Related Experiment Videos
Immunologic quantitation of tissue factors.
B Parhami-Seren1, S Butenas, J Krudysz-Amblo
1Department of Biochemistry, College of Medicine, University of Vermont, Burlington, VT 05446-0068, USA. parhami-seren@uvm.edu
Journal of Thrombosis and Haemostasis : JTH
|August 2, 2006
Summary
Conflicting reports on circulating tissue factor (TF) levels may stem from assay limitations. A new, sensitive immunoassay reveals TF concentrations in plasma are lower than previously reported, impacting disease association studies.
Area of Science:
- Biochemistry
- Immunology
- Clinical Diagnostics
Background:
- Conflicting reports on circulating tissue factor (TF) concentrations create uncertainty about its role in hemostasis and disease.
- Existing TF quantitation assays may contribute to these discrepancies.
- Accurate measurement of TF is crucial for understanding its clinical relevance.
Purpose of the Study:
- To develop and validate a highly sensitive and specific immunoassay for TF antigen quantitation.
- To investigate the impact of different TF species and assay methodologies on measured TF concentrations.
- To determine the concentration of TF antigen in healthy human plasma using the developed assay.
Main Methods:
- Development of a double monoclonal antibody fluorescence-based immunoassay integrated with the Luminex Multi-Analyte Platform.
- Quantitation of recombinant and natural TF species using physiologically relevant standards and specificity controls.
- Comparison of TF antigen levels in plasma from 91 healthy individuals using the new assay.
Main Results:
- The new immunoassay demonstrated high sensitivity (quantifiable at < or = 2 pM) and specificity.
- Differential recognition of various TF species (full-length and truncated) by antibodies was observed.
- Plasma TF antigen concentrations were found to be substantially lower than generally reported, with 79% of individuals below the assay's limit of quantification.
Conclusions:
- Assay methodology significantly impacts TF quantitation, contributing to previous reporting discrepancies.
- The developed immunoassay provides a more accurate assessment of plasma TF antigen levels.
- Lower observed TF levels necessitate re-evaluation of its association with hemostasis and disease.