Electron-Impact Induced Molecular Fluoresence Spectroscopy

Abstract

By the bombardment of gaseous molecules with monoenergetic electrons the molecules can be electronically excited and/or fragmented, depending upon the energy supplied by the electrons. Any fragments which may be produced also have the possibility of being produced in electronically (as well as vibrationally and rotationally) excited states. The electronically excited molecules and fragments both possess the possibility of radiative decay (fluorescence). The measurement of this fluorescence as a function of both electron energy and photon wavelength is termed “electron-impact induced fluorescence spectroscopy.” A plot of fluorescence intensity as a function of the electron energy at a given photon wavelength is generally referred to as an “excitation function,” which is exactly analogous to the excitation spectrum which can be obtained in conventional (optically excited) fluorescence spectrometry. A plot of the fluorescence intensity as a function of photon wavelength at a given electron energy is termed the “fluorescence spectrum.” A three dimensional plot of the fluorescence intensity as a function of both the photon wavelength and the electron energy may also be constructed, and is referred to as an “excitation contour plot.”     

Optically sectioned fluorescence lifetime imaging using a Nipkow disk microscope and a tunable ultrafast continuum excitation source

We demonstrate an optically sectioned fluorescence lifetime imaging microscope with a wide-field detector, using a convenient, continuously tunable (435-1150 nm) ultrafast source for fluorescence imaging applications that is derived from a visible supercontinuum generated in a microstructured fiber.     

Wide-field fluorescence sectioning with hybrid speckle and uniform-illumination microscopy

We describe a method of obtaining optical sectioning with a standard wide-field fluorescence microscope. The method involves acquiring two images, one with nonuniform illumination (in our case, speckle) and another with uniform illumination (in our case, randomized speckle). An evaluation of the local contrast in the speckle-illumination image provides an optically sectioned image with low resolution. This is complemented with high-resolution information obtained from the uniform-illumination image. A fusion of both images leads to a full resolution image that is optically sectioned across all spatial frequencies. This hybrid illumination method is fast, robust, and generalizable to a variety of illumination and imaging configurations.     

散斑照明宽场荧光层析显微成像技术研究

介绍了一种新的宽场荧光层析显微方法.在传统宽场显微镜中引入散斑图案照明样品,控制散斑图案的动态变化,利用CCD相机记录对应的一系列荧光图像.由于焦平面内强度变化远比焦平面外强度变化剧烈,通过合适的算法能够获得焦平面的层析分辨的荧光显微图像.标定了系统参数,并研究了不同的图像重建算法对系统性能的影响,获得了不同生物组织样品的层析图像.实验表明,该显微方法能用于组织光学切片成像,在临床医学中具有实际应用价值. 关键词： 荧光 散斑照明 荧光显微 层析     

Fine-structure spectroscopy of 2-methylnaphthalene cooled in a supersonic jet

The fluorescence and fluorescence excitation spectra of 2-methylnaphthalene molecules cooled in a supersonic jet are measured. The frequencies of vibrations in the S ₀ and S ₁ states, as well as the relative intensities of electronic-vibrational transitions in the fluorescence and fluorescence excitation spectra, are calculated with the semiempirical MO/M8ST method. The intensities are calculated in the Franck-Condon approximation taking into account the mixing of all the 38 totally symmetric normal vibrations. Based on the calculations, most observed spectral lines are assigned. It is shown that the calculation accuracy of the method is high enough for it to be used to interpret the spectra of molecules of aromatic compounds such as substituted naphthalenes. It is found that the main contribution to the fluorescence spectrum is made by four optically active vibrations.     

Image formation in structured illumination wide-field fluorescence microscopy

We present a theoretical analysis of the image formation in structured illumination wide-field fluorescence microscopy (SIWFFM). We show that the optically sectioned images obtained with this approach possess the optical sectioning strengths comparable to those obtained with the confocal microscope. We further show that the transfer function behaviour is directly comparable to that of the true confocal instrument. The theoretical considerations are compared with and confirmed by experimental results.     

Two-photon imaging to a depth of 1000 microm in living brains by use of a Ti:Al2O3 regenerative amplifier

It is shown that two-photon fluorescence images can be obtained throughout almost the entire gray matter of the mouse neocortex by using optically amplified femtosecond pulses. The achieved imaging depth approaches the theoretical limit set by excitation of out-of-focus fluorescence.     

Calculation of Transfer Functions of Multilayer Biotissues in the Problems of Correction of Their Fluorescence Spectra

A method for rapid calculation of a flux of stimulated fluorescence of a multilayer optically dense medium with inhomogeneous distribution of the fluorophore has been developed. The light field in the medium at the excitation wavelength of fluorescence is represented by a superposition of incident collimated, incident diffuse, and reflected diffuse fluxes. A two-stream approximation is used to describe the light field in the medium at the wavelength of emission of the fluorescence. Fluxes in adjacent elementary layers of the medium and on its surface are connected by simple matrix operators that are obtained using a combination of engineering approaches of radiation-transfer theory and single-scattering approximation. The calculations of fluorescence fluxes of a four-layer biotissue that are excited and recorded at 400–800 nm are compared with their Monte Carlo simulation with a discrepancy of 1%. The effect of the propagation medium on the fluorescence spectra of 5-ALA-induced protoporphyrin IX that are recorded from human skin was studied, and a technique for their correction that is based on measurements and quantitative analysis of the diffuse reflectance spectrum of skin was proposed.     

基于跃迁偶极矩取向特征研究乙醚溶液中分子的各向异性度

利用偏振荧光光谱和偏振激发光谱研究了乙醚溶液中荧光分子跃迁偶极距的取向特征.实验结果表明,在垂直线偏振光照射下,乙醚溶液发射出峰值位于305 nm的荧光谱,对应的最佳激励光波长为256 nm.由偏振激发光谱分析得到荧光体的吸收跃迁偶极矩和发射跃迁偶极矩间夹角α的变化规律,揭示了荧光去偏振过程：在粘性溶液中的荧光分子具有一定的偶极取向,α随激发光波长发生变化,当激发波长接近最佳激发光波长时,吸收跃迁偶极矩和发射跃迁偶极矩间趋于平行,荧光的退偏效果较弱,偏振度最大.研究结果能为分子空间取向特征的理论研究提供参考.     

Electron impact excitation of gas-phase thymine molecules

The electron impact excitation of gas-phase thymine molecules is studied optically in crossed electron and molecular beams. The emission spectrum of molecular bands is obtained and the bands are identified. The excitation functions of the most strong bands are measured.     

基于激光诱导荧光的油类污染物检测系统研究

油类污染物具有破坏海洋生态系统和间接污染大气及土壤的危害,快速、准确地检测污染物的成分及其浓度具有重要意义。由于油类污染物光谱重叠严重,因此难以通过传统荧光分析准确加以区分。本文基于激光诱导荧光技术,以氙灯作为激发光设计荧光光谱检测系统,并对0#柴油、92#汽油和煤油进行扫描和检测,从而获得激发/发射光谱以及最佳激发/发射波长。并对该系统的软件算法部分进行改进,运用Savitzky-Golay卷积平滑直接获得更加精确的激发/发射光谱,更能全面、准确地反映油类物质的荧光特性信息。并与传统的荧光光谱仪得到的光谱图进行对比,经实验验证激光诱导荧光技术的荧光光谱检测系统的有效性,对油类污染物的荧光光谱信号的检测具有更高的灵敏度。     

Optical sectioning in wide-field microscopy obtained by dynamic structured light illumination and detection based on a smart pixel detector array

Optical sectioning in wide-field microscopy is achieved by illumination of the object with a continuously moving single-spatial-frequency pattern and detecting the image with a smart pixel detector array. This detector performs an on-chip electronic signal processing that extracts the optically sectioned image. The optically sectioned image is directly observed in real time without any additional postprocessing.     

Autowave instability in refractive crystals

The intensities of vibronic lines are experimentally measured in fluorescence and fluorescence excitation spectra of jet-cooled anthracene. An original method is developed for calculating geometrical parameters of benzene hydrocarbons in the ground and excited electronic states. Using these parameters, the intensities of vibronic lines in fluorescence and absorption spectra of anthracene are calculated in the Franck-Condon approximation taking into account the mixing of all the twelve normal coordinates of totally symmetric vibrational modes. After correction for the quantum yield of fluorescence, good agreement is obtained between the calculated line intensities in the absorption spectrum and the measured line intensities in the fluorescence excitation spectrum. Based on these data, a new assignment of the lines in the fluorescence excitation spectrum corresponding to totally symmetric modes 7 and 8 is suggested.     

Agitation-free multiphoton microscopy of oblique planes

The scanning two-photon fluorescence microscope produces optically sectioned images from the focal plane. It is sometimes desirable to acquire images from other planes of the specimen that are inclined with respect to the focal plane. In this Letter, we discuss the issues concerned with acquiring such images together with the effects of the inclination angle on image resolution and sectioning strength. To obtain images from oblique planes at high speed, a two-photon system was built wherein a novel optical system is used to provide aberration-free scanning.     

Experimental determination and interpretation of the fluorescence and fluorescence excitation spectra of chrysene cooled in a supersonic jet

The fluorescence and fluorescence excitation spectra of jet-cooled chrysene are measured. The frequencies of in-plane vibrations in the ground and first excited singlet electronic states, as well as the relative intensities of transitions between them, are calculated with the MO/M8ST method. Based on these data, experimental spectra are interpreted. In the fluorescence excitation spectrum, the position of the line of the 0–0 transition (28 195 ± 1 cm^?1), which is the most intense, is determined. In the experimental fluorescence excitation spectrum, 21 lines correspond to fundamental vibrations (altogether, 37 lines are attributed). This supports our assignment and is consistent with the group-theoretical analysis of vibronic interactions. Upon excitation at the frequency of the 0–0 transition, 10 lines corresponding to the excitation of fundamental vibrations are detected, and all 17 lines observed are attributed. In the fluorescence excitation spectrum, the standard deviation between the calculated and measured frequencies of attributed fundamental vibrations is 19 cm^?1, while that in the fluorescence spectrum is 15 cm^?1.     

Simultaneous optically sectioned fluorescence and optical coherence microscopy with full-field illumination

Full-field optical coherence microscopy (FF-OCM) and optically sectioned fluorescence microscopy are two imaging techniques that are implemented here in a novel dual modality instrument. The two imaging modalities use a broad field illumination to acquire the entire field of view without raster scanning. Optical sectioning is achieved in both imaging modalities owing to the coherence gating property of light for FF-OCM, and a structured illumination setup for fluorescence microscopy. Complementary image data are provided by the dual modality instrument in the context of biological tissue screening. FF-OCM imaging modality shows the tissue microarchitecture, while fluorescence microscopy highlights specific tissue features with cellular-level resolution by using targeting contrast agents. Complementary tissue morphology and biochemical features could potentially improve the understanding of cellular functions and disease diagnosis.     

Fast-electron optical spectrum of a two-dimensional electron gas in the presence of the Rashba effect

We examine how the Rashba spin-orbit interaction (SOI) affects the fast-electron optical spectrum of a two-dimensional electron gas (2DEG). It is found that for a spin-split 2DEG, the spectrum of optical absorption is mainly induced by plasmon excitation via inter-SO electronic transition. From the width and position of the spectrum, the Rashba spin-splitting can be identified optically and, therefore, important spintronic properties can be measured though optical experiments.     

自由基型光引发剂的瞬态及稳态荧光特性研究

利用荧光光谱技术研究了不同自由基型光引发剂的瞬态及稳态荧光光谱特性,从分子结构出发分析了共轭结构对光引发剂荧光光谱的影响.实验结果表明随共轭效应的增强,荧光激发与发射峰波长逐渐增大;瞬态荧光谱的衰减受电子基团的影响较为明显,含有吸电子基团的光引发剂荧光衰减快,而含有给电子基团的光引发剂荧光衰减慢.通过对溶剂极性及粘度研...     

碘分子单频及双频多光子光谱

采用多光子离化谱和荧光激发谱技术，对碘分子单频和双频五光子离化谱和三光子荧光谱进行了研究和标识．对OODR五光子离化谱中一组间隔为72cm^-1的振动序列进行了归属，并确定了其激发、发射和离化通道 关键词：     

X射线管激发X荧光光谱连续本底扣除方法研究

X射线管是目前X射线荧光光谱分析中最常采用的激发源,它所产生的原级谱成为了X荧光光谱中本底成分的主要来源,在对这种光谱进行进一步的分析处理之前需要对其本底进行扣除,对本底估计的准确性直接影响后续处理步骤的效果。对射线管激发X荧光光谱的成分进行了分析,针对其本底特点构造了一种本底强度的估计方法,并根据实测谱线构建了理论测试谱线以便对光谱处理算法的效果进行评价。该方法利用测得X射线荧光光谱中不包含特征峰的谱段对X射线管原级谱造成的本底成分进行估计,使用只包含连续本底的谱段对整个测量谱段进行插值,从而避免了谱线特征峰重叠或对半高宽估计不当时所产生的影响。利用构建的测试光谱对SNIP法、傅里叶变换法和本文的本底估计方法的使用效果进行了比较,使用该方法估计的本底与理论本底更加接近。结果表明使用的方法对X射线管激发的X荧光光谱的本底估计准确,可以采用这种方法对连续本底进行扣除,在对实际测得的X射线荧光光谱的本底扣除中取得了较好的应用效果。