Thioglucose-stabilized gold nanoparticles as a novel platform for colorimetric bioassay based on nanoparticle aggregation.
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-β-D-glucose as a reducing and stabilizing agent, and their application to colorimetric bioassay was demonstrated using the carbohydrate-lectin system. p-Aminophenyl α-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 Kd values for high- and low-affinity binding of 11.3 and 66.5 pM, respectively, which was significantly lower than that for methyl-α-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 (R2=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.
Watanabe
Shigeru
S
Department of Applied Science, Faculty of Science, Kochi University, 2-5-1 Akebono-cho, Kochi 780-8520, Japan. watanabe@kochi-u.ac.jp
Yoshida
Kazuma
K
Shinkawa
Keitarou
K
Kumagawa
Daisuke
D
Seguchi
Hideki
H
D001681:Biological Assay / Q000379:methods*; D003124:Colorimetry / Q000379:methods*; D003208:Concanavalin A / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay; D005947:Glucose / Q000031:analogs & derivatives* / Q000737:chemistry; D006046:Gold / Q000737:chemistry*; D053768:Metal Nanoparticles / Q000737:chemistry*; D009942:Organometallic Compounds / Q000138:chemical synthesis / Q000737:chemistry*; D010316:Particle Size; D020349:Surface Plasmon Resonance; D013499:Surface Properties
D009942:Organometallic Compounds; D003208:Concanavalin A; D006046:Gold; C018399:1-thioglucose; D005947:Glucose