Gold nanoparticles (GNPs) are attractive alternative optical probes and good biocompatible materials due to their special physical and chemical properties. However, GNPs have a tendency to aggregate particularly in the presence of high salts and certain biological molecules such as nucleic acids and proteins. How to improve the stability of GNPs and their bioconjugates in aqueous solution is a critical issue in bioapplications. In this study, we first synthesized 17 nm GNPs in aqueous solution and then modified them with six thiol compounds, including glutathione, mercaptopropionic acid (MPA), cysteine, cystamine, dihydrolipoic acid, and thiol-ending polyethylene glycol (PEG-SH), via a Au-S bond. We systematically investigated the effects of the thiol ligands, buffer pH, and salt concentrations of the solutions on the colloidal stability of GNPs using UV-vis absorption spectroscopy. We found that GNPs modified with PEG-SH were the most stable in aqueous solution compared to other thiol compounds. On the basis of the above results, we developed a simple and efficient approach for modification of GNPs using a mixture of PEG-SH and MPA as ligands. These biligand-modified GNPs were facilely conjugated to antibody using 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide and N-hydroxysulfosuccinimide as linkage reagents. We conjugated GNPs to epidermal growth factor receptor antibodies and successfully used the antibody-GNP conjugates as targeting probes for imaging of cancer cells using the illumination of a dark field. Compared to current methods for modification and conjugation of GNPs, our method described here is simple, has a low cost, and has potential applications in bioassays and cancer diagnostics and studies.

Fluorescence resonance energy transfer (FRET) is a powerful technique for probing very small changes in the distance between donor and acceptor fluorophores, and is ideal for the sensitive detection of molecular binding events and changes in protein conformation in response to interactions with a particular target molecule or to changes in the solution environment. In recent years, luminescent semiconductor nanocrystals (also called quantum dots, QDs) have been favorably adopted in the FRET-based studies, such as deriving QD-protein-conjugate configuration, [1] QD-protein sensing assemblies, [2,3] photochromic switching, [4] photodynamic medical therapy, [5] and probing DNA replication and telomerization.[6] This success is due to the unique sizedependent physical and chemical characteristics of QDs. QDs were applied to FRET as both donors [7][8][9][10] and acceptors, [11][12][13] and some of the advantages offered compared to conventional dyes include sharp and symmetrical emission spectra, high quantum yield (QY), good chemical and photo stability, and size-dependent emission-wavelength tunability. Recently, QDs as acceptors were successfully used in bioluminescence resonance energy transfer (BRET).[14]As with any fluorescence technique, however, photobleaching and autofluorescence limit the usefulness of FRET. Herein we report a resonance energy transfer between chemiluminescent donors and QDs as acceptors, which is called chemiluminescence (CL) resonance energy transfer (CRET), and is similar to BRET. [14][15][16][17][18][19] CRET involves nonradiative (dipole-dipole) transfer of energy from a chemiluminescent donor to a suitable acceptor molecule. In contrast to FRET, CRET occurs by the oxidation of a luminescent substrate without an excitation source. In our system, we chose the luminol/hydrogen peroxide CL reaction catalyzed by horseradish peroxidase (HRP) because this is one of the most sensitive CL reactions. In capillary electrophoresis with CL detection, the detection limit of HRP was below 10 À19 mol

In this article, we reported a new and sensitive method for characterizing rapid rotational and translational diffusion of gold nanoparticles (GNPs) and gold nanorods (GNRs) by resonance light scattering correlation spectroscopy (RLSCS). The RLSCS is a new single nanoparticle method, and its principle is based on measuring the resonance light scattering fluctuations in a highly focused volume due to Brownian motion of single particles, which resembles fluorescence correlation spectroscopy (FCS). On the basis of the theory of FCS, we first developed a model for rotational and translational diffusion and aspect ratio of nanoparticles in the RLSCS system. Then, we investigated the effects of certain factors such as the wavelength of illumination light and viscosity of solution using GNPs and GNRs as model samples and discovered that the polarization anisotropy and the scattering light intensity of GNPs and GNRs were significantly dependent on the wavelengths of illumination light. Using the 632.8 nm He-Ne laser as a light source, which was close to the resonance scattering band, we successfully obtained the translational and rotational diffusion coefficients and aspect ratios of anisotropic nanoparticles by the RLSCS method. The results obtained by this new method were in good agreement with transmission electron microscopy and theoretical calculation. Furthermore, the homogeneous sandwich immunoreaction was investigated using the antibody-modified GNPs as the probes. The changes in translational diffusion behaviors and aspect ratios of GNPs in immunoreaction were observed by the RLSCS method. By these changes, we can develop a new homogeneous immunoassay. Our preliminary results illustrated that the RLSCS method was a powerful tool for characterizing rapid rotational and translational diffusion behaviors of anisotropic nanoparticles in solution. We believe that the RLSCS method exhibits the wide applications in biological science especially in vivo study on the interaction of nanoparticles and biomolecules.

Fluorescence resonance energy transfer (FRET) is a powerful technique for probing very small changes in the distance between donor and acceptor fluorophores, and is ideal for the sensitive detection of molecular binding events and changes in protein conformation in response to interactions with a particular target molecule or to changes in the solution environment. In recent years, luminescent semiconductor nanocrystals (also called quantum dots, QDs) have been favorably adopted in the FRET-based studies, such as deriving QD-protein-conjugate configuration, [1] QD-protein sensing assemblies, [2,3] photochromic switching, [4] photodynamic medical therapy, [5] and probing DNA replication and telomerization.[6] This success is due to the unique sizedependent physical and chemical characteristics of QDs. QDs were applied to FRET as both donors [7][8][9][10] and acceptors, [11][12][13] and some of the advantages offered compared to conventional dyes include sharp and symmetrical emission spectra, high quantum yield (QY), good chemical and photo stability, and size-dependent emission-wavelength tunability. Recently, QDs as acceptors were successfully used in bioluminescence resonance energy transfer (BRET).[14]As with any fluorescence technique, however, photobleaching and autofluorescence limit the usefulness of FRET. Herein we report a resonance energy transfer between chemiluminescent donors and QDs as acceptors, which is called chemiluminescence (CL) resonance energy transfer (CRET), and is similar to BRET. [14][15][16][17][18][19] CRET involves nonradiative (dipole-dipole) transfer of energy from a chemiluminescent donor to a suitable acceptor molecule. In contrast to FRET, CRET occurs by the oxidation of a luminescent substrate without an excitation source. In our system, we chose the luminol/hydrogen peroxide CL reaction catalyzed by horseradish peroxidase (HRP) because this is one of the most sensitive CL reactions. In capillary electrophoresis with CL detection, the detection limit of HRP was below 10 À19 mol

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