Label-free colorimetric detection of picomolar thrombin in blood plasma using a gold nanoparticle-based assay
Journal
Biosensors and Bioelectronics
Journal Volume
25
Journal Issue
8
Pages
1922-1927
Date Issued
2010
Author(s)
Abstract
We unveil a novel, label-free, colorimetric assay - using fibrinogen (Fib) and gold nanoparticles (Au NPs) - for the highly selective and sensitive detection of thrombin. Addition of fibrinogen to a solution of Au NPs (average diameter: 56 nm) led to ready conjugation, forming Fib-Au NPs through electrostatic and hydrophobic interactions. Introduction of thrombin (a serine protease) into the Fib-Au NPs solutions in the presence of excess fibrinogen induced the formation of insoluble fibrillar fibrin-Au NPs agglutinates through the polymerization of the unconjugated and conjugated fibrinogen. After centrifugation, the absorbance at 532 nm of the supernatants decreased upon increasing the concentration of thrombin. This Fib-Au NP probe provides high sensitivity [limit of detection (LOD): 0.04 pM] for thrombin, with remarkable selectivity over other proteins and proteases. The range of linearity for the absorbance against the thrombin concentration was 0.1-10 pM (R2 = 0.96). This approach provides an LOD for thrombin that is lower than those obtainable using other nanomaterial- and aptamer-based detection methods. We validated the utility of this Fib-Au NP probe through separate analyses of thrombin and Factor Xa at picomolar levels in plasma samples-without the need for sample pretreatment. This technique appears to have practical potential in the diagnosis of diseases associated with coagulation abnormalities and cancers (e.g., pulmonary metastasis). ? 2010 Elsevier B.V. All rights reserved.
Subjects
Biosensor; Colorimetric detection; Enzyme; Gold nanoparticles; Thrombin
SDGs
Other Subjects
Absorbances; Aptamers; Average diameter; Blood plasma; Colorimetric assays; Colorimetric detection; Concentration of; Detection methods; Factor Xa; Gold nanoparticle; Gold Nanoparticles; High sensitivity; Hydrophobic interactions; Label free; Nanomaterial; Picomolar levels; Plasma samples; Sample pretreatment; Sensitive detection; Separate analysis; Serine protease; Supernatants; Amino acids; Biosensors; Blood; Centrifugation; Coagulation; Color; Colorimetry; Diagnosis; Enzymes; Gold alloys; Hydrophobicity; Labels; Nanoparticles; Phosphatases; Probes; Gold; aptamer; blood clotting factor 10a; fibrin; fibrinogen; gold nanoparticle; nanomaterial; thrombin; adult; article; biosensor; centrifugation; colorimetry; controlled study; human; male; molecular interaction; normal human; particle size; polymerization; protein analysis; sensitivity and specificity; validation process; Biosensing Techniques; Blood Chemical Analysis; Colorimetry; Equipment Design; Equipment Failure Analysis; Gold; Humans; Nanoparticles; Reproducibility of Results; Sensitivity and Specificity; Staining and Labeling; Thrombin
Type
journal article