Raman Spectroscopy Imaging Research Articles

Probing Deuteration-Induced Phase Separation in Supported Lipid Monolayers using Hyperspectral TERS Imaging.

In this study, we investigate the impact of deuteration on the formation of phase-separated domains in supported lipid monolayers using hyperspectral Tip-Enhanced Raman Spectroscopy (TERS) imaging. The intricate organization of biological membranes plays a crucial role in cellular functions. Various factors that influence domain formation have been identified in previous studies such as lipid tail length and cholesterol concentration. Deuterium labeling of lipids has proven useful for probing cellular structures and dynamics, but its impact on lipid phase separation remains underexplored. By examining 1:1 mixed monolayers of dipalmitoylphosphatidylcholine (DPPC) and deuterated DPPC on Au(111) surfaces, we reveal partial segregation of domains rich in deuterated and nondeuterated lipids. This study addresses a gap in knowledge by examining the impact of deuteration on lipid tail behavior, offering new insights into how even subtle structural modifications can influence phase behavior. Furthermore, it demonstrates that TERS can be a powerful, nondestructive, and label-free nanoanalytical tool for analyzing lipid membranes and advance the field of membrane biophysics.

Nanoscale Chemical Imaging of Amyloid Fibrils in Water Using Total-Internal-Reflection Tip-Enhanced Raman Spectroscopy.

Total-internal-reflection tip-enhanced Raman spectroscopy (TIR-TERS) imaging of amyloid-β (Aβ1-42-L34T) fibrils is performed with nanoscale spatial resolution in water, using TERS tips fabricated by bipolar electrodeposition. Ideal experimental parameters are corroborated by both theoretical simulations and TIR-TERS measurements. TIR-TERS imaging reveals the predominant parallel β-sheet secondary structure of Aβ1-42-L34T fibrils as well as the nanoscale spatial distribution of tyrosine, histidine, and phenylalanine aromatic amino acids. Their proportion in TERS spectra can be qualitatively explained by the combined effect of their localization in the Aβ1-42-L34T fibril structure and their molecular orientation with respect to the excitation laser light polarization. Conclusions drawn from the TERS experiments in water corroborate and significantly enrich our previous study in ambient air, thus confirming that hydration has only a marginal impact on the structure of such amyloid fibrils. This first TIR-TERS study in liquid opens fascinating perspectives for future applications in biology.

A Novel HER2 Protein Identification Methodology in Breast Cancer Cells Using Raman Spectroscopy and Raman Imaging: An Analytical Validation Study.

Conventional assays such as immunohistochemistry (IHC) and in situ hybridization (ISH) used in clinical procedures for quantification of the human epidermal growth factor receptor-2 (HER2) status in breast cancer have many limitations. In the current study, we have used HER2 expression in a broad range of breast cancer phenotypes to explore the potential utility of a novel immunodetection technique using Raman spectroscopy and Raman imaging combined with artificial intelligence models. The correlations between the Raman method and conventional HER2 testing methodologies (IHC and ISH) have been tested. Raman measurements showed a strong linear correlation (p = 0.05, R2=0,9816) with IHC analysis in the studied breast cell lines: MCF-10A, MCF-7, MDA-MB-231, HTB-30 (SK-BR-3), and AU-565 represent normal, nontumorigenic epithelial cells, triple-positive breast carcinoma, and triple-negative breast cancer cell lines. Analytic testing of Raman spectroscopy and Raman imaging demonstrated that this method may offer advantages over the currently used diagnostic methodologies.

Oxidative weathering of pyrite in acidic environments: Data-driven experimental evaluation coupled with Raman hyperspectral imaging

This study deals with the identification of the factors that affect pyrite oxidation in acid mine drainage conditions. For this scope, weathering experiments have been carried at laboratory scale based on the design of experiments methodology to evaluate the effect of factors such as major ion concentrations, crystal size, and humic acids presence over the amount of elemental sulfur produced due to the involved weathering reactions. In particular, metal and anionic concentrations in solution were quantified by inductively coupled plasma-atomic emission spectroscopy and ion-chromatography techniques, respectively, whereas the amount of elemental sulfur was quantified with a high-performance liquid chromatography with diode-array detection technique after proper extraction procedure. A partial least squares regression was calculated to establish a quantitative relationship between the considered factors and the amount of elemental sulfur. After evaluation of the model, ferric iron, crystal size and the presence of humic acids were identified as the relevant factors for pyrite oxidation under acidic conditions. In addition, the surface of the samples was characterized by Raman imaging spectroscopy and subsequently analyzed by explorative hyperspectral analysis methods to assess the spatial distribution of the elemental sulfur as the main weathering product, resulting in a homogenous distribution.

A tutorial on optical photothermal infrared (O-PTIR) microscopy.

This tutorial reviews the rapidly growing field of optical photothermal infrared (O-PTIR) spectroscopy and chemical imaging. O-PTIR is an infrared super-resolution measurement technique where a shorter wavelength visible probe is used to measure and map infrared (IR) absorption with spatial resolution up to 30× better than conventional techniques such as Fourier transform infrared and direct IR laser imaging systems. This article reviews key limitations of conventional IR instruments, the O-PTIR technology breakthroughs, and their origins that have overcome the prior limitations. This article also discusses recent developments in expanding multi-modal O-PTIR approaches that enable complementary Raman spectroscopy and fluorescence microscopy imaging, including wide-field O-PTIR imaging with fluorescence-based detection of IR absorption. Various practical subjects are covered, including sample preparation techniques, optimal measurement configurations, use of IR tags/labels and techniques for data analysis, and visualization. Key O-PTIR applications are reviewed in many areas, including biological and biomedical sciences, environmental and microplastics research, (bio)pharmaceuticals, materials science, cultural heritage, forensics, photonics, and failure analysis.

Unsupervised and interpretable discrimination of lithium-bearing minerals with Raman spectroscopy imaging

As lithium-bearing minerals become critical raw materials for the field of energy storage and advanced technologies, the development of tools to accurately identify and differentiate these minerals is becoming essential for efficient resource exploration, mining, and processing. Conventional methods for identifying ore minerals often depend on the subjective observation skills of experts, which can lead to errors, or on expensive and time-consuming techniques such as Inductively Coupled Plasma Mass Spectrometry (ICP-MS) or Optical Emission Spectroscopy (ICP-OES). More recently, Raman Spectroscopy (RS) has emerged as a powerful tool for characterizing and identifying minerals due to its ability to provide detailed molecular information. This technique excels in scenarios where minerals have similar elemental content, such as petalite and spodumene, by offering distinct vibrational information that allows for clear differentiation between such minerals. Considering this case study and its particular relevance to the lithium-mining industry, this manuscript reports the development of an unsupervised methodology for lithium-mineral identification based on Raman Imaging. The deployed machine-learning solution provides accurate and interpretable results using the specific bands expected for each mineral. Furthermore, its robustness is tested with additional blind samples, providing insights into the unique spectral signatures and analytical features that enable reliable mineral identification.

Miniaturized confocal Raman and Laser-induced breakdown spectroscopy imaging system based on micro-electro-mechanical mirror

Detection of the elemental and molecular structural distribution with high resolution and miniaturization of unknown minerals is a main bottleneck in deep space exploration and geology analysis. The aim is to enhance the accuracy of the chemical analysis of micro-samples by combining the distribution information from Raman spectroscopy and laser-induced breakdown spectroscopy (LIBS). The existing Raman-LIBS imaging methods are difficult to balance the imaging performance and system volume. There is an urgent need to develop a Raman-LIBS imaging method with miniaturization, and high imaging performance. A miniaturized Raman-LIBS imaging instrument based on the micro-electro-mechanical (MEMS) mirror has been developed to overcome these challenges. The instrument utilizes dual 2D MEMS mirror scanning technology to shorten the optical length of the system and improve the detection efficiency of hybrid spectral signals. The optical probe measures 94 mm × 66 mm, and has an axial focusing ability of approximately 40 nm, with a lateral resolution of approximately 700 nm for Raman maps and 9.5 μm for LIBS maps. As a proof experiment, 3D high-resolution Raman-LIBS hybrid spectral distribution maps of meteorite Tisserlitine 001 were obtained. The attainment of high imaging performance and miniaturization in hybrid spectral imaging is crucial for on-site chemical analysis. The proposed instrument enables in-situ spectrum and multispectral imaging with miniaturization, high spatial resolution, and high stability. The instrument is a powerful tool for composition and structural information characterization in the fields of space exploration and geological exploration.

Different methods for single molecule detection

Abstract The molecule is the smallest unit that can exist independently and maintain the physical and chemical properties of the substance, so single molecule detection is a critical technology in biology, chemistry, and other disciplines. Today, different instruments and methods for single-molecule detection exist. Each single molecule detection method’s principle, operation, and application scope differ. Systematic analysis and summary are needed. This research will outline the principles and recent achievements of several single-molecule detection methods, from microscopes to spectrometers and biosensors. The development of atomic force microscopy (AFM) is mainly introduced in microscopy. First, the fundamentals of AFM will be introduced, followed by high-speed atomic AFM (Hs-AFM) and its integrated green laser irradiation of single molecules for dynamics, Later, we will introduce the recently developed Localization AFM (LAFM) with higher resolution and the Single-molecule localization microscopy (SMLM) with similar principle. Raman spectroscopy and Raman imaging are mainly introduced in spectroscopy. In the last aspect of biosensors, the principle and development of nanopore and enzyme-linked immunosorbent assay (ELISA) are mainly introduced.

Raman spectroscopic imaging – practical applications in industrial pharmacy and medical diagnostics: a review.

Currently, Raman imaging as a non-destructive technique providing knowledge on structural characterization of the investigated samples is gaining more and more interest among industrial formulation and process scientists. In the pharmaceutical sector, the creation of novel products and formulations necessitates flexible and adaptable analysis, and increasingly also imaging capabilities. In this paper, the attention is directed toward the research progress, that has occurred in the application of non-invasive Raman chemical imaging techniques in industrial pharmacy and medical diagnostics in recent years. This work aims to explore the potential of Raman imaging as a knowledge discovery method for the non-destructive evaluation of pharmaceutical formulations as well as human cells to support medical diagnostics in clinical practice. A review of the latest scientific papers indicates that the applications of Raman imaging in pharmaceutical sciences and medical diagnostics are diverse and far-reaching from ensuring the quality of pharmaceutical formulations to studying complex drug delivery systems, monitoring manufacturing processes, and medical diagnostics of cancers and other diseases. As technology continues to evolve, the integration of Raman imaging into pharmaceutical and clinical practices is expected to expand, providing new insights and solutions to the challenges faced by the pharmaceutical industry and medical diagnostics.

A Miniaturized Endoscopic Device Integrating Raman Spectroscopy and Laser Speckle Technology Via an Image Fusion Algorithm for Intraoperative Identification and Functional Protection of Parathyroid Glands

Abstract The importance of parathyroid glands (PGs) protection is increasingly recognized by thyroid surgeons in the field of minimally invasive video-assisted thyroidectomy (MIVAT). However, current techniques for intra-operative identification and vascularity assessment of the PGs are contentious and complex. This study presents the design and validation of a miniaturized device for fusion near-infrared autofluorescence (NIRAF) based on Raman spectroscopy and laser speckle contrast imaging (LSCI) via an image fusion algorithm for functional protection of PGs in the MIVAT. Our light source components include an integrated light emitting diode (LED) light source fiber, an NIRAF optical fiber, an acquisition optical fiber, and a laser speckle optical fiber in the endoscopic system to achieve identification and vascularity assessment of PGs using a single endoscopic probe. It has been validated in ex vivo tissue experiments that the fluorescence intensity detected by the device was equivalent to that of the marker indocyanine green (ICG) in visual images and superior to that of the thyroid and all other tissues in the neck. Based on clinical studies, MIVAT using functional fluorescence endoscopy, compared with the White light pattern group, the NIRAF combined with LSCI modality group increased the number of intra-operative confirmations of the PGs (P < 0.001), declined the decrease in parathyroid hormone (PTH) (P < 0.05) and calcium levels (P < 0.05) on the first postoperative day, and reduced the incidence of symptomatic hypocalcemia (P < 0.05). Our device may reduce the incidence of postoperative permanent hypoparathyroidism. Application of an miniaturized functional fluorescence endoscope for real-time and label-free PGs identification and vascularity assessment in MIVAT could be realized.

O21 Raman spectroscopy for detection of squamous cell carcinoma in recessive dystrophic epidermolysis bullosa skin

Abstract Introduction and aims Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic skin disorder caused by biallelic pathogenic variants in COL7A1, which encodes type VII collagen (C7). Lack of C7 leads to blistering, chronic wounds, scarring and early onset of life-shortening squamous cell carcinomas (SCCs). Currently, the suspicion of SCC is mostly determined by clinical observation of visible changes in chronic wounds, and often requires multiple biopsies for histological diagnosis. This project aims to improve clinical care by developing a new noninvasive means of detecting SCC. To achieve this, we used Raman spectroscopy, a nondestructive analytical technique to study the changes occurring in RDEB skin when SCC is present, with the goal of creating a bedside test that can aid clinicians in the early diagnosis of SCC. Methods Raman spectroscopic imaging was performed on 7-µm frozen skin sections from RDEB (n = 3), RDEB SCC (n = 3) and normal skin (n = 3) to identify signalling changes between nonlesional and lesional states. An in-depth molecular composition analysis of the samples was performed using multivariate analysis with the goal of highlighting changes in abundance of specific molecules when SCC is present. Results For both nonmalignant and RDEB SCC skin sites we found specific Raman peaks associated with protein (1004 cm−1, 1650 cm−1) or lipids (1450 cm−1). RDEB SCC indicated a relative increase in DNA and protein. High-quality Raman spectroscopic images could be acquired that showed clear correlation with histological appearances. Current work is under way to determine which specific biological compounds contribute to the Raman spectra observed. Conclusions Raman spectroscopic imaging can provide comprehensive and noninvasive molecular tissue mapping with resolution comparable to histological sections, enabling molecular-based detection of SCC. Future work will focus on characterization of the SCC molecular profile, in addition to determining whether fibre optic Raman spectroscopy can be used to guide the biopsy/resection of RDEB lesions.

Single‐Walled Carbon Nanotube‐Based Optical Nano/Biosensors for Biomedical Applications: Role in Bioimaging, Disease Diagnosis, and Biomarkers Detection

AbstractThe convergence of advanced nanotechnology with disease diagnosis has ushered in a transformative era in healthcare, empowering early and accurate detection of diseases and paving the way for timely interventions, improved treatment outcomes, and enhanced patient well‐being. The development of novel materials is frequently the impetus behind significant advancements in sensor technology. Among them, single‐walled carbon nanotubes (SWCNTSs) have emerged as promising nanomaterials for developing biosensors. Their unique optical, electrical, and biocompatibility properties make them promising candidates for enhancing the sensitivity and real‐time monitoring capabilities of biosensors, as well as for enabling various bioimaging techniques. Recent studies have demonstrated the utility of SWCNTS‐based biosensors in the real‐time monitoring of biological analytes, such as nitric oxide and hydrogen peroxide (H2O2), with potential implications for disease understanding and therapeutic response assessment. Moreover, SWCNTSs have shown promise in bioimaging applications, including fluorescence, Raman spectroscopy, and photoluminescence imaging of biological samples. This article delves into the core principles, design strategies, and operational mechanisms that underpin SWCNTS‐bioimaging techniques‐based biosensors. It emphasizes on their unique properties and versatile functionalization of carbon nanotubes, laying the foundation for their integration into biosensor platforms and applications aimed at diagnosing a wide spectrum of diseases including infectious diseases, cancer, neurological disorders, and metabolic conditions.

Isotope labeled 3D-Raman confocal imaging and atomic force microscopy study on epithelial cells interacting with the fungus Candida albicans

The human pathogenic fungus Candida albicans damages epithelial cells during superficial infections. Here we use three-dimensional-sequential-confocal Raman spectroscopic imaging and atomic force microscopy to investigate the interaction of C. albicans wild type cells, the secreted C. albicans peptide toxin candidalysin and mutant cells lacking candidalysin with epithelial cells. The candidalysin is responsible for epithelial cell damage and exhibits in its deuterated form an identifiable Raman signal in a frequency region distinct from the cellular frequency region. Vibration modes at 2100–2200 cm−1 attributed to carbon‑deuterium bending and at 477 cm−1, attributed to the nitrogen‑deuterium out-of-plane bending, found around the nucleus, can be assigned to deuterated candidalysin. Atomic force microscopy visualized 100 nm deep lesions on the cell and force-distance curves indicate the higher adhesion on pore surrounding after incubation with candidalysin. Candidalysin targets the plasma membrane, but is also found inside of the cytosol of epithelial cells during C. albicans infection.

Técnicas de Imagenología Óptica Aplicadas a Dermatología

The use of optical imaging technology in recent years has improved diagnostics in the dermatology area. The techniques that can be named are optical coherence tomography, Raman spectroscopy, and optoacoustic imaging. Each technique has applications in general medicine and dermatology but with their respective limitations and focus. This work reviews the imaging techniques in general, prioritizing the theoretical background that allows the technique to operate, and then an image example obtained with the technique is shown. Then, an extensive review of the techniques of hyperspectral imaging and hyperspectral polarized dermatoscope is made. Finally, we talk about new trends in optical imaging future.

Portable spectroscopy, digital imaging colorimetry and multivariate statistical tools in contaminant identification: A case study of mint (Mentha) and basil (Ocimum basilicum)

The advent of portable Fourier-Transform Infrared (FTIR) and Raman spectrometers has revolutionized analysis capabilities, presenting the possibility of on-site contaminant identification without the need for specialized laboratory settings. Compared to laboratory instrumentation, portable spectroscopy is more prone to noise, and appropriate spectral processing procedures need to be established. This paper introduces a comprehensive methodology that integrates acquisition techniques, spectral analysis, and mathematical tools necessary for utilizing handheld spectrometers to diagnose plant contamination. It focuses on determining the efficacy of handheld FTIR, Raman spectroscopy, and digital imaging for detecting contaminants in two food plants, Basil (Ocimum basilicum) and Mint (Mentha). The study examines the impact of three pollutants: iron (II) sulphate (FeSO4), zinc (II) sulphate (ZnSO4), and copper (II) sulphate (CuSO4), on these plants, but also the necessary amount of measurements to spot the pollutants' effects. Measurements were conducted at the start, after 24 hours, and after 48 hours of exposure, on both fresh and dried plant leaves, as well as in solution. Spectral effects of each of the pollutants were identified with the use of multivariate statistical process control techniques. With the help of the developed methodologies, researchers can identify in-situ contaminant effects, exposure times and run diagnostics directly on the leaf both in alive and dried plants.

Precise regulation of surface plasmonic hotspots in an Au split nanoring coupled system

The signal enhancement in plasmon-enhanced spectroscopy is primarily contributed by hotspots generated through the surface plasmon resonance effect. Precise control of hotspot positions within nanostructures is of great significance in surface-enhanced Raman spectroscopy (SERS) and high spatial resolution imaging. However, predicting the positions of hotspots is challenging due to the small nanogaps and complex plasmonic interactions. In this study, we theoretically investigate the localized surface plasmonic properties of split ring nanostructure. The effects of variations in the inner radius, gap distance and thickness of the split ring on the hotspot distribution at the nanogaps are systematically analyzed using a three-dimensional finite element method. The results demonstrate that precise control over the positions and quantities of hotspots can be achieved by choosing the excitation wavelength and the gap size of the split rings. The calculated maximum Raman enhancement obtained at the hotspots can reach nine orders of magnitude. Our findings not only contribute to a deeper understanding of the SERS enhancement mechanism, but also provide theoretical insights for further development of surface-enhanced fluorescence, plasmonic sensors, and super-resolution imaging in the field of interface spectroscopy.

Characterization of individual spores of two biological insecticides, Bacillus thuringiensis and Lysinibacillus sphaericus, in response to glutaraldehyde using single-cell optical approaches.

Bacillus thuringiensis (Bt) and Lysinibacillus sphaericus (Ls) are the most widely used microbial insecticides. Both encounter unfavorable environmental factors and pesticides in the field. Here, the responses of Bt and Ls spores to glutaraldehyde were characterized using Raman spectroscopy and differential interference contrast imaging at the single-cell level. Bt spores were more sensitive to glutaraldehyde than Ls spores under prolonged exposure: <1.0% of Bt spores were viable after 10min of 0.5% (v/v) glutaraldehyde treatment, compared to ~ 20% of Ls spores. The Raman spectra of glutaraldehyde-treated Bt and Ls spores were almost identical to those of untreated spores; however, the germination process of individual spores was significantly altered. The time to onset of germination, the period of rapid Ca2+-2,6-pyridinedicarboxylic acid (CaDPA) release, and the period of cortex hydrolysis of treated Bt spores were significantly longer than those of untreated spores, with dodecylamine germination being particularly affected. Similarly, the germination of treated Ls spores was significantly prolonged, although the prolongation was less than that of Bt spores. Although the interiors of Bt and Ls spores were undamaged and CaDPA did not leak, proteins and structures involved in spore germination could be severely damaged, resulting in slower and significantly prolonged germination. This study provides insights into the impact of glutaraldehyde on bacterial spores at the single cell level and the variability in spore response to glutaraldehyde across species and populations.

Glutamic Acid Modified Gold Nanorod Sensor for the Detection of Calcium ions in Neuronal Cells.

Calcium (Ca2+) ions play a crucial role in the functioning of neurons, governing various aspects of neuronal activity such as rapid modulation and alterations in gene expression. Ca2+ signaling has a significant impact on the development of diseases and the impairment of neuronal functions. Herein, the study reports a Ca2+ ion sensor in neuronal cells using a gold nanorod. The gold nanorod (GA-GNR) conjugated glutamic acid developed in the study was used as a nano-bio probe for the experimental and in vitro detection of calcium. The nanosensor is colloidally stable, preserves plasmonic properties, and shows good viability in neuronal cells, as well as promoting neuron cell line growth. The cytotoxicity and cell penetration of the nanosensor are studied using Raman spectroscopy, brightfield and darkfield microscopy imaging, and MTT assays. The quantification of Ca2+ ions in neuronal cells is determined by monitoring the surface plasmon resonance (SPR) of the GA-GNR. The change in the intensity profile in the presence of Ca2+ incubated neurons was effectively used to develop a portable prototype of an optical Ca2+ sensor, proposing it as a tool for neurodegenerative disease diagnosis and neuromodulation evaluation.

Food vs packaging: Dynamics of oil migration from particle systems into fibrous material

Paper packaging for compacted or tabletted foods is seen as a key sustainable packaging of the future. Yet its fibrous structure is susceptible to absorb oils, fats, greases and other small molecules from the contacting food. Underlying phenomena associated with oil release from compacted food on fibre-based packaging, such as viscous liquid flow, capillarity, and gravity from compacted particle systems into fibre networks are not fully understood yet. As such, oil stain mitigation on packaging remains a challenge. Using model food tablets of 95% 500 and 50 μm particle size with 5% sunflower oil as the liquid phase, this work employed for the first-time quantitative Raman spectroscopic chemical imaging (RCI) coupled with automated image quantification for comparison of oil flow dynamics between food compacts and contacting paper packaging. The extent of de-oiling from the compact and imbibed into paper showed similar exponential decay with time for both porous systems. For the first time, oil migration dynamics on the food compact surface was 2D visualised via Raman spectroscopy and showed markedly different trends with varying environment climatic conditions and compact microstructure. The larger particle system leaked up to 50% of oil into paper, whereas the 50 μm system retained 100% of its oil, creating an effective internal oil barrier. This novel technique opens the way for further understanding liquid transfer between porous food media and harnessing microstructure engineering to increase food and packaging performance.

Advanced Microspatially Offset Raman Spectroscopy for Noninvasive Imaging of Concealed Texts and Figures Using Raman Signal, Fluorescence Emission, and Overall Spectral Intensity.

This study explores the possibility of using microspatially offset Raman spectroscopy (micro-SORS) imaging to reconstruct noninvasively letters and figures hidden by opaque layers. Micro-SORS experiments were conducted on mockup samples that mimic real situations encountered in the cultural heritage field, such as sealed letters with inaccessible text and original documents. Subsurface images were obtained using both the characteristic Raman bands of the hidden compounds and their different optical properties from the remaining matrix. In the latter case, contrast obtained through observing a difference in the overall spectral intensity and fluorescence profile rather than any specific Raman bands were used to track the images within the hidden layer. This approach opens new prospects for the use of micro-SORS in heritage science, with applications in the field that include the study of objects covered by opaque overlayers not only through their Raman signatures but also through differences in their optical properties (e.g., fluorescence emission, absorption).