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4 protocols using d dimer

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Coagulation Assays in BSL-3 Setting

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Prothrombin time (PT) and activated partial thromboplastin time (APTT) were measured using a BCS-XP coagulation analyzer (Siemens Healthcare Diagnostics) according to the instructions of the manufacturer. Clotting was initiated with Thromborel S (PT) and Pathrombin SL (APTT). VWF ristocetin cofactor activity was also determined on the BCS-XP with reagents of the manufacturer, and was expressed as percentage of normal pooled human plasma. Thrombin-antithrombin complexes (TAT, Siemens Healthcare Diagnostics) and D-dimer levels (Asserachrom, Roche, The Netherlands) were measured using enzyme-linked immunosorbent assay. All these assays were carried out within the BSL-3 setting after careful calibration and validation.
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Multiplex Cytokine and Coagulation Biomarker Assay

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Interleukin (IL)-6, IL-10, IL-12p70, interferon (IFN)-γ, monocyte-chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) were measured by cytometric bead array (CBA) multiplex assay (BD Biosciences, San Jose, CA) in accordance with the manufacturers' recommendations. Thrombin-antithrombin complexes (TATc; Siemens Healthcare Diagnostics, Marburg, Germany) and D-dimer (Asserachrom D-dimer, Roche Woerden, the Netherlands) were measured with commercially available ELISA kits. Protein levels in BALF were measured using a Bradford-based protein assay (Bio-Rad Laboratories, Hercules, CA). Aspartate aminotranspherase (ASAT) and alanine aminotranspherase (ALAT) were determined with commercial available kits (Sigma-Aldrich, St. Louis, MO), using a Hitachi analyzer (Boehringer Mannheim, Mannheim, Germany) according to the manufacturers' instructions.
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Coagulation Assays Validation Across Analyzers

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The study was conducted between January 2017 and April 2017 at three European teaching hospitals (Inselspital University Hospital, Bern, Switzerland; Erasmus University Medical Center, Rotterdam, The Netherlands; Sheffield Haemostasis and Thrombosis Centre, UK). Anonymized residual sodium citrate (3.2%/0.109 mol/l) plasma samples were evaluated on the cobas t 711 (high-throughput; max. 390 tests/h: all three centers) and cobas t 511 (mid-throughput; max. 195 tests/h: UK center only) analyzers, using a number of coagulation assays (aPTT, aPTT Lupus, aPTT Screen, AT, d-Dimer, Fibrinogen, PT-derived Fibrinogen, PT Owren, PT Rec and TT; Roche Diagnostics, Switzerland) described in detail previously [27 (link),29 (link)]. All assays and analyzers were used according to manufacturers’ instructions.
Independent ethics committee approval was obtained before study initiation and the study was performed according to the principles of the Declaration of Helsinki and International Council for Harmonisation Good Clinical Practice guidelines.
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Biomarker Profiling in Clinical Study

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All biomarkers were measured on the study entry specimens. Vitamin D was measured as 25(OH)D with the 25(OH)D iodine 125 radioimmunoassay (Diasorin, Saluggia, Italy). The following soluble biomarkers were measured at the Diabetes Research and Training Center Radioimmunoassay Core Laboratory (Washington University School of Medicine): IL-8 (BD Biosciences, San Jose, CA) using an ezyme-linked immunosorbent assay (ELISA) based assay, as well as hsCRP (Kamiya Biomedical Company, Seattle, WA) and D-dimer (Roche Diagnostics, Indianapolis, IN) using immunoturbidometric assays on a Hitachi 917 analyzer. We measured sCD14 using an ELISA based assay (R&D Systems, Minneapolis MN). Tumor necrosis factor (TNF)-α was measured using chemiluminescence on the Immulite 1000 (Erlangen, Germany). The following biomarkers were measured at the National Institutes of Allergy and Infectious Diseases: IL-6 (Meso Scale Discovery, Rockville, MD) using electrochemiluminescence immunoassays and sCD163 (Aviscera Bioscience, Inc., Santa Clara, CA) using an ELISA based assay.
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