Compact conical beam shaper and freeform segmented reflector for SERS analysis.

We present a Raman spectroscopy setup containing a conical beam shaper in combination with a freeform segmented reflector for surface enhanced Raman scattering (SERS) analysis. The freeform segmented reflector and the conical beam shaper are designed by numerical approaches and fabricated by means of ultra-precision diamond tooling. The segmented reflector has a numerical aperture of 0.984 and a working distance of 1mm for SERS measurements. We perform systematic simulations using non-sequential ray tracing to assess the detecting abilities of the designed SERS-based system. We implement a proof-of-concept setup and demonstrate the confocal behavior by measuring the SERS signal of 10µM rhodamine B solution. The experimental results agree well with the simulations concerning the misalignment tolerances of the beam shaper with respect to the segmented reflector and the misalignment tolerances of the collecting fiber. In addition, we conduct benchmark SERS measurements by using a 60× objective lens with a numerical aperture of 0.85. We find that the main Raman intensity of rhodamine B at 1502 cm-1 obtained by our segmented reflector working together with the conical beam shaper is approximately 30% higher compared to the commercial objective lens.

Label-Free Quantification of Anticancer Drug Imatinib in Human Plasma with Surface Enhanced Raman Spectroscopy.

Therapeutic drug monitoring (TDM) for anticancer drug imatinib has been suggested as the best way to improve the treatment response and minimize the risk of adverse reactions in chronic myelogenous leukemia (CML) and gastrointestinal stromal tumor (GIST) patients. TDM of oncology treatments with standard analytical methods, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS) is, however, complex and demanding. This paper proposes a new method for quantitation of imatinib in human plasma, based on surface enhanced raman spectroscopy (SERS) and multivariate calibration using partial least-squares regression (PLSR). The best PLSR model was obtained with three latent variables in the range from 123 to 5000 ng/mL of imatinib, providing a standard error of prediction (SEP) of 510 ng/mL. The method was validated in accordance with international guidelines, through the estimate of figures of merit, such as precision, accuracy, systematic error, analytical sensitivity, limits of detection, and quantitation. Moreover, the feasibility and clinical utility of this approach have also been verified using real plasma samples taken from deidentified patients. The results were in good agreement with a clinically validated LC-MS/MS method. The new SERS method presented in this preliminary work showed simplicity, short analysis time, good sensitivity, and could be considered a promising platform for TDM of imatinib treatment in a point-of-care setting.

Quantitative Detection of Trace Level Cloxacillin in Food Samples Using Magnetic Molecularly Imprinted Polymer Extraction and Surface-Enhanced Raman Spectroscopy Nanopillars.

There is an increasing demand for rapid, sensitive, and low cost analytical methods to routinely screen antibiotic residues in food products. Conventional detection of antibiotics involves sample preparation by liquid-liquid or solid-phase extraction, followed by analysis using liquid chromatography-mass spectrometry (LC-MS), capillary electrophoresis (CE), or gas chromatography (GC). The process is labor-intensive, time-consuming, and expensive. In this study, we developed a new analytical method that combines magnetic molecularly imprinted polymer (MMIP)-based sample preparation with surface-enhanced Raman spectroscopy (SERS)-based detection for quantitative analysis of cloxacillin in pig serum. MMIP microspheres were synthesized using a core-shell technique. The large loading capacity and high selectivity of the MMIP microspheres enabled efficient extraction of cloxacillin, while the magnetically susceptible characteristics greatly simplified sample handling procedures. Low cost and robust SERS substrates consisting of vertical gold capped silicon nanopillars were fabricated and employed for the detection of cloxacillin. Quantitative SERS was achieved by normalizing signal intensities using an internal standard. By coherently combining MMIP extraction and silicon nanopillar-based SERS biosensor, good sensitivity toward cloxacillin was achieved. The detection limit was 7.8 pmol. Cloxacillin recoveries from spiked pig plasma samples were found to be more than 80%.

Surface Enhanced Raman Scattering for Quantification of p-Coumaric Acid Produced by Escherichia coli.

The number of newly developed genetic variants of microbial cell factories for production of biochemicals has been rapidly growing in recent years, leading to an increased need for new screening techniques. We developed a method based on surface-enhanced Raman scattering (SERS) coupled with liquid-liquid extraction (LLE) for quantification of p-coumaric acid (pHCA) in the supernatant of genetically engineered Escherichia coli (E. coli) cultures. pHCA was measured in a dynamic range from 1 µM up to 50 µM on highly uniform SERS substrates based on leaning gold-capped nanopillars, which showed an in-wafer signal variation of only 11.7%. LLE using dichloromethane as organic phase was combined with the detection in order to increase selectivity and sensitivity by decreasing the effect of interfering compounds from the analytes of interest. The difference in pHCA production yield between three genetically engineered E. coli strains was successfully evaluated using SERS and confirmed with high-performance liquid chromatography. As this novel approach has potential to be automated and parallelized, it can be considered for high-throughput screening in metabolic engineering.

Lithography-Free Fabrication of Silica Nanocylinders with Suspended Gold Nanorings for LSPR-Based Sensing.

Tunable plasmonic platforms are important for a variety of applications such as photovoltaics, LED's, optoelectronics, medical research, and biosensors. In particular, development of label-free plasmonic biosensors is one of the key research areas that utilizes plasmonic nanostructures for detection of biologically relevant molecules at low concentrations. The authors have developed a cost-effective, fast, and lithography-free method to fabricate transparent fused silica nanocylinders. The technique allows tuning of nanocylinder height, diameter, and density and can be scaled to large surface areas, such as 8 in. wafers. The authors demonstrate that gold coated nanocylinders support localized surface plasmon resonances (LSPR) from visible to near infrared wavelengths. The plasmonic platform can be characterized as suspended gold nanorings and exhibits a sensitivity of 658 nm RIU-1 with a figure-of-merit of 10, comparable to other state-of-the-art LSPR sensing platforms that utilize more complex nanofabrication pathways. It was observed that the LSPR peak positions can be controlled by varying the geometry of the nanocylinders. The authors illustrate surface functionalization, biosensing, and surface regeneration properties of the platform using thiols and detection of bovine serum albumin (BSA). The observed LSPR shifts for 11-mercaptoundecanoic acid and BSA was 12 and 26 nm, respectively.

Plasmon resonances of Ag capped Si nanopillars fabricated using mask-less lithography.

Localized surface plasmon resonances (LSPR) and plasmon couplings in Ag capped Si Nanopillar (Ag NP) structures are studied using 3D FEM simulations and dark-field scattering microscopy. Simulations show that a standalone Ag NP supports two LSPR modes, i.e. the particle mode and the cavity mode. The LSPR peak position of the particle mode can be tuned by changing the size of the Ag cap, and can be hybridized by leaning of pillars. The resonance position of the cavity resonance mode can be tuned primarily via the diameter of the Si pillar, and cannot be tuned via leaning of Ag NPs. The presence of a substrate dramatically changes the intensity of these two LSPR modes by introducing constructive and destructive interference patterns with incident and reflected fields. Experimental scattering spectra can be interpreted using theoretical simulations. The Ag NP substrate displays a broad plasmonic resonance band due to the contribution from both the hybridized particle LSPR and the cavity LSPR modes.

Selective surface-enhanced Raman scattering detection of Tabun, VX and Cyclosarin nerve agents using 4-pyridine amide oxime functionalized gold nanopillars.

We have earlier demonstrated sensitive detection of low the volatile nerve agents Tabun, Cyclosarin and VX by using handheld Raman instrumentation in conjunction with surface-enhanced Raman scattering (SERS) attained with gold and silver coated Si nanopillar substrates. In the present proof-of-concept study, the gold substrates chemically are functionalized to realize selectivity towards organophosphorus compounds (OPs) with high sensitivity. A potential capturer and reporter molecule, chemical nerve agent antidote, 4-pyridine amide oxime, is evaluated due to its high Raman cross section, high chemical affinity towards gold, and binding specificity to the target substances Tabun, VX and Cyclosarin via the oxime group. Upon selective and covalent binding, the SERS probe undergoes structural changes which are reflected in the spectral SERS responses, making it suitable for indirect monitoring of nerve agents in aqueous solution. With the probe attached to the hotspots of Au-coated Si nanopillars, the SERS signals distinctly discriminate between specific and non-specific analyte binding of Tabun, Cyclosarin and VX down to sub ppm levels. SERS spectrum of 4-PAO is measured after microliter drop coating of aqueous sample solution onto the functionalized substrates and subsequent water evaporation from surfaces. This binding assay is complemented by letting functionalized substrates being immersed into sample solutions 1 h before measuring. Binding specific SERS response decreases in following order: Tabun > VX > Cyclosarin. Overall, the concept looks promising, as expected the candidate probe 4-PAO introduces selectivity to the nanopillar gold substrates without loss of sensitivity.

Nanopillar Filters for Surface-Enhanced Raman Spectroscopy.

We present a simple, robust, and automated molecule extraction technique based on a centrifugal microfluidic platform. Fast and facile extraction of a food adulterant (melamine) from a complex sample medium (milk) on a SERS substrate is demonstrated. The unique characteristic of the detection method is the obtained "filter paper/chromatographic" effect which combines centrifugal force and wetting properties of the SERS substrate. The work addresses issues related to SERS-based detection of analytes in complex media, which is important for realizing next generation SERS platforms applicable for a broad variety of real-life applications.

Hand-Held Femtogram Detection of Hazardous Picric Acid with Hydrophobic Ag Nanopillar SERS Substrates and Mechanism of Elasto-Capillarity.

Picric acid (PA) is a severe environmental and security risk due to its unstable, toxic, and explosive properties. It is also challenging to detect in trace amounts and in situ because of its highly acidic and anionic character. Here, we assess sensing of PA under nonlaboratory conditions using surface-enhanced Raman scattering (SERS) silver nanopillar substrates and hand-held Raman spectroscopy equipment. The advancing elasto-capillarity effects are explained by molecular dynamics simulations. We obtain a SERS PA detection limit on the order of 20 ppt, corresponding attomole amounts, which together with the simple analysis methodology demonstrates that the presented approach is highly competitive for ultrasensitive analysis in the field.

Large-Scale, Lithography-Free Production of Transparent Nanostructured Surface for Dual-Functional Electrochemical and SERS Sensing.

In this work, we present a dual-functional sensor that can perform surface-enhanced Raman spectroscopy (SERS) based identification and electrochemical (EC) quantification of analytes in liquid samples. A lithography-free reactive ion etching process was utilized to obtain nanostructures of high aspect ratios distributed homogeneously on a 4 in. fused silica wafer. The sensor was made up of three-electrode array, obtained by subsequent e-beam evaporation of Au on nanostructures in selected areas through a shadow mask. The SERS performance was evaluated through surface-averaged enhancement factor (EF), which was ∼6.2 × 105, and spatial uniformity of EF, which was ∼13% in terms of relative standard deviation. Excellent electrochemical performance and reproducibility were revealed by recording cyclic voltammograms. On nanostructured electrodes, paracetamol (PAR) showed an improved quasi-reversible behavior with decrease in peak potential separation (ΔEp ∼ 90 mV) and higher peak currents (Ipa/Ipc ∼ 1), compared to planar electrodes (ΔEp ∼ 560 mV). The oxidation potential of PAR was also lowered by ∼80 mV on nanostructured electrodes. To illustrate dual-functional sensing, quantitative evaluation of PAR ranging from 30 µM to 3 mM was realized through EC detection, and the presence of PAR was verified by its SERS fingerprint.

Detection of nerve gases using surface-enhanced Raman scattering substrates with high droplet adhesion.

Threats from chemical warfare agents, commonly known as nerve gases, constitute a serious security issue of increasing global concern because of surging terrorist activity worldwide. However, nerve gases are difficult to detect using current analytical tools and outside dedicated laboratories. Here we demonstrate that surface-enhanced Raman scattering (SERS) can be used for sensitive detection of femtomol quantities of two nerve gases, VX and Tabun, using a handheld Raman device and SERS substrates consisting of flexible gold-covered Si nanopillars. The substrate surface exhibits high droplet adhesion and nanopillar clustering due to elasto-capillary forces, resulting in enrichment of target molecules in plasmonic hot-spots with high Raman enhancement. The results may pave the way for strategic life-saving SERS detection of chemical warfare agents in the field.

Non-labeling multiplex surface enhanced Raman scattering (SERS) detection of volatile organic compounds (VOCs).

In this paper, we report multiplex SERS based VOCs detection with a leaning nano-pillar substrate. The VOCs analyte molecules adsorbed at the tips of the nano-pillars produced SERS signal due to the field enhancement occurring at the localized surface plasmon hot spots between adjacent leaning nano-pillars. In this experiment, detections of acetone and ethanol vapor at different concentrations were demonstrated. The detection limits were found to be 0.0017 ng and 0.0037 ng for ethanol and acetone vapor molecules respectively. Our approach is a non-labeling method such that it does not require the incorporation of any chemical sensing layer for the enrichment of gas molecules on sensor surface. The leaning nano-pillar substrate also showed highly reproducible SERS signal in cyclic VOCs detection, which can reduce the detection cost in practical applications. Further, multiplex SERS detection on different combination of acetone and ethanol vapor was also successfully demonstrated. The vibrational fingerprints of molecular structures provide specific Raman peaks for different VOCs contents. To the best of our knowledge, this is the first multiplex VOCs detection using SERS. We believe that this work may lead to a portable device for multiplex, specific and highly sensitive detection of complex VOCs samples that can find potential applications in exhaled breath analysis, hazardous gas analysis, homeland security and environmental monitoring.

Surface-enhanced Raman spectroscopy based quantitative bioassay on aptamer-functionalized nanopillars using large-area Raman mapping.

Surface-enhanced Raman spectroscopy (SERS) has been used in a variety of biological applications due to its high sensitivity and specificity. Here, we report a SERS-based biosensing approach for quantitative detection of biomolecules. A SERS substrate bearing gold-decorated silicon nanopillars is functionalized with aptamers for sensitive and specific detection of target molecules. In this study, TAMRA-labeled vasopressin molecules in the picomolar regime (1 pM to 1 nM) are specifically captured by aptamers on the nanostructured SERS substrate and monitored by using an automated SERS signal mapping technique. From the experimental results, we show concentration-dependent SERS responses in the picomolar range by integrating SERS signal intensities over a scanning area. It is also noted that our signal mapping approach significantly improves statistical reproducibility and accounts for spot-to-spot variation in conventional SERS quantification. Furthermore, we have developed an analytical model capable of predicting experimental intensity distributions on the substrates for reliable quantification of biomolecules. Lastly, we have calculated the minimum needed area of Raman mapping for efficient and reliable analysis of each measurement. Combining our SERS mapping analysis with an aptamer-functionalized nanopillar substrate is found to be extremely efficient for detection of low-abundance biomolecules.

Large area fabrication of leaning silicon nanopillars for surface enhanced Raman spectroscopy.

Using a simple two step fabrication process substrates with a large and uniform Raman enhancement, based on flexible free standing nanopillars can be manufactured over large areas using readily available silicon processing equipment.