Quantitative Surface-enhanced Raman Spectroscopy Research Articles

Absolute Quantification of Serum Exosomes in Patients with an SERS-Lateral Flow Strip Biosensor for Noninvasive Clinical Cancer Diagnosis.

Exosomes (exos) widely existing in body fluids show great potential for noninvasive cancer diagnosis. Quantitative analysis of exos is traditionally performed by targeting specific exosomal surface proteins, but it is often imprecise due to the common expression of exosomal proteins and subtle expression differences between different cancer subtypes. Herein, we report quantitative surface-enhanced Raman spectroscopy (SERS) of serum exos through a combination of a paper-based lateral flow strip (LFS) biosensor with multivariate spectral unmixing analysis rather than simply quantifying exosomal proteins. Our SERS-LFS biosensor enables absolute quantification of two different serum exos with a limit of detection down to ∼106 particles/mL for both exos. We further exemplify the application of this strategy in quantitative dual-plex detection of serum exos from breast cancer patients. We find that human epidermal growth factor receptor 2+ (HER2+) and luminal A breast cancer patients undergoing no surgery are enriched in serum exos derived from SKBR-3 cells and MCF-7 cells (denoted as SKBR and MCF exos), respectively. The surgical treatment of these breast cancer patients accompanies an obvious decrease of either SKBR or MCF exos in the serum. These results suggest the great potential of the combination of the SERS-LFS biosensor and multivariate spectral unmixing for breast cancer subtyping and therapeutic surveillance with the powerful quantitative capability of exos in clinical samples.

Engineered Au@CuO Nanoparticles for Wide-Range Quantitation of Sulfur Ions by Surface-Enhanced Raman Spectroscopy.

Efficient detection of sulfide ions (S2-), especially in a wide quantitative range, is of significance but faces challenges. This work strategizes and fabricates Au@CuO nanoparticles for quantitative surface-enhanced Raman spectroscopy (SERS) detection of the S2- ions based on the S2- concentration-dependent ion-solid interactions. We have achieved fast and quantitative S2- detection in a wide range from 5 ppb to 64,000 ppm (saturation concentration of the S2- source). We also demonstrated that the optimal CuO shell thickness for the detection is about 7 nm and that the detection can be further improved by prolonging the soaking duration. Moreover, this detection method has also shown the merits of reusable substrates (especially for low S2- concentrations) and good anti-interference ability to many common anions (Cl-, NO3-, OH-, HCOO-, CO32-, and SO42-). Finally, the high feasibility of this detection in actual water (tap water and pond water) has also been demonstrated. This work provides efficient S2- detection with great potential in practical use and also inspires the design of quantifiable SERS substrates for detecting more small inorganic molecules and ions.

Quantitative Surface-Enhanced Raman Spectroscopy for Field Detections Based on Structurally Homogeneous Silver-Coated Silicon Nanocone Arrays.

Practical application of surface-enhanced Raman spectroscopy (SERS) is greatly limited by the inaccurate quantitative analyses due to the measuring parameter’s fluctuations induced by different operators, different Raman spectrometers, and different test sites and moments, especially during the field tests. Herein, we develop a strategy of quantitative SERS for field detection via designing structurally homogeneous and ordered Ag-coated Si nanocone arrays. Such an array is fabricated as SERS chips by depositing Ag on the template etching-induced Si nanocone array. Taking 4-aminothiophenol as the typical analyte, the influences of fluctuations in measuring parameters (such as defocusing depth and laser powers) on Raman signals are systematically studied, which significantly change SERS measurements. It has been shown that the silicon underneath the Ag coating in the chip can respond to the measuring parameters’ fluctuations synchronously with and similar to the analyte adsorbed on the chip surface, and the normalization with Si Raman signals can well eliminate the big fluctuations (up to 1 or 2 orders of magnitude) in measurements, achieving highly reproducible measurements (mostly, <5% in signal fluctuations) and accurate quantitative SERS analyses. Finally, the simulated field tests demonstrate that the developed strategy enables quantitatively analyzing the highly scattered SERS measurements well with 1 order of magnitude in signal fluctuation, exhibiting good practicability. This study provides a new practical chip and reliable quantitative SERS for the field detection of real samples.

Plasmonic Electronic Raman Scattering as Internal Standard for Spatial and Temporal Calibration in Quantitative Surface-Enhanced Raman Spectroscopy.

Ultrasensitive surface-enhanced Raman spectroscopy (SERS) still faces difficulties in quantitative analysis because of its susceptibility to local optical field variations at plasmonic hotspots in metallo-dielectric nanostructures. Current SERS calibration approaches using Raman tags have inherent limitations due to spatial occupation competition with analyte molecules, spectral interference with analyte Raman peaks, and photodegradation. Herein, we report that plasmon-enhanced electronic Raman scattering (ERS) signals from metal can serve as an internal standard for spatial and temporal calibration of molecular Raman scattering (MRS) signals from analyte molecules at the same hotspots, enabling rigorous quantitative SERS analysis. We observe a linear dependence between ERS and MRS signal intensities upon spatial and temporal variations of excitation optical fields, manifesting the |E|4 enhancements for both ERS and MRS processes at the same hotspots in agreement with our theoretical prediction. Furthermore, we find that the ERS calibration's performance limit can result from orientation variations of analyte molecules at hotspots.

Roughened silver microtubes for reproducible and quantitative SERS using a template-assisted electrosynthesis approach

The synthesis and application of silver microtube substrates for quantification by surface enhanced Raman spectroscopy (SERS) is presented in this work. The roughened silver microtubes were prepared by template assisted electrosynthesis method. Using rhodamine 6G as a Raman probe molecule, the silver microtubes were tested for SERS determination, demonstrating a high performance for quantitative analysis with high precision and low data dispersion (%RSD < 3), which is the limiting step in quantitative SERS. This performance can be explained by obtaining a single silver microtube substrate highly homogeneous in SERS activity in the whole surface and with high reproducibility between different synthesis batches.

Quantitative Surface-Enhanced Raman Spectroscopy Analysis through 3D Superlattice Arrays of Au Nanoframes with Attomolar Detection.

This paper reports a methodology for synthesizing and ordering gold nanoframes into three-dimensional (3D) arrays with a controlled thickness, leading to homogeneous plasmonic superstructures, with which quantitative analysis via surface-enhanced Raman spectroscopy (SERS) has been successfully demonstrated. Because this preparation method allows for systematic control of nanoframe film thickness and the resulting 3D plasmonic superstructure, which exhibits a unique nanoporous network of hot-spots, detection limits down to 10-18 M, corresponding to ≈6000 molecules, have been measured. Compared to analogous solid nanoparticle superstructures, the nanoframe superstructures with their unique nanoporous architecture effectively dissipate the heat inevitably generated by laser excitation during measurement, effectively suppressing the formation of carbonaceous materials and therefore their accompanying fluorescence interference, especially important for low concentration detection.

Quantitative surface-enhanced Raman spectroscopy chemical analysis using citrate as an in situ calibrant.

Direct detection, or inferring the presence of illicit substances, is of great forensic and toxicological value. Surface-enhanced Raman spectroscopy (SERS) has been shown capable of detecting such molecules in a quick and sensitive manner. Herein we describe an analysis strategy for quantitation of low concentrations of three analytes (methamphetamine, cocaine, and papaverine) by SERS analysis using the citrate capping agent that initially saturates the silver nanoparticles' surface as an in situ standard. The citrate is subsequently displaced by the analyte to an extent dependent on the analyte's concentration in the analyte solution. A general model for the competitive adsorption of citrate and a target analyte was developed and used to determine the relative concentrations of the two species coexisting on the surface of the silver nanoparticles. To apply this model, classical least squares (CLS) was used to extract the relative SERS contribution of each of the two species in a given SERS spectrum, thereby accurately determining the analyte concentration in the sample solution. This approach, in essence, transforms citrate into a local standard against which the concentration of an analyte can be reliably determined.

Quantitative Surface-Enhanced Raman Spectroscopy through the Interface-Assisted Self-Assembly of Three-Dimensional Silver Nanorod Substrates.

The realization of surface-enhanced Raman spectroscopy (SERS) to be a reliable quantitative analytical technique requires sensitive and reproducible enhancing substrates. Here, uniform three-dimensional (3D) Ag nanorod (AgNR) substrates with well-defined interlayer spacings are prepared through the air-liquid interface-assisted self-assembly of AgNR in a layer-by-layer manner. The correlation of the SERS performance with the 3D AgNR structures is performed by SERS mapping the substrates. SERS mapping reveals the excellent enhancement uniformity of the 3D substrates with the relative standard deviation (RSD) less than 10%. It finds that both of the number of layers (NL) and the length of the AgNR have effects on the SERS performance of the 3D AgNR substrates. It is demonstrated that the intergaps between layers contribute much to the SERS intensity of the 3D AgNR by creating the interlayer (out-of-plane) plasmonic coupling. The impact of the excitation wavelengths (532, 633, and 785 nm) on SERS performance is also determined. The optimal 3D AgNR structures achieved by the correlation study is further used to detect a set of related molecules (l-tryptophan (Trp), l-phenylalanine (Phe), urea, and melamine). The 3D AgNR SERS of the analytes exhibits linear responses over wide concentration ranges. The sensitivity of the 3D AgNR SERS is proved by comparing to that of the current methods. Moreover, the 3D AgNR substrates maintain the performance stability during 4 weeks of storage.

Quantitative SERS analysis based on multiple-internal-standard embedded core-shell nanoparticles and spectral shape deformation quantitative theory

Due to the practical impossibility in controlling the number and distribution of "hot spots" on or close to the surfaces of enhancing substrates, internal standards are generally used to improve the accuracy and precision of quantitative surface-enhanced Raman spectroscopy (SERS). Traditional internal standard methods based on either univariate ratiometric models or conventional multivariate calibration models can use the information of only one characteristic SERS peak of only one internal standard, which weakens the efficiency of traditional internal standard methods in improving the quantitative results of SERS assays. In this contribution, the concept of using multiple internal standards was introduced for the first time to quantitative SERS assays by incorporating a unique spectral shape deformation (SSD) quantitative theory with multiple-internal-standard embedded core-shell nanoparticles. The quantification of phosmet residues on apple skins by SERS technique with Au-core/Ag-shell nanoparticles embedded with both 2-MB and PATP as enhancing substrate was used to illustrate the applicability of the proposed strategy. Experimental results demonstrated that the proposed multiple-internal-standard strategy based on SSD achieved much more accurate and precise concentration predictions for phosmet residues on apple skin than traditional single-internal-standard methods based on either univariate ratiometric calibration models or conventional multivariate calibration models. The SERS quantitative results of the multiple-internal-standard strategy were comparable to those obtained by LC-MS/MS. The average relative deviation between the quantitative results of the proposed method and those of ICP-MS was about 8%. The good accuracy of the proposed method makes it a promising alternative for quantitative SERS assays.

Characterization method for relative Raman enhancement for surface-enhanced Raman spectroscopy using gold nanoparticle dimer array

In this paper, a characterization method for Raman enhancement for highly sensitive and quantitative surface-enhanced Raman spectroscopy (SERS) is reported. A particle dimer shows a marked electromagnetic enhancement when the particle connection direction is matched to the polarization direction of incident light. In this study, dimers were arrayed by nanotrench-guided self-assembly for a marked total Raman enhancement. By measuring acetonedicarboxylic acid, the fabricated structures were characterized for SERS depending on the polarization angle against the particle connection direction. This indicates that the fabricated structures cause an effective SERS enhancement, which is dominated by the electromagnetic enhancement. Then, we measured 4,4′-bipyridine, which is a pesticide material, for quantitative analysis. In advance, we evaluated the enhancement of the particle structure by the Raman measurement of acetonedicarboxylic acid. Finally, we compared the Raman intensities of acetonedicarboxylic acid and 4,4′-bipyridine. Their intensities showed good correlation. The advantage of this method for previously evaluating the enhancement of the substrate was demonstrated. This developed SERS characterization method is expected to be applied to various quantitative trace analyses of molecules with high sensitivity.

In Situ Quantitative Graphene-Based Surface-Enhanced Raman Spectroscopy

Quantitative surface-enhanced Raman spectroscopy (SERS) with ultrahigh sensitivity will significantly promote its practical application in many fields, such as environment monitoring, food safety, and drug detection. However, the challenges that remain unresolved, particularly in the low concentration levels, arise from the instability of the SERS spectra and the uncertainty of the number of detected molecules. Herein, a graphene-based, flexible, and transparent substrate for SERS quantification is reported, wherein the 2D single-crystalline nature of graphene promises the homogeneous adsorption of molecules, facilitating the determination of the number of molecules, the separation of molecules from metal, which ensures the stability of the Raman signals, and an internal standard for the calibration of SERS intensities. The in situ quantification of probe molecules is demonstrated in an aqueous solution down to the detection limit of 10−8m, and the real-time, in situ monitoring of the release process of rhodamine B molecules, which mimics practical application, for example, the controlled release of medicine, is shown. The results open up an avenue for reliable SERS quantification for practical applications with high efficiency and low cost.

Self-assembled activated carbon nanoparticles for reliable time-discretized quantitative surface-enhanced Raman spectroscopy

Quantitative SERS is achieved by the saturated adsorption time of extraction film. Perturbations caused by performance change of substrates are overcome.

Quantitative surface-enhanced Raman spectroscopy of single bases in oligodeoxynucleotides.

To address the question of whether the SERS signals of ss-DNA are simply combinations of the signals from the individual bases that comprise the sequence, SERS spectra of unmodified ss-DNA sequences were obtained using a hydroxylamine-reduced Ag colloid aggregated with MgSO4. Initially, synthetic oligodeoxynucleotides with systematic structural variations were used to investigate the effect of adding single nucleobases to the 3' terminus of 10-mer and 20-mer sequences. It was found that the resulting SERS difference spectra could be used to identify the added nucleobases since they closely matched reference spectra of the same nucleobase. Investigation of the variation in intensity of an adenine probe which was moved along a test sequence showed there was a small end effect where nucleobases near the 3' terminus gave slightly larger signals but the effect was minor (30%). More significantly, in a sample set comprising 25-mer sequences where A, T or G nucleobases were substituted either near the centres of the sequences or the 5' or 3' ends, the SERS difference spectra only matched the expected form in approximately half the cases tested. This variation appeared to be due to changes in secondary structure induced by altering the sequences since uncoiling the sequences in a thermal pre-treatment step gave difference spectra which in all cases matched the expected form. Multivariate analysis of the set of substitution data showed that 99% of the variance could be accounted for in a model with just three factors whose loadings matched the spectra of the A, T, and G nucleobases and which contained no positional information. This suggests that aside from the differences in secondary structure which can be eliminated by thermal pre-treatment, the SERS spectra of the 25-mers studied here are simply the sum of their component parts. Although this means that SERS provides very little information on the primary sequence it should be excellent for the detection of post-transcription modifications to DNA which can occur at multiple positions along a given sequence.

Quantitative analysis of surface enhanced Raman spectroscopy of Rhodamine 6G using a composite graphene and plasmonic Au nanoparticle substrate

A novel type of substrate for quantitative surface enhanced Raman spectroscopy (SERS) composed of chemical vapor deposition (CVD) graphene and in-situ fabricated rounded gold nanoparticles (AuNPs) was designed. SERS was measured on samples of different concentrations of Rhodamine 6G (R6G) on the AuNPs/graphene substrates using a low power 632.8 nm laser. Finite element simulations were carried out for a system of two gold hemiellipsoids under various conditions such as with R6G analyte covering the surface and with graphene underneath the nanoparticles. Graphene or R6G being present between the two nanoparticles caused a redshift in the plasmonic resonance frequency, and the graphene dampened the electric field of the surface. Regardless of the weakened electric field, the synergy of the AuNPs and graphene still enhanced the Raman signature of R6G to a greater extent than the nanoparticles or graphene alone could, which is attributed to the charge transferring mechanism effect of graphene on SERS. The lowest concentration of solid phase R6G deposited in this manner that could be detected was 8 × 10−7 M. Higher analyte concentrations and the characteristic peak intensities of the analyte showed a logarithmic relation as anticipated from the plasmonically enhanced Raman scattering.

Fabrication of gold nanoparticle-coated paper and its use as a sensitive substrate for quantitative SERS analysis

This paper describes a rapid method for fabrication of a paper substrate coated with gold nanoparticles (GNPs) that results in the formation of a large number of hot spots on the surface and allows an adequate control of the active area. The resulting substrate is shown to be a viable material for use in quantitative surface-enhanced Raman spectroscopy (SERS) analysis. The influence of the amount of GNPs on the SERS signals (using crystal violet as a sample analyte) was correlated with field-emission scanning electron microscopy, UV-visible and theoretical studies. The use of this substrates results in larger enhancement of Raman signals and in comparably repeatability when compared to commercially available substrates. The substrate was applied to SERS-based determination of nicotine and uric acid in aqueous solution, and the respective limits of detection are 20 and 30 μg L−1. The results indicate that the SERS substrates may be applied to the quantification of a wide variety of molecules.

Quantitative surface-enhanced Raman spectroscopy for kinetic analysis of aldol condensation using Ag-Au core-shell nanocubes.

Surface-enhanced Raman spectroscopy (SERS) is a powerful tool with high potential for multiplexed detection of dilute analytes. However, quantitative SERS of kinetic assays can be difficult due to the variation in enhancement factors caused by changing reaction conditions. We report a method for quantitative SERS kinetic analysis using colloidal Ag-Au core-shell nanocubes (Ag@AuNCs) as the SERS substrate. This substrate is mass producible, possesses large SERS enhancement, and is resistant to degradation in most environments. The SERS enhancement of the Ag@AuNCs was evaluated both experimentally and computationally. Quantitation was achieved by covalently attaching a non-reactive internal standard (IS) to substrate surfaces and normalizing SERS spectra to the IS signal. We demonstrated that IS normalization corrects for temporal variations in enhancement factor and particle concentration. Quantitation was demonstrated by monitoring the base-catalyzed aldol condensation of surface-bound 4-(methylthio)benzaldehyde with free acetone. The kinetic model of this reaction was fitted to IS normalized SERS data, resulting in kinetic parameters that agreed well with published values. This SERS platform is a robust and sensitive method for quantitative analysis of kinetic assays, with potential applications in many fields.

Optimization of Ag-Coated Polystyrene Nanosphere Substrates for Quantitative Surface-Enhanced Raman Spectroscopy Analysis

The optical properties of Ag-coated polystyrene nanospheres can be varied by changing the vapor deposition angle θ. When θ 55°, broad band absorbance is achieved. The difference in optical absorbance is due to the coverage and shape change of the Ag coating on nanospheres with multiple oriented domains, as demonstrated by finite-difference time-domain simulations. The localized surface plasmon resonance of substrates coated at small θ can be tuned closely to the excitation wavelength, and these samples can serve as good surface-enhanced Raman scattering (SERS) substrates. The polystyrene vibrational peak at Δν = 1004 cm–1, which is inherent to the substrate, can be used as an internal standard. By normalizing the SERS peak of an analyte by this peak intensity, the SERS intensity ratio is independent of the substrate fabrication conditions. Such an internal standard is also shown to be effective for revealing the adsorption kinetics for rhodamine ...

Large-Scale Hot Spot Engineering for Quantitative SERS at the Single-Molecule Scale.

Quantitative surface enhanced Raman spectroscopy (SERS) requires precise control of Raman enhancement factor and detection uniformity across the SERS substrate. Here, we show that alkanethiolate ligand-regulated silver (Ag) nanoparticle films can be used to achieve quantitative SERS measurements down to the single-molecule level. The two-dimensional hexagonal close-packed superlattices of Ag nanoparticles formed in these films allow for SERS detection over a large area with excellent uniformity and high Raman enhancement factor. In particular, the SERS signal from the thiolate ligands on Ag nanoparticle surfaces can be utilized as a stable internal calibration standard for reproducible quantitative measurements. We demonstrate the capability of quantitative SERS by measuring the areal densities of crystal violet molecules embedded in an ultrathin spin-on-glass detection "hot zone", which is a planar and uniformly enhanced region several nanometers above the Ag nanoparticles. The Raman measurement results exhibit a linear response over a wide dynamic range of analyte concentration.

Quantitative SERS studies by combining LOC-SERS with the standard addition method.

Here, we report on a proof-of-concept study highlighting a new approach for quantitative surface enhanced Raman spectroscopy (SERS) measurements. This has been achieved by implementing the standard addition method (SAM) within a lab-on-a-chip (LOC) system. The approach has been successfully tested to quantify congo red as a model analyte even in the presence of the chemically related molecule methyl red. Thus, the developed concept demonstrates its potential to quantify analytes via SERS in the presence of other SERS active molecules. Graphical Abstract Congo red was quantified by means of the standard addition method implemented in the lab-on-a-chip device. Due to the developed approach, a direct detection out of the sample and in the presence of an interfering substance was possible.

Quantitative surface-enhanced Raman spectroscopy based on the combination of magnetic nanoparticles with an advanced chemometric model

Surface-enhanced Raman spectroscopy (SERS) is a leading non-destructive technique with single-molecule sensitivity and has great potential for application in various fields. However, quantitative analysis of analytes using SERS is still quite challenging, since SERS signals are significantly affected by the physical properties of SERS enhancing substrates. Here, we report the detection of rhodamine 6G (R6G) and thiram (a dithiocarbamate fungicide) by SERS technique through the combination of a recently proposed multiplicative effects model (MEM) and magnetic nanoparticles (i.e. Fe3O4@Au and Fe3O4@Ag). Experimental results showed that R6G and thiram can be accurately determined with average relative prediction error of 3.2% and 7.5%, respectively. It looks promising that SERS technique based on the combination of multiplicative effects model and magnetic nanoparticles can be applied to the quantitative assay of many other analytes.