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We present a high-throughput screening Raman spectroscopy (HTS-RS) platform for a rapid and label-free macromolecular fingerprinting of tens of thousands eukaryotic cells. The newly proposed label-free HTS-RS platform combines automated imaging microscopy with Raman spectroscopy to enable a rapid label-free screening of cells and can be applied to a large number of biomedical and clinical applications. The potential of the new approach is illustrated by two applications. (1) HTS-RS-based differential white blood cell count. A classification model was trained using Raman spectra of 52 218 lymphocytes, 48 220 neutrophils, and 7 294 monocytes from four volunteers. The model was applied to determine a WBC differential for two volunteers and three patients, producing comparable results between HTS-RS and machine counting. (2) HTS-RS-based identification of circulating tumor cells (CTCs) in 1:1, 1:9, and 1:99 mixtures of Panc1 cells and leukocytes yielded ratios of 55:45, 10:90, and 3:97, respectively. Because the newly developed HTS-RS platform can be transferred to many existing Raman devices in all laboratories, the proposed implementation will lead to a significant expansion of Raman spectroscopy as a standard tool in biomedical cell research and clinical diagnostics. [ABSTRACT FROM AUTHOR]

A single platform, integrated by a laser-induced breakdown spectroscopy detector and a Raman spectroscopy sensor, has been designed to remotely (5 m) and simultaneously register the elemental and molecular signatures of rocks under Martian surface conditions. From this information, new data fusion architecture at decisions level is proposed for the correct categorization of the rocks. The approach is based on a decision-making process from the sequential checking of the spectral features representing the cationic and anionic counterparts of the specimen. The scrutiny of the LIBS response by using a moving-window algorithm informs on the diversity of the elemental constituents. The output rate of emission lines allows projecting in a loop the elements as the cationic counterpart of the rock. In parallel, the Raman response of the unknown is compared with all the molecular counterparts of the hypothesized cation that are stored in a spectral library. The largest similarity rate unveils the final identity of the unknown. The identification capabilities of the architecture have been underscored through blind tests of 10 natural rocks with different origins. The great majority of forecasts have matched with the real identities of the inspected targets. The strength of this platform to simultaneously acquire the multielemental and the molecular information from a specimen by using the same laser events greatly enhances the "on-surface" missions for the surveillance of mineralogy. [ABSTRACT FROM AUTHOR]

Electrochemical deposition of aromatic organic molecules by reduction of diazonium reagents enables formation of molecular layers with sufficient integrity for use in molecular electronic junctions of interest to microelectronics. Characterization of organic films with thicknesses in the 1-10 nm range is difficult with Raman spectroscopy, since most molecular structures of electronic interest have Raman cross sections which are too small to observe as either thin films on solid electrodes or within intact molecular junctions. Layer formation on a 10 nm thick Ag island film on a flat carbon surface (eC/Ag) permitted acquisition of structural information using surface enhanced Raman spectroscopy (SERS), in many cases for molecules with weak Raman scattering. Raman spectra obtained on eC/Ag surfaces were indistinguishable from those on carbon without Ag present, and the spectra of oligomeric molecular layers were completely consistent with those of the monomers. Layer growth was predominantly linear for cases where such growth was sterically allowed, and linear growth correlated strongly with the line width and splitting of the C?C phenyl ring stretches. Molecular bilayers made by successive reduction of different diazonium reagents were also observable and will be valuable for applications of 1-20 nm organic films in molecular electronics. [ABSTRACT FROM AUTHOR]

Polarization-resolved Raman spectroscopy provides much more information than its conventional counterpart. However, it usually either requires a complicated setup with two spectrographs and detectors or two measurements must be performed sequentially. This study presents a simple and straightforward approach to recording both polarization components simultaneously with a single spectrograph and detector. The vertically and a horizontally polarized laser beam exiting a Wollaston prism are focused into the sample with a small spatial separation. The scattered light from both beams is imaged onto the slit of an imaging spectrograph as two spatially separated signals, i.e., the polarized and the depolarized Raman signal. Eventually, both spectra are acquired on a single CCD chip simultaneously. Experimental data of ethanol and dimethyl sulfoxide are shown as proof-of-concept. The new method has a number of advantages, for example, laser intensity fluctuations and the polarization dependence of the diffraction grating do not play a role. The proposed approach will be useful for an improved structural analysis and it will be the enabling technology for temporally resolved enantioselective Raman (esR) spectroscopy. [ABSTRACT FROM AUTHOR]

Product identification is a critical and required analysis for biotheraputics. In addition to regulatory requirements for identity testing on final drug products, in-process identity testing is implemented to reduce business risks associated with fill operations and can also be used as a tool against counterfeiting. Biotherapeutics, in particular monoclonal antibodies, represent a challenging cohort for identity determination because of their similarity in chemical structure. Traditional methods used for product identification can be time and labor intensive, creating a need for quick, inexpensive and reliable methods of drug identification. Here, driven by its molecular-specific and nonperturbative nature, we present Raman spectroscopy as an alternate analytical tool for identity testing. By exploiting subtle differences in vibrational modes of the biologics, we have developed partial least-squares-discriminant analysis derived decision algorithms that offer excellent differentiation capability using spontaneous Raman spectra as well as label-free plasmon-enhanced Raman spectra. Coupled with the robustness to spurious correlations due to its high information content, our results highlight the potential of Raman spectroscopy as a powerful method for rapid, on-site biotherapeutic product identification. [ABSTRACT FROM AUTHOR]

Controlling the packing structure and revealing the intermolecular interaction of self-assembled monolayers (SAMs) on solid surfaces are crucial for manipulating its properties. We utilized tip-enhanced Raman spectroscopy (TERS) to address the challenge in probing the subtle change of the intermolecular interaction during the assembly of a pyridine-terminated aromatic thiol on the single crystal Au (lll) surface that cannot produce enhanced Raman signal, together with electrochemical methods to study the charge transfer properties of SAM. We observed that the aromatic C = C bond stretching vibration can be a marker to monitor the strength of the intermolecular interaction of SAMs, because this Raman peak is very sensitive to the intermolecular π-π stacking. Our results indicate that the SAM experiences a surface restructuring after the formation of a densely packed monolayer. We propose that the intermolecular electrostatic repulsion governs the restructuring when the packing density is high. The correlated TERS and electrochemical studies also suggest that the intermolecular interaction may have some impact on the charge transfer properties of SAM. This study provides a molecular-level insight into understanding and exploiting the intermolecular interactions toward better control over the assembling process and tuning the electrical properties of aromatic thiols. [ABSTRACT FROM AUTHOR]

Here we describe enhanced Raman scattering at Au electrode 1 (E1)/Ag nanowire (NW)/4-aminothiophenol (4- ATP)/Au electrode 2 (E2) nanojunctions fabricated by combining self-assembly and metal electrodeposition at microgap electrodes (E1 and E2). In this method we assemble the 4-ATP on electrode E2 and electrodeposit Ag on the opposite electrode El of an Au interdigitated array (IDA) electrode device. The electrodeposited Ag grows in the form of NWs on E1 and makes nanoscale contact to E2 to form the junctions. The presence of the Ag NW leads to strong Raman scattering of the 4-ATP molecules within the nanojunction leading to estimated enhancement factors ranging from 103 to 106. Scanning electron microscopy (SEM) images provide insight into the morphology of the junctions. The magnitude of the Raman enhancement depends on the extent of contact between the Ag NW and the 4-ATP self-assembled monolayer (SAM). With this approach we could detect 4-ATP molecules diluted by a factor of 1000 with hexanethiol molecules within the junctions. Our approach is simple and fast with the potential to correlate electronic measurements of molecules with Raman spectroscopy data of the same molecules in a nanoscale junction for molecular electronics or chemiresistive sensing applications. [ABSTRACT FROM AUTHOR]

The first use of surface enhanced Raman spectroscopy (SERS) to detect trace organic and/or inorganic species in ambient atmospheric aerosol particles is presented. This new analytical method provides direct, spectroscopic detection of species present at attogram to femtogram levels in individual submicrometer atmospheric particles. An array of spectral features resulting from organic functional groups in secondary organic aerosol (SOA) material were observed in individual particles impacted on silver nanoparticle-coated substrates. The results demonstrate the complexity of organic and inorganic species in SOA formed by oxidation of biogenic volatile organic compounds (BVOCs) at the single particle level. While SOA composition is frequently assumed to be homogeneous between and within individual particles, substantial particle-to-particle variability in SOA composition and changes on scales <1 μm were observed. The observations obtained with this new method demonstrate the power of SERS to probe difficult to detect inter- and intraparticle variability in ambient SOA particles. [ABSTRACT FROM AUTHOR]

The dynamic properties of phospholipid (PL) membranes (phase state and phase transition) play crucial roles in biological systems. However, highly sensitive, direct analytical methods that shed light on the nature of lipids and their assemblies have not been developed to date. Here, we describe the analysis of PL-modified Au nanoparticles (Au@PL) using membrane surface-enhanced Raman spectroscopy (MSERS) and report the properties of the self-assembled PL membranes on the Au nanoparticle. The Raman intensity per PL concentration increased by 50-170 times with Au@PL, as compared to large unilamellar vesicles (LUVs) at the same PL concentration. The phase state and phase transition temperature of the PL membrane of Au@PL were investigated by analyzing the Raman peak ratio (R = I2882/I2930). The enhancement at 714 cm-1 (EF(714)) varied with the hydrocarbon chain length of the PLs and the assembled degree of Au@PLs. In calculation, the EF(714),assembled was estimated to be 111-142 when the distance between AuNPs was 7.0-7.5 nm, which was correlated to the speculative enhancement factor, suggesting that the assembly of the Au@PLs contributed to the MSERS. [ABSTRACT FROM AUTHOR]

A new, novel, rapid method to detect and direct readout of drugs in human urine has been developed using dynamic surface-enhanced Raman spectroscopy (D-SERS) with portable Raman spectrometer on gold nanorods (GNRs) and a classification algorithm called support vector machines (SVM). The high-performance GNRs can generate gigantic enhancement and the SERS signals obtained using D-SERS on it have high reproducibility. On the basis of this feature of DSERS, we have obtained SERS spectra of urine and urine containing methamphetamine (MAMP). SVM model was built using these data for fast identified and visual results. This general method was successfully applied to the detection of 3, 4-methylenedioxy methamphetamine (MDMA) in human urine. To verify the accuracy of the model, drug addicts' urine containing MAMP were detected and identified correctly and rapidly with accuracy more than 90%. The detection results were displayed direcdy without analysis of their SERS spectra manually. Compared with the conventional method in lab, the method only needs a 2 pL sample volume and takes no more than 2 min on the portable Raman spectrometer. It is anticipated that this method will enable rapid, convenient detection of drugs on site for the police. [ABSTRACT FROM AUTHOR]

Hair is one of the most common types of physical evidence found at a crime scene. Forensic examination may suggest a connection between a suspect and a crime scene or victim, or it may demonstrate an absence of such associations. Therefore, forensic analysis of hair evidence is invaluable to criminal investigations. Current hair forensic examinations are primarily based on a subjective microscopic comparison of hair found at the crime scene with a sample of suspect's hair. Since this is often inconclusive, the development of alternative and more-accurate hair analysis techniques is critical. In this study, we utilized surface-enhanced Raman spectroscopy (SERS) to demonstrate that artificial dyes can be directly detected on hair. This spectroscopic technique is capable of a confirmatory identification of analytes with single molecule resolution, requires minimal sample, and has the advantage of fluorescence quenching. Our study reveals that SERS can (1) identify whether hair was artificially dyed or not, (2) determine if a permanent or semipermanent colorants were used, and (3) distinguish the commercial brands that are utilized to dye hair. Such analysis is rapid, minimally destructive, and can be performed directly at the crime scene. This study provides a novel perspective of forensic investigations of hair evidence. [ABSTRACT FROM AUTHOR]

Although surface-enhanced Raman spectroscopy (SERS) has been featured by high sensitivity, additional signal enhancement is still necessary for trace amount of biomolecules detection. In this paper, a SERS amplified approach, featuring "ions-mediated cascade amplification (IMCA)", was proposed by utilizing the dissolved silver ions (Ag+) from silver nanoparticles (AgNPs). We found that using Ag+ as linkage agent can effectively control the gaps between neighboring 4-aminobenzenethiol (4-ABT) encoded gold nanoparticles (AuNPs@4-ABT) toform "hot spots" and thus produce SERS signal output, in which the SERS intensity was proportional to the concentration of Ag+. Inspired by this finding, the IMCA was utilized for ultrasensitive detection of single nucleotide polymorphism in human mitochondrial DNA (16189T → C). Combining with the DNA ligase reaction, each target DNA binding event could successfully cause one AgNP introduction. By detecting the dissolved Ag+ from AgNPs using IMCA, low to 3.0 × 10(-5) fm/μL targeted DNA can be detected, which corresponds to extractions from 200 nL cell suspension containing carcinoma pancreatic β-cell lines from diabetes patients. This IMCA approach is expected to be a universal strategy for ultrasensitive detection of analytes and supply valuable information for biomedical research and clinical early diagnosis. [ABSTRACT FROM AUTHOR]

Interesting properties of water with distinguishable hydrogen-bonding structure on interfacial phase or in confined environment have drawn wide attentions. However, these unique properties of water are only found within the interfacial phase and confined environment, thus, their applications are limited. In addition, quantitative evaluation on these unique properties associating with the enhancement of water's physical and chemical activities represents a notable challenge. Here we report a practicable production of free-standing liquid water at room temperature with weak hydrogen-bonded structure naming Au nanoparticles (NPs)-treated (AuNT) water via treating by plasmon-induced hot electron transfer occurred on resonantly illuminated gold NPs (AuNPs). Compared to well-known untreated bulk water (deionized water), the prepared AuNT water exhibits many distinct activities in generally physical and chemical reactions, such as high solubilities to NaCl and O2. Also, reducing interaction energy within water molecules provides lower overpotential and higher efficiency in electrolytic hydrogen production. In addition, these enhanced catalytic activities of AuNT water are tunable by mixing with deionized water. Also, most of these tunable activities are linearly proportional to its degree of nonhydrogen-bonded structure (DNHBS), which is derived from the O-H stretching in deconvoluted Raman spectrum. [ABSTRACT FROM AUTHOR]

Surface-enhanced Raman spectroscopy (SERS) is generally performed on planar surfaces, which can be difficult to prepare and may limit the interaction of the sensing surface with targets in large volume samples. We propose that nanocomposite materials can be configured that both include SERS probes and provide a high surface area-to-volume format, i.e., fibers. Thiol-yne nanocomposite films and fibers were fabricated using exposure to long-wave ultraviolet light after the inclusion of gold nanoparticles (AuNPs) functionalized with thiophenol. A SERS response was observed that was proportional to the aggregation of the AuNPs within the polymers and the amount of thiophenol present. Overall, this proof-of-concept fabrication of SERS active polymers indicated that thiol-yne nanocomposites may be useful as durable film or fiber SERS probes. Properties of the nanocomposites were evaluated using various techniques including UV-vis spectroscopy, μ-Raman spectroscopy, dynamic mechanical analysis, differential scanning calorimetry, therm ogravimetric analysis, and transmission electron microscopy. [ABSTRACT FROM AUTHOR]

Understanding cellular metabolism is a major challenge in current systems biology and has triggered extensive metabolomics research, which in most cases involves destructive analysis. However, the information obtainable only in a nondestructive manner will be required for accurately mapping the global structure of the organism's metabolic network at a given instant. Here we report that metabolic pathways can be explored in vivo by mixed stable isotope-labeled Raman microspectroscopy in conjunction with multivariate curve resolution analysis. As a model system, we studied ergosterol biosynthesis in single living fission yeast cells grown in mixtures of normal and 13C-labeled glucose as the sole carbon source. The multivariate spectral data analysis of space-resolved Raman spectra revealed the intrinsic spectra and relative abundances of all isotopomers of ergosterol whose carbon atoms in the 5,7-diene moiety of the sterol skeleton are either partly or fully substituted with 13C. Our approach is applicable to other metabolites and will earn a place in the toolbox of metabolomic analysis. [ABSTRACT FROM AUTHOR]

Total internal reflection (TIR) Raman and reflectivity spectra were collected for nonresonant analytes as a function of incident angle at sapphire or sapphire/smooth 50 rim gold interfaces using 785 nm excitation. For both interfaces, the Raman signal as a function of incident angle is well-modeled by the calculated interfacial mean square electric field (MSEF) relative to the incident field times the thickness of the layer being probed in the Ransan measurement (Das). The Raman scatter was reproducibly enhanced at the interface containing a gold film relative to the sapphire interface by a factor of 4.3-4.6 for aqueous pyridine or 2.2-3.7 for neat nitrobenzene, depending on the analyzed vibrational mode. The mechanism for the increased Raman signal is the enhanced MSEF at incident angles where propagating surface plasmons are excited in the metal film. The background from the TIR prism was reduced by 89-95% with the addition of the gold film, and the percent relative uncertainty in peak area was reduced from 15 to 1.7% for the 1347 cm-1 mode of nitrobenzene. Single monolayers of benzenethiol (S/N = 6.8) and 4-mercaptopyridine (S/N = 16.5) on gold films were measured by TW Raman spectroscopy with 785 rim excitation (210 mW) without resonant enhancement in 1 mm. [ABSTRACT FROM AUTHOR]

The ability to unequivocally identify a gunshot residue (GSR) when a firearm is discharged is a very important and crucial part of crime scene investigation. To date, the great majority of the analyses have focused on the inorganic components of GSR, but the introduction of "lead-free" or "nontoxic" ammunitions makes it difficult to prevent false negatives. This study introduces a fast methodology for the organic analysis of GSR using Raman spectroscopy. Six different types of ammunition were fired at short distances into cloth targets, and the Raman spectra produced by the GSR were measured and compared with the spectra from the unfired gunpowder ammunition. The GSR spectrum shows high similarity to the spectrum of the unfired ammunition, allowing the GSR to be traced to the ammunition used. Additionally, other substances that might be found on the victim's, shooter's, or suspect's clothes and might be confused with GSR, such as sand, dried blood, or black ink from a common ballpoint pen, were analyzed to test the screening capability of the Raman technique. The results obtained evidenced that Raman spectroscopy is a useful screening tool when fast analysis is desired and that little sample preparation is required for the analysis of GSR evidence. [ABSTRACT FROM AUTHOR]

In this work, we establish the use of surface-enhanced Raman scattering (SERS) as a label-free analytical technique for the direct detection of G-quadruplex formation. In particular, we demonstrate that SERS analysis allows the evaluation of the relative stability of G quadruplexes that differ for the number of G tetrads and investigate several structural features of quadruplexes, such as the orientation of glycosidic bonds, the identification of distortions in the sugar-phosphate backbone, and the degree of hydrogen-bond solvation. Herein, the fluctuation of the SERS spectra, due to the specific interaction of vibrational modes with the SERS-active substrate, is quantitatively analyzed before and after quadruplex formation. The results of this study suggest a perpendicular orientation of the quadruplexes (with or without the 3'-tetra end linker) with respect to the silver colloidal surface, which opens new perspectives for the use of SERS as a label-free analytical tool for the study of the binding mode between quadruplexes and their ligands. [ABSTRACT FROM AUTHOR]

Protein-based biopharmaceuticals are becoming increasingly widely used as therapeutic agents, and the characterization of these biopharmaceuticals poses a significant analytical challenge. In particular, monitoring posttranslational modifications (PTMs), such as glycosylation, is an important aspect of this characterization because these glycans can strongly affect the stability, immunogenicity, and pharmacokinetics of these biotherapeutic drugs. Raman spectroscopy is a powerful tool, with many emerging applications in the bioprocessing arena. Although the technique has a relatively rich history in protein science, only recently has Raman spectroscopy been investigated for assessing posttranslational modifications, including phosphorylation, acetylation, trimethylation, and ubiquitination. In this investigation, we develop for the first time Raman spectroscopy combined with multivariate data analyses, including principal components analysis and partial least-squares regression, for the determination of the glycosylation status of proteins and quantifying the relative concentrations of the native ribonuclease (RNase) A protein and RNase B glycoprotein within mixtures. [ABSTRACT FROM AUTHOR]

We report implementation of two powerful characterization tools, in situ secondary ion mass spectrometry (SIMS) and ex situ surface enhanced Raman spectroscopy (SERS), in analyzing surfaces modified by ion soft landing (SL). Cations derived from Rhodamine fiG are soft landed Onto Raman-active silver colloidal substrates and detected using SERS. Alternatively and more conveniently, high-quality SERS data are obtained by spin coating a silver colloidal solution over the modified surface once SL is complete. Well-defined SERS features are observed for Rhodainine 6G in as little as 15 mm of ion deposition. Deposition of ∼3 pmol gave high-quality SERS spectra with characteristic spectroscopic responses being derived from just ∼0.5 fmol of material. Confocal SERS imaging allowed the enhancement to be followed in different parts of deposited dried droplets on surfaces. Characteristic changes in Raman spectral features occur when Rhodamme 6G is deposited under conditions that favor gas-phase ion fragmentation. Simultaneous deposition of both the intact dye and its fragment ion occurs and is confirmed by SIMS analysis. The study was extended to other Raman active surfaces, induding Au nanostar and Au coated Ni nanocarpet surfaces and to SL of other molecules induding fluorescein and methyl red. Overall, the results suggest that combination of SERS and SIMS measurements are effective in the characterization of surfaces produced by ion SL with significantly enhanced molecular specificity. [ABSTRACT FROM AUTHOR]

We report on the characterization and detection of single metallic nanoparticles using a combination of con-elation spectroscopy and surface-enhanced resonance Raman spectroscopy (SERRS). Minimizing the number of independent characterization steps is critical to efficiently perform such an analysis. In this article, we improve upon conventional diffusion-limited approaches by implementing a flow-based system with high temporal resolution detection. The benefits of flow over diffusion measurements allow for a higher throughput of detected events resulting in shorter analysis times. The nanoparticles are sized using their rotational diffusion time calculated with a modified autocorrelation function. Experiments are performed using Au nanoparticles labeled with the reporter molecule malachite green isothiocyanate on a custom-built Raman spectrometer. [ABSTRACT FROM AUTHOR]

Confocal Raman microscopy is often used for optical sectioning but is problematic when the sample plane of interest has a weak Raman cross-section/signal relative to areas that are out-of-focus. This is especially true for clinical samples in pathology, which consist of a thin tissue (5 μm) sample placed on a thick glass slide. Here, we recognize that the problem is the result of the extent of the illumination at the confocal plane being larger than the size of the sample and propose a dark field illumination scheme to efficiently reject substrate signals. The ability of several optical configurations in rejecting out-of-plane signal is investigated for two model systems: SU-8 photo resist over Teflon and SU-8 photo resist over polystyrene. The proposed reflective dark field approach, in which excitation converged to a focal point slightly above the focal plane of the collection optics, was found to be most effective in recording data from the sample. The proposed approach is validated by the rejection of substrate response (fluorescence) in spectra acquired from 4 μm of breast tissue on a glass microscope slide. The proposed approach is easy to implement on existing confocal systems, has a straightforward optimization in acquiring data, and is not expected to result in loss of lateral resolution in mapping experiments. [ABSTRACT FROM AUTHOR]

This letter discusses the potential of deep Raman spectroscopy, surface enhanced spatially offset Raman spectroscopy (SESORS and its variants), for noninvasively detecting small, deeply buried lesions using surface enhanced resonance Raman scattering (SERRS) active nanoparticles. An experimental demonstration of this concept is performed in transmission Raman geometry. This method opens prospects for in vivo, noninvasive, specific detection of molecular changes associated with disease up to depths of several centimeters representing significant improvement over traditionally detected Raman signals by 2 orders of magnitude. The disease specific signals can be achieved using uniquely tagged nanoparticles conjugated to target molecules, e.g., antibodies for production of the SERRS signal. This provides the molecular specific signal which is many orders of magnitude greater than normal biological Raman signals and can be easily multiplexed. To date, there have been no studies demonstrating the viability of deep Raman spectroscopy coupled to surface enhanced techniques for detecting low concentrations of molecules of interest at depths of greater than 5.5 mm in tissue. Such a breakthrough would open a host of new applications in medical diagnoses. Here we propose to facilitate such capability by combining SERRS (as a probe for disease specific changes) with deep Raman spectroscopy techniques. This permits noninvasive measurement of Raman signatures from conjugated SERRS nanoparticles at clinically relevant concentrations through tissues of between 15 and 25 mm thick. [ABSTRACT FROM AUTHOR]

Raman chemical imaging microspectroscopy is evaluated as a technology for waterborne pathogen and bioaerosol detection. Raman imaging produces a three-dimensional data cube consisting of a Raman spectrum at every pixel in a microscope field of view. Binary and ternary mixtures including combinations of polystyrene beads, Gram- positive Bacillus anthracis, B. thuringiensis, and B. atrophaeus spores, and B. cereus vegetative cells were investigated by Raman imaging for differentiation and characterization purposes. Bacillus spore aerosol sizes were varied to provide visual proof for corroboration of spectral assignments. Conventional applications of Raman imaging consist of differentiating relatively broad areas of a sample in a microscope field of view. The spectral angle mapping data analysis algorithm was used to compare a library spectrum with experimental spectra from pixels in the microscope field of view. This direct one-to-one matching is straightforward, does not require a training set, is independent of absolute spectral intensity, and only requires univariate statistics. Raman imaging is expanded in its capabilities to differentiate and distinguish between discrete 1-6 μm size bacterial species in single particles, clusters of mixed species, and bioaerosols with interference background particles. [ABSTRACT FROM AUTHOR]

Self-renewable, pluripotent human embryonic stem cells (hESCs) can be differentiated into cardiomyocytes (CMs), providing an unlimited source of cells for transplantation therapies. However, unlike certain cell lineages such as hematopoietic cells, CMs lack specific surface markers for convenient identification,, physical separation, and enrichment. Identification by immunostaining of cardiac- specific proteins such as troponin requires permeabilization, which renders the cells unviable and nonrecoverable. Ectopic expression of a reporter protein under the transcriptional control of a heart-specific promoter for identifying hESC-derived CMs (hESC-CMs) is useful for research but complicates potential clinical applications. The practical detection and removal of undifferentiated hESCs in a graft, which may lead to tumors, is also critical. Here, we demonstrate a nondestructive, label-free optical method based on Raman scattering to interrogate the intrinsic biochemical signatures of individual hESCs and their cardiac derivatives, allowing cells to be identified and classified. By combination of the Raman spectroscopic data with multivariate statistical analysis, our results indicate that hESCs, human fetal left ventricular CMs, and hESC-CMs can be identified by their intrinsic biochemical characteristics with an accuracy of 96%, 98%, and 66%, respectively. The present study lays the groundwork for developing a systematic and automated method for the noninvasive and label-free sorting of (i) high-quality hESCs for expansion and (ii) ex vivo CMs (derived from embryonic or adult stem cells) for cell-based heart therapies. [ABSTRACT FROM AUTHOR]

The functionalization of silver nanoparticles (Ag NPs) by viologen dications (VGDs) is reported in this work as well as their applications in the surface-enhanced Raman scattering (SERS) detection of polycyclic aromatic hydrocarbons (PAHs). VGDs are able to form intermolecular cavities at interparticle junctions (SERS hot spots) where the analyte can be allocated. This leads to a giant intensification of the Raman emission of the target molecule. This effect was applied in the detection of PAHs, one of the most widespread and dangerous group of pollutants existing in the atmosphere and waters. A comparison between sensing-systems based on different VGDs (lucigenin, diquat, and paraquat) was done for the detection of two PAHs (pyrene and benzo[c]phenanthrene). The functionalization with lucigenin (LG) provided the most powerful and stable VGD-NPs sensor, which allowed the SERS detection of pyrene (PYR) down to 10-9 M in the macro setup and in the zeptomole range for spectra obtained by single NPs aggregates (micro setup). Besides, SERS spectra afforded important structural information about the interaction mechanism of VGD and PAHs, revealing the formation of a CT complex between the VGD and PYR and changes in the host conformation. The position of the ν(Ag—Cl) band and the plasmon resonance contribution assigned to Ag dimers were also used as spectral markers to monitor the host-guest interaction. [ABSTRACT FROM AUTHOR]

We demonstrate experimentally the feasibility of monitoring the depth of optically thick layers within turbid media using spatially offset Raman spectroscopy (SORS) in combination with multivariate analysis. The method uses the deep penetration capability of SORS to characterize significantly thicker (by at least a factor of 2) layers than possible with conventional Raman spectroscopy. Typical relative accuracies were between 5 and 10%. The incorporation of depth information into a SORS experiment as an additional dimension allows pure spectra of each individual layer to be resolved using three-dimensional multivariate techniques (parallel factor analysis, PARAFAC) to accuracies comparable with the results of a two-dimensional analysis. [ABSTRACT FROM AUTHOR]

Surface-enhanced Raman spectroscopy (SERS) can be made an attractive approach for identification of Raman-active compounds and biological materials (i.e., toxins, viruses, or intact bacterial cells/spores) through development of reproducible, spatially uniform SERS-active substrates. Recently, reproducible (from substrate-to-substrate), spatially homogeneous (over large areas) SERS-active substrates have been commercialized and are now available in the marketplace. We have utilized these patterned surfaces to acquire SERS spectral signatures of intact bovine papular stomatitis, pseudocowpox, and Yaba monkey tumor viruses. Salient spectral signature features make it possible to discriminate among these genetically distinct Poxviridae-Chordopoxvirinae virions. In addition, partial least-squares, a multivariate calibration method, has been used to develop personal computer-borne algorithms useful for classification of unknown Parapoxvirus (e.g., bovine papular stomatitis virus and pseudocowpox virus) samples based solely on SERS spectral signatures. To our knowledge, this is the first report detailing application of these commercial-off-the-shelf (COTS) SERS-active substrates to identification of intact poxviruses. [ABSTRACT FROM AUTHOR]

In situ Raman spectroelectrochemistiy has been used to distinguish between thin single-wall carbon nanotubes (SWCNT) and the inner tubes of double-wall carbon nanotubes (DWCN1'). The spectroelectrochemical method is based on the different change of the electronic structure of the inner tube in DWCNT and that of SWCNT during electrochemical charging, which is reflected in the Raman spectra. During electrochemical charging the inner tubes of DWCNT exhibit a delayed attenuation of the intensities of their Raman modes as referred to the behavior of SWCNT of similar diameter. The changes are pronounced for the radial breathing mode (RBM), and thus, these modes are diagnostic for the distinction of inner tubes of DWCNT from the thin SWCNT. The different sensitivities of inner and outer tubes to the applied electrochemical charging is a simple analytical tool for differentiation of SWCNT and DWCNTin a mixture. The significance of the proposed method is demonstrated on a commercial DWCNT sample. [ABSTRACT FROM AUTHOR]

The irreversible loss of surface enhanced Raman spectroscopy (SERS) intensity derived from electrostatically immobilized Au colloidal substrate is studied. The total intensity of the SERS scattering from crystal violet molecules adsorbed onto the substrate was monitored for up to 240 s under continuous laser irradiation. The rate of signal decay was found to depend upon the thickness of the liquid layer over the coated substrate. On the basis of this observation, we propose a plausible mechanism in which surface plasmon related heating induced a small but significant lateral particle diffusion in closely spaced Au particles. The result of the shifting is an increase in the average interparticle distance, which subsequently reduces the electromagnetic coupling between the Au nanoparticles, and in turn causes a reduction in the SERS activity. Finally, a three-state model is proposed and shown to satisfactorily describe the observed decays. [ABSTRACT FROM AUTHOR]

We present a Raman spectroscopic method for the non-invasive detection of liquid explosives within bottles, and other packaging, of substantially higher sensitivity and wider applicability than that currently available via conventional Raman spectroscopy. The approach uses a modification of the spatially offset Raman spectroscopy (SORS) concept, which permits the interrogation of a wide range of containers, including transparent, colored, and diffusely scattering plastic and glass beverage, medicine, and cosmetic bottles, with no change in experimental geometry. The enhanced sensitivity is achieved by the technique's inherent ability to effectively suppress fluorescence and Raman contributions originating from the wall of the container. The application is demonstrated on the noninvasive detection of hydrogen peroxide solution, a critical component of a number of liquid explosives. In contrast to conventional Raman speciroscopy, the modified SORS concept enables the detection of concealed hydrogen peroxide solution in all the studied cases. [ABSTRACT FROM AUTHOR]

The aim of the present study was to propose a strategy for the implementation of a Process Analytical Technology system in freeze-drying processes. Mannitol solutions, some of them supplied with NaCl, were used as models to freeze-dry. Noninvasive and in-line Raman measurements were continuously performed during lyophilization of the solutions to monitor real time the mannitol solid state, the end points of the different process steps (freezing, primary drying, secondary drying), and physical phenomena occurring during the process. At-line near-infrared (NIR) and X-ray powder diffractometry (XRPD) measurements were done to confirm the Raman conclusions and to find out additional information. The collected spectra during the processes were analyzed using principal component analysis and multivariate curve resolution. A two-level full factorial design was used to study the significant influence of process (freezing rate) and formulation variables (concentration of mannitol, concentration of NaCl, volume of freeze-dried sample) upon freeze-drying. Raman spectroscopy was able to monitor (i) the mannitol solid state (amorphous, α, β, δ, and hemihydrate), (ii) several process step end points (end of mannitol crystallization during freezing, primary drying), and (iii) physical phenomena occurring during freeze-drying (onset of ice nucleation, onset of mannitol crystallization during the freezing step, onset of ice sublimation). NIR proved to be a more sensitive tool to monitor sublimation than Raman spectroscopy, while XRPD helped to unravel the mannitol hemihydrate in the samples. The experimental design results showed that several process and formulation variables significantly influence different aspects of lyophilizalion and that both are interrelated. Raman spectroscopy (in-line) and NIR spectroscopy and XRPD (at-line) not only allowed the real-time monitoring of mannitol freeze-drying processes but also helped (in combination with experimental design) us to understand the process. [ABSTRACT FROM AUTHOR]

In situ Raman spectra of solution-phase electrogenerated species have been recorded in a channel-type electrochemical cell incorporating a flat optically transparent window placed parallel to the channel plane that contains the embedded working electrode. A microscope objective with its main axis (Z) aligned normal to the direction of flow was used to focus the excitation laser beam (λexc = 532 nm) in the solution and also to collect the Raman scattered light from species present therein. Judicious adjustment of the cell position along Z allowed the depth of focus to overlap the diffusion boundary layer to achieve maximum detection sensitivity. Measurements were performed using a Au working electrode in iron hexacyanoferrate(II), [Fe(CN)6]4-, and nitrite, NO2-, containing aqueous solutions as a function of the applied potential, E. linear correlations were found between both the gain and the loss of the integrated Raman intensity, IR, of bands, attributed to [Fe(CN)6]3- and [Fe(CN)6]4-, respectively, recorded downstream from the edge of the working electrode, and the current measured at the Au electrode as a function of E. The same overall trend was found for the gain in the IR of the NO3- band in the nitrite solution. Also included in this work is a ray trace analysis of the optical system. [ABSTRACT FROM AUTHOR]

An ultrasonic standing wave around 2 MHz has been used for trapping and concentration of suspended micrometer-size particles in a flow cell, whereas Raman microspectroscopy was used as a nondestructive technique to provide molecular information about the trapped particles. With this approach, detection and discrimination of different polymer microparticles based on their characteristic Raman spectra was performed. Dextran, poly(vinyl alcohol), and melamine resin-based beads, with and without functionalization, were used for this purpose. Furthermore, taking advantage of the flow-through characteristics of the cell and the versatility of the employed flow system, full control over the media surrounding the trapped particles was achieved. This allowed us to perform chemical reactions on the trapped particles and to monitor spectral changes in real time. Here retention of cation-exchanger beads loaded with silver ions and subsequent reduction of the silver ions was demonstrated. In this way, surface-enhanced Raman (SER) active beads were prepared and retained in the focus of the Raman microscope by means of the ultrasonic field. Injection of analytes in the flow system thus allowed recording of their SER spectra. Using 9-aminoacridine, a linear dependence of the found SER signal in the range from 1 to 10 μM has been achieved. The repeatability in the recorded SER intensities was on the order of 4–5%. This included bead retention, surface-enhanced Raman layer synthesis, and analyte detection. [ABSTRACT FROM AUTHOR]

A simple surface-enhanced Raman speciroscopy (SERS) microflow cell was developed to investigate distributions of scytonemin pigment within cyanobacteria from samples of rock collected from an arctic desert that contained endolithic cyanobacteria. The assay, which has future potential use in a variety of applications, including astrobiology and analysis of microorganisms in remote environments, involved studying SERS spectra of bacteria from within geological samples. By using a dispersed colloidal substrate in the microfluidic device, surface enhancement of the order > 105 was obtained for the determination of the pigment in the microorganisms when compared to the native Raman spectra. The SERS assay, which had a nM sensitivity for scytonemin, showed that the concentration of pigment was highest in samples that had experienced the highest stress environments, as a result of high doses of UV irradiation. [ABSTRACT FROM AUTHOR]

Lactate production under anaerobic conditions is indicative of human performance levels, fatigue, and hydration. Elevated lactate levels result from several medical conditions including congestive heart failure, hypoxia, and diabetic ketoacidosis. Real-time detection of lactate can therefore be useful for monitoring these medical conditions, posttrauma situations, and in evaluating the physical condition of a person engaged in strenuous activity. This paper represents a proof-of-concept demonstration of a lactate sensor based on surface-enhanced Raman spectroscopy (SERS). Furthermore, it points the direction toward a multianalyte sensing platform. A mixed decanethiol/mercaptohexanol partition layer is used herein to demonstrate SERS lactate sensing. The reversibility of the sensor surface is characterized by exposing it alternately to aqueous lactate solutions and buffer without lactate. The partitioning and departitioning time constants were both found to be ∼30 s. In addition, physiological lactate levels (i.e., 6–240 mg/dL) were quantified in phosphate-buffered saline medium using multivariate analysis with a root-mean-square error of prediction of 39.6 mg/dL Finally, reversibility was tested for sequential glucose and lactate exposures. Complete partitioning and departitioning of both analytes was demonstrated. [ABSTRACT FROM AUTHOR]

The article surveys pharmaceutical analysis and related methodology in analytical chemistry. This primarily directs on analyzing techniques in pharmaceuticals in unformulated and dosage forms, as well as assesses the procedures of evaluating their quality, purity, strength and stability. Some of these methods relate to the analysis of a particular compound or class of compounds, determine the volatile impurities in pharmaceuticals, evaluate the role of water in the physical stability of solid dosage formulations, predict the chemical stability from accelerated aging studies, identify topical drugs and characterize Internet pharmaceuticals. Further details of these techniques are provided which include liquid chromatography, capillary electrophoresis and Raman spectroscopy.

The article discusses the potential ability of Raman spectroscopy to detect ailments such as cancer and osteoporosis and other medical conditions as varied as simple dental cavities. According to Nicholas Stone of Gloucestershire Royal Hospital, He see Raman spectroscopy as the thing that gives a person the best answer as to what's going on in the tissue, because it is purely based upon the molecules present in the cells. Identification of cancerous versus noncancerous tissue from the brain, cervix, bladder and other organs, is one of the hottest areas of research in tissue Raman. According to researchers, the direct advantage of Raman is that one do not need to do anything with the tissue, the only thing is to shine light on the tissue and collect the light that comes back.

Raman spectroscopy has become a powerful tool for microscopic analysis of organic and biological materials. When combined with optical tweezers (Raman tweezers), it allows investigating single, selected micrometric particles in their natural environment, therefore, reducing unwanted interferences from the cover plate. A general problem affecting both Raman spectrometers and Raman tweezers systems is the background caused by the environment surrounding the sample under investigation. In this paper, we report on a novel method that allows acquiring Raman spectra of a single trapped particle (polystyrene microspheres) free from any background contribution. The method is based on the use of two collinear and copropagating laser beams: the first is devoted to trapping (trap laser), while the second one is used to excite the Raman transitions (pump laser). The trap laser moves the trapped particle periodically, by means of a galvomirror, back and forth across the pump laser. The back-scattered photons are analyzed by a spectrometer and detected by a photomultiplier; finally, the resulting signal is sent to a lock-in amplifier for phase-sensitive detection. The purpose of the present work is to give a detailed description of our method and to supply a systematic study concerning the formation of the Raman signal. We trap polystyrene beads and study the dependence of the Raman signal on several parameters, such as height from the coverslip surface, the bead size, the modulation amplitude, and the pump laser intensity. Our results establish a direct and practical approach for background suppression in the spectroscopic analysis of optical trapped microsized samples. [ABSTRACT FROM AUTHOR]

There is a continued interest in the development of new on-chip protocols for the determination of the causes of infectious disease. In this paper, we demonstrate the use of surface-enhanced resonance Raman scattering (SERRS) for detecting the clinically relevant nucleic acid sequences of Chlamydia trachomatis in a bead-based lab-on-a-chip format, incorporating a solid-phase sample clean-up on-chip. The assay uses streptavidinated polymer microspheres to capture a biotinylated PCR product of the oligonucleotide sequence, which was subsequently hybridized against a complementary rhodamine-labeled, Raman active probe. Central to the assay is an in-channel integrated microfilter, which was used to retain the microspheres, enabling the bound target to be separated from the rest of the sample as part of a solid-phase cleanup (thereby removing any contributions from the background). After washing, the bound Rhodamine labeled detection probe was released thermally from the microspheres by heating and was subsequently mixed on-chip with a stream of silver nanoparticles. The signal was detected downstream using a Raman spectrometer to collect the SERRS response. The assay offers several advantages over traditional laboratory methods, including: the speed of the assay on-chip, the potential for sample clean-up; and the low volume of sample required. [ABSTRACT FROM AUTHOR]

Here, it is demonstrated that similar chemical species within a multicomponent sample can be distinguished, down to the single-molecule level, by means of their surface-enhanced vibrational fingerprints. Surface-enhanced resonance Raman scattering spectra and 2D spatial intensity maps are recorded from thin Ag nanoparticle films coated with fatty acid Langmuir-Blodgett monolayers containing one-to-one binary mixtures, at varying concentrations, of two dye molecules of similar absorption and scattering cross section (n-pentyl-5-sali-cylimidoperylene and octadecylrhodamine B). The results reveal the change in the distribution of the two dyes within the monolayer, and the breakdown of ensemble spectral averaging, which occur as the single-molecule regime is approached. It is found that the unimolecular level is reached when 1-10 molecules of each dye occupy the 1-μm² scattering areas probed by the laser. These signals are attributed to the rare spatial coincidence of isolated target analyte molecules and localized electromagnetic hot spots in the nanostructured metal film. The bianalyte nature of the samples provides strong corroborative support for the attribution of spectra to single molecules at high dilution, while the effect of domain formation/aggregation is found to be important at higher concentrations. [ABSTRACT FROM AUTHOR]

The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6,20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+ Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 µmol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., β-sheets, α-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to β-sheets and α-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement. [ABSTRACT FROM AUTHOR]

We demonstrate the use of spatially offset Raman spectroscopy (SORS) in the identification of counterfeit pharmaceutical tablets and capsules through different types of packaging. The technique offers a substantially higher sensitivity than that available from conventional backscattering Raman spectroscopy. The approach is particularly beneficial in situations where the conventional Raman backscattering method is hampered or fails because of excessive surface Raman or fluorescence signals emanating from the packaging, capsule shell, or tablet coating contaminating the much weaker subsurface Raman signals of the active pharmaceutical ingredients and excipients held in the product. It is demonstrated that such interfering signals can be effectively suppressed by SORS. [ABSTRACT FROM AUTHOR]

The great capabilities of UV Raman imaging have been demonstrated on the three Martian meteorites: Sayh al Uhaymir, Dar al Gani, and Zagami. Raman spectra without disturbing fluorescence and with high signal-to-noise-ratios and full of spectral features were derived. This result is of utmost importance for the development of powerful instruments for space missions. By point scanning the surfaces of the meteorite samples, it was possible for the first time to construct UV-Raman images out of the array of Raman spectra. Deep-UV Raman images are to the best of our knowledge presented for the first time. The images were used for a discussion of the chemical- mineralogical composition and texture of the meteorite surfaces. Comparative Raman studies applying visible and NIR Raman excitation wavelengths demonstrate a much better performance for UV Raman excitation. This comparative study of different Raman excitation wavelengths at the same sample spots was done by constructing a versatile, robust sample holder with a fixed micro-raster. The overall advantages of UV resonance Raman spectroscopy in terms of sensitivity and selectivity are demonstrated and discussed. Finally the application of this new technique for a UV Raman instrument for envisaged astrobiological focused space missions is suggested. [ABSTRACT FROM AUTHOR]

UV resonance Raman microspectroscopy was applied for a localization of the antiplasmodial naphthylisoquinoline alkaloid dioncophylline A in very low concentrations in different parts of the samples (e.g., in the roots) of the tropical liana Triphyophyllum peltatum. The application of resonance Raman microspectroscopy was characterized by a very high sensitivity and selectivity. It was possible to assign the resonance Raman spectra of dioncophylline A, dioncophylline C, and dioncopeltine A by means of a combination of NIR Raman spectroscopy and DFT calculations. The UV resonance Raman spectra of T. peltatum are very well resembled by the spectra of dioncophylline A, while they can be clearly distinguished from the spectra of dioncophylline C and dioncopeltine A. This distinction between the various naphthylisoquinolines was possible by the two modes at 1356 and 1613 cm-1. These two modes were assigned to CC stretching and CII bending vibrations. The presented results of a highly sensitive and selective in situ localization of the active agent dionco-phylline A in different parts of the plant material of T. peltatum are of high importance for the acquisition of new antimalarials and for plant science in general. [ABSTRACT FROM AUTHOR]

A novel method for pH measurements between pH 7.5 and 10.4 is reported in this paper. The method combines Raman spectroscopy and the automated sequential injection analysis system (SIA) and makes use of the acid-base properties of a commercially available water dispersion of polyaniline (PANI) nanoparticles with a mean particle size of 46 nm. The useful pH range of the PANI nanoparticles is broader than for conventional acid-base indicators, due to the 1:2 reaction stoichiometry of the proton-ligand complex of the nanoparticles. The pH measurements were conducted with the 633-nm laser excitation wavelength by calculating the difference between the Raman intensities of the primary and reference wavenumbers. In this study, the pH-sensitive CH=CH stretching band at 1422 cm-1 and C-H in-plane bending band of the quinoid form at 1163 cm-1 were chosen as the primary wavenumbers. The presented method is fast and allows pH to be measured with a precision of 0.2 pH unit. The advantage of the proposed method is that the hysteresis effect, which is usually observed for PANI films, can be overcome with the PANI nanoparticles, because a fresh nanoparticle solution is used in each measurement. It should be pointed out, that this work is a fundamental study showing the applicability of Raman spectroscopy in combination with the SIA technique for pH measurements in specific applications, where the conventional glass pH electrode cannot be used. [ABSTRACT FROM AUTHOR]

Accurate targeting of diseased and healthy tissue has significantly been improved by MM/CT-based navigation systems. Recently, intraoperative MM navigation systems have proven to be powerful tools for the guidance of the neurosurgical operations. However, the widespread use of such systems is held back by the costs, the time consumption during operation, and the need for MR- compatible surgical devices. Raman spectroscopy is a nondestructive optical technique that enables real-time tissue identification and classification and has proved to be a powerful diagnostic tool in a large number of studies. In the present report, we have investigated the possibility of distinguishing different brain structures by using a single fiber-optic probe to collect Raman scattered light in the high-wavenumber region of the spectrum. For the Raman measurements, 7 pig brains were sliced in the coronal plain and Raman spectra were obtained of 11-19 anatomical structures. Adjacent brain structures could be distinguished based on their Raman spectra, reflecting the differences in their biochemical composition and illustrating the potential Raman spectroscopy holds as a guidance tool during neurosurgical procedures. [ABSTRACT FROM AUTHOR]

Visualization of three-dimensional distribution of drug molecules and subsequent changes during the release process is critical for understanding drug delivery mechanisms as well as designing tailor-made release profiles. This study utilized coherent anti-Stokes Raman scattering (CARS) imaging to examine paclitaxel distribution in various polymer films with lateral resolution of 0.3 μm and depth resolution of 0.9 μm. Raman bands in the CH stretch vibration and fingerprint regions were used to distinguish paclitaxel from the polymers. The detection sensitivity was measured to be 29 mM by imaging paclitaxel molecules dissolved in N,N-dimethylformamide solution. Release of paclitaxel from a polymer matrix was monitored at an acquisition speed of 1 frame/s. Our results show that CARS microscopy can be used effectively for in situ imaging of native drug molecules in a delivery system. [ABSTRACT FROM AUTHOR]

We have applied Raman spectroscopy to discriminate between nontumor and tumor bladder tissue and to determine the biochemical differences therein. Tissue samples from 15 patients were collected, and frozen sections were made for Raman spectroscopy and histology. Twenty-five pseudocolor Raman maps were created in which each color represents a cluster of spectra measured on tissue areas of similar biochemical composition. For each cluster, the cluster-averaged spectrum (CAS) was calculated and classified as tumor and nontumor in accordance to pathohistology. Unguided hierarchical clustering was applied to display heterogeneity between and within groups of nontumor and tumor CM. A linear discriminant analysis model was developed to discriminate between CM from tumor and nontumor. The model was tested by a leave-one-patient-out validation, 84 of the 90 CM (93%) were correctly classified with 94% sensitivity and 92% specificity. Biochemical differences between tumor and nontumor CM areas were analyzed by fitting spectra of pure compounds to the CM. Nontumor CM showed higher collagen content while tumor CM were characterized by higher lipid, nucleic acid, protein, and glycogen content. Raman spectroscopy enabled effective discrimination between tumor and nontumor bladder tissue based on characterized biochemical differences, despite heterogeneity expressed in both tumor and nontumor CM. [ABSTRACT FROM AUTHOR]

This paper describes a label-free assay for measuring endogenous caspase protease activities in cell lysates. The assay format, termed SAMDI-MS (self-assembled mono-layers for matrix assisted laser desorption ionization time-of-flight mass spectrometry), is based on the enzymatic modification of peptides immobilized to monolayer substrates, followed by direct detection of the products with mass spectrometry. Monolayers presenting peptide substrates for either caspase-3 or -8 were treated with lysates from Jurkat cells that were stimulated with staurosporine and SKW6.4 cells that were stimulated with LzCD95L. In both cases, the SAMDI assays reported on the activation of endogenous caspase enzymes with levels of detection that are similar to those observed using the commonly employed fluorogenic assays. The use of longer peptide substrates, which are not compatible with the fluorogenic assays, provided for a better resolution of the two caspase activities. This work is significant because it demonstrates that the SAMDI assay can be used to measure endogenous enzyme activities and because it avoids the loss of activity and specificity that often accompany label-dependent assay formats. [ABSTRACT FROM AUTHOR]