Recent advances in Raman spectroscopy for skin diagnosis

The skin is the largest organ in humans.It comprises about 16%of our body.Many diseases originate from the skin,including acne vulgaris,skin cancer,fungal skin disease,etc.As a common skin cancer in China,melanoma alone grows at year rate of nearly 4%.Therefore,it is crucial to develop an objective,reliable,accurate,non-invasive,and easy-to-use diagnostic method for skin diseases to support clinical decision-making.Raman spectroscopy is a highly specic imaging technique,which is sensitive,even to the single-cell level in skin diagnosis.Raman spectroscopy provides a pattern of signals with narrow bandwidths,making it a common and essential tool for researching individual characteristics of skin cells.Raman spectroscopy already has a number of clinical applications,including in thyroid,cervical and colorectal cancer.This review will introduce the advantages and recent developments in Raman spectroscopy,before focusing on the advances in skin diagnosis,including the advantages,methods,results,analysis,and notications.Finally,we discuss the current limitations and future progress of Raman spectroscopy in the context of skin diagnosis.

Assessing pathological features of breast cancer via the multimodal information of multiphoton and Raman imaging

For unveiling the pathological evolution of breast cancer, nonlinear multiphoton microscopic(MPM) and confocal Raman microspectral imaging(CRMI) techniques were both utilized to address the structural and constitutional characteristics of healthy(H), ductal carcinoma in situ(DCIS), and invasive ductal carcinoma(IDC) tissues. MPM-based techniques,including two-photon excited fluorescence(TPEF) and second harmonic generation(SHG), visualized label-free and the fine structure of breast tissue. Meanwhile, CRMI not only presented the chemical images of investigated samples with the K-mean cluster analysis method(KCA), but also pictured the distribution of components in the scanned area through univariate imaging. MPM images illustrated that the cancer cells first arranged around the basement membrane of the duct,then proliferated to fill the lumens of the duct, and finally broke through the basement membrane to infiltrate into the stroma.Although the Raman imaging failed to visualize the cell structure with high resolution, it explained spectroscopically the gradual increase of nucleic acid and protein components inside the ducts as cancer cells proliferated, and displayed the distribution pattern of each biological component during the evolution of breast cancer. Thus, the combination of MPM and CRMI provided new insights into the on-site pathological diagnosis of malignant breast cancer, also ensured technical support for the development of multimodal optical imaging techniques for precise histopathological analysis.

Raman Micro-Imaging of the Coexistence of sⅠ and sⅡ Hydrates Formed from a Mixed Methane-Propane Gas in a Confined Space

Natural gas hydrate contains a specific amount of heavy hydrocarbons, such as ethane, propane, etc., aside from the primary guest gas of methane. Although the coexistence of two or even three hydrate structures has been discovered at several hydrate sites, the requisite formation mechanism is still not well understood. In-situ observation of the formation process of mixed methane-propane hydrate in a confined space was conducted using confocal Raman imaging microscopy. The Raman imaging results reveal that sI methane hydrate and sII mixed methane-propane hydrate are formed and coexist in the reaction system. In the confined space, the sI hydrate originates from the dissolved gas in water, while the sII hydrate is formed from free gas. The results obtained can help explain the coexistence of sI and sII hydrates found in natural hydrate samples, as well as providing insights into a possible dynamic scenario of hydrate reservoirs in geological history.

Wide-Field Vibrational Phase Contrast Imaging Based on Coherent Anti-Stokes Raman Scattering Holography

We propose and implement a wide-field vibrational phase contrast detection to obtain imaging of imaginary components of third-order nonlinear susceptibility in a coherent anti-Stokes Raman scattering （CARS） microscope with full suppression of the non-resonant background. This technique is based on the unique ability of recovering the phase of the generated CARS signal based on holographic recording. By capturing the phase distributions of the generated CARS field from the sample and from the environment under resonant illumination, we demonstrate the retrieval of imaginary components in the CARS microscope and achieve background free coherent Raman imaging.

3D Confocal Raman Imaging of Oil-Rich Emulsion from Enzyme-Assisted Aqueous Extraction of Extruded Soybean Powder

The understanding of the structure morphology of oil-rich emulsion from enzyme-assisted extraction processing(EAEP)was a critical step to break the oil-rich emulsion structure in order to recover oil.Albeit EAEP method has been applied as an alternative way to conventional solvent extraction method,the structure morphology of oil-rich emulsion was still unclear.The current study aimed to investigate the structure morphology of oil-rich emulsion from EAEP using 3 D confocal Raman imaging technique.With increasing the enzymatic hydrolysis duration from 1 to 3 h,the stability of oil-rich emulsion was decreased as visualized in the 3 D confocal Raman images that the protein and oil were mixed together.The subsequent Raman spectrum analysis further revealed that the decreased stability of oil-rich emulsion was due to the protein aggregations via SS bonds or protein-lipid interactions.The conformational transfer in protein indicated the formation of a compact structure.

Stimulated Raman scattering tomography for rapid three-dimensional chemical imaging of cells and tissue

Three-dimensional(3D)imaging is essential for understanding intricate biological and biomedical systems,yet live cell and tissue imaging applications still face challenges due to constrained imaging speed and strong scattering in turbid media.Here,we present a unique phase-modulated stimulated Raman scattering tomography(PM-SRST)technique to achieve rapid label-free 3D chemical imaging in cells and tissue.To accomplish PM-SRST,we utilize a spatial light modulator to electronically manipulate the focused Stokes beam along the needle Bessel pump beam for SRS tomography without the need for mechanical z scanning.We demonstrate the rapid 3D imaging capability of PM-SRST by real-time monitoring of 3D Brownian motion of polystyrene beads in water with 8.5 Hz volume rate,as well as the instant biochemical responses to acetic acid stimulants in MCF-7 cells.Further,combining the Bessel pump beam with a longer wavelength Stokes beam(NIR-II window)provides a superior scattering resilient ability in PM-SRST,enabling rapid tomography in deeper tissue areas.The PM-SRST technique providestwofold enhancement in imaging depth in highly scattering media(e.g.,polymer beads phantom and biotissue like porcine skin and brain tissue)compared with conventional point-scan SRS.We also demonstrate the rapid 3D imaging ability of PM-SRST by observing the dynamic diffusion and uptake processes of deuterium oxide molecules into plant roots.The rapid PM-SRST developed can be used to facilitate label-free 3D chemical imaging of metabolic activities and functional dynamic processes of drug delivery and therapeutics in live cells and tissue.

Investigating Microplastics and Nanoplastics Released from a Rubber Band Used for Orthodontic Treatment with Improved Raman Imaging Algorithms

Most teenagers experience orthodontic treatment,but we do not know the possible adverse effect of the released microplastics and nanoplastics that are recently categorized as emerging contaminants.Herein,we test the rubber band that has been employed to improve the biting of teeth during the orthodontic process to confirm the release of microplastics and nanoplastics.We improve the characterization of microplastics and nanoplastics by(i)Raman imaging,to extract and map the signal from the scanning spectrum matrix or the hyperspectral matrix and to enhance the signal-to-noise ratio statistically.To effectively extract the signal,(ii)chemometrics such as principal component analysis(PCA)are explored to convert the hyperspectral matrix to an image with an increased certainty.The nonsupervised PCA is intentionally corrected,via(iii)the algebra-based algorithm,to further increase the certainty to image the microplastics and nanoplastics.Once the signal is weak,(iv)an additional algorithm of image reconstruction via deconvolution is developed to average the background noise and smooth the image.By doing so,we estimate that millions of microplastics and nanoplastics are released daily in potential from a rubber band applied in a teenager's mouth,which might be a big concern.Overall,our approach provides a suitable option to characterize the microplastics and nanoplastics from a complex background.

Raman Spectroscopy and Imaging of Graphene

Graphene has many unique properties that make it an ideal material for fundamental studies as well as for potential applications.Here we review recent results on the Raman spectroscopy and imaging of graphene.We show that Raman spectroscopy and imaging can be used as a quick and unambiguous method to determine the number of graphene layers.The strong Raman signal of single layer graphene compared to graphite is explained by an interference enhancement model.We have also studied the effect of substrates,the top layer deposition,the annealing process,as well as folding(stacking order)on the physical and electronic properties of graphene.Finally,Raman spectroscopy of epitaxial graphene grown on a SiC substrate is presented and strong compressive strain on epitaxial graphene is observed.The results presented here are highly relevant to the application of graphene in nano-electronic devices and help in developing a better understanding of the physical and electronic properties of graphene.

Design of SERS nanoprobes for Raman imaging:materials, critical factors and architectures

Raman imaging yields high specificity and sensitivity when compared to other imaging modalities, mainly due to its fingerprint signature. However, intrinsic Raman signals are weak, thus limiting medical applications of Raman imaging. By adsorbing Raman molecules onto specific nanostructures such as noble metals, Raman signals can be significantly enhanced, termed surfaceenhanced Raman scattering(SERS). Recent years have witnessed great interest in the development of SERS nanoprobes for Raman imaging. Rationally designed SERS nanoprobes have greatly enhanced Raman signals by several orders of magnitude, thus showing great potential for biomedical applications.In this review we elaborate on recent progress in design strategies with emphasis on material properties,modifying factors, and structural parameters.

Versatile Graphene-Isolated AuAg-Nanocrystal for Multiphase Analysis and Multimodal Cellular Raman Imaging

Surface-enhanced Raman spectroscopy(SERS)-based bioanalytical technique involves the interaction of SERS-active substrate with complex environment,which has aroused intensive research interests.Compared to the commonly used Au SERS substrates,Ag nanocrystals have larger optical absorption cross section and acceptable price,but they possess poor oxidation resistance and potential biotoxicity,and the occurrence of unnecessary chemical reactions is inevitable due to the direct contact with probe molecules.Herein,we report a graphene-isolated AuAg nanocrystal(GIAAN)with the SERS-active AuAg core confined in a nanospace of few-layer graphene shell,which possesses unique Raman peaks,high SERS activity,excellent stability,superior fluorescence quenching performance and good biocompatibility.Based on the limited solubility of GIAAN in water and organic solvents,it is able to spontaneously generate interfacial self-assembled GIAAN(ISA-GIAAN)film at immiscible two-phase interfaces without any inducer,and multiphase Raman analysis of both water-and lipid-soluble drug model molecules is further achieved.Moreover,the GIAAN is further non-covalently functionalized with polyoxyethylenestearyl ether(C18-PEG)to acquire GIAAN@PEG with good water-solubility for SERS quantitative analysis in homogeneous system and multimodal Raman imaging of MCF-7 cells.We expect the versatile GIAAN holds great potential to monitor drug metabolism and guide intended drug delivery in clinic trials.

Raman imaging-assisted customizable assembly of MOFs on cellulose aerogel

Because of a weak interface-bonding force between metal–organic frameworks(MOFs)and substrates and the loss of customization in structural designs owing to the lack of the regulation of ion sites,MOFs tend to escape from the constructed composite template.In this study,the as-prepared 2,2,6,6-tetramethylpiperidyl-1-oxyl(TEMPO)-oxidized algae cellulose nanofibers(TACFs)were used to chelate metal ions at controllable sites and subsequently firmly entangle the assembled MOF crystals.The distribution of ions and synthesized MOFs inside the gel was monitored using Raman imaging technology,which provided an intuitive approach for visually observing the ions and MOF distribution.Using this technology,the synthesized customizable TACFs@ZIF-67 aerogels exhibited a high specific surface area(734.7 m^(2)/g),low density(6.18 mg/cm^(3)),controlled particle distribution,good underwater structural stability,and excellent adsorption of dyes.This study provides a way for solving the dispersion problem of MOFs in nanofibrous aerogels using Raman imaging technology–assisted microcosmic fixed-point design.

Design and implementation of the galvanometer scanning system for reflectance confocal and stimulated Raman scattering microscopy

We report on two strategies to design and implement the galvanometer-based laser-scanning mechanisms for the realization of reflectance confocal microscopy(RCM) and stimulated Raman scattering(SRS) microscopy systems. The RCM system uses a resonant galvanometer scanner driven by a home-built field-programmable gate array circuit with a novel dual-trigger mode and a home-built high-speed data acquisition card. The SRS system uses linear galvanometers with commercially available modules. We demonstrate video-rate high-resolution imaging at 11 frames per second of in vivo human skin with the RCM system and label-free biomolecular imaging of cancer cells with the SRS system. A comparison of the two strategies for controlling galvanometer scanners provides scientific and technical reference for future design and commercialization of various laser-scanning microscopes using galvanometers.

Visualization of Ferroelectric Domains in Thin Films of Molecular Materials Using Confocal Micro-Raman Spectroscopy

Ferroelectrics are an important class of functional materials.Among all their unique properties,the study of their ferroelectric domains and domain walls is of great interest due to their importance in ferroelectric applications.There are many methods to characterize ferroelectric domains,namely,scanning probe microscopy,optical microscopy,electron microscopy,etc.Currently,newly emerged molecular ferroelectrics are attracting much attention from chemists,physicists and researchers in material sciences due to their structural flexibility,light mass,simple fabrication,etc.However,for the characterization of molecular ferroelectric domains,most conventional methods require either a complicated preparation process or direct contact between physical probes and material surfaces,limiting the development of molecular ferroelectric materials.In this report,we have demonstrated that confocal micro-Raman spectroscopy,as a nondestructive and noncontact in-situ method,is very suitable for studying the ferroelectric polarization and structures of domains in molecular ferroelectrics.Taking recently reported molecular ferroelectric trimethylchloromethyl ammonium trichlorocadmium(II)(TMCM-CdCl_(3))as an example,the non-180°domains have been characterized and visualized at different temperatures.Such a simple and extendable method requires minimum sample preparation,which would further benefit the research of molecular ferroelectric domain engineering and promote the miniaturization and integration of molecular ferroelectric films.

Silicone matrices for controlled dexamethasone release:toward a better understanding of the underlying mass transport mechanisms

Dexamethasone-loaded silicone matrices offer an interesting potential as innovative drug delivery systems,e.g.for the treatment of inner ear diseases or for pacemakers.Generally,very long drug release periods are targeted:several years/decades.This renders the development and optimization of novel drug products cumbersome:experimental feedback on the impact of the device design is obtained very slowly.A better understanding of the underlying mass transport mechanisms can help facilitating research in this field.A variety of silicone films were prepared in this study,loaded with amorphous or crystalline dexamethasone.Different polymorphic drug forms were investigated,the film thickness was altered and the drug optionally partially/completely exchanged by much more water-soluble dexamethasone‘phosphate’.Drug release studies in artificial perilymph,scanning electron microscopy,optical microscopy,differential scanning calorimetry,X-ray diffraction and Raman imaging were used to elucidate the physical states of the drugs and polymer,and of the systems’structure as well as dynamic changes thereof upon exposure to the release medium.Dexamethasone particles were initially homogeneously distributed throughout the systems.The hydrophobicity of the matrix former very much limits the amounts of water penetrating into the system,resulting in only partial drug dissolution.The mobile drug molecules diffuse out into the surrounding environment,due to concentration gradients.Interestingly,Raman imaging revealed that even very thin silicone layers(<20µm)can effectively trap the drug for prolonged periods of time.The physical state of the drug(amorphous,crystalline)did not affect the resulting drug release kinetics to a noteworthy extent.

Live-Cell Imaging of NADPHProduction from Specific Pathways

As anconversion of essential cofactor for lipid biosynthesisand antioxidant defense, reduced nicotinamideadenine dinucleotide phosphate (NADPH) isproduced via various pathways, including the oxidativepentose phosphate pathway (oxPPP) and themalic enzyme 1 (ME1)-catalyzed conversion of malateto pyruvate. Live-cell detection of NADPH productionroutes remains challenging. Here, we reporttracing hydrides into lipid droplets (THILD), achemical imaging strategy for the detection ofpathway-specific NADPH generation in live cells. Thisstrategy exploits deuterium (2H)-labeled glucose([2H]Glc) tracers that transfer deuterides to NADPHvia specific pathways. The NADP^(2)H, in turn, transfersdeuterides to lipids, resulting in accumulation of C-2Hbonds in lipid droplets, which can be visualized bybioorthogonal stimulated Raman scattering (SRS)microscopy. We used this concept to demonstratethe imaging of oxPPP-produced NADPH using theoxPPP-specific tracer, [3-2H]Glc. Furthermore, the“switch on” of NADPH production by ME1 in differentiatingadipocytes was imaged by [4-2H]Glc. Finally,comparison of [3-2H]Glc and [4-2H]Glc THILDimaging of adipocytes showed that hypoxia inducessuppression of ME1-mediated NADPH productionand oxPPP-produced NADPH becomes the mainsource.

Tracking the Ti^(4+)substitution in phlogopite by spectroscopic imaging:A tool for unravelling the growth of micas at HP-HT conditions

Phlogopite solid-solutions have a wide pressure-temperature(P-T)stability field and are ubiquitous in a wide variety of geological settings,from deep lithosphere magmatic environments to upper crust metamorphic domains.Phlogopite composition represents therefore a valuable physical-chemical archive and may provide important information regarding its crystallization and the petrogenesis of the host-rock.In this paper we examine the phlogopite phenocrysts from the well-known Fort Regent mica-bearing lamprophyre minette from St.Helier(Island of Jersey,UK).Phlogopite phenocrystals from lamprophyres generally show normal-step and continuous compositional zoning,however those from the Fort Regent minette show a peculiar texture characterized by dark brown high-Ti(average TiO_(2)≈8.5 wt.%)cores enveloped by euhedral low-to mid-amplitude zonation due to oscillatory contents in Ti,Fe and Mg.Thermo-barometry modelling based on biotite-only composition yields relatively high P-T estimates(T≈970±54℃at P≈0.73±0.13 GPa)for cores whereas lower values(T≈790±54℃at P≈0.29±0.13 GPa)are obtained for the outer rims.Comparable temperatures(T≈1075±54℃)but extremely high and anomalous pressure values(P≈1.82±0.13 GPa)are obtained for the yellowish inner rims.The combination of electron micro probe(EMP)analysis and single-crystal infra-red(FTIR)imaging in the OH-stretching region shows that the exceptional and oscillatory Ti contents are due to the Ti-vacancy substitution,typical of crystallization and growth processes of HP/HT environments.Raman imaging provides additional insight for this process,confirming the dominant dioctahedral nature for the Ti-Fe-rich cores and outer rims.Interpretation of thermobaric estimates obtained from the phlogopite compositiononly model,based on the fine-scale compositional evolution,shows that pressure-temperature values from low-Ti high-Mg domains should be carefully evaluated because the substitution mechanisms during the dark mica growth are not univocally related to pressure-temperature variation of the crystallizing environment.Our results demonstrate how a multidisciplinary approach based on the combination of chemical investigations and vibrational spectroscopies could represent a valuable tool to evaluate pressure-temperature estimates from biotite composition-only thermo-barometry models and therefore to correctly unravel HP/HT petrogenetic processes at a very fine scale。