human thrombomodulin duoset elisa  (R&D Systems)

Journal: Virulence

Article Title: Staphylococcus aureus toxins mediate endothelial Thrombomodulin release during severe invasive infections

doi: 10.1080/21505594.2025.2605767

Figure Lengend Snippet: Assessments of mechanisms leading to increased sTM levels following stimulation with S. aureus proteins. HUVEC monolayers were stimulated for 22 h with supernatant of overnight cultures of clinically isolated strains of S. aureus (M37, P08) and α-toxin. sTM was assessed in the host cell culture supernatant of stimulated cells (A). Thrombomodulin gene (THBD) expression in HUVECs after stimulation was assessed by qPCR (B) and total TM levels were measured after cell lysis of stimulated cells (C). LDH and TM were measured in cell culture supernatants after stimulations, with and without addition of 1300 nM broad-spectrum MMP inhibitor Marimastat (MMPI) or 10,6 µM ADAM10 selective inhibitor GI254023X (A10I) (D-E). LDH release measurements indicate cytotoxicity. Quantification of sTM and LDH are given in corrected optical density values. Asterisks indicate statistically significant differences (* = p < 0.05, ** = p < 0.01, *** = p < 0.001) according to ANOVA followed by Fisher’s least significant difference (LSD) post hoc test (A, B, C) or student t-test (D, E). Colors indicate different stimuli and symbols indicate different biological replicates. Direct cleavage of 40 ng rTM by 100 ng rADAM10 (rA10) was evaluated by western blot after 2 h incubation at 37 °C, with or without MMPI or A10I (F). Cleavage products are indicated by arrows at approximately 60 and 40 kDa.

Article Snippet: The Human Thrombomodulin DuoSet ELISA (R&D Systems, DY3947) was used according to the manufacturer’s instructions to measure sTM levels.

Techniques: Isolation, Cell Culture, Expressing, Lysis, Western Blot, Incubation

Journal: Frontiers in Immunology

Article Title: Thromboinflammation Supports Complement Activation in Cancer Patients With COVID-19

doi: 10.3389/fimmu.2021.716361

Figure Lengend Snippet: Comparison of plasma markers of endothelial cell dysfunction (ICAM 1, Thrombomodulin, P-Selectin). (A) shows results for patients with positive (+) or negative (-) COVID-19 PCR test results. (B) further stratifies results based on low (<1.0 ug/ml FEU) or high (>1.0 ug/ml FEU) D-dimer concentration. Numerical results are summarized in (C) .

Article Snippet: Selected biomarkers of endothelial cell dysfunction, including ICAM-1 (Quantikine ELISA, Human ICAM-1/CD54 -specific immunoassay, R&D Systems, Inc., Minneapolis, MN), and thrombomodulin (Quantikine ELISA, Human Thrombomodulin/BDCA-3 immunoassay, R&D Systems, Inc., Minneapolis, MN), as well as platelet and endothelial cell activation, P-selectin (Quantikine ELISA, Human P-Selectin/CD62P immunoassay, R&D Systems, Inc., Minneapolis, MN), were evaluated.

Techniques: Comparison, Clinical Proteomics, Concentration Assay

Journal: EClinicalMedicine

Article Title: Upregulation of pulmonary tissue factor, loss of thrombomodulin and immunothrombosis in SARS-CoV-2 infection

doi: 10.1016/j.eclinm.2021.101069

Figure Lengend Snippet: Biomarkers of endothelial cell activation in SARS-CoV-2 infection and controls. (A) vWF:Ag, (B) vWF:RCo, (C) ADAMTS13, (D) thrombomodulin, (E) sEPCR, (F) TFPI, (G) Angiopoietin-1, (H) Angiopoietin-2 and (I) P -selectin plasma concentrations were determined by ELISA in healthy controls and COVID-19 patients. Each symbol correspond one patient. Shaded bars indicate the means, and the lines show the SEM. NS, non-significant; * p < 0.05; ** p < 0.01, *** p < 0.001, **** p < 0.0001 (Kruskall-Wallis test, or Mann-Whitney U test).

Article Snippet: DUOSet ELISA (enzyme-linked immunosorbent assay) was performed for human angiopoietin-1 (DY623), angiopoietin-2 (DY923), elastase (DY9167), P-selectin/CD62P (DY137), soluble endothelial protein C receptor (sEPCR, DY2245), plasminogen activation inhibitor-1 (PAI-1, DY1786), Tissue Factor Pathway Inhibitor (TFPI, DY2974), thrombomodulin (DY3947), tissue-type plasminogen activator (t-PA, DY7449) (R&D, Minneapolis, MN).

Techniques: Activation Assay, Infection, Clinical Proteomics, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: EClinicalMedicine

Article Title: Upregulation of pulmonary tissue factor, loss of thrombomodulin and immunothrombosis in SARS-CoV-2 infection

doi: 10.1016/j.eclinm.2021.101069

Figure Lengend Snippet: Univariate and multivariate logistic regression of association of covariates with severe disease.

Article Snippet: DUOSet ELISA (enzyme-linked immunosorbent assay) was performed for human angiopoietin-1 (DY623), angiopoietin-2 (DY923), elastase (DY9167), P-selectin/CD62P (DY137), soluble endothelial protein C receptor (sEPCR, DY2245), plasminogen activation inhibitor-1 (PAI-1, DY1786), Tissue Factor Pathway Inhibitor (TFPI, DY2974), thrombomodulin (DY3947), tissue-type plasminogen activator (t-PA, DY7449) (R&D, Minneapolis, MN).

Techniques:

Journal: EClinicalMedicine

Article Title: Upregulation of pulmonary tissue factor, loss of thrombomodulin and immunothrombosis in SARS-CoV-2 infection

doi: 10.1016/j.eclinm.2021.101069

Figure Lengend Snippet: Tissue Factor (TF), thrombomodulin and EPCR in the lung of SARS-CoV-2 infection and controls. (A) Control case #1. H&E shows preserved alveolar structures. (B) COVID-19 case #1. H&E shows loss of normal lung architecture and changes associated with the disease. (C) Control case #1. TF expression is minimally detected in the alveolar epithelium. (D) COVID-19 case #1. Marked upregulation of TF in epithelial cells. (E) Control case #1. Preserved thrombomodulin expression in the endothelial cells of alveolar capillaries. (F) COVID-19 case #1. Markedly attenuated staining of thrombomodulin. (G) Control case#1. Preserved EPCR expression in the endothelial cells of larger vessels. (H) COVID-19 case #1. Variable/preserved staining of EPCR in large vessels. The yellow arrows show 3 vessels in consecutive sections of COVID-19 case #1, allowing comparison of similar areas for all immunostains. The red arrows indicate expression of the proteins of interest. All images (x200). Bar represents 100 μm (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.).

Article Snippet: DUOSet ELISA (enzyme-linked immunosorbent assay) was performed for human angiopoietin-1 (DY623), angiopoietin-2 (DY923), elastase (DY9167), P-selectin/CD62P (DY137), soluble endothelial protein C receptor (sEPCR, DY2245), plasminogen activation inhibitor-1 (PAI-1, DY1786), Tissue Factor Pathway Inhibitor (TFPI, DY2974), thrombomodulin (DY3947), tissue-type plasminogen activator (t-PA, DY7449) (R&D, Minneapolis, MN).

Techniques: Infection, Control, Expressing, Staining, Comparison

Journal: EClinicalMedicine

Article Title: Upregulation of pulmonary tissue factor, loss of thrombomodulin and immunothrombosis in SARS-CoV-2 infection

doi: 10.1016/j.eclinm.2021.101069

Figure Lengend Snippet: Immunothrombosis and Virchow`s triad in SARS-CoV-2 infection. Multiple mechanisms ( e.g. viral cytopathic, complement activation, NETs, hypoxia, pyroptosis, “cytokine storm”) lead to TF expression in the epithelial cells and possibly other cell types. It also promotes activation of endothelial cells resulting in immunothrombosis with release of NETs (Netosis). Given the tropism of SARS-CoV-2, these events are particularly intense in (but not restricted to) the lungs and creates a marked inflammatory response. This process is dysregulated resulting in an exacerbated coagulation-inflammation-angiogenesis cycle. Excessive thrombin generation overcomes inhibition by local anticoagulants whose functions are impaired due to endothelial cell damage with loss of thrombomodulin and TFPI. As a result fibrin deposition and platelet thrombi occur. Accordingly, 3 component of the Virchow's triad, namely hypercoagulability, endothelium activation, and blood stasis are present in most patients and may predispose to thrombosis. Excess thrombin in the lung may also reach the systemic circulation and is likely neutralized by natural anticoagulants, preventing a consumption coagulopathy, unless superimposed with bacterial sepsis and/or liver dysfunction. This process is typical of a compensated coagulopathy (non-overt DIC) [ , ], with compensatory fibrinolysis explaining elevated D -dimers secondary to plasmin degradation of fibrin. In extreme conditions, impaired microcirculation results in organ failure, high morbidity and mortality.

Article Snippet: DUOSet ELISA (enzyme-linked immunosorbent assay) was performed for human angiopoietin-1 (DY623), angiopoietin-2 (DY923), elastase (DY9167), P-selectin/CD62P (DY137), soluble endothelial protein C receptor (sEPCR, DY2245), plasminogen activation inhibitor-1 (PAI-1, DY1786), Tissue Factor Pathway Inhibitor (TFPI, DY2974), thrombomodulin (DY3947), tissue-type plasminogen activator (t-PA, DY7449) (R&D, Minneapolis, MN).

Techniques: Infection, Activation Assay, Expressing, Coagulation, Inhibition