham Respiratory Research Unit, Nottingham City Hospital). The HBECs were incubated under growth-factor free conditions for 4-24 h with TNF-α (5 ng/ml) or sIL-6R ± IL-6 prior to collection of supernatants for measurement. For the serum stimulation experiments 65% serum (v/v) was added to each well to obtain a final sIL-6R concentration as close to 50 ng/ml in the C/C donors.
4
Quantification of Serum OPG Levels
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Serum OPG was quantified by an enzyme immunoassay using commercially available matched antibodies (R&D Systems, Minneapolis, MN). The intra-assay and interassay coefficients of variation were 3.6% and 10.6%, respectively.24 (link)
5
Biomarker Assessment in Post-PCI Remodeling
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A prospective objective of the POSTEMI trial was to study specific biomarkers in relation to LV remodeling and function. Blood samples for biobanking and analyses of OPG were drawn before and immediately after the PCI procedure, at Day 1 (median 14.7 hours after PCI), and at 4-month follow-up. Blood was centrifuged after coagulation (within 1 hour) at 2,500g for 10 minutes and serum was stored at -80°C in multiple aliquots until analyses. Serum levels of OPG were quantified by enzyme immunoassay using matched antibodies from R&D Systems (Minneapolis, Minnesota, USA) as previously described and validated [21 (link)]. The intra-assay and inter-assay coefficients of variation (CV) were <10%. The sensitivity was calculated to be 15 pg/ml. C-reactive protein (CRP) was determined by routine laboratory high-sensitivity assay, peak CRP was defined as the maximum value measured during hospitalization. Levels of N-terminal pro-B-type natriuretic peptide (NT-proBNP) on admission were determined with Elecsys proBNP sandwich immunoassay on Elecsys 2010 (Roche Diagnostics). Serum cardiac-specific troponin T (TnT) was measured by electrochemiluminescence technology for quantitative measurement (Elecsys 2010, Roche, Mannheim, Germany). The peak TnT level was defined as the maximum value measured during hospitalization. Inter-assay and intra-assay CV% were <7% for all assays.
6
Serum Biomarkers of Inflammation Measurement
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Peripheral venous blood was drawn into pyrogen-free tubes without any additives. After clotting at room temperature, the tubes were centrifuged at 1,500g for 10 minutes, and serum was stored at -70°C. Serum levels of sCD14 (product no. DY383/intra-assay CV: 2.2% /inter-assay CV: 7.3%), sCD163 (DY1607/2.9%/3.9%), sCD25 (DY223/3.1%/8.9%), sCD166 (DY656/3.6%/6.4%), Gal3BP (DY2226/6.5%/9.5%) and hsCRP (DY1707/3.0%/9.1%) were measured in duplicate by enzyme immune-assay with matched antibodies obtained from R&D Systems (Minneapolis, MN) in a 384 format using the combination of a SELMA (Jena, Germany) pipetting robot and a BioTek (Winooski, VT, USA) dispenser/washer. Absorption was read at 450 nm with wavelength correction set to 540 nm using an ELISA plate reader (Bio-Rad, Hercules, CA, USA)
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