Chunyuan Song

Highly sensitive immunoassay based on Raman reporter-labeled immuno-Au aggregates and SERS-active immune substrate

A highly sensitive immunoassay based on surface enhanced Raman scattering (SERS) has been developed with a novel immune marker named as Raman reporter-labeled immuno-Au aggregates on a SERS-active immune substrate. The features of those immune aggregates were characterized by UV-vis extinction spectra, TEM images, SEM pictures and SERS spectra. It is found that stable gold aggregates in appropriate morphologies can be induced by mixing proper amounts of reporter molecules with gold nanoparticles. Based on those reporter-labeled Au aggregates, the immune aggregates with high stability can be prepared successfully by immobilizing antibody to the surface of the aggregates. Using those easily prepared aggregates showing strong SERS activity and high bio-specificity, a highly effective SERS-based immunoassay has been performed. Moreover, a SERS-active immune substrate has been introduced to replace a typical immune substrate without any SERS activity. Our experimental results indicate that the SERS-active immune substrate makes a certain contribution to the highly sensitive immunoassay. As a result, with this proposed immunoassay structure, the concentration detection of human-IgG was performed and a calibration curve was obtained in the range from 100ngmL(-1) to 100fgmL(-1). This work opens a new avenue for sensitive immunoassay and other biochemical analysis based on SERS.

Distinguishing breast cancer cells using surface-enhanced Raman scattering

AbstractThe detection and identification of epidermal growth factor receptor 2 (HER2)-positive breast cancer cells is crucial for the clinic therapy of breast cancer. For the aim of the detection, a novel surface-enhanced Raman scattering (SERS) probe for distinguishing breast cancers at different HER2 statuses is reported in this paper. In such a probe, anti-HER2 antibody-conjugated silver nanoparticles have been synthesized for specific targeting of HER2-positive breast cancer cells. More importantly, different from the previously reported SERS probe for targeting cancer cells, p-mercaptobenzoic acid is utilized as both the Raman reporter and the conjugation agent for attaching antibody molecules, which leads to a much simplified structure. For investigating the ability of such a probe to distinguish breast cancer cells, SKBR3 and MCF7 cells were chosen as two model systems, which are HER2-positive- and HER2-negative-expressing cells, respectively. The experimental results reveal that SKBR3 cells exhibit much stronger SERS signals than MCF7 cells, indicating that the probe could be utilized to distinguish breast cancer cells at different HER2 statuses. This kind of SERS probe holds a potential for a direct detection of living breast cancer cells with the advantages of easy fabrication, high SERS sensitivity, and biocompatibility. FigureSERS spectra of the probe in SKBR3 cells and in MCF7 cells

One-step functionalized gold nanorods as intracellular probe with improved SERS performance and reduced cytotoxicity

An intracellular SERS probe based on gold nanorods (GNRs) is successfully presented with improved SERS performance and greatly reduced cytotoxicity. Unlike the previous layer-by-layer methods to reduce the obvious cytotoxicity of GNRs, the CTAB-stabilized GNRs only need to be treated with hydrochloric acid (HCl) without any other steps. The aqueous solution of the resultant nanorods exhibited an aggregation with limited extend but remained stable for more than 6 months with no sediment being found. Interestingly, the aggregation helps to yield an improved surface enhanced Raman scattering (SERS) performance in comparison with those stabilized by CTAB bilayers. More importantly, the resultant gold nanorods achieve high cell viability as more than 95%, a level comparable to that obtained by traditional layer-by-layer coating method. The greatly increased cell viability suggests that the cytotoxicity of CTAB-stabilized GNRs is mainly caused by free or loosely surrounded CTAB rather than those tightly bound on the surfaces of GNRs. Moreover, when such gold nanorods were incorporated into living cells, they exhibited strong SERS signal as well as very low cytotoxicity. The result suggests that HCl-treated gold nanorods are potentially very feasible substrates for use as highly sensitive SERS probes in biosensing and biomedical diagnostics, for which the fabrication protocol is advantageous in its simplicity and reproducibility.

Tracking multiplex drugs and their dynamics in living cells using the label-free surface-enhanced Raman scattering technique.

Label free and real time detection of nonfluorescent drugs inside living cells has been realized by using surface-enhanced Raman scattering (SERS). For the first time, the characteristics of 6-mercapotopurine and methimazole, two different drugs, were monitored simultaneously by SERS in living cells. Particularly, the processes of diffusion and metabolism of drugs occurring in the intracellular matrix were investigated. The results indicate that the metabolism speed of 6-mercapotopurine in living HeLa cells is much faster than that of methimazole. Moreover, the detection sensitivity of intracellular drugs has also been checked and a low detection limit of 1 nM was obtained of drug 6-mercapotopurine in a single HeLa cell.

A straightforward route to the synthesis of a surface-enhanced Raman scattering probe for targeting transferrin receptor-overexpressed cells.

A tumor cell targeting surface-enhanced Raman scattering (SERS) probe has been successfully synthesized by using p-mercaptobenzoic acid (pMBA) as both the SERS reporter and the conjugation agent for attaching transferrin molecules, which shows experimentally the targeting ability for transferrin receptor-overexpressed HeLa cells and exhibits strong SERS signals when being incubated inside cells. To prove that the uptake of such a SERS probe is through a Tf-receptor-mediated endocytosis process, two control experiments: (1) HeLa cells being incubated with the probe at 4 degrees C and (2) HeLa cells being pre-blocked with free transferrin at 37 degrees C, were employed. The difference of SERS intensity between the transferrin-overexpressed HeLa cells and transferrin-pre-blocked HeLa cells indicates that the probe has the potential to selectively target tumor cells.

Surface‐Enhanced Fluorescence from Fluorophore‐Assembled Monolayers by Using Ag@SiO2 Nanoparticles

A new and simple procedure to enhance the fluorescence of analytes on the surfaces of a solid substrate is demonstrated based on Ag@SiO(2) nanoparticles. Two kinds of silver-silica core-shell nanoparticles with shell thicknesses of around 3 and 15 nm have been prepared and used as enhancing agents, respectively. By simply pipetting drops of the enhancing agents onto substrate surfaces with Rose Bengal monolayers, an enhancement of about 27 times, compared with the control sample, is achieved by using the Ag@SiO(2) nanoparticles with shells of about 3 nm, whereas an enhancement of around 11.7 times is obtained when using those with thicker shells. The effects of shell thickness and surface density of the enhancing agents on the enhancement have been investigated experimentally. The results show that this method can be potentially helpful in fluorescence-based surface analysis.

Immunoassays Based on Surface-Enhanced Fluorescence using Gap-Plasmon-Tunable Ag Bilayer Nanoparticle Films

A novel gap-plasmon-tunable Ag bilayer nanoparticle film for immunoassays is demonstrated. Different from a traditional Ag monolayer nanoparticle film, a desired number of polyelectrolyte (PEL) layers are deposited on the nanoparticles before the self-assembly of a second Ag nanoparticle layer. Interestingly, by controlling the number of the PEL interlayers, the gap plasmon between the two Ag nanoparticle layers can be tuned across the visible spectral range. The ability of the presented Ag bilayer nanoparticle films in fluorescence enhancement has been examined experimentally. A maximal enhancement of around 15.4 fold was achieved when 7 layers of polyelectrolyte were used. When this optimal Ag bilayer nanoparticle film was applied to fluorescence immunoassay, a performance with approximately 3.3-fold enhancement was obtained compared with that performed on a traditional glass substrate. The experimental results suggest that the presented gap-plasmon tunable Ag bilayer nanoparticle films have great potential in fluorescence-based immunoassays. The method of the bilayer-film construction presented here also provides new insights into the rational design of the plasmonic substrates.

Preparation of 2-mercaptobenzothiazole-labeled immuno-Au aggregates for SERS-based immunoassay

A detailed method for preparation of 2-mercaptobenzothiazole (MBT)-labeled immuno-Au aggregates as a novel Raman probe for SERS-based immunoassay has been proposed in this paper. The formation kinetics of gold aggregates induced by MBT and the impact of the amount of labeled reporter on the aggregation, as well as the fabrication of MBT-labeled immuno-Au aggregates were characterized comprehensively by UV-vis spectrophotometer. Meanwhile, a chain-like morphology of aggregates was monitored by TEM images. Experimental results show that this Raman tag can act as an efficient label and an enhanced Raman signal of SERS-based immunoassay can be obtained successfully with this novel probe. This work shows the method for the preparation of immune probe for SERS-based immunoassay using MBT as reporter and provides a reference for the preparation of immune probe with other Raman markers for multi-channel SERS-based immunoassay.

Interaction between antitumor drug and silver nanoparticles: combined fluorscence and surface enhanced Raman scattering study

Optical methods and MTT method are used to characterize the antiproliferation effect of antitumor drug 9-aminoacridine (9AA) with and without silver nanoparticles. Intracellular surface enhanced Raman scattering (SERS) spectra and fluorescent spectra of 9AA indicate the form of 9AA adsorbed on the surface of silver nanoparticles. Although both silver nanoparticles and antitumor drug can inhibit the growth of Hela cells, silver nanoparticles can slow down the antiproliferation effect on Hela cells at low concentration of antitumor drugs. Our experimental results suggest that silver nanoparticles may serve as slow-release drug carriers, which is important in antitumor drug delivery.