A thorough understanding of biological species and emerging nanomaterials requires, among other efforts, their in-depth characterization through optical techniques capable of nanoresolution. Nanoscopy techniques based on tip-enhanced optical effects have gained tremendous interest over the past years, given their potential to obtain optical information with resolutions limited only by the size of a sharp probe interacting with focused light, irrespective of the illumination wavelength. Although their popularity and number of applications is rising, tip-enhanced nanoscopy (TEN) techniques still largely rely on probes that are not specifically developed for such applications, but for atomic force microscopy. This limits their potential in many regards, e.g., in terms of signal-to-noise ratio, attainable image quality, or extent of applications. We take the first steps toward next-generation TEN by demonstrating the fabrication and modeling of specialized TEN probes with known optical properties. The proposed framework is highly flexible and can be easily adjusted to be used with diverse TEN techniques, building on various concepts and phenomena, significantly augmenting their function. Probes with known optical properties could potentially enable faster and more accurate imaging via different routes, such as direct signal enhancement or facile and ultrafast optical signal modulation. We consider that the reported development can pave the way for a vast number of novel TEN imaging protocols and applications, given the many advantages that it offers.
Scattering-type Scanning Near Field Optical Microscopy (s-SNOM) has been demonstrated as a valuable tool for revealing important properties of materials at nanoscale. Recent proof-of-concept experiments have shown that, among others, s-SNOM can provide quantitative information on the real and imaginary parts of the dielectric function, and hence of intrinsic optical properties of materials and biological samples. In this work we further explored these capabilities in several experiments dealing with microcapsules for drug delivery, ultra-thin optical coatings with tunable color properties, and two types of nanoparticles with important applications in energy storage and conversion, or biosensing and theranostics.
Autofluorescence, diffuse-reflectance and transmission spectral, and microscopic measurements were made on different cutaneous neoplastic lesions, namely basal cell carcinoma, squamous cell carcinoma, malignant melanoma, and dysplastic and benign lesions related. Spectroscopic measurements were made on ex vivo tissue samples, and confocal microscopy investigations were made on thin tissue slices.
Fluorescence spectra obtained reveal statistically significant differences between the different benign, dysplastic and malignant lesions by the level of emission intensity, as well by spectral shape, which are fingerprints applicable for differentiation algorithms. In reflectance mode the most significant differences are related to the influence of skin pigments – melanin and hemoglobin. Transmission spectroscopy mode gave complementary optical properties information about the tissue samples investigated to that one of reflectance and absorption spectroscopy.
Using autofluorescence detection of skin lesions we obtain very good diagnostic performance for distinguishing of nonmelanoma lesions. Using diffuse reflectance and transmission spectroscopy we obtain significant tool for pigmented pathologies differentiation, but it is a tool with moderate sensitivity for non-melanoma lesions detection. One could rapidly increase the diagnostic accuracy of the received combined “optical biopsy” method when several spectral detection techniques are applied in common algorithm for lesions’ differentiation.
Specific spectral features observed in each type of lesion investigated on micro and macro level would be presented and discussed. Correlation between the spectral data received and the microscopic features observed would be discussed in the report.
The standard procedure for cancer detection includes rigorous biopsy protocols, which are costly and time consuming; also the accuracy of the current diagnostic procedure relays entirely on the physician’s experience and it is limited by the high probability of miss rates. Therefore new sensitive diagnostic modalities for analysis of biopsy tissue samples or on site, in vivo microscopy tissue examination, are necessary.
In this study we present an investigation using nonlinear microscopy techniques for histological sections from biopsy tissue samples analysis. The samples were routinely processed for histological analysis and during the standard sampling the tissue slices were stained with hematoxylin and eosin dyes.
The application of nonlinear microscopy techniques, such as two photon excitation fluorescence microscopy and second harmonic generation microscopy in biomedical research for cancer diagnosis has been vastly expanding in the last few years. Two-photon excitation fluorescence microscopy is based on a non-linear optical effect of simultaneously absorption of two photons, thus achieves excited state of the absorbing molecule with energy corresponding to the sum of the energies of two incident photons. This method allows for using an excitation wavelength which is double the typically required one for excitation of diagnostically valuable endogenous fluorophores. This results in more efficient depth penetration of the longer wavelength light in the tissue. The second harmonic generation microscopy is based on the principle of the non-linear susceptibility in noncentrosymmetric structures; such structures in the tissue are formed mainly by the collagen fibers. After excitation with near-infrared photons with wavelength λ of the collagen structures, photons with wavelength ½ λ are emitted — this corresponding to the second harmonic of the excitation beam’s frequency.
The applied nonlinear microscopy techniques are suitable for detection and quantification of the morphological changes associated with stroma and epithelial transformation in colon cancer, providing complementary information about the tissue microstructure and displaying distinctive patterns between normal and malignant human colon tissues.
Proper calibration of any instrument is vital to an investigator's ability to compare laboratory experiments, as well as to
draw quantitative relations between experimental results and the real objects. Traceability is a term used to certify an
instrument's accuracy relative to a known standard. Because traceability to meter is a very expensive and complicated
process, accurate and traceable calibration of lateral and vertical standards (e.g. 1D and 2D gratings) is a basic
metrological task for nano- and micro- technology. On the other hand laser interferometry is the de facto method to
transfer the meter standard to practical measurement. In this lecture, we describe interferometric vertical and lateral
calibration of a grating used to quantify the parameters necessary for proper translation of AFM data into physically
meaningful data.
Some dental ceramics were coated with a bioactive glass and resulted the formation of a stable and well bonded
with the ceramic substrate thin layer. After immersion in a solution with ion concentrations similar to those of
human blood plasma the development of hydroxy carbonate apatite layer on the surface of bioactive glass may be
observed. The objective of this study was to investigate structural surface changes of bioactive glass, after
exposure in a simulated body fluid for a different number of days. The roughness and topography of the
hydroxyapatite surface were investigated by Confocal Scanning Laser Microscopy. The chemical composition
was analyzed by Energy Dispersive Spectroscopy measurements.
The compound Mercuric Bromoiodide (HgBrxI2-x) belongs to the HgI2-HgBr2 system. It is promising for the same applications as HgI2, that is as solid state X and (gamma) detector, operating at room temperature since it exhibits a number of properties which are necessary for such applications, i.e. large atomic number, large band gap (2.4 - 3.4 eV) and high resistivity. Using a Confocal Laser Scanning Microscope we made some investigations regarding topographical distribution of impurities or related defects. Analysis of chemical composition of HgBrxI2-x crystals were performed on a SEM model Phillips 515 equipped with Energy Dispersive X-ray Analysis.
A scanning laser computer assisted microscope in which the sample is scanned electromagnetic is used to produce images from semiconductor devices using photo induced current. We obtained 2D and 3D images which were analyzed in connection with current-reverse voltage (I-VR) characteristics of the semiconductor devices. Images are stored in digital memories in real time and then processed with the different programs in order to obtain the required information. Our scanner offers the possibility 2D or 3D images ('map' or graphs) and pixelated images too. The localization of the different defects of the semiconductor devices was made.
Scanning optical microscopy computer assisted has the advantages that a wide range of imaging modes is possible. We have constructed a new computer assisted laser scanning which offers the possibility to obtain 2D and 3D images. The paper presents the principal hardware and software characteristics of the scanner.
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