Journal
COLLOIDS AND SURFACES B-BIOINTERFACES
Volume 81, Issue 2, Pages 570-577Publisher
ELSEVIER
DOI: 10.1016/j.colsurfb.2010.07.061
Keywords
Gold nanoparticle; Lectin; Carbohydrate; Colorimetric bioassay; Thioglucose; Surface plasmon resonance
Funding
- Japan Society for the Promotion of Science(JSPS)
- Ministry of Education, Culture, Sports, Science and Technology, Japan [18101005, 21550084]
- Kochi University
- Grants-in-Aid for Scientific Research [21550084, 18101005] Funding Source: KAKEN
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Gold nanoparticles stabilized with thioglucose (TGlu-AuNPs), which have carboxyl groups on the particle surface as anchoring sites for covalent immobilization of biomolecules, were prepared by the chemical reduction of HAuCl4 using 1-thio-beta-D-glucose as a reducing and stabilizing agent, and their application to colorimetric bioassay was demonstrated using the carbohydrate-lectin system. p-Aminophenyl alpha-D-mannose (Man-NH2) was covalently attached by a conventional method to the activated carboxyl groups on the TGlu-AuNPs. On addition of Con A to the Man-AuNPs, multiple binding events occurred between Con A and the mannoses immobilized on the particle surface. This Con A-induced aggregation resulted in a significant red shift in local surface plasmon resonance. The binding isotherm showed a sigmoidal curve, indicating cooperativity in the binding of Con A and the Man-AuNPs. In addition, Hill plots showed two nonequivalent binding modes, with the K-d values for high- and low-affinity binding of 11.3 and 66.5 pM, respectively, which was significantly lower than that for methyl-alpha-D-mannose binding to Con A. The enhanced binding affinity between Man-AuNPs and Con A involves the cluster effect of the carbohydrate groups on the AuNPs. A linear correlation curve was obtained in the range 10-100 nM (R-2 =0.983). The limit of detection (LOD) for Con A was 9.0 nM in aqueous buffer, which is comparable to that of other conventional methods such as ELISA. (C) 2010 Elsevier B.V. All rights reserved.
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