Laser scanning gradient index optics microscope: submicrometre scale spectroscopy and imaging at cryogenic temperatures

A laser scanning far-field optical microscope for low-temperature imaging and spectroscopy based on gradient index optics is presented. A rod-shaped gradient index microlens is used as a zero-working-distance solid immersion objective lens. The obtained lateral resolution is 310 nm of the FWHM at a wavelength of 545 nm. A laser scanning mechanism located outside an optical cryostat enables one to achieve large scanning ranges independent of temperature. The use of the microscope for submicrometre-scale spectroscopy and low-temperature photochemistry performed on molecular J aggregates in thin polymer films is presented.     

Fluorescence imaging with a laser trapping scanning near-field optical microscope

We investigated fluorescence imaging using a near-field scanning optical microscope which uses a laser-stabilized gold nanoparticle as a near-field probe. This microscope is suitable for observations of biological specimens in aqueous solutions because the probe particle is held by a noncontact force exerted by a laser beam. Theoretical calculations based on Mie scattering theory are presented to evaluate the near-field enhancement by a gold particle of 40 nm diameter. We also present fluorescence images of a single fluorescent bead and discuss the near-field contribution to the fluorescence image in this type of microscope.     

Fluorescence imaging with a laser trapping scanning near-field optical microscope

We investigated fluorescence imaging using a near-field scanning optical microscope which uses a laser-stabilized gold nanoparticle as a near-field probe. This microscope is suitable for observations of biological specimens in aqueous solutions because the probe particle is held by a noncontact force exerted by a laser beam. Theoretical calculations based on Mie scattering theory are presented to evaluate the near-field enhancement by a gold particle of 40 nm diameter. We also present fluorescence images of a single fluorescent bead and discuss the near-field contribution to the fluorescence image in this type of microscope.     

Analysis of fluorescence from algae fossils of the Neoproterozoic Doushantuo formation of China by confocal laser scanning microscope

Chinese algae fossils can provide unique information about the evolution of the early life. Thin sections of Neoproterozoic algae fossils, from Guizhou, China, were studied by confocal laser scanning microscopy, and algae fossils were fluorescenced at different wavelengths when excited by laser light of 488 nm, 476 nm, and 568 nm wavelength. When illuminated by 488 nm laser light, images of the algae fossils were sharper and better defined than when illuminated by 476 nm and 568 nm laser light. The algae fossils fluoresce at a wide range of emission wavelengths. The three-dimensional images of the fluorescent algae fossils were compared with the transmission images taken by light microscope. We found that the fluorescence image of the confocal laser scanning microscope in a single optical section could pass for the transmission image taken by a light microscope. We collected images at different sample depths and made a three-dimensional reconstruction of the algae fossils. And on the basis of the reconstruction of the three-dimensional fluorescent images, we conclude that the two algae fossils in our present study are red algae.     

An evanescent-field optical microscope

The classic diffraction limit of resolution in optical microscopy (～γ/2) can be overcome by detecting the diffracted field of a submicrometre-size probe in its near field. The present stage of this so-called scanning near-field optical microscopy (SNOM) is reviewed. An evanescent-field optical microscope (EFOM) is presented in which the near-field regime is provided by the exponentially decaying evanescent field caused by total internal reflection at a refractive-index transition. A sample placed in this field causes a spatial variation of the evanescent field which is characteristic for the dielectric and topographic properties of the sample. The evanescent field is frustrated by a dielectric probe and thus converted into a radiative field. In our case the probe consists either of an etched optical fibre or of a highly sharpened diamond tip. The probe is scanned over the sample surface with nanometre precision using a piezo-electric positioner. The distance between probe and sample is controlled by a feedback on the detected optical signal. The resolution of the microscope is determined by both the gradient of the evanescent field and the sharpness of the tip. Details of the experimental set-up are discussed. The coupling of the evanescent field to the submicrometre probe as a function of probe-sample distance, angle of incidence and polarization has been characterized quantitatively. The observed coupling is generally in agreement with presented theoretical calculations. Microscopy has been performed on a regular latex sphere structure, which clearly demonstrates the capacity of the evanescent-field optical microscope for nanometre-scale optical imaging. Resolution is typically 100 nm laterally and 10 nm vertically. The technique is promising for biological applications, especially if combined with optical spectroscopy.     

Shear force imaging of DNA in a near-field scanning optical microscope (NSOM)

A near-field scanning optical module has been constructed as an accessory for a Nanoscope IIIa commercial scanning probe microscope. Distance feedback and topographic registration are accomplished with an uncoated optical fibre scanning tip by implementation of the shear force technique. The tip is driven by a piezoelectric actuator at a resonance frequency of 8–80 kHz. A laser diode beam is scattered by the tip and detected by a split photodiode, with lock-in detection of the difference signal. The amplitude ( r ) and phase (τ) responses were characterized as a function of the calibrated tip–sample separation. Using an r cos τ feedback signal, imaging of pUC18 relaxed circular plasmid DNA spread on mica precoated with cetylpyridinium chloride was achieved. The apparent width (28 ± 5 nm) was approximately four times that achieved by scanning force measurements with the same instrument; the apparent height of the DNA (0.6 ± 0.3 nm) was similar with the two techniques. These results demonstrate the applicability of the shear force signal for imaging biological macromolecules according to topography and in conjunction with the optical signals of a near-field scanning optical microscope (NSOM).     

Unique laser-scanning optical microscope for low-temperature imaging and spectroscopy

Low-temperature optical characterization of single quantum nanostructures can reveal detailed information on structure-dependent properties of these materials. We describe the development of a unique laser-scanning optical microscope capable of low-temperature single molecule/particle imaging and spectroscopy. Making use of the magnification of a microscope objective, the laser- scanning scheme of the present microscope allows for high-repeatability imaging over large sample areas. The microscope is utilized to measure the low-temperature Raman scattering spectra of individual single-walled carbon nanotubes and single molecule fluorescence spectra of conjugated polymers. The developed low-temperature microscope can be applied to study a wide array of nanomaterials at a single particle level.     

Real-time phase microscopy using a phase-lock interferometer

A phase-lock interference microscope (PLIM) has been designed and constructed. The instrument measures optical phase in real time with analog output voltages proportional to phase and position. The PLIM has been used to measure gradient index profiles in optical fibers, wall thicknesses of microballoons for laser induced fusion, the phase of biological specimens, and the surface finish of optical surfaces. The peak-to-peak noise of the instrument is lambda/50.     

A versatile multipurpose scanning probe microscope

A combined scanning probe microscope has been developed that allows simultaneous operation as a non‐contact/tapping mode atomic force microscope, a scattering near‐field optical microscope, and a scanning tunnelling microscope on conductive samples. The instrument is based on a commercial optical microscope. It operates with etched tungsten tips and exploits a tuning fork detection system for tip/sample distance control. The system has been tested on a p‐doped silicon substrate with aluminium depositions, being able to discriminate the two materials by the electrical and optical images with a lateral resolution of 130 nm.     

Fibre-optic two-photon scanning fluorescence microscopy

Two geometries of a novel two‐photon fluorescence microscope incorporating single‐mode fibre optics for the delivery of ultrashort‐pulsed illumination to a remote sample are characterized. First, a 785 nm single‐mode optical fibre is implemented in a scanning microscope, which demonstrates that an improvement in axial resolution is achieved due to the non‐linear response of the fibre to intense ultrashort‐pulsed light. Second, a 785 nm single‐mode optical fibre coupler is adapted, in which case spectral broadening and blue shifting of the ultrashort‐pulsed laser beam caused by the non‐linear response of the fibre to ultrashort‐pulsed illumination are experimentally characterized. An investigation into the impact of temporal broadening of the ultrashort‐pulsed beam on the systems is also considered. The coupling efficiency of both geometries for various illumination wavelengths is also presented. The introduction of the fibre coupler to the system has significant advantages, including an improved optical sectioning effect and a reduction in the number of bulk optical components resulting in a low‐cost, compact instrument. Sets of three‐dimensional images of fluorescent polymer microspheres and biological material confirm these features.     

An alternative flat scanner and micropositioning method for scanning probe microscope

An alternative flat scanner used for combining a scanning probe microscope with an inverted optical microscope is presented. The scanner has a novel structure basically consisting of eight identical piezoelectric tubes, metal flexure beams, and one sample mount. Because of the specially designed structure, the scanner is able to carry a sample of more than 120 g during imaging. By applying voltages of ±150 V, scanning range of more than 30 μm in three dimensions can be achieved. To improve the reliability of the stick-slip motion, a new method for sample micropositioning is proposed by applying a pulsed voltage to the piezotubes to produce a motion in the z-axis. Reliable translation of the sample has been thus accomplished with the step length from ～700 nm to 9 μm over a range of several millimeters. A homemade scanning probe microscope-inverted optical microscope system based on the scanner is described. Experimental results obtained with the system are shown.     

An optical spectrometer for a confocal scanning laser microscope

An optical spectrometer for the visible range has been developed for the confocal scanning laser microscope (CSLM) Phoibos 1000. The spectrometer records information from a single point or a user-defined region within the microscope specimen. A prism disperses the spectral components of the recorded light over a linear CCD photodiode array with 256 elements. A regulated cooling unit cools the diode array, thereby reducing the detector dark current to a level, which allows integration times of up to 60 s. The spectral resolving power, λ/Δλ, ranges from 400 at λ = 375 nm to 100 at λ = 700 nm. Since the entrance aperture of the spectrometer has the same diameter as the detector aperture of the CSLM, the three-dimensional spatial resolution for spectrometer readings is equivalent to that of conventional confocal scanning, that is, down to 0.2 μm lateral and 0.8 μm axial resolution with an N.A.=1.3 objective.     

A novel light source for SICM–SNOM of living cells

We have developed a novel light source for use in a scanning near‐field optical microscope (SNOM or NSOM) based on a nanopipette whose distance from the sample surface is controlled using scanning ion conductance microscopy. The light source is based on the general principle of the chemical reaction between a fluorophore in the pipette and ligand in the bath, to produce a highly fluorescent complex that is continually renewed at the pipette tip. In these experiments we used fluo‐3 and calcium, respectively. This complex is then excited with an Ar⁺ laser, focused on the pipette tip, to produce the light source. This method overcomes the transmission problem of more traditional SNOM probes and has been used to acquire simultaneous high‐resolution topographic and optical images of biological samples in physiological buffer. A resolution of ～220 nm topographic and ～190 nm optical was determined through imaging fixed sea‐urchin sperm flagella. Live A6 cells were also imaged, demonstrating the potential of this system for SNOM imaging of living cells.     

面向微纳材料的激光扫描二维成像系统

在对微纳材料光学特性表征中,需要获得分辨率更高的波长和强度的荧光图像。普通的显微镜无法满足测试的要求,因此将普通的成像显微镜、光谱仪以及纳米移动台组成激光扫描显微镜成像系统,并利用LabVIEW开发了一套完整的集二维扫描采集与信号图像处理一体的系统上位机软件。扫描采集过程使用了低通滤波等数字信号处理方法消除光谱仪信号噪声的影响。利用本系统测量硒化镉纳米带、单层二硫化钼得到了荧光强度图像以及荧光峰值波长图像,能分辨出最小波长为0.03 nm的荧光。将采集长度与实际长度进行比较并分析荧光强度差异,取得了较好的效果。     

The tetrahedral tip as a probe for scanning near-field optical microscopy at 30 nm resolution

The tetrahedral tip is introduced as a new type of a probe for scanning near-field optical microscopy (SNOM). Probe fabrication, its integration into a scheme of an inverted photon scanning tunnelling microscope and imaging at 30 nm resolution are shown. A purely optical signal is used for feedback control of the distance of the scanning tip to the sample, thus avoiding a convolution of the SNOM image with other simultaneous imaging modes such as force microscopy. The advantages of this probe seem to be a very high efficiency and its potential for SNOM at high lateral resolution below 30 nm.     

Mirrorlike pulsed laser deposited tungsten thin film

Mirrorlike tungsten thin films on stainless steel substrate deposited via pulsed laser deposition technique in vacuum (10(-5) Torr) is reported, which may find direct application as first mirror in fusion devices. The crystal structure of tungsten film is analyzed using x-ray diffraction pattern, surface morphology of the tungsten films is studied with scanning electron microscope and atomic force microscope. The film composition is identified using energy dispersive x-ray. The specular and diffuse reflectivities with respect to stainless steel substrate of the tungsten films are recorded with FTIR spectra. The thickness and the optical quality of pulsed laser deposition deposited films are tested via interferometric technique. The reflectivity is approaching about that of the bulk for the tungsten film of thickness ～782 nm.     

A confocal beam scanning white-light microscope

We report on a confocal beam scanning microscope utilizing a continuous Xe short-arc lamp operating in the visible spectrum with unprecedented radiance. Measurements of lateral and vertical resolution will be presented and compared with those of an equivalent scanning laser microscope. Resolution of the white-light microscope is equivalent to that of the scanning laser microscope. White-light microscope images positively stand out from those of the scanning laser microscope by their lack of artefacts caused by interference.     

Simple fiber-optic confocal microscopy with nanoscale depth resolution beyond the diffraction barrier

A novel fiber-optic confocal approach for ultrahigh depth-resolution (相似文献   

Fabrication of near-field optical apertures in aluminium by a highly selective corrosion process in the evanescent field

A simple, one‐step process to fabricate high‐quality apertures for scanning near‐field optical microscope probes based on aluminium‐coated silicon nitride cantilevers is presented. A thin evanescent optical field at a glass–water interface was used to heat the aluminium at the tip apex due to light absorption. The heat induced a breakdown of the passivating oxide layer and local corrosion of the metal, which selectively exposed the front‐most part of the probe tip from the aluminium. Apertures with a protruding silicon nitride tip up to 72 nm in height were fabricated. The height of the protrusion was controlled by the extent of the evanescent field, whereas the diameter depended on the geometry of the probe substrate. The corrosion process proved to be self‐terminating, yielding highly reproducible tip heights. Near‐field optical resolution in a transmission mode of 85 nm was demonstrated.     

Confocal laser scanning photoelasticity: Improvement of precision for quantitative photoelastic studies on model composite materials

Photoelastic studies on model composite materials with different model materials and load applications have been performed to understand the various failure mechanisms in real composites. Optical investigations of the stress fields have been conducted using a polarising, confocal laser scanning microscope (Zeiss LSM 410, Zeiss, Jena, Germany), where the phase of the isochromatic fringes has been determined from the data. Two different methods are presented to improve the quantitative evaluation of the isochromatic phase. By combining manual and automatic evaluation of fringes, even samples with a high stress gradient can be analysed. A method has been developed for the correction of the integrated through‐thickness birefringence, in the case of rotationally symmetric stress fields. This technique permits the inverse calculation of stress at various depths in the sample, in particular in the case of a nonlinear dependence of optical birefringence on stress. Both methods are demonstrated in the evaluation of tensile tests on multifibre model samples (glass fibres in polycarbonate).