Chemical Image Contrast Research Articles

Structural Evolution in an Annealed (Eu, Tb)-Doped ZnO/Si Nanoscale Junction: Implication for Red LED Development

Codoped (Tb,Eu) ZnO films grown by magnetron sputtering on a silicon substrate and annealed up to 1200 °C showed intense photoluminescence (PL) emission from Eu3+ ions. The high-temperature annealing led to diffusion and segregation of rare earth (RE) elements toward the bottom of the film, which induced the formation of nanometric Zn-free inclusions responsible for remarkable PL emission intensity. Combined electron diffraction, chemical contrast imaging, and optical studies of these nanometric phases have been carried out. Large inclusions of zinc silicates and RE silicates a few hundreds of nanometers in size were observed, embedded in a silica phase. The structural determination of these RE-rich inclusions was carried out by combining atomic Z contrast imaging (high-angle annular dark-field imaging) and precession electron diffraction data. Upon annealing at 1200 °C, it appeared that the structure was related to an F-type disilicate structure. Energy-dispersive X-ray spectroscopy and electron energy loss spectroscopy experiments were carried out to determine the ratios between elements and the oxidation states of the RE elements in the abovementioned inclusions. A (Tb,Eu)2Si2O7 formulation is proposed from dynamical precession electron diffraction tomography refinements, leading to a +III valence state for the RE species in agreement with spectroscopic results. PL modeling is also in good agreement with the experimental data. These results complete those obtained at 1100 °C for which the inclusions were identified as some RE10–x(SiO4)6O2–x oxyapatite structures and pointed out a combined structural and PL properties’ evolution between 1100 and 1200 °C annealing temperatures.

Wide-Field Mid-Infrared Hyperspectral Imaging by Snapshot Phase Contrast Measurement of Optothermal Excitation.

Vibrational microscopy methods based on Raman scattering or infrared absorption provide a label-free approach for chemical-contrast imaging, but employ point-by-point scanning and impose a compromise between the imaging speed and field-of-view (FOV). Optothermal microscopy has been proposed as a promising imaging modality to avoid this compromise, although at restrictively small FOVs capable of imaging only few cells. Here, we present wide-field optothermal mid-infrared microscopy (WOMiM) for wide-field chemical-contrast imaging based on snapshot pump–probe detection of optothermal signal, using a custom-made condenser-free phase contrast microscopy to capture the phase change of samples after mid-infrared irradiation. We achieved chemical contrast for FOVs up to 180 μm in diameter, yielding 10-fold larger imaging areas than the state-of-the-art, at imaging speeds of 1 ms/frame. The maximum possible imaging speed of WOMiM was determined by the relaxation time of optothermal heat, measured to be 32.8 μs in water, corresponding to a frame rate of ∼30 kHz. This proof-of-concept demonstrates that vibrational imaging can be achieved at an unprecedented imaging speed and large FOV with the potential to significantly facilitate label-free imaging of cellular dynamics.

Identification of (Tb,Eu)9.43(SiO4)6O2-δ Oxy-Apatite Structures as Nanometric Inclusions in Annealed (Eu,Tb)-Doped ZnO/Si Junctions: Combined Electron Diffraction and Chemical Contrast Imaging Studies.

(Tb,Eu)-doped ZnO-annealed films at 1100 °C showed intense photoluminescense (PL) emission from Eu and Tb ions. The high-temperature annealing led to a chemical segregation and a secondary Zn-free phase formation that is suspected to be responsible for the high PL intensity. Large faceted inclusions of rare-earth (RE) silicates of a size of few hundred nanometers were observed. Owing to various advanced electron microscopy techniques, a detailed microstructural study of these nanometric inclusions combining atomic Z contrast imaging (STEM) and precession electron diffraction tomography (PEDT) data was carried out and resulted in the determination of a hexagonal P63/m-type (Tb,Eu)9.43(SiO4)6O2-δ structure related to an oxy-apatite structure. Chemical analyses from spectroscopic data (energy-dispersive X-ray mapping and electron energy loss spectroscopy) at the atomic scale showed that both RE elements sitting on two independent (4f) and (6h) atomic sites have three-fold oxidation states, while refinements of their occupancy sites from PEDT data have evidenced preferential deficiency for the first one. The deduced RE-O distances and their corresponding bond valences are listed and discussed with the efficient energy transfer from Tb3+ toward Eu3+.

Simultaneous Scanning Electron Microscope Imaging of Topographical and Chemical Contrast Using In-Lens, In-Column, and Everhart-Thornley Detector Systems.

The scanning electron microscope provides a platform for subnanometer resolution characterization of material morphology with excellent topographic and chemical contrast dependent on the used detectors. For imaging applications, the predominantly utilized signals are secondary electrons (SEs) and backscattered electrons (BSEs) that are emitted from the sample surface. Recent advances in detector technology beyond the traditional Everhart-Thornley geometry have enabled the simultaneous acquisition and discrimination of SE and BSE signals. This study demonstrates the imaging capabilities of a recently introduced new detector system that consists of the combination of two in-lens (I-L) detectors and one in-column (I-C) detector. Coupled with biasing the sample stage to reduce electron-specimen interaction volumes, this trinity of detector geometry allows simultaneous acquisition of signals to distinguish chemical contrast from topographical changes of the sample, including the identification of surface contamination. The I-C detector provides 4× improved topography, whereas the I-L detector closest to the sample offers excellent simultaneous chemical contrast imaging while not limiting the minimization of working distance to obtain optimal lateral resolution. Imaging capabilities and contrast mechanisms for all three detectors are discussed quantitatively in direct comparison to each other and the conventional Everhart-Thornley detector.

Quantitative metrics for assessment of chemical image quality and spatial resolution.

Currently objective/quantitative descriptions of the quality and spatial resolution of mass spectrometry derived chemical images are not standardized. Development of these standardized metrics is required to objectively describe the chemical imaging capabilities of existing and/or new mass spectrometry imaging technologies. Such metrics would allow unbiased judgment of intra-laboratory advancement and/or inter-laboratory comparison for these technologies if used together with standardized surfaces. Two image metrics, viz., "chemical image contrast" (ChemIC) based on signal-to-noise related statistical measures on chemical image pixels and "corrected resolving power factor" (cRPF) constructed from statistical analysis of mass-to-charge chronograms across features of interest in an image, were developed. These metrics, quantifying chemical image quality and spatial resolution, respectively, were used to evaluate chemical images of a model photoresist patterned surface collected using a laser ablation/liquid vortex capture mass spectrometry imaging system under different instrument operational parameters. The calculated ChemIC and cRPF metrics determined in an unbiased fashion the relative ranking of chemical image quality obtained with the laser ablation/liquid vortex capture mass spectrometry imaging system. These rankings were used to show that both chemical image contrast and spatial resolution deteriorated with increasing surface scan speed, increased lane spacing and decreasing size of surface features. ChemIC and cRPF, respectively, were developed and successfully applied for the objective description of chemical image quality and spatial resolution of chemical images collected from model surfaces using a laser ablation/liquid vortex capture mass spectrometry imaging system. Published in 2016. This article is a U.S. Government work and is in the public domain in the USA.

Super-Resolution Chemical Imaging with Plasmonic Substrates

We demonstrate super-resolution chemical imaging with plasmonic nanoholes via surface-enhanced Raman spectroscopy (SERS). Due to large field enhancements, blinking behavior of SERS hot spots was observed and processed using a stochastic optical reconstruction microscopy (STORM) algorithm. This enabled localization to within 10 nm and high-resolution imaging. However, illumination of the sample with a static laser beam produced only SERS hot spots in fixed locations, leaving noticeable gaps in the final images. By randomly altering the phase profile of the incident beam with a simple optical diffuser, the hot spots were shifted across the plasmonic surface to illuminate different areas of the sample, thereby rendering a final image without the gaps. A tunable band-pass filter was used to preserve spectral information, allowing chemical contrast imaging. Images were then compared to those obtained with a scanning electron microscope. Finally, we show that super-resolution SERS images can also be obtained wi...

Stimulated Raman photoacoustic spectroscopy for chemical-contrast imaging of a sample deeply buried in scattering media.

The development of a stimulated Raman scattering photoacoustic (SRS-PA) spectrometer is presented. In the apparatus, a molecular vibrational mode is excited by the SRS process. The vibrational excitation energy is converted to heat by vibrational relaxation. The volumes around the excited molecules including the surrounding solvent molecules expand by heating, resulting in the generation of an ultrasonic wave. The ultrasonic wave can be used as a molecular-selective signal. Because the ultrasonic wave is scarcely scattered by media, SRS-PA is expected to be applied for obtaining molecular-selective signals from deeply buried samples. In the present study, a SRS-PA spectrometer was developed and applied to obtain molecular-selective signals from test samples. The SRS-PA signals from water and lipid, which are important components in biological systems, were first obtained. The SRS-PA signal from a polystyrene film buried in a highly light-scattering intralipid suspension was also measured. We succeeded in obtaining the signal from the film when it was buried with a depth of up to 1.8 mm. The results indicate that SRS-PA can be effectively applied for the chemical-contrast imaging of deeply buried samples.

X-ray absorption near-edge structure (XANES) spectroscopy identifies differential sulfur speciation in corneal tissue

The chemical composition of tissues can influence their form and function. As a prime example, the lattice-like arrangement of collagen fibrils required for corneal transparency is controlled, in part, by sulfated proteoglycans, which, via core proteins, bind to the collagen at specific locations along the fibril axis. However, to date, no studies have been able to directly identify and characterize sulfur (S) in the cornea as a function of tissue location. In this study, X-ray absorption near-edge structure spectroscopy and micro-beam X-ray fluorescence (μ-XRF) chemical contrast imaging were employed to probe the nature of the mature (bovine) cornea as a function of position from the anterior sub-epithelial region into the deep stroma. Data indicate an inhomogeneity in the composition of S species in the first ≈50μm of stromal depth. In μ-XRF chemical contrast imaging, S did not co-localize with phosphorous (P) in the deep stroma where sulfates are prominent. Rather, P is present only as isolated micrometric spots, presumably identifiable as keratocytes. This study lends novel insights into the elemental physiology of mature cornea, especially in relation to its S distribution; future studies could be applied to human tissues. Moreover, it defines an analytical protocol for the interrogation of S species in biological tissues with micrometric resolution.

TOF-SIMS analysis of polystyrene/polybutadiene blend using chemical derivatization and multivariate analysis

Chemical imaging with high spatial resolution is one of the features of TOF-SIMS. However, degradation of the sample due to primary ion bombardment becomes problematic when the analysis area is small. Although polystyrene (PS) and polybutadiene (PB) separately show relatively distinct spectra, observation of their phase separation in PS/PB blends is difficult when the analysis area is small because degradation of both polymers and especially PS leads to disappearance of their characteristic peaks, resulting in low chemical image contrast. We therefore investigated the application of various forms of multivariate analysis (MVA) to the TOF-SIMS image data to improve the chemical image contrast. PCA, MCR, and the other forms of MVA provided improvement in contrast, but the images were still obscure and observation of phase separation remained difficult. Chemical derivatization using osmium tetroxide was also investigated, and found to give clear images of phase separation in the PS/PB blend. In quantitative determinations with MVA and chemical derivatization, PLS demonstrated the best predictive capability and chemical derivatization resulted in large deviations from both the bulk chemical composition and the determinations with MVA, particularly in regions of low PB content.

Chemical Patterning and Chemical Contrast Imaging with the Tip of an Atomic Force Microscope

Extended abstract of a paper presented at MC 2007, 33rd DGE Conference in Saarbrücken, Germany, September 2 – September 7, 2007.

A Spectral Identity Mapper for Chemical Image Analysis

Generating chemically relevant image contrast from spectral image data requires multivariate processing algorithms that can categorize spectra according to shape. Conventional chemometric techniques like inverse least squares, classical least squares, multiple linear regression, principle component regression, and multivariate curve resolution are effective for predicting the chemical composition of samples having known constituents, but they are less effective when a priori information about the sample is unavailable. We have developed a multivariate technique called spectral identity mapping (SIM) that reduces the dependence of spectral image analysis on training datasets. The qualitative SIM method provides enhanced spectral shape specificity and improved chemical image contrast. We present SIM results of spectral image data acquired from polymer-coated paper substrates used in the manufacture of pressure sensitive adhesive tapes. In addition, we compare the SIM results to results from spectral angle mapping (SAM) and cosine correlation analysis (CCA), two closely related techniques.

Chemical analysis by high-angle hollow cone illumination

The high-angle hollow cone illumination realized in a conventional transmission microscope is known to suppress coherent contrast mechanisms. It is shown that under suitable experimental conditions, orientation and strain contrasts are sufficiently suppressed to obtain a Z-contrast image. Various factors, influencing the image intensity, are determined by a quantitative image analysis of pure metals. Based on these measurements an incoherent multiscattering model is generalized to binary alloys in order to describe the chemical image contrast and to determine the local composition of the specimen from an intensity measurement. To test the concept, the early stages of interdiffusion between thin films of Cu and Au are investigated. The results demonstrate that a useful chemical analysis is possible, even in the case of nanocrystalline specimens. Complete composition profiles are determined on a length scale of 10 nm. The spatial and chemical resolution of the method are discussed.

Raman Chemical Imaging of Breast Tissue

Raman chemical imaging of lipid and protein distribution in breast is performed without the use of invasive contrast agents. Instead, tissue component discrimination is based on the unique vibrational spectra intrinsic to lipids and proteins. Component discrimination is possible using ratiometric image analysis techniques. However, chemical image contrast is enhanced when techniques are employed that use the entire Raman spectrum, including peak shift analysis and multivariate image analysis. Visualization of breast tissue components is an essential step in the development of a quantitative Raman ‘optical biopsy’ technique suitable for the non-invasive detection and classification of breast cancer. A high spatial/spectral resolution Raman imaging microscope is used that employs a liquid crystal tunable filter (LCTF). The imaging performance of the Raman microscope is quantitated with the measurement of the imaging system’s contrast transfer function (CTF). A comparison between the theoretical (diffraction limited) CTF and the measured CTF establishes that the LCTF does not produce appreciable image degradation. © 1997 by John Wiley & Sons, Ltd.

Raman Chemical Imaging: Noninvasive Visualization of Polymer Blend Architecture

A generally applicable methodology for routine, noninvasive chemical image characterization of materials is described. The methodology is based on Raman imaging microscopy employing a solid-state acoustooptic electronically tunable imaging spectrometer integrated within a laser-excited optical microscope. Multichannel charge-coupled device detection is employed to capture the Raman images. Chemical imaging with electronically tunable filters can be performed automatically under computer control, making analysis of unknowns routine. The Raman chemical imaging system is applied to the visualization of polypropylene/polyurethane blends. Raman chemical imaging is complementary to electron microscopy for polymer blend characterization ; however, Raman imaging does not require as extensive sample preparation. Raman imaging provides images having molecular specificity at the spatial resolution of the optical microscope, without the use of dyes or stains. Chemical image contrast is provided by the intrinsic vibrational signature of the polymer constituents in the blend.