Three-dimensional total internal reflection microscopy

We investigate the inverse-scattering problem that arises in total internal reflection microscopy. An analytic solution to this problem within the weak-scattering approximation is used to develop a novel form of three-dimensional microscopy with subwavelength resolution.     

Two-photon fluorescence scanning near-field microscopy based on a focused evanescent field under total internal reflection

We present two-photon fluorescence near-field microscopy based on an evanescent field focus produced by a ring beam under total internal reflection. The evanescent field produced by this method is focused by a high-numerical-aperture objective, producing a tightly confined volume that can effectively induce two-photon excitation. The imaging system is characterized by the two-photon-excited images of the nanocrystals, which show that the focused evanescent field is split into two lobes because of the enhancement of the longitudinal polarization component at the focus. This feature is confirmed by the theoretical prediction. Unlike other two-photon near-field probes, this method does not have the heating effect and requires no control mechanism of the distance between a sample and the probe.     

Optimization of a total internal reflection lens by using a hybrid Taguchi-simulated annealing algorithm

In this paper, we propose a new method for optimization of a total internal reflection (TIR) lens by using a hybrid Taguchi-simulated annealing algorithm. The conventional simulated annealing (SA) algorithm is a method for solving global optimization problems and has also been used in non-imaging systems in recent years. However, the success of SA depends heavily on the annealing schedule and initial parameter setting. In this study, we successfully incorporated the Taguchi method into the SA algorithm. The new hybrid Taguchi-simulated annealing algorithm provides more precise search results and has lower initial parameter dependence.     

Subdiffraction resolution in total internal reflection fluorescence microscopy with a grating substrate

We propose a fluorescence surface imaging system that presents a power of resolution beyond that of the diffraction limit without resorting to saturation effects or probe scanning. This is achieved by depositing the sample on an optimized periodically nanostructured substrate in a standard total internal reflection fluorescence microscope. The grating generates a high-spatial-frequency light grid that can be moved throughout the sample by changing the incident angle. An appropriate reconstruction procedure permits one to recover the fluorescence amplitude from the images obtained for various incidences. Simulations of this imaging system show that the resolution is not limited by diffraction but by the period of the grating.     

Surface‐sensitive Raman microscopy with total internal reflection illumination

A Raman microscope using a total internal reflection (TIR) annular illumination geometry through a ZnSe solid immersion lens (SIL) is described. Spectra of a thin‐film sample of the transparent organic conductor poly(3,4‐ethylenedioxythiophene) poly(styrenesulfonate) (PEDOT:PSS) on a polyethylene terephthalate (PET) substrate are presented and compared with those from a conventional confocal Raman configuration. These spectra demonstrate a significant increase in surface selectivity upon the use of TIR illumination, as the decay length of the evanescent excitation field limits the depth of sample probed in this configuration. Spectral interference from the underlying PET substrate layer is thus greatly reduced. An increase in surface selectivity is also demonstrated for spectra acquired through the SIL with uniform illumination. Raman images of a micropatterned PEDOT:PSS film acquired with TIR illumination are also reported. Enhanced lateral resolution is realized in this configuration because of the immersion effect of the SIL, and the sampling depth is limited to 150 nm by the choice of illumination geometry. This results in analysis volumes on the order of tens of femtoliters, nearly two orders of magnitude smaller than typically achieved in conventional confocal Raman microscopes. This approach yields Raman spectra and images with surface selectivity significantly greater than can be achieved in confocal Raman, and provides a valuable tool for the microanalysis of thin surface films. Published in 2010 by John Wiley & Sons, Ltd.     

Effects of using a metal layer in total internal reflection fluorescence microscopy

This proceeding examines the characteristics of imaging through a metal-coated glass cover slip using total internal reflection fluorescence microscopy (TIRFM). Through back and front focal plane imaging of sub-diffraction-limited fluorescent beads, the experimental characteristics of the emission are compared with theoretical simulations. Furthermore, at the angle at which surface plasmon resonance occurs, we show that the anisotropic emission of the fluorescent beads collected through the metal layer results in a irregular point spread function that has a donut-like structure with multiple concentric rings. PACS 42.30.Va; 73.20.Mf     

Adsorption of firefly luciferase at interfaces studied by total internal reflection fluorescence spectroscopy

The adsorption of luciferase onto silica surfaces was studied by total internal reflection fluorescence (TIRF) spectroscopy. Two model surfaces were used: hydrophilic and hydrophobic silica. Luciferase adsorbed differently on these two surfaces. Initial kinetics of luciferase adsorption onto the hydrophilic surface showed that luciferase adsorbs over an adsorption energy barrier of 3 kT The quantum yield of luciferase fluorescence decreased at the hydrophilic silica surface, which indicated that the protein conformation was altered during adsorption. Luciferase adsorption onto the hydrophobic silica surface proceeded with a small adsorption energy barrier and the fluorescence efficiency of adsorbed protein remained unchanged after adsorption. The affinity of luciferase for luciferin was measured using quenching of luciferase fluorescence with luciferin. The binding constant of the adsorbed luciferase-luciferin complex at the hydrophilic silica surface was two orders of magnitude smaller than the respective binding constant in the solution. Adsorbed luciferase showed an absence of ATP-dependent visible luminescence, indicating that the adsorbed enzyme was not active at either of the two silica surfaces.     

Structured illumination in total internal reflection fluorescence microscopy using a spatial light modulator

In wide-field fluorescence microscopy, illuminating the specimen with evanescent standing waves increases lateral resolution more than twofold. We report a versatile setup for standing-wave illumination in total internal reflection fluorescence microscopy. An adjustable diffraction grating written on a phase-only spatial light modulator controls the illumination field. Selecting appropriate diffraction orders and displaying a sheared (tilted) diffraction grating allows one to tune the penetration depth in very fine steps. The setup achieves 91 nm lateral resolution for green emission.     

Measurement of the thickness of thin polymer layers by infrared frustrated total internal reflection spectroscopy

A method is proposed for determining the thickness of thin (0.1–5.0 μm) polymer layers and coatings by means of infrared frustrated total internal reflection (FTIR) spectroscopy. This method is based on an analytic expression derived for the dependence of the intensities of absorption bands in IR FTIR spectra on the thickness of a polymer layer. The method is tested on model samples consisting of a thick film of polyethylene terephthalate with a layer of different thicknesses of polystyrene deposited on it. The advantage of this method is the ability to determine the thickness of thin polymer layers deposited on bulk or opaque polymer substrates. Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 75, No. 6, pp. 881–885, November–December, 2008.     

X-ray absorption spectroscopy in total electron yield mode of scanning photoelectron microscopy

Total electron yield (TEY) measurement was applied to the scanning photoelectron microscopy (SPEM). The resultant image showed the thickness variation of a zinc overlayer deposited on an iron substrate with 18 nm nominal thickness. The contrast and signal-to-noise ratio of the image were much higher than those of the images obtained by the conventional SPEM. When the order-sorting aperture (OSA) located between the Fresnel zone plate and the sample was biased to +80 V relative to the sample, the contrast of the image was further improved. It also made it possible to perform microscopic X-ray absorption spectroscopy in TEY mode by preventing the unwanted photoelectrons emitted from the OSA, which can cause false TEY, from reaching the sample. Oxidized areas of the iron substrate under the thick zinc overlayer were clearly identified.     

Local thermal conductivity of polycrystalline AlN ceramics measured by scanning thermal microscopy and complementary scanning electron microscopy techniques

The local thermal conductivity of polycrystalline aluminum nitride (AlN) ceramics is measured and imaged by using a scanning thermal microscope (SThM) and complementary scanning electron microscope (SEM) based techniques at room temperature.The quantitative thermal conductivity for the AlN sample is gained by using a SThM with a spatial resolution of sub-micrometer scale through using the 3ω method.A thermal conductivity of 308 W/m·K within grains corresponding to that of high-purity single crystal AlN is obtained.The slight differences in thermal conduction between the adjacent grains are found to result from crystallographic misorientations,as demonstrated in the electron backscattered diffraction.A much lower thermal conductivity at the grain boundary is due to impurities and defects enriched in these sites,as indicated by energy dispersive X-ray spectroscopy.     

Local thermal conductivity of polycrystalline AlN ceramics measured by scanning thermal microscopy and complementary scanning electron microscopy techniques

The local thermal conductivity of polycrystalline aluminum nitride (AlN) ceramics is measured and imaged by using a scanning thermal microscope (SThM) and complementary scanning electron microscope (SEM) based techniques at room temperature. The quantitative thermal conductivity for the AlN sample is gained by using a SThM with a spatial resolution of sub-micrometer scale through using the 3ω method. A thermal conductivity of 308 W/m·K within grains corresponding to that of high-purity single crystal AlN is obtained. The slight differences in thermal conduction between the adjacent grains are found to result from crystallographic misorientations, as demonstrated in the electron backscattered diffraction. A much lower thermal conductivity at the grain boundary is due to impurities and defects enriched in these sites, as indicated by energy dispersive X-ray spectroscopy.     

Extended resolution wide-field optical imaging: objective-launched standing-wave total internal reflection fluorescence microscopy

Standing-wave total-internal-reflection fluorescence (SW-TIRF) microscopy uses a super-diffraction-limited standing evanescent wave to extract the high-spatial-frequency content of an object through a diffraction-limited optical imaging system. The effective point-spread function is better than a quarter of the emission wavelength. With a 1.45 numerical aperture objective and 532 nm excitation wavelength, a Rayleigh resolution of approximately 100 nm can be achieved, which is better than twice the resolution of conventional TIRF microscopy. This first experimental realization of SW-TIRF in an objective-launched geometry demonstrates the potential for extended resolution imaging at high speed by using wide-field microscopy.     

全内反射荧光显微术应用于单分子荧光的纵向成像

利用这种显微术中激发场的纵向强度呈指数衰减的分布特性，测得的荧光强 度将强烈依赖渗透深度（强度衰减到1/e时的距离），从而快速直接的确定单分子荧光 团间的纵向间隔、每个荧光团的纵向绝对位置和荧光团的半径大小，即实现荧光分子三维分 布的重构.在整个重构的过程中，只需要改变一次入射角的大小，即只需探测两幅荧光分子 的全内反射成像. 关键词： 全内反射荧光显微术 纵向重构     

Frustrated total internal reflection from thin-layer structures with a metal film

Optics and Spectroscopy - Formulas of the model of a conducting surface for calculating the reflection coefficient and phase shift of light reflected from a thin metal film at the interface between...     

Determining optical constants by frustrated total internal reflection spectrophotometry

The possibility of using the method of frustrated total internal reflection (the two-media method) to determine the optical constants n and k in the value ranges 1. 3相似文献