共查询到20条相似文献,搜索用时 140 毫秒
1.
2.
3.
4.
激光扫描共聚焦显微镜技术在材料学研究中的应用 总被引:1,自引:0,他引:1
激光扫描共聚焦显微镜技术(laser scanning confocal microscopy,LSCM)有效地排除了非焦平面信息,提高了分辨率和对比度,使图像更为精确清晰,与计算机及相应的软件技术组合,LSCM实现了连续光学切片,广泛应用于生物三维结构重组及动态分析.LSCM是一种无损深层形态结构分析的重要方法,可以对材料进行深层形貌观察;本文主要综述了LSCM的成像原理以及LSCM在高分子材料和生物工程材料方面的应用. 相似文献
5.
随着生物医学技术的发展,组织样本经常被多种荧光标记物标记,需要通过光谱成像的方法区分出样本中不同的成分。本文在共聚焦显微镜基础上,介绍了一种由精密丝杠和步进电机控制的狭缝机构实现光谱成像的方法,讨论了狭缝缝片的具体设计和狭缝运动精度对光谱带宽和波长准确度的影响。 相似文献
6.
传统光学显微镜与近场光学显微镜 总被引:3,自引:0,他引:3
近场光学显微镜是对于常规光学显微镜的革命。它不用光学透镜成像 ,而用探针的针尖在样品表面上方扫描获得样品表面的信息。分析了传统光学显微镜与近场光学显微镜成像原理的物理本质和两种显微镜系统结构的异同点。介绍了光纤探针的制作方法。重点讨论了近场探测原理、光学隧道效应及非辐射场的性质 相似文献
7.
8.
概述了光学宽视场显微镜、共聚焦显微镜、超分辨率显微镜中所应用的现代显微成像技术,对各种传统和先进的显微成像原理进行了总结。光学宽视场显微镜最常用的显微技术有明场成像、暗场成像、相衬成像、偏光成像、微分干涉(DIC)成像、调制对比成像和荧光成像。相衬成像中根据不同的成像结构还有切趾相衬成像。微分干涉除了传统的偏振光照明还有圆偏振光照明(C-DIC)和专用于塑料的微分干涉(PlasDIC)。共聚焦显微镜随着计算机技术和制造技术的发展而有了巨大的发展。除了传统的共聚焦荧光显微镜以外,还有连续反斯托克斯拉曼散射(CARS)共聚焦、多光子共聚焦和白光共聚焦。超分辨率显微镜中主要介绍了受激辐射淬灭(STED)技术和紧随基态淬灭显微技术的单分子返回(GSDIM)技术。 相似文献
9.
10.
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。 相似文献
11.
The imaging performance in single-photon (1-p) and two-photon (2-p) fluorescence microscopy is described. Both confocal and conventional systems are compared in terms of the three-dimensional (3-D) point spread function and the 3-D optical transfer function. Images of fluorescent sharp edges and layers are modelled, giving resolution in transverse and axial directions. A comparison of the imaging properties is also given for a 4Pi confocal system. Confocal 2-p 4Pi fluorescence microscopy gives the best axial resolution in the sense that its 3-D optical transfer function has the strongest response along the axial direction. 相似文献
12.
K. OGOH R. AKIYOSHI MAY‐MAW‐THET T. SUGIYAMA S. DOSAKA Y. HATTA‐OHASHI H. SUZUKI 《Journal of microscopy》2014,253(3):191-197
Bioluminescence from cells is so dim that bioluminescence microscopy is performed using an ultra low‐light imaging camera. Although the image sensor of such cameras has been greatly improved over time, such improvements have not been made commercially available for microscopes until now. Here, we customized the optical system of a microscope for bioluminescence imaging. As a result, bioluminescence images of cells could be captured with a conventional objective lens and colour imaging camera. As bioluminescence microscopy requires no excitation light, it lacks the photo‐toxicity associated with fluorescence imaging and permits the long‐term, nonlethal observation of living cells. Thus, bioluminescence microscopy would be a powerful tool in cellular biology that complements fluorescence microscopy. 相似文献
13.
Phase evolution in cholesterol/DPPC monolayers: atomic force microscopy and near field scanning optical microscopy studies 总被引:1,自引:0,他引:1
A combination of atomic force microscopy (AFM) and near field scanning optical microscopy has been used to study domain formation in dipalmitoylphosphatidylcholine (DPPC)/cholesterol monolayers with cholesterol concentrations ranging from 0 to 50%. The results show a clear evolution from a mixture of liquid expanded and liquid condensed phases for cholesterol concentrations < 10% to a mixture of liquid expanded and two cholesterol‐containing phases at intermediate concentrations, and finally to a single homogeneous liquid ordered phase for 33% cholesterol. Mixtures of the various phases are clearly identified by height differences in AFM and in some cases by fluorescence imaging for samples containing 0.5% BODIPY dye, which localizes preferentially in the more fluid phase. Note that fluorescence imaging, at least with the dye used here, is unable to distinguish between the cholesterol‐rich and cholesterol‐poor phases detected at intermediate cholesterol concentrations. The combination of fluorescence and AFM imaging provides a more complete picture of the phase evolution for cholesterol/DPPC monolayers than could be obtained by either technique alone, and presents substantial advantages over conventional fluorescence microscopy in that submicrometre‐sized domains can be readily detected. 相似文献
14.
Fibre-optic nonlinear optical microscopy and endoscopy 总被引:1,自引:0,他引:1
Nonlinear optical microscopy has been an indispensable laboratory tool of high‐resolution imaging in thick tissue and live animals. Rapid developments of fibre‐optic components in terms of growing functionality and decreasing size provide enormous opportunities for innovations in nonlinear optical microscopy. Fibre‐based nonlinear optical endoscopy is the sole instrumentation to permit the cellular imaging within hollow tissue tracts or solid organs that are inaccessible to a conventional optical microscope. This article reviews the current development of fibre‐optic nonlinear optical microscopy and endoscopy, which includes crucial technologies for miniaturized nonlinear optical microscopy and their embodiments of endoscopic systems. A particular attention is given to several classes of photonic crystal fibres that have been applied to nonlinear optical microscopy due to their unique properties for ultrashort pulse delivery and signal collection. Furthermore, fibre‐optic nonlinear optical imaging systems can be classified into portable microscopes suitable for imaging behaving animals, rigid endoscopes that allow for deep tissue imaging with minimally invasive manners, and flexible endoscopes enabling imaging of internal organs. Fibre‐optic nonlinear optical endoscopy is coming of age and a paradigm shift leading to optical microscope tools for early cancer detection and minimally invasive surgery. 相似文献
15.
Combined AFM and confocal fluorescence microscope for applications in bio-nanotechnology 总被引:4,自引:0,他引:4
R. KASSIES K. O. VAN DER WERF† A. LENFERINK† C. N. HUNTER‡ J. D. OLSEN‡ V. SUBRAMANIAM† & C. OTTO† 《Journal of microscopy》2005,217(1):109-116
We present a custom-designed atomic force fluorescence microscope (AFFM), which can perform simultaneous optical and topographic measurements with single molecule sensitivity throughout the whole visible to near-infrared spectral region. Integration of atomic force microscopy (AFM) and confocal fluorescence microscopy combines the high-resolution topographical imaging of AFM with the reliable (bio)-chemical identification capability of optical methods. The AFFM is equipped with a spectrograph enabling combined topographic and fluorescence spectral imaging, which significantly enhances discrimination of spectroscopically distinct objects. The modular design allows easy switching between different modes of operation such as tip-scanning, sample-scanning or mechanical manipulation, all of which are combined with synchronous optical detection. We demonstrate that coupling the AFM with the fluorescence microscope does not compromise its ability to image with a high spatial resolution. Examples of several modes of operation of the AFFM are shown using two-dimensional crystals and membranes containing light-harvesting complexes from the photosynthetic bacterium Rhodobacter sphaeroides. 相似文献
16.
The effects of fluorescence saturation on imaging in confocal microscopy have been studied. To include saturation it was necessary to deviate from the widely assumed linear relationship between the fluorescence and the illumination intensity. The lateral response for a point-like object, as well as the optical sectioning power, decreases depending on the degree of saturation. For very high illumination intensities the response for a saturated point object approached that of a conventional fluorescence microscope in which the fluorescence was not saturated. The decrease in the axial confocal response has been confirmed qualitatively by experiment. 相似文献
17.
A. H. Buist M. Müller E. J. Gijsbers G. J. Brakenhoff T. S. Sosnowski T. B. Norris & J. Squier 《Journal of microscopy》1997,186(3):212-220
It is demonstrated that fluorescence lifetimes in the nanosecond and picosecond time-scale range can be observed with the recently proposed double-pulse fluorescence lifetime imaging technique (Müller et al. , 1995, Double-pulse fluorescence lifetime imaging in confocal microscopy. J. Microsc 177, 171–179).
A laser source with an optical parametric amplifier (OPA) system is used to obtain short pulse durations needed for high time resolution, wavelength tunability for selective excitation of specific fluorophores and high pulse energies to obtain (partial) saturation of the optical transition.
It is shown that fluorescence lifetimes can be determined correctly also with nonuniform saturation conditions over the observation area.
A correction scheme for the effect on the measurements of laser power fluctuations, which are inherently present in OPA systems, is presented. Measurements on bulk solutions of Rhodamine B and Rhodamine 6G in different solvents confirm the experimental feasibility of accessing short fluorescence lifetimes with this technique.
Because signal detection does not require fast electronics, the technique can be readily used for fluorescence lifetime imaging in confocal microscopy, especially when using bilateral scanning and cooled CCD detection. 相似文献
A laser source with an optical parametric amplifier (OPA) system is used to obtain short pulse durations needed for high time resolution, wavelength tunability for selective excitation of specific fluorophores and high pulse energies to obtain (partial) saturation of the optical transition.
It is shown that fluorescence lifetimes can be determined correctly also with nonuniform saturation conditions over the observation area.
A correction scheme for the effect on the measurements of laser power fluctuations, which are inherently present in OPA systems, is presented. Measurements on bulk solutions of Rhodamine B and Rhodamine 6G in different solvents confirm the experimental feasibility of accessing short fluorescence lifetimes with this technique.
Because signal detection does not require fast electronics, the technique can be readily used for fluorescence lifetime imaging in confocal microscopy, especially when using bilateral scanning and cooled CCD detection. 相似文献
18.
Petty HR 《Microscopy research and technique》2007,70(8):687-709
Cutting-edge biophysical technologies including total internal reflection fluorescence microscopy, single molecule fluorescence, single channel opening events, fluorescence resonance energy transfer, high-speed exposures, two-photon imaging, fluorescence lifetime imaging, and other tools are becoming increasingly important in immunology as they link molecular events to cellular physiology, a key goal of modern immunology. The primary concern in all forms of microscopy is the generation of contrast; for fluorescence microscopy contrast can be thought of as the difference in intensity between the cell and background, the signal-to-noise ratio. High information-content images can be formed by enhancing the signal, suppressing the noise, or both. As improved tools, such as ICCD and EMCCD cameras, become available for fluorescence imaging in molecular and cellular immunology, it is important to optimize other aspects of the imaging system. Numerous practical strategies to enhance fluorescence microscopy experiments are reviewed. The use of instrumentation such as light traps, cameras, objectives, improved fluorescent labels, and image filtration routines applicable to low light level experiments are discussed. New methodologies providing resolution well beyond that given by the Rayleigh criterion are outlined. Ongoing and future developments in fluorescence microscopy instrumentation and technique are reviewed. This review is intended to address situations where the signal is weak, which is important for emerging techniques stressing super-resolution or live cell dynamics, but is less important for conventional applications such as indirect immunofluorescence. This review provides a broad integrative discussion of fluorescence microscopy with selected applications in immunology. 相似文献
19.
Scanning near-field optical microscopy (SNOM) has been employed to simultaneously acquire high-resolution fluorescence images along with shear-force atomic force microscopy from cell membranes. Implementing such a technique overcomes the limits of optical diffraction found in standard fluorescence microscopy and also yields vital topographic information. The application of the technique to investigate cell-cell adhesion has revealed the interactions of filopodia and their functional relationship in establishing adherens junctions. This has been achieved via the selective tagging of the cell adhesion protein, E-cadherin, by immunofluorescence labelling. Two labelling routes were explored; Alexa Fluor 488 and semiconductor quantum dots. The quantum dots demonstrated significantly enhanced photostability and high quantum yield making them a versatile alternative to the conventional organic fluorophores often used in such a study. Analysis of individual cells revealed that E-cadherin is predominantly located along the cell periphery but is also found to extend throughout their filopodia. We have demonstrated that with a fully optimised sample preparation methodology, quantum dot labelling in conjunction with SNOM imaging can be successfully applied to interrogate biomolecular localisation within delicate cellular membranes. 相似文献
20.
The bilateral scanning approach to confocal microscopy is characterized by the direct generation of the image on a two-dimensional (2-D) detector. This detector can be a photographic plate, a CCD detector or the human eye, the human eye permitting direct visualization of the confocal image. Unlike Nipkow-type systems, laser light sources can be used for excitation. A design called a carousel has been developed, in which the bilateral confocal scan capability can be added to an existing microscope so that rapid exchange and comparison between confocal and non-confocal imaging conditions is possible. The design permits independent adjustment of confocal sectioning properties with lateral resolutions better than, or, in the worst case equivalent to, those available in conventional microscopy. The carousel can be considered as a stationary optical path in which certain imaging conditions, such as confocality, are defined and operate on part of the imaging field. The action of the bilateral scan mirror then extends this image condition over the whole field. A number of optical arrangements for the carousel are presented which realize various forms of confocal fluorescence and reflection imaging, with point, multiple point or slit confocal detection arrangements. Through the addition of active elements to the carousel direct stereoscopic, ratio, time-resolved and other types of imaging can be achieved, with direct image formation on a CCD, eye or other 2-D detectors without the need to modify the host microscope. Depending on the photon flux available, these imaging modes can run in real-time or can use a cooled CCD at (very) low light level for image integration over an extended period. 相似文献