Confocal three‐dimensional scanning laser Raman–SERS–fluorescence microprobe. Spectral imaging and high‐resolution applications

Abstract

A multi-purpose instrument which allows the recording of confocal micro-Raman, micro-SERS and microfluorescence spectral images of sample areas from 5 x 5 to 150 x 150 pm with a lateral resolution of cu. 0.3 pm and an axial resolution of cu. 1 pm was developed. The instrument is a combination of two spectrometers both coupled to the same microscope, motorized sample stage, confocal entrance chamber, macro-sample chamber and CCD detector. The first spectrometer includes a double monochromator coupled with a spectrograph and exhibits the properties of a typical high-resolution Raman instrument permitting measurements of the low-wavenumber regions of the spectra. The second spectrometer includes a Notch filter and a spectrograph equipped with two interchangeable low-dispersion gratings and exhibits the properties of a high-luminosity spectrometer, suitable for low resolution, over a wide spectral range and highly sensitive micro-Raman and microfluorescence measurements. The choice of spectrometer most suitable for a particular application can be made automatically without additional prealignment of the system. The system of optical scanners operating in the confocal mode and two-dimensional CCD detection allow the accumulation of spectra from hundreds of points of the sample under the microscope simultaneously. A computercontrolled scanning sample stage and a system using a 'scanning line' of the laser beam allow fast recording of well resolved confocal spectral images (CSI) without sample degradation. Conventional images of species could be recorded with a TV-CCD camera through the microscope optics. The software supports all stages of CSI recording and allows the combined treatment of conventional and spectral images including their spatial calibration and conversion of spectral image into a conventional image, to assign point-bypoint the spectral data to the conventional image. The applicability of the instrumentation and techniques to the study of polymeric materials, industrial samples and fluid inclusions in minerals was demonstrated. Spectral images based on the micro-SERS analysis of an antitumour drug adsorbed on the hydrosol were recorded and are discussed in terms of their application to the micro-SERS imaging studies of living cells.

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“…For pure vaterite, no EPR signals were evidenced. All the signals NASSRALLAH-ABOUKAÏS: VATERITE observed on copper/vaterite samples correspond, without ambiguity, to isolated and clusters of Cu 2+ ions [15][16][17][18]. For the untreated sample, an EPR signal with the EPR parameter values g 1 =2.35, g ⊥ 1 =2.06, g iso1 =2.16, A 1 =134 G and A ⊥ 1 =15 G was obtained.…”

Advanced Optical Imaging of Endocytosis

“…Spectral imaging has been implemented in a variety of optical modalities for biological applications including visible reflectance (Zuzak, et al, 2002), fluorescence (Michalet, et al, 2003) and vibrational spectroscopies such IR absorption (Levin & Bhargava, 2005), Raman scattering (Christensen & Morris, 1998), and surface-enhanced Raman (SERS) (Sharonov, et al, 1994)), as well as in non-optical methods like mass spectrometry (Fletcher, et al, 2008). In practice, higher degrees of multiplexing, higher accuracy, and lower detection limits are achievable with spectral imaging due to the ability to implement multivariate analysis methods to identify and/or classify spectral signatures even in the presence of high degrees of spectral overlap from other labels and cellular autofluorescence (Mansfield, et al, 2005).…”

Geochemistry Features of Oil-Gas Inclusions and UV-VIS Micro-spectra Analysis of Single Oil-Gas Inclusions in Bai 95 Oil Well

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