Preparation of highly stable oligo (ethylene glycol) derivatives-functionalized gold nanoparticles and their application in LSPR-Based detection of PSA/ACT complex

A sandwich immunoassay for PSA/ACT complex detection based on gold nanoparticle aggregation using two probes was developed. The functionalized colloidal gold nanoparticles (AuNPs) showed highly stable not only in the presence of high ionic strength but also in a wide pH range. The functionalized AuNPs were tagged with PSA/ACT complex monoclonal antibody and goat PSA polyclonal antibody and served as the probes to induce aggregation of the colloidal particles. As a result, PSA/ACT complex was detected at concentrations as low as 1 ng/ml. This is the first time that a new aggregation sandwich-immunoassay technique using two gold probes has been used, and the results are generally applicable to other LSPR-based immunoassays.

Preparation of Highly Stable OEG derivatives-functionalized AuNPs and Its Application in LSPR-based Detection of PSA/ACT Complex

The zero-length crosslinker EDC has been widely used to make amide bonds between carboxylic acid and amine groups for bioconjugation because no residues remain in the crosslinked protein. During the conjugation process, EDC activates the carboxyl groups (negatively charged) and forms an unstable amine-reactive intermediate (positively charged). However, the process turns to be a problematic issue if it is applied to modify carboxyl-functionalized and –stabilized Au nanoparticles (AuNPs) due to the fact that the negatively repulsive forces which help to stabilize the AuNPs were disrupted leading to the colloid aggregation. Therefore, to modify the negatively carboxyl-terminated AuNPs while their stability can be maintained yet, we assume that functionalization of the AuNPs using 02 kinds of negatively charged groups which one serves as a linking agent, and the other one plays a role of negative charge maintainer could overcome the impediment.

In this study, the colloidal gold nanoparticles were synthesized by Turkevitch’s method, and then their surface was rationally functionalized with different molar ratios of HS(CH2)11(OCH2CH2)6OCH2COOH and HS(CH2)11(OCH2CH2)3OH (OEG6-COOH/OEG3-OH) by self assembling technique. As a result, the most appropriate molar ratio was found to be 1:10, and the AuNP aggregation was prevented not only in the activation process by EDC but also in the present of high concentration of NaCl as well as over in a wide pH range. This is the first time that extremely stable OEG derivatives-functionalized Au nanoparticles for protein bioconjugation using EDC chemistry is reported, and the results open the door for covalent bioconjugation of AuNPs in biological applications.

Resonant Rayleigh light scattering response of individual Au nanoparticles to antigen--antibody interaction

A proof-of-concept study was reported on analysis of antigen–antibody recognition based on resonant Rayleigh scattering response of single Au nanoparticles in an imaging chamber. As benefited by a traditional dark-field microscope and a spectrograph, individual Au nanoparticles (30 nm) were observed with high signal-to-noise ratio and they were effectively utilized to monitor changes in refractive index induced by specific binding of the adsorbates. Using PSA antigen as a model, a LSPR ?max shift of about 2.85 nm was recorded for a molecular binding corresponding to 0.1 pg ml-1 of the protein biomarker. This result successfully demonstrates a non-labeling detection system for proteins as well as thousands of different chemical or biological species, and it possesses a great potential as a sensitive, on-chip and multiplexing detection.

A new method for non-labeling attomolar detection of diseases based on an individual gold nanorod immunosensor

Herein, we present the use of a single gold nanorod sensor for detection of diseases on an antibody-functionalized surface, based on antibody–antigen interaction and the localized surface plasmon resonance (LSPR) ?max shifts of the resonant Rayleigh light scattering spectra. By replacing the cetyltrimethylammonium bromide (CTAB), a tightly packed self-assembled monolayer of HS(CH2)11(OCH2CH2)6OCH2COOH(OEG6) has been successfully formed on the gold nanorod surface prior to the LSPR sensing, leading to the successful fabrication of individual gold nanorod immunosensors. Using prostate specific antigen (PSA) as a protein biomarker, the lowest concentration experimentally detected was as low as 111 aM, corresponding to a 2.79 nm LSPR ?max shift. These results indicate that the detection platform is very sensitive and outperforms detection limits of commercial tests for PSA so far. Correlatively, its detection limit can be equally compared to the assays based on DNA biobarcodes. This study shows that a gold nanorod has been used as a single nanobiosensor to detect antigens for the first time; and the detection method based on the resonant Rayleigh scattering spectrum of individual gold nanorods enables a simple, label-free detection with ultrahigh sensitivity.

Au Nanoparticles for Applications in Analysis of Cellular and Biomolecular Recognitions

Au nanoparticles (AuNPs) have attracted a great interest in fabrication of various biosensor systems for analysis of cellular and biomolecular recognitions. In conjunction with vast conjugation chemistry available, the materials are easily coupled with biomolecules such as nucleic acids, antigens or antibodies in order to achieve their many potential applications as ligand carriers or transducing platforms for preparation, detection and quantification purposes. Furthermore, the nanoparticles possess easily tuned and unique optical/ physical/ chemical characteristics, and high surface areas, making them ideal candidates to this end. In this topic, sensing mechanisms based on localized surface plasmon resonance (LSPR), particle aggregation, catalytic property, and Fluorescence Resonance Energy Transfer (FRET) of AuNPs as well as barcoding technologies including DNA biobarcodes will be discussed.

METHOD OF DETECTING BIOPRODUCTS USING LOCALIZED SURFACE PLASMON RESONANCE SENSOR OF GOLD NANOPARTICLES

(EN)Disclosed is a method of detecting bioproducts using Localized Surface Plasmon Resonance (LSPR) of gold nanoparticles, which can diagnose bioproducts based on changes in the maximum wavelength occurred by an antigen-antibody reaction after immobilization of the gold nanoparticles onto a glass panel. A sensor using such method exhibits high sensitivity, is low in price, and makes quick diagnosis possible, thereby being applicable to various biological fields associated with environmental contaminants, pathogens and the like, as well as diagnosis of diseases. Further, it provides a technology for manufacturing a sensor having higher sensitivity, low price and quick performance, as compared to conventional methods using SPR.

PROCÉDÉ DE DÉTECTION DE BIOPRODUITS À L'AIDE D'UN DÉTECTEUR PAR RÉSONANCE PLASMONIQUE DE SURFACE LOCALISÉE DE NANOPARTICULES D'OR

L'invention porte sur un procédé consistant à détecter des bioproduits à l'aide d'une résonance plasmonique de surface localisée (LSPR) de nanoparticules d'or. Ledit procédé peut diagnostiquer des bioproduits sur la base de changements de la longueur d'onde maximale produite par une réaction antigène-anticorps après immobilisation des nanoparticules d'or sur un panneau en verre. Un détecteur utilisant un tel procédé présente une sensibilité élevée, est peu coûteux, permet un diagnostic rapide, et il peut ainsi être appliqué à divers domaines biologiques associés aux contaminants environnementaux, pathogènes et similaires, ainsi qu'au diagnostic de maladies. L'invention porte en outre sur une technologie pour fabriquer un détecteur dont la sensibilité est élevée, le prix faible et la performance rapide, par comparaison avec des procédés classiques utilisant SPR.

The ultrasensitive detection of prostate cancer biomarker based on resonant Rayleigh light-scattering response of single Au nanoparticle

A proof-of-concept study was reported on analysis of antigen-antibody recognition based on resonant Rayleigh scattering response of single Au nanoparticles on a microimaging chamber. As benefited by a traditional dark-field microscope and a spectrograph, tiny 30 nm Au nanoparticles were effectively used as nanosensors to monitor changes in refractive index induced by every single binding of the adsorbates. The individual Au nanoparticles were observed with very high signal-to-noise ratio, and a LSPR ?max shift of about 2.5 nm accounting for the detection of PSA antigen with concentration as low as 0.1 pg ml-1 was recorded. This resulted in the successful demonstration of a non-labelling detection system for proteins as well as thousands of different chemical or biological species with possibility of miniaturization and multiplexing scheme.

One-Pot, High-Yield Synthesis of Convex Au Octahedra by H2O2 - Mediated Process In Solution Phase

A simple, non-seeding and high-yield synthesis of convex gold octahedra with size of ca. 50 nm in aqueous solution is described. The octahedral nanoparticles were systematically prepared by reduction of HAuCl4 using ascorbic acid (AA) in the presence of cetyltrimethylammonium bromide (CTAB) as the stabilizing surfactant while concentrations of Au3+ were fixed. The synthesizing process is especially different to other wet synthesis of metallic nanoparticles because it is mediated by H2O2. Mechanism of the H2O2 – mediated process will be described in details. The gold octahedra were shown to be single crystals with all 8 faces belonging to {111} family. Moreover, the single crystalline particles also showed attractive optical properties towards LSPR that should find uses as labels for microscopic imaging, materials for colorimetric biosensings, or nanosensor developments.

Label-free Analysis of Conformational Dynamics and Molecular Interaction of Biomolecules by Vis-NIR Metasensor

Analysis of binding recognition and conformation of biomolecules is of paramount important in understanding of their vital functions in complex biological systems. By enabling sub-wavelength light localization and strong local field enhancement, plasmonic biosensors have become dominant tools used for such analysis owing to their label-free and real-time attributes1,2. However, the plasmonic biosensors are not well-suited to provide information regarding conformation or chemical fingerprint of biomolecules. Here, we show that plasmonic metamaterials, consisting of periodic arrays of artificial split-ring resonators (SRRs)3, can enable capabilities of both sensing and fingerprinting of biomolecules. We demonstrate that by engineering geometry of individual SRRs, localized surface plasmon resonance (LSPR) frequency of the metamaterials could be tuned to visible-near infrared regimes (Vis-NIR) such that they possess high local field enhancement for surface-enhanced Raman scattering spectroscopy (SERS). This will provide the basis for the development of a dual mode label-free conformational-resolving and quantitative detection platform. We present here the ability of each sensing mode to independently detect binding adsorption and to identify different conformational states of Guanine (G)-rich DNA monolayers in different environment milieu. Also shown is the use of the nanosensor for fingerprinting and detection of Arginine-Glycine-Glycine (RGG) peptide binding to the G-quadruplex aptamer. The dual-mode nanosensor will significantly contribute to unraveling the complexes of the conformational dynamics of biomolecules as well as to improving specificity of biodetection assays that the conventional, population-averaged plasmonic biosensors cannot achieve.