Use of a geometry optimized fiber-optic surface-enhanced Raman scattering sensor in trace detection

A novel SERS (surface-enhanced Raman scattering) sensor has been recently developed; its peculiar geometry is able to increase considerably both the SERS active surface and the number of internal reflections at the interface between silica and silver, thus allowing an increase of the signal intensity. The aim of this work is to demonstrate that this sensor could be efficiently used to detect some molecules such as illegally used veterinary medicine (crystal violet and malachite green) below the ppb detection limit. The advantages of this sensor with respect to other detection techniques are not only the higher sensitivity, but also the fast response and the possibility of coupling with a portable Raman spectrometer for "on-field" measurements. The ability of the sensor to work under real environmental conditions in the presence of many cationic and anionic species has been tested both in solutions containing sodium and chlorine ions and in water coming from the aqueduct of Milan and from the (normally polluted) river Serio.     

Nanoparticle-functionalized porous polymer monolith detection elements for surface-enhanced Raman scattering

The use of porous polymer monoliths functionalized with silver nanoparticles is introduced in this work for high-sensitivity surface-enhanced Raman scattering (SERS) detection. Preparation of the SERS detection elements is a simple process comprising the synthesis of a discrete polymer monolith section within a silica capillary, followed by physically trapping silver nanoparticle aggregates within the monolith matrix. A SERS detection limit of 220 fmol for Rhodamine 6G is demonstrated, with excellent signal stability over a 24 h period. The capability of the SERS-active monolith for label-free detection of biomolecules was demonstrated by measurements of bradykinin and cytochrome c. The SERS-active monoliths can be readily integrated into miniaturized micrototal-analysis systems for online and label-free detection for a variety of biosensing, bioanalytical, and biomedical applications.     

Simplified protocol for detection of protein-ligand interactions via surface-enhanced resonance Raman scattering and surface-enhanced fluorescence

A simple and effective protocol for detections of protein-protein and protein-small molecule interactions has been developed. After interactions between proteins and their corresponding ligands, we employed colloidal silver staining for producing active substrates for surface-enhanced Raman scattering (SERS) and surface-enhanced fluorescence (SEF). Tetramethylrhodamine isothiocyanate (TRITC) and Atto610 were used for both Raman and fluorescent probes. We detected interactions between human IgG and TRITC-anti-human IgG, and those between avidin and Atto610-biotin by surface-enhanced resonance Raman scattering (SERRS) and SEF. The detection limits of the proposed SERRS-based method are comparable to those of the proposed SEF-based one, 0.9 pg/mL for anti-human IgG and 0.1 pg/mL for biotin. This protocol exploits several advantages of simplicity over other SERS and SEF-based related methods because of the protein staining-based strategy for silver nanoparticle assembling, high sensitivity from SERRS and SEF, and high stability in photostability comparing to fluorescence-based protein detections. Therefore, the proposed method for detection of protein-ligand interactions has great potential in high-sensitivity and high-throughput chip-based protein function determination.     

Single-molecule detection using surface-enhanced resonance Raman scattering and Langmuir-Blodgett monolayers

The Langmuir-Blodgett (LB) technique has been used to obtain spatially resolved surface-enhanced resonance Raman scattering (SERRS) spectra of single dye molecules dispersed in the matrix of a fatty acid. The experimental results presented here mimic the original electrochemical surface-enhanced Raman scattering (SERS) work where the background bulk water did not interfere with the detection of the SERS signal of molecules adsorbed onto the rough silver electrode. LB monolayers of the dye in fatty acid have been fabricated on silver island films with a concentration, in average, of one probe molecule per micrometer square. The properties of single-molecule spectroscopy were investigated using micro-Raman including mapping and global images. Blinking of the SERRS signal was also observed.     

Low-level detection of viral pathogens by a surface-enhanced Raman scattering based immunoassay

The need for rapid, highly sensitive, and versatile diagnostic tests for viral pathogens spans from human and veterinary medicine to bioterrorism prevention. As an approach to meet these demands, a diagnostic test employing monoclonal antibodies (mAbs) for the selective extraction of viral pathogens from a sample in a chip-scale, sandwich immunoassay format has been developed using surface-enhanced Raman scattering (SERS) as a readout method. The strengths of SERS-based detection include its inherent high sensitivity and facility for multiplexing. The capability of this approach is demonstrated by the capture of feline calicivirus (FCV) from cell culture media that is exposed to a gold substrate modified with a covalently immobilized layer of anti-FCV mAbs. The surface-bound FCVs are subsequently coupled with an extrinsic Raman label (ERL) for identification and quantification. The ERLs consist of 60-nm gold nanoparticles coated first with a layer of Raman reporter molecules and then a layer of mAbs. The Raman reporter molecule is strategically designed to chemisorb as a thiolate adlayer on the gold nanoparticle, to provide a strong and unique spectral signature, and to covalently link a layer of mAbs to the gold nanoparticle. The last feature provides a means to selectively tag substrate-bound FCV. This paper describes the development of the assay, which uses cell culture media as a sample matrix and has a linear dynamic range of 1 x 10(6)-2.5 x 10(8) viruses/mL and a limit of detection of 1 x 10(6) viruses/mL. These results reflect the findings from a detailed series of investigations on the effects of several experimental parameters (e.g., salt concentration, ERL binding buffer, and sample agitation), all of which were aimed at minimizing nonspecific binding and maximizing FCV binding efficiency. The performance of the assay is correlated with the number of captured FCV, determined by atomic force microscopy, as a means of method validation.     

Preparation of gold nano-cones as surface-enhanced Raman scattering sensors for molecule detection

Surface-enhanced Raman spectroscopy (SERS) is a powerful novel analytical tool which integrates high levels of sensitivity for trace analysis of chemical and biomolecular species due to the massive enhancement of Raman signals by using nanometre-sized metal particles. However, SERS can be envisaged as an analytical tool only if substrates with strong, predictable and reproducible SERS enhancement can be produced. Here we have developed one simple Ar+ ions sputtering technology to prepare gold nano-cones array on silicon substrates as surface-enhanced Raman scattering (SERS)-active substrates. The tip of the gold cone-structures exhibited an extremely sharp curvature with an apex diameter of 20 nm and the interior apex angle of the nanocones was around 20 degrees. These samples were evaluated as potential SERS substrates using Rhodamine 6G molecules as molecule probe and exhibited SERS enhancement factor of greater than 10, originated from the localized electron field enhancement around the apex of cones and the surface plasmon coupling of periodic structures.     

Preparation of silver nanoparticles by photo-reduction for surface-enhanced Raman scattering

A substrate for surface-enhanced Raman scattering (SERS) has been developed. Based on the surface-catalyzed reduction of Ag⁺ by citrate on the silver nanoparticles surface under light irradiation, small silver seeds on a quartz slide can be enlarged. The optical properties and characteristics of the silver films have been investigated by ultraviolet-visible spectroscopy, scan electron microscope and atomic force microscopy (AFM). The results indicate that the particle size and shape are different at different reduction time. At the first 3 h, some triangular and hexagonal nanoparticles formed; with the reduction proceeding, the shape of the silver particles became irregular and the size became larger. The silver films obtained are very suitable as SERS active substrate. The relationship between SERS intensity and the reduction time has been investigated for 1,4-bis[2-(4-pyridyl)ethenyl]-benzene molecule adsorbed on the silver film. The SERS intensity reached a maximum at 8 h reduction. The AFM measurements indicate that roughness features with an average size of 100 nm are present on the surface, which yielded the strongest SERS signal. Pyridine was used as a probe molecule to investigate the enhancement factor (EF) of the silver films. According to the formalism of Tian and co-workers, the EF of the silver films is estimated to be 3.4 × 10⁵. The silver film that can remain active for more than 50 days would seem to be suitable for various analytical applications.     

Comparison of surface-enhanced resonance Raman scattering and fluorescence for detection of a labeled antibody

A comparison is made of the quantitative detection of a labeled antibody by surface-enhanced resonance Raman scattering (SERRS) and by fluorescence using the same instrument with the same laser excitation source. The area under the curve for the fluorescence band is greater than for any single peak in the SERRS spectrum, but the broad fluorescence band is more difficult to discriminate from the background at low concentrations. Using the peak height of one SERRS band and the peak height at the fluorescence maximum, the detection limit for SERRS was lower (1.19 x 10-11 mol.dm-3) than that obtained using fluorescence (3.46 x 10-10 mol.dm-3). The SERRS detection limit is calculated for the concentration of the sample added, but compared to fluorescence, there is an additional dilution step due to the addition of the colloid and the extent of this dilution is dependent on assay format. For comparison with the detection limits found earlier with labeled oligonucleotides, SERRS was remeasured with a 10 s accumulation time, and the final concentration in the cuvette after colloid addition and before any adsorption to the silver was used to calculate a detection limit of 2.79 x 10-13 mol.dm-3. This is comparable to the detection limit found using a similar SERRS procedure for an oligonucleotide labeled with the same dye. This experiment is dependent on many parameters that could affect this result, including the nature of the SERRS substrate, the excitation wavelength, and the dye chosen. However, the result indicates that SERRS can give assay sensitivities comparable or better than fluorescence for quantitative direct assay determination, suggesting that the much greater potential for multiple analyte detection could be exploited.     

Highly reproducible solid-phase extraction membrane for removal and surface-enhanced Raman scattering detection of antibiotics

A disposable solid-phase extraction (SPE) membrane was developed for rapid removal and sensitive surface-enhanced Raman scattering (SERS) detection of antibiotics in water samples. The membrane was fabricated on commercially available SPE column by filtration of the activated carbon modified with silver nanoparticles (Ag NPs/AC). The prepared SPE membrane exhibited outstanding preconcentration ability due to the high adsorption properties of AC, and excellent ability to enhance Raman signal resulting from “hot spots” between the embedded Ag NPs, improving the sensitivity of SERS detection. A detection limit (LOD) of 5.0?×?10^?11 and 1.6?×?10^?10 M was achieved for rhodamine 6G and p-aminothiophenol. In addition, the membrane exhibited high reproducibility with spot-to-spot variation in SERS spectral intensity less than 15%. Based on the membrane, the qualitative and quantitative analysis of the antibiotics in aqueous solution was accomplished with the LOD at nM level, demonstrating the feasibility of the disposable SPE membrane for in situ rapid preconcentration and detection.     

Optimizing detection sensitivity on surface-enhanced Raman scattering of transition-metal electrodes with confocal Raman microscopy

Some points on how to improve the detection sensitivity of confocal Raman microscopy for the study of surface-enhanced Raman scattering (SERS) of transition-metal electrodes are discussed, including the careful design of the spectroelectrochemical cell, proper selection of the thickness of the solution layer, the binning of charge-coupled device (CCD) pixels, and appropriate setting of the notch filter. Various roughening methods for the Pt, Rh, Fe, Co, and Ni electrode surfaces have been introduced in order to obtain SERS-active surfaces. It has been shown that the appropriate roughening procedure and the optimizing performance of the confocal Raman microscope are the two most important factors to directly generate and observe SERS on net transition-metal electrodes.     

Analytical technique for label-free multi-protein detection based on Western blot and surface-enhanced Raman scattering

We have developed a new analytical procedure for label-free protein detection designated "Western SERS", consisting of protein electrophoresis, Western blot, colloidal silver staining, and surface-enhanced Raman scattering (SERS) detection. A novel method of silver staining for Western blot that uses a silver colloid, an excellent SERS-active substrate, is first proposed in the present study. During the process of silver staining, interactions between proteins and silver nanoparticles result in the emergence of SERS of proteins. In the present study, we use myoglobin (Mb) and bovine serum albumin (BSA) as model proteins. From different protein bands on a nitrocellulose (NC) membrane, we have observed surface-enhanced resonance Raman scattering (SERRS) spectra of Mb and SERS spectra of BSA. The proposed technique offers dual advantages of simplicity and high sensitivity. On one hand, after the colloidal silver staining, we can detect label-free multi-proteins directly on a NC membrane without digestion, extraction, and other pretreatments. On the other hand, the detection limit of the Western SERS is almost consistent with the detection limit of colloidal silver staining, and the SERRS detection limit of Mb is found to be 4 ng/band. This analytical method, which combines the technique of protein separation with SERS, may be a powerful protocol for label-free protein detection in proteomic research.     

Quantitative analysis of mitoxantrone by surface-enhanced resonance Raman scattering

Mitoxantrone is an anticancer agent for which it is important to know the concentration in blood during therapy. Current methods of analysis are cumbersome, requiring a pretreatment stage. A method based on surface-enhanced resonance Raman scattering (SERRS) has been developed using a flow cell and silver colloid as the SERRS substrate. It is simple, sensitive, fast, and reliable. Both blood plasma and serum can be analyzed directly, but fresh serum is preferred here due to reduced fluorescence in the clinical samples available. Fluorescence is reduced further by the dilution of the serum in the flow cell and by quenching by the silver of surface-adsorbed material. The effectiveness of the latter process is dependent on the contact time between the serum and the silver. The linear range encompasses the range of concentrations detected previously in patient samples using HPLC methods. In a comparative study of a series of samples taken from a patient at different times, there is good agreement between the results obtained by HPLC and SERRS with no significant difference between them at the 95% limit. The limit of detection in serum using the final optimized procedure for SERRS was 4.0 x 10(-11) M (0.02 ng/mL) mitoxantrone. The ease with which the SERRS analysis can be carried out makes it the preferred choice of technique for mitoxantrone analysis.     

Label-free probing of G-quadruplex formation by surface-enhanced Raman scattering

In this work, we establish the use of surface-enhanced Raman scattering (SERS) as a label-free analytical technique for the direct detection of G-quadruplex formation. In particular, we demonstrate that SERS analysis allows the evaluation of the relative stability of G quadruplexes that differ for the number of G tetrads and investigate several structural features of quadruplexes, such as the orientation of glycosidic bonds, the identification of distortions in the sugar-phosphate backbone, and the degree of hydrogen-bond solvation. Herein, the fluctuation of the SERS spectra, due to the specific interaction of vibrational modes with the SERS-active substrate, is quantitatively analyzed before and after quadruplex formation. The results of this study suggest a perpendicular orientation of the quadruplexes (with or without the 3'-tetra end linker) with respect to the silver colloidal surface, which opens new perspectives for the use of SERS as a label-free analytical tool for the study of the binding mode between quadruplexes and their ligands.     

Coupling reaction-based ultrasensitive detection of phenolic estrogens using surface-enhanced resonance Raman scattering

Studies have shown that many adverse health effects are associated with human exposure to dietary or environmental estrogens. Therefore, the development of rapid and highly sensitive detection methods for estrogens is very important and necessary to maintain hormonal concentration below the safety limit. Herein, we demonstrate a simple and rapid approach to detect trace amounts of phenolic estrogen based on surface-enhanced resonance Raman scattering (SERRS). Because of a coupling reaction between diazonium ions and the phenolic estrogens, azo compounds are formed with strong SERRS activity, which allows phenolic estrogen recognition at subnanomolar levels in solution. The proposed protocol has multiplexing capability, because each SERRS fingerprint of the azo dyes specifically corresponds to the related estrogen. Moreover, it is universal and highly selective, not only for phenolic estrogens but also for other phenolic molecules, even in complex systems.     

Silver nanorod arrays as a surface-enhanced Raman scattering substrate for foodborne pathogenic bacteria detection

Surface-enhanced Raman scattering (SERS) using novel silver nanorod array substrates has been used for the detection of pathogenic bacteria. The substrate consists of a base layer of 500 nm silver film on a glass slide and a layer of silver nanorod array with a length of approximately 1 microm produced by the oblique angle deposition method at a vapor incident angle of 86 degrees . Spectra from whole cell bacteria, Generic Escherichia coli, E. coli O157:H7, E. coli DH 5alpha, Staphylococcus aureus, S. epidermidis, and Salmonella typhimurium, and bacteria mixtures have been obtained. This SERS active substrate can detect spectral differences between Gram types, different species, their mixture, and strains. Principal component analysis (PCA) has been applied to classify the spectra. Viable and nonviable cells have also been examined, and significantly reduced SERS responses were observed for nonviable cells. SERS detection of bacteria at the single cell level, excited at low incident laser power (12 micro W) and short collection time (10 s), has also been demonstrated. These results indicate that the SERS-active silver nanorod array substrate is a potential analytical sensor for rapid identification of microorganisms with a minimum of sample preparation.     

Silver-doped sol-gel film as a surface-enhanced Raman scattering substrate for detection of uranyl and neptunyl ions

A surface-enhanced Raman scattering (SERS) substrate containing silver particles was prepared by an acid-catalyzed sol-gel method. Silver nitrate was first doped into the sol-gel film followed by chemical reduction of the silver ions with sodium borohydride to produce silver particles. This silver-doped sol-gel substrate exhibits strong enhancement of Raman scattering from adsorbed uranyl ions with a detection limit of 8.5 x 10(-8) M, which is comparable to existing methods of uranyl detection such as spectrophotometry, fluorometry, and a SERS method based on ligand-modified solution silver colloids. However, in the present method, no preconcentration steps, chromogens, or complexing ligands are needed. Compared with the SERS method using Ag colloidal sols, the silver-doped sol-gel film has the advantage that the silver particles trapped in the sol-gel matrix are much more stable than Ag colloids in liquid media. Furthermore, porous silica sol-gel materials are known to have affinities toward many inorganic and organic molecules. The enhanced adsorption affinities could also lead to the increased SERS sensitivity. The performance of the new silver-doped sol-gel substrate was evaluated with uranyl ions and compared to that of a SERS substrate based on silver-coated silica beads prepared by vacuum deposition. The detection limit for the silver-doped sol-gel film was 104 times lower than that for the silver-coated silica beads. The sol-gel substrate was further used to obtain, for the first time, the surface-enhanced Raman spectrum of neptunyl ions in dilute aqueous solutions.     

Structural engineering of transition-metal nitrides for surface-enhanced Raman scattering chips

Noble-metal-free surface-enhanced Raman scattering(SERS)substrates have attracted great attention for their abundant sources,good signal uniformity,superior bio...     

Evaluation of surface-enhanced resonance Raman scattering for quantitative DNA analysis

The labeling of biological species using dyes has become common practice to aid in their detection, and immediate positive identification of specific dyes in high dilution is a key requirement. Here the detection by surface-enhanced resonance Raman scattering (SERRS) of eight commercially available dye labels (ROX, rhodamine 6G, HEX, FAM, TET, Cy3, Cy5, TAMRA) attached to oligonucleotide strands is reported. Each of the eight labels was easily detected by using the SERRS from silver nanoparticles to produce a unique, molecularly specific spectrum. The conditions were optimized to obtain the best signal enhancement, and linear concentration graphs at low oligonucleotide concentrations were obtained. At higher concentrations (above approximately 10(-)(8) mol dm(-)(3)), curvature was introduced into the concentration graphs with the exception of rhodamine 6G, TET, and FAM, which gave linearity over the entire concentration range studied. Detection limits as low as 0.5 fmol were obtained, with lower possible if a smaller sample was analyzed. Investigation was also carried out into the effect of a Tris-HCl buffer containing the surfactant Tween 20 to aid in the prevention of surface adhesion of the oligonucleotides to the sample vessels at ultralow concentrations. The Tween 20 allowed lower detection limits to be obtained for each of the labels studied. This study shows that the different dyes commonly used with oligonucleotides can give quantitative SERRS at concentration levels not possible when the same dyes are used with fluorescence detection.