大视场结构光照明显微技术（特邀）

结构光照明显微（SIM）具有成像速度快、对样品损伤小,以及对荧光标记物和标记程序无特殊要求等优点,成为研究活细胞结构和动态过程的重要成像手段之一。介绍了一种大视场、双模式（条纹/点阵）SIM显微技术。该技术结合二维光栅投影产生条纹/点阵结构光,空间光调制器选择条纹方向和实现相移,打破传统SIM中条纹数量受数字投影设备像素个数限制的瓶颈,同时保持了传统SIM成像速度快的优势。实验结果表明：该技术在20×/0.75数值孔径物镜下可获得690μm×517μm成像视场和1.8倍空间分辨率提升,其空间带宽积是传统SIM的3倍。该技术有望被应用于生物学、化学和工业等领域。     

平场复用多焦点结构光照明超分辨显微成像

多焦点结构光照明显微技术(multifocal structured illumination microscopy, MSIM)能在50μm的成像深度内和1Hz的成像速度下实现两倍于衍射极限分辨率的提升,相比传统的宽场结构光照明显微技术,具有较大的成像深度和层析能力,更适合应用于厚样品的长时程三维超分辨成像.然而, MSIM存在成像速度慢、图像处理过程复杂等问题.本文提出了一种基于平场复用多焦点结构光照明的快速超分辨显微成像方法和系统(flat-field multiplexed MSIM, FM-MSIM),通过在照明光路中插入光束整形器件,将高斯光束转变为均为分布的平顶光束,提高激发点阵的强度均匀性和扩大视场;通过将每个衍射受限的激发点沿y方向延长,形成新的多路复用多焦点阵照明图案,提高能量利用率,减少扫描步数,进而提高成像速度和信噪比;结合基于多重测量矢量模型的稀疏贝叶斯学习图像重构算法,简化图像重构步骤,在保证空间分辨率的同时实现至少4倍于传统MSIM的成像速度.在此基础上,利用搭建的FM-MSIM系统进行了BSC细胞微管样片和小鼠肾切片标准样片的超分辨成像实验,实验结果证明...     

随机扫描多光子荧光显微成像系统

多光子荧光显微成像是生物学研究的有力手段,但目前的成像速度难以满足神经成像中快事件检测的需要。针对这一问题,提出了一套随机扫描快速多光子荧光显微成像系统。系统采用二维声光偏转器快速扫描飞秒激发光束,能够以每点10μs的速度对特定的感兴趣区域进行跳跃式扫描,即随机扫描,使得有效的扫描速度大为提高。引入单棱镜补偿方法解决应用声光偏转器带来的色散问题。以170 nm荧光小球为样品,测得系统的横向分辨力为0.3μm,纵向分辨力为1.3μm。给出了随机扫描系统和商品化多光子荧光显微镜对同一个荧光细胞的成像结果,证明了新系统的成像能力。     

基于双螺旋点扩散函数工程的多焦点图像扫描显微

在传统共聚焦显微技术的基础上,图像扫描显微技术使用面阵探测器来代替单点探测器,结合虚拟数字针孔并利用像素重定位和解卷积图像重构算法将传统宽场显微镜的分辨率提高一倍,实现了高信噪比的超分辨共焦成像.但是,由于采用逐点扫描的方式,三维成像速度相对较慢,限制了其在活体样品成像中的应用.为了进一步提高图像扫描显微术的成像速度,本文提出了一种基于双螺旋点扩散函数工程的多焦点图像扫描显微成像方法和系统.在照明光路中,利用高速数字微镜器件产生周期分布的聚焦点阵对样品进行并行激发和快速二维扫描;在探测光路中,利用双螺旋相位片将激发点荧光信号的强度分布转换为双螺旋的形式;最终,利用后期数字重聚焦处理,从单次样品扫描数据中重构出多个样品层的超分辨宽场图像.在此基础上,利用搭建的系统分别对纤维状肌动蛋白和海拉细胞线粒体进行成像实验,证明了该方法的超分辨能力和快速三维成像能力.     

基于时间相关单光子计数的荧光寿命成像技术

采用时域法中的时间相关单光子计数方法记录荧光寿命,时间相关单光子计数采用多波长通道同时记录荧光光子数,可以提高计数效率和信息量,还可以在稳态图像中分离不同荧光团,形成4维图像。并采用多光子激发技术,利用长波长光源发出的两个或多个光子可以激发出一个短波长的光子。多个光子必须几乎同时到达激发点,才能提供被激发分子足够的能量以产生荧光。多光子激发波长较长,生物组织对其散射减小,因而可以穿透到更深层的组织,从而提高荧光成像深度和空间分辨力,并减少对活体样品的损伤。     

大视场高分辨HiLo光切片显微成像系统

现代生物学和生物医学领域迫切需要研制兼顾大视场、高分辨率的显微成像技术和仪器以对生物样品实现跨尺度观测,满足重大科学问题的研究需求。受限于系统的空间带宽积,传统商业显微镜无法满足这一需求,且现有高空间带宽积显微成像系统存在体积庞大、实施成本高昂等问题。本文基于HiLo光切片技术和自主设计的大视场高分辨显微物镜,研发了具有高空间带宽积特点的大视场高分辨HiLo光切片显微成像系统,测试了系统的成像视场和分辨率。应用该系统对小鼠脑切片开展了白光照明明场成像实验,并与OLYMPUS商业显微镜成像结果做了对比;对小麦种子荧光切片开展了光切片成像和宽场荧光成像对比实验。实验结果表明,大视场高分辨HiLo光切片显微成像系统的成像视场达到4.8 mm×3.6 mm (对角视场为6.0 mm),横向分辨率达到0.74μm,轴向分辨率达到4.16μm。大视场高分辨HiLo光切片显微成像系统兼有大视场和高分辨率成像的优势和快速光切片成像的能力,能够对大体积生物样本开展快速三维成像,将为胚胎发育、脑成像、数字病理诊断等研究提供有力的技术支撑。     

基于多光子脉冲内干涉相位扫描法对飞秒激光脉冲进行相位测量和补偿的研究

研制了一套基于多光子脉冲内干涉相位扫描方法的可以同时对飞秒激光脉冲进行相位测量和补偿的实验系统装置.实验中,通过自主研发的LabVIEW程序控制液晶空间光调制器和光纤光谱仪,对待测飞秒激光脉冲施加相位扫描,并同时记录受到调制的飞秒激光脉冲的倍频光谱,得到了多光子脉冲内干涉相位扫描(MIIPS)轨迹图.通过MIIPS轨迹图的三次测量和迭代运算,还原出了经过预先啁啾调制的中心波长约为810 nm、重复频率为1 kHz的飞秒激光脉冲的光谱相位,测量精度在0.1 rad以内.根据测量结果,利用液晶空间光调制器对该飞秒激光脉冲进行相位补偿,得到了近似傅里叶变换极限的飞秒激光脉冲.这一装置将在多光子显微成像、脉冲整形、飞秒激光光谱学等众多领域发挥重要作用.     

新型小型化牙科扫描仪探头的光学系统设计

为解决牙科扫描探头小型化问题,基于结构光三维重建算法,创新性地设计出一种投影与成像共孔径设置的小型化牙科三维扫描仪探头光学系统。系统采用分视场共孔径技术,部分视场用于实现投影功能,另一部分视场用于实现成像功能,投影区域和成像区域存在公共区域。仿真实验给出了在扫描范围为46×35mm2内的系统性能,借助LightTools软件做了照明分析仿真,证明了系统光路的可行性。     

基于空间光调制器的多层图像的构建与可视化

为解决实时直观地观察多层再现图像的问题,提出利用发光材料实现其可视化.针对由沿光轴方向的二维图层组成的空间图像,利用高效、快速的三维Gerchberg-Saxton算法,得到位相型计算全息图,并通过计算机进行了数字模拟再现.阐述了三维Gerchberg-Saxton算法的流程,并搭建了基于液晶空间光调制器的位相全息图光学再现与可视化光路.利用液晶空间光调制器的灰度-位相曲线把计算全息图转换为灰度图,加载在液晶空间光调制器上,再现出高质量的三维光场,同时利用量子点材料的荧光特性实现了图像的可视化.实验结果表明,光学再现与计算机模拟结果较吻合.该技术在医学、军事、三维显示、微加工以及显微技术等领域有重要应用价值.     

基于空间光调制器的多层图像的构建与可视化研究

 为解决实时直观地观察多层再现图像的问题,提出利用发光材料实现其可视化.针对由沿光轴方向的二维图层组成的空间图像,利用高效、快速的三维Gerchberg-Saxton算法,得到位相型计算全息图,并通过计算机进行了数字模拟再现.阐述了三维Gerchberg-Saxton算法的流程,并搭建了基于液晶空间光调制器的位相全息图光学再现与可视化光路.利用液晶空间光调制器的灰度-位相曲线把计算全息图转换为灰度图,加载在液晶空间光调制器上,再现出高质量的三维光场,同时利用量子点材料的荧光特性实现了图像的可视化.实验结果表明,光学再现与计算机模拟结果较吻合.该技术在医学、军事、三维显示、微加工以及显微技术等领域有重要应用价值.     

Ultrafast, large-field multiphoton microscopy based on an acousto-optic deflector and a spatial light modulator

We present an ultrafast, large-field multiphoton excitation fluorescence microscope with high lateral and axial resolutions based on a two-dimensional (2-D) acousto-optical deflector (AOD) scanner and spatial light modulator (SLM). When a phase-only SLM is used to shape the near-infrared light from a mode-locked titanium:sapphire laser into a multifocus array including the 0-order beam, a 136 μm × 136 μm field of view is achieved with a 60× objective using a 2-D AOD scanner without any mechanical scan element. The two-photon fluorescence image of a neuronal network that was obtained using this system demonstrates that our microscopy permits observation of dynamic biological events in a large field with high-temporal and -spatial resolution.     

Addressable microlens array for parallel laser microfabrication

Parallel processing in femtosecond-laser-based microfabrication is demonstrated using a microlens array in conjunction with a liquid-crystal spatial light modulator (SLM). A portion of the SLM is mapped onto each individual lenslet in the array and can be used to effectively switch foci on and off for fabrication. In addition, the technique allows for homogenizing the intensity of the array of foci and translating spots relative to their natural focus. The technique demonstrates the potential for high efficiency processing of aperiodic structures.     

采用液晶空间光调制器的可控性阵列菲涅耳波带片

利用液晶空间光调制器的光学调制特性,实现阵列数与焦长可调控的阵列菲涅耳波带片,并给出了理论分析和实验结果,该方法操作简单、具有实时可调控的优点.     

Autofocus technique for three-dimensional imaging, direct-detection laser radar using Geiger-mode avalanche photodiode focal-plane array

An autofocus technique is proposed for a three-dimensional imaging, direct-detection laser radar system that uses a Geiger-mode avalanche photodiode focal plane array (GmAPD-FPA). This technique is implemented by pointing laser pulses on a target of interest and observing its scattered photon distribution on a GmAPD-FPA. Measuring the standard deviation of the photon distribution on a GmAPD-FPA enables the best focus condition to be found. The feasibility of this technique is demonstrated experimentally by employing a 1 × 8 pixel GmAPD-FPA. It is shown that the spatial resolution improves when the GmAPD-FPA is located in the best focus position found by the autofocus technique.     

基于SiPM单光子探测器的荧光光谱仪研究

为了解决激光诱导荧光检测系统存在的光学结构复杂、体积大、成本高以及灵敏度不足等主要问题,研制了一种高灵敏、小型化的荧光光谱仪。该光谱仪以349 nm半导体激光器作为激发光源,采用正交型光路,将4×4窄带滤光片阵列与具有单光子灵敏度的硅光电倍增器（SiPM）阵列耦合,可实现光谱信息的多通道探测,具备结构紧凑、成本较低以及稳定性好等优点。以荧光素钠为测试样品,对光谱仪性能进行了评估。实验结果表明,光谱仪的检测限优于5×10?11 mol·L?1,在5×10?11 mol·L?1到1×10?9 mol·L?1的溶液浓度范围内,被测溶液浓度与检测所得荧光强度满足良好的线性关系,线性相关系数为0.998 39。此外,光谱仪还具备良好的重复性,荧光峰值强度的相对标准偏差小于10%。因此,该光谱仪兼具灵敏度高、线性度好、重复性与可靠性强等优点,可以满足现场实时检测的需求。     

Parallel Raman microspectroscopy using programmable multipoint illumination

We present a novel parallel Raman microspectroscopy scheme for simultaneously collecting Raman spectra from multiple points. This scheme is realized by projectinga multiple-point laser illumination pattern using a spatial light modulator (SLM) and wide-field Raman imaging collection. We demonstrate the performance of this scheme using uniform samples, trapped polymer microparticles and fixed polymer microparticles with mixed molecular composition within a 50×50 μm(2) field of view. This scheme enables the acquisition of Raman spectra from as many as 40 points simultaneously using a single illumination pattern and detector recording frame without scanning.     

稀土镧离子对1,10-二氮杂菲紫外荧光的增强效应

测量了１,１０－二氮杂菲和硝酸［二－（１,１０－二氮杂菲）合镧］甲醇溶液的单光子和双光子激发紫外荧光谱。研究表明：镧离子的配位对１,１０－二氮杂菲的紫外荧光效率具有显著的增强作用,镧离子的桥梁作用增强了１,１０－二氮杂菲之间的耦合,并引起１,１０－二氮杂菲紫外荧光谱的红移。     

Interaction-Free Effects Between Distant Atoms

A Gedanken experiment is presented where an excited and a ground-state atom are positioned such that, within the former’s half-life time, they exchange a photon with 50% probability. A measurement of their energy state will therefore indicate in 50% of the cases that no photon was exchanged. Yet other measurements would reveal that, by the mere possibility of exchange, the two atoms have become entangled. Consequently, the “no exchange” result, apparently precluding entanglement, is non-locally established between the atoms by this very entanglement. This quantum-mechanical version of the ancient Liar Paradox can be realized with already existing transmission schemes, with the addition of Bell’s theorem applied to the no-exchange cases. Under appropriate probabilities, the initially-excited atom, still excited, can be entangled with additional atoms time and again, or alternatively, exert multipartite nonlocal correlations in an interaction free manner. When densely repeated several times, this result also gives rise to the Quantum Zeno effect, again exerted between distant atoms without photon exchange. We discuss these experiments as variants of interaction-free-measurement, now generalized for both spatial and temporal uncertainties. We next employ weak measurements for elucidating the paradox. Interpretational issues are discussed in the conclusion, and a resolution is offered within the Two-State Vector Formalism and its new Heisenberg framework.     

Beam characterization and spatial coherence measurement of F2 laser using Lumilass glass fluorescence

This paper presents the spectral emission and saturation properties of a Lumilass G9 fluorescence glass excited using a 157 nm laser emitting ∼10 ns duration pulses. This fluorescence glass can be applied in conjunction with a CCD array for beam profiling and divergence measurements. This method can also be used for deriving information on the spatial coherence function of the F₂ laser. This constitutes one of the major contributions of the paper and is also discussed.     

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.”