基于液体变焦透镜和振镜的三维光片显微成像系统

搭建了一种基于液体变焦透镜和振镜的三维光片显微成像系统,设计了振镜、液体变焦透镜、相机的同步控制采集成像系统,通过调谐振镜和液体变焦透镜,使得光片激发样品和成像同步,获得样品不同切面的图像堆栈并实现样品的三维重建。当采用数值孔径为0.3、放大倍率为10的成像物镜时,该系统的轴向扫描范围为507μm,横向视场达到1970μm×1300μm,横向分辨率为1.32μm,轴向分辨率可达12.75μm。在轴向扫描过程中,系统的放大倍率保持恒定,可以用于对一定尺寸生物样品的成像实验和相关研究,并通过对斑马鱼胚胎进行成像验证所提系统对厚生物样品成像的可行性。     

充液型可调焦液态透镜

可调焦液态透镜因具备体积小、成像好、调焦方便等优点而备受瞩目.使用透明的弹性薄膜和光学平板玻璃将一定体积的液体封装在圆柱形的空腔内,当空腔内的液体体积变化时,透镜的曲率半径随之而改变,实现了透镜的可调焦.实验表明,该可调焦液态透镜有较好的成像效果,在一定液体体积变化范围内符合薄透镜成像的规律,在通讯、国防、医疗等方面具...     

基于线扫描成像的光片荧光显微镜

设计了一种基于线扫描成像(LSI)的光片荧光显微镜(LSFM),旨在通过抑制样本散射达到提高成像质量的目的 .该显微镜将数字扫描光片荧光显微镜(DSLM)与LSI两种方法结合,以前者为基础,在探测光路上增加了一个用于解扫描的扫描振镜.在控制过程中,将照明光路的扫描振镜与解扫描振镜同步,使得均匀运动的图像在相机前固定于同...     

结合虚拟单像素成像解卷积的双边照明光片荧光显微技术

光片荧光显微术(light-sheet fluorescence microscopy,LSFM)采用薄片光束从侧面激发样品,在垂直于光片方向上进行成像,具有成像速度快、光学层析能力强以及光漂白和光毒性低等优点,适用于对较大活体生物样品进行高质量、长时间三维动态观测.然而,传统高斯光束LSFM存在分辨率低和成像视场小的...     

用于多尺度高分辨率三维成像的双环光片荧光显微技术

本文提出了一种无衍射光片荧光显微成像技术,可以方便地对从微米到厘米各种尺寸的多种生物样本进行多尺度三维荧光成像。提出一种双环调控方法以解决传统贝塞尔光片旁瓣过重的问题,该方法能够可调节地产生0.4～5μm之间不同厚度类型的无衍射光片,同时其旁瓣占比被压低到30%以下。基于这种新的调控手段设计搭建了一套多尺度光片荧光显微成像系统。该系统展示出适用性极强的多尺度成像能力,例如：活细胞双色三维动态成像、膨胀细胞三维超分辨成像以及介观尺度下全器官的高通量三维成像。证明该多尺度成像模式能够显著提升光片荧光显微镜的成像效率,由此可以推进细胞生物学、组织病理学、神经科学等多种生物医学相关研究的发展。     

基于简单透镜列阵的可调焦激光均匀辐照光学系统研究

本文对基于简单透镜列阵的大口径激光均匀辐照光学系统的调焦能力进行了研究, 结果表明, 改变透镜列阵与靶镜之间的距离即可方便地改变靶面上光斑的大小. 文中详细分析了相关参数对调焦能力的影响, 并在此基础上设计出一个实用的光学系统. 用数值方法模拟了激光束通过光学系统后的传输, 发现尺寸不同的靶面光斑具有基本一致的强度结构特征. 定量地分析了光斑内部散斑间隔、调制对比度、顶部不均匀度及能量集中度等描述光斑均匀辐照质量的指标, 并研究了它们随靶面离焦量的变化关系.     

无透镜显微成像的重构算法研究

针对单幅图像进行了无透镜显微成像的重构算法研究，介绍了无透镜显微成像系统实验装置和ASM(angle spectrum method)、改编后的L-R(Lucy-Richardson)两种重构算法。对比两种算法重构后的USAF分辨率板图像的分辨率，利用瑞利判据得出ASM获得的振幅图分辨率最高(即3.10 μm)，且计算用时最少(即0.9 s)，证明了ASM为最佳的单幅无透镜显微重构算法。其次，利用无透镜显微成像系统结合ASM重构的方法，进行细胞成像实验。该无透镜成像视场为5×显微镜的4.4倍，且分辨率介于5×及10×光学显微镜之间，统计学优势明显，在生物医学领域具有广阔的应用前景。     

基于ICCD的快速荧光显微成像技术及在活细胞研究中的初步应用

利用像增强型CCD、氩离子激光器和氙灯等建立了一套快速荧光显微成像系统,并初步应用于活细胞研究。实时观测和拍摄了大鼠脑微血管内皮细胞增殖分裂过程中细胞内钙敏感荧光探针Fluo-3标记的钙离子浓度分布快速变化的图像,并选取动态图像中四个典型的点给出荧光强度灰度值的变化曲线。该系统可用于高灵敏实时记录活细胞内基于荧光显微成像的快速变化过程,为活细胞的研究提供了一种很好的观察和分析手段。     

荧光寿命显微成像技术及应用的最新研究进展

由于荧光寿命不受探针浓度、激发光强度和光漂白效应等因素影响,荧光寿命显微成像技术(fluorescence lifetime imaging microscopy, FLIM)在监测微环境变化、反映分子间相互作用方面具有高特异性、高灵敏度、可定量测量等优点,近年来已被广泛应用于生物医学等领域.然而,尽管FLIM的发明和发展已历经数十年时间,其在实际应用中仍然面临着许多挑战.例如,其成像分辨率受衍射极限限制,而其成像速度与成像质量和寿命测量精度则存在相互制约的关系.近几年来,相关硬件和软件的快速发展及其与其他光学技术的结合,极大地推动了FLIM技术及其应用的新发展.本文简要介绍了基于时域和频域的不同寿命探测方法的FLIM技术的基本原理及特点,在此基础上概述了该技术的最新研究进展,包括其成像性能的提升和在生物医学应用中的研究现状,详细阐述了近几年来研究者们通过硬件和软件算法的改进以及与自适应光学、超分辨成像技术等新型光学技术的结合来提升FLIM的成像速度、寿命测量精度、成像质量和空间分辨率等方面所做的努力,以及FLIM在生物医学基础研究、疾病诊断与治疗、纳米材料的生物医学研究等方面的应用,最后对其未来发展趋势进行了展望.     

High-speed 3D imaging based on structured illumination and electrically tunable lens

In this Letter, we present a high-speed volumetric imaging system based on structured illumination and an electrically tunable lens(ETL), where the ETL performs fast axial scanning at hundreds of Hz. In the system,a digital micro-mirror device(DMD) is utilized to rapidly generate structured images at the focal plane in synchronization with the axial scanning unit. The scanning characteristics of the ETL are investigated theoretically and experimentally. Imaging experiments on pollen samples are performed to verify the optical cross-sectioning and fast axial scanning capabilities. The results show that our system can perform fast axial scanning and threedimensional(3D) imaging when paired with a high-speed camera, presenting an economic solution for advanced biological imaging applications.     

超半球形固体浸没透镜系统的亚表面显微

应用矢量衍射理论和薄膜光学方法,研究了有禁带存在时超半球形固体浸没透镜(SIL)系统的亚表面显微。模拟计算结果显示,光斑的强度,边瓣强度和光斑大小随禁带厚度发生振荡。光斑的大小和光斑的强度几乎是反相变化的。当禁带材料的折射率与固体浸没透镜(SIL)的折射率相匹配时,光斑的强度较大,分辨率较高。随着离开样品表面的距离的增加,光斑的强度不断减小,光斑的大小和边瓣强度逐渐增加。     

High frequency acoustic microscopy with Fresnel zoom lens

The acoustic field distributions and the convergent beams generated by the planar-structure Fresnel zone transducers on solid surface are investigated. Because only 0 and 180 degree phase transducers are used, an imaging system with the Fresnel zoom lens could work at very high frequency, which overcomes the frequency limit of the traditional phased array acoustic imaging system. Simulation results are given to illustrate the acoustic field distributions along the focal axis and the whole plane as well. Based on the principle of scanning of the focus with the change of frequency for the excited signal, an experimental imaging system is also built. Acoustic Fresnel zone transducers are fabricated at center frequency of 400 MHz. Measurements and detections of the known hole flaws at different depths of the fused quartz sample are presented to show that the imaging system with Fresnel zoom lens could move its focus by only changing the frequency of the excited signal.     

Dispersion effect in optical microscopy systems with a supersphere solid immersion lens

This paper studies the dispersion effect of the supersphere solid immersion lens (SIL) on a near field optical microscopy system by using the vector diffraction theory. Results show that when a real non-monochromatic beam illuminates a supersphere SIL microscopy, the dispersion effect of the SIL has an important influence on the image quality. As the wavelength bandwidth of the non-monochromatic beam increases, the size of the focused spot increases and its intensity decreases in near-field microscopy systems with a supersphere SIL.     

大变焦范围电调谐液晶变焦透镜的研究

基于向列相液晶指向矢随施加电场作用发生变化的特性, 结合几何光学和液晶理论提出了一种液晶透镜结构模型, 研究了电极大小、隔垫物厚度等液晶透镜参数对液晶透镜焦距的影响. 通过优化参数, 得到结构简单、变焦范围较大的新型液晶透镜样品, 在驱动电压0 V_RMS–250 V_RMS下可调的焦距范围为75–230 mm, 达到155 mm.     

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.     

Nonlinear optical microscopy for drug delivery monitoring and cancer tissue imaging

A multimodal nonlinear optical microscope that combines coherent anti‐Stokes Raman scattering (CARS), two‐photon excitation fluorescence (TPEF), second‐harmonic generation (SHG) and sum‐frequency generation (SFG) was developed and applied to image breast cancer tissue and MCF‐7 cells as well as monitoring anticancer drug delivery in live cells. TPEF imaging showed that drugs are preferentially localized in the cytoplasm and the nuclear envelope in resistant cells. Moreover, the extracellular matrix was observed by TPEF signals arising from elastin's autofluorescence and SHG signals from collagen fibrils in breast tissue sections. Additionally, CARS signals arising from proteins and (PO₂)⁻ allowed identification of tumors. Label‐free imaging with chemical contrast of significant components of cancer cells and tissue suggests the potential of multimodal nonlinear optical microscopy for early detection and diagnosis of cancer. Copyright © 2010 John Wiley & Sons, Ltd.     

四光路透镜成像系统研究

针对现有膜厚监控系统的双平行光路系统和监测光直接照射单色仪入射狭缝的结构，提出了四光路透镜成像系统，同时采用光纤传光。实验表明它提高了膜厚监的稳定性和精度，并有较高的性能价格比，此外从理论上对多光路透镜成像系统的设计和调校进行了定量分析。