ADVANCES IN RAMAN TECHNIQUES Laser requirements and advances for Raman techniques Andreas Isemann Laser Quantum GmbH, 78467 Konstanz, Germany INTRODUCTION 473 nm and 1064 nm, a narrow Raman scattering as a probe of bandwidth output of few tens of GHz vibrational transitions has made or below 1 MHz if needed within the leaps and bounds since its discovery, linewidth of vibrational transitions and various schemes based on this for high resolution, low noise (less phenomenon have been developed than 0.02%) and excellent beam with great success. quality (fundamental transversal Applications range from basic electromagnetic mode TEM00) scientific research, to medical and provides optimised performance industrial instrumentation. Some for the resolution of the Raman schemes utilise linear Raman measurement needed. scattering, whilst others take advantage The wavelength is chosen based of high peak-power fields to probe on the sample under investigation, nonlinear Raman responses. with 532 nm being commonly used This article intends to provide a for the necessary virtual electronic brief overview of the differences and transition. In the following section, benefits, together with the laser source four examples from different areas of requirements and the advancements Raman applications show the diverse in techniques enabled by recent applications of linear Raman and what developments in lasers. advances have been achieved. An example of studying a real-world LINEAR RAMAN application, the successful control of Figure 1 An example of the RR microfluidic device counting of The advent of the laser in providing a food quality using Raman spectroscopy photosynthetic microorganisms. As the cells of the model strain high-intensity coherent light source and multivariate analysis, is described Synechocystis sp. PCC 6803 flow through the Raman detection area 1 of the microfluidic device, RR spectra were acquired continuously has helped to make Raman scattering by Jernshøj et al. Examining the about 27 times every second. The v1 RR band was used to a useful method for spectroscopy quality of meat and vegetables is differentiate 12 C- and 13 C-cells. The intensity of this band was by increasing signal levels of the discussed, together with the source plotted against the temporal axis and displayed in green and red for spontaneous event to enable use of specification, when constructing a 12 C- and 13 C-cells, respectively. A part of the figure near 23.3 s was readily available detectors. system for conditions outside of a enlarged to show a 13 C- and 12 C-cell passed through the Raman detection area sequentially only about 0.1 s apart from each other; The most widely used method to laboratory environment. the untreated RR spectra of those two cells show a distinctive red shift date is linear Raman, which takes For real-world applications, the of all of the carotenoids RR bands. advantage of commercial continuous- laser source needs not only to work Reprinted (adapted) with permission from Li et al2. Copyright (2012) American Chemical Society wave lasers. The choice of excitation at the right output parameters, but wavelength depends on the sample also it necessitates a robust design used, where in general, a shorter making it portable and highly efficient wavelength would yield a higher for integration into instruments and efficiency yet suffers from higher being able to interface with operating scattering and may induce damage software. in biological samples within the UV The two next experiments benefit region. from the high resolution that a narrow A diode pumped solid state (DPSS) linewidth laser source offers as well as laser with a wavelength between its high beam quality. MicroscopyandAnalysis | July/August 2017 19 AM ADVANCES IN RAMAN TECHNIQUES Application of a single longitudinal enhanced Raman scattering (SERS) or mode (SLM) laser at 532 nm with as a subvariant, tip enhanced Raman a linewidth of 1 MHz enables scattering (TERS) have been developed. high resolution resonance Raman In SERS, structured surfaces are used (RR) spectra from the excitation to increase the Raman signal level of carotenoids to differentiate with the electrical field enhancements 13 12 between CO2 and CO2, used for due to those structures. Recently, high throughput profiling of cell also hybrid structures making use of populations2, see fig. 1. The cells nano-sized semiconductors, including analysed are from photosynthetic magnetic materials, have attracted microorganisms that are of interest due to their excellent SERS extraordinary value in the area of performance, as described by Prakash renewable energy and biotechnology. and colleagues5. In this research, a high throughput In TERS, the excitation light is Raman-activated cell sorting (RACS) brought onto the sample via an system has been developed, to perform atomic force microscope (AFM) tip Raman-activated cell counting. using plasmonic effects. This helps Figure 2 Single-molecule TERS of a porphyrin derivate examined Being able to tightly focus the laser to overcome the diffraction limit of with UHV-STM at low temperatures. (A) The Raman spectrum from beam provides for necessary spatial the laser spot, increasing the spatial powder represents many orientations, whereas (B) TERS was able to resolution in the order of 0.5 – 1 µm to resolution, as well as giving a field resolve a molecule tilt, and (C) also shows single-molecule resolution look at individual cells. enhancement due to the tip geometry. on a flatly oriented molecule. Especially the differences between TERS spectra (B) and (C) are remarkable. Adapted with permission from A novel approach for precise In some cases, depending on the Xhang, et al.8 quantitative measurements of optical sample, the increase of the signal due Copyright Macmillan Publishers Ltd. 2013 mobility in atomically thin films based to the field enhancement overcomes on high resolution Raman is presented the decrease due to a lower number by Ji et al.3. Using a large change in of oscillators (molecules) inside the the intensity of the Raman mode with excitation volume. An excellent laser an applied strong magnetic field that beam quality with a high degree of occurs in one plane, but not the out polarization enables it to focus down of plane Raman mode. It stems from to a micrometre onto the AFM needle a magnetic field induced symmetry as well as providing efficient excitation breaking. of the polarization dependent surface A quantitative analysis of the field plasmon, the mechanism responsible dependent Raman intensity allows the for electric field enhancement at determination of the optical mobility. the tip of the needle. The articles by For the quantitative measurements, the Kumar6 and Langelüddecke7 provide low noise and high power stability are an overview of the method and variety important. The excellent beam quality of applications. of the SLM laser used (Laser Quantum As always with input coupling into torus), enables confocal micro-Raman microscope setups, a superb beam in the measurement setup. pointing stability with fast warmup a The more commonly used linewidth times on the order of a few minutes in continuous wave (CW) Raman is help with reliable day to day operation a few 10s of GHz, where most robust to concentrate on the experiment. lasers are readily available. In cancer In the following, two examples of research, screening cells for potentially achievements using TERS are given, cancerous behaviour calls for an before turning to non-linear Raman. automated analysis system. Such a The high spatial resolution of TERS system based on laser tweezers Raman has enabled the visualisation of the b spectroscopy (LTRS) has been presented tilt of a single molecule by means of by Casabella et al.4. An excellent beam measuring a spectrum, see figure 2 c quality of TEM00 ensuring a tight from the paper by Zhang et al.8 Also, focus as well as a long-term stable the authors were able to resolve the beam pointing are needed for trapping ring system of a single porphyrin the individual cells in the microfluidic derivative with a spatial resolution of channels and for the Raman 0.5 nm, see fig. 2 (right side). measurement. In another fascinating experiment, Two lasers of different wavelength TERS has been able to reveal spin were used (1064 nm for trapping, waves in iron oxide nanoparticle, as 532 nm for Raman, both from the described by Rodriguez et al.9 and Figure 3 a) Tip-enhanced Raman spectra in a line scan along the Laser Quantum ventus series), which depicted in fig. 3 using micro-Raman nano-particles/glass interface. The stacking representation shows integrated well into the overall system experiments. several first order modes below 700 cm−1 , the 2LO mode around 1323 cm−1 and the spin-wave mode around 1587 cm−1 . The step size control by use of their remote control used to obtain this set of spectra was 1 nm. In the inset is shown the capability. NON-LINEAR RAMAN AFM image acquired during the TERS line scan (b) intensity profiles Some samples allow increased To drastically increase the efficiency of of three modes along the line scan showing the sharp increase in the efficiency of the scattering when the the Raman process, various coherent spin-wave intensity when the TERS tip is on the nanoparticle. This is Raman transition happens to occur (nonlinear) Raman processes have been followed by a decrease in the intensity of all modes when the tip was retracted (tip off). (c) A sketch of the tip–nanoparticle configuration close to a real transition. However, studied. The Nonlinear Raman process shows the anisotropy between the two sides of the particle with not all samples of interest would offer uses at least two different wavelengths, respect to the tip. Crystallographic facets and edges are set according this benefit. To increase the sensitivity but uses the same Raman active to Rodriguez et al.27 and spatial resolution of linear vibrational mode as the linear Raman Reprinted (adapted) with permission from Rodriguez et al.9.