[go: up one dir, main page]

CN108801863A - The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained - Google Patents

The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained Download PDF

Info

Publication number
CN108801863A
CN108801863A CN201810353723.8A CN201810353723A CN108801863A CN 108801863 A CN108801863 A CN 108801863A CN 201810353723 A CN201810353723 A CN 201810353723A CN 108801863 A CN108801863 A CN 108801863A
Authority
CN
China
Prior art keywords
femtosecond
sample
optical
tested
fluorescence
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CN201810353723.8A
Other languages
Chinese (zh)
Inventor
马渊
杨京法
赵江
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Institute of Chemistry CAS
University of Chinese Academy of Sciences
Original Assignee
Institute of Chemistry CAS
University of Chinese Academy of Sciences
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Institute of Chemistry CAS, University of Chinese Academy of Sciences filed Critical Institute of Chemistry CAS
Priority to CN201810353723.8A priority Critical patent/CN108801863A/en
Publication of CN108801863A publication Critical patent/CN108801863A/en
Pending legal-status Critical Current

Links

Classifications

    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/01Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials specially adapted for biological cells, e.g. blood cells

Landscapes

  • Chemical & Material Sciences (AREA)
  • Dispersion Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)

Abstract

本发明涉及一种可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统,其特征在于该系统包括:用于作为飞秒捕获激光为待测样品提供稳定捕获光阱的飞秒光镊捕获激光单元;用于探测被捕获待测样品在相对于光阱中心做受限布朗运动时位置分布的位置探测单元;用于作为激发光照射待测样品进行荧光激发的激发光源单元;用于将飞秒捕获激光收集到位置探测单元,且将激发光源单元出射的激发光引入到位于捕获光阱中的待测样品,使得待测样品的荧光染料分子受激发产生荧光信号的光学显微单元;用于收集待测样品产生的荧光信号,完成对荧光染料包被的单个胶体粒子的实时荧光成像,获得待测样品在不同粘弹性特征体系中的单个胶体粒子的动力学信息的单分子荧光成像单元。

The invention relates to a femtosecond optical tweezers system capable of obtaining dynamics and imaging information of colloidal particles in a solution, which is characterized in that the system includes: a femtosecond light used as a femtosecond capture laser to provide a stable capture optical trap for a sample to be tested A tweezers capture laser unit; a position detection unit used to detect the position distribution of the captured sample to be measured when doing limited Brownian motion relative to the center of the optical trap; an excitation light source unit used to irradiate the sample to be measured with excitation light for fluorescence excitation; The optical microscope is to collect the femtosecond capture laser light into the position detection unit, and introduce the excitation light emitted by the excitation light source unit into the sample to be measured in the capture light trap, so that the fluorescent dye molecules of the sample to be measured are excited to generate fluorescence signals Unit; used to collect the fluorescent signal generated by the sample to be tested, complete the real-time fluorescence imaging of a single colloidal particle coated with a fluorescent dye, and obtain the single molecule of the kinetic information of a single colloidal particle of the sample to be tested in different viscoelastic characteristic systems Fluorescence Imaging Unit.

Description

可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统Femtosecond optical tweezers system capable of obtaining dynamics and imaging information of colloidal particles in solution

技术领域technical field

本发明是关于一种可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统,涉及胶体科学以及生物物理基础研究领域。The invention relates to a femtosecond optical tweezers system capable of acquiring dynamics and imaging information of colloidal particles in a solution, and relates to the fields of colloid science and basic biophysics research.

背景技术Background technique

光镊技术是一种可以捕获和操控微小粒子以及进行力学测量的重要单分子技术,自美国贝尔实验室的阿什金于1986年发明,经过三十多年的发展已经日益成熟,在理论以及应用方面都做了大量的研究工作,由单光镊逐步发展为双光镊、多光镊、飞秒光镊以及涡旋光镊等各类光镊,这一技术被广泛应用于生物物理、纳米加工、胶体科学以及物理学等多个领域,并且光镊由于易与其他一些技术相结合,例如单分子荧光成像,荧光共振能量转移等,使其成为相对于单一技术或测量手段的一大优势。飞秒光镊,是人们将光镊的捕获光源用飞秒激光代替连续波激光,飞秒激光具有飞秒的脉宽以及高重复频率,使其在满足连续波激光捕获要求的同时,还具备连续波激光不具备的性质,在较低的平均功率下就可实现捕获。毛方林等人2004年在理论上首次提出并计算了飞秒光镊的横向光学力,同年B.Agate等人用飞秒光镊实现了双光子荧光激发以及原位控制,发现其捕获所需的激光平均光强明显低于连续波激光;降雨强等在2010年首次发现由于强聚焦飞秒光镊的非线性捕获形成的光阱劈裂的现象;Goswami等人基于新的飞秒光镊的微流变方法在一个微小体积内直接原位测量了固-液界面处的温度等。相较于传统光镊,飞秒激光自身的复杂特性使得飞秒光镊的相关实验研究都还很有限,因此,将飞秒光镊与全内反射荧光成像相结合,一种可获取溶液中胶体粒子的动力学信息测量的高灵敏度高精度的成像测量方法的建立是非常有必要的。Optical tweezers technology is an important single-molecule technology that can capture and manipulate tiny particles and perform mechanical measurements. It was invented by Ashkin of Bell Laboratories in the United States in 1986. After more than 30 years of development, it has become increasingly mature. In theory and A lot of research work has been done in terms of application, and the single optical tweezers have gradually developed into various types of optical tweezers such as dual optical tweezers, multi-optical tweezers, femtosecond optical tweezers, and vortex optical tweezers. This technology is widely used in biophysics, nanoscale Processing, colloid science and physics and other fields, and optical tweezers are easy to combine with other technologies, such as single-molecule fluorescence imaging, fluorescence resonance energy transfer, etc., making it a great advantage over a single technology or measurement method . Femtosecond optical tweezers is the use of femtosecond laser instead of continuous wave laser as the capture light source of optical tweezers. Femtosecond laser has femtosecond pulse width and high repetition rate, which makes it meet the requirements of continuous wave laser capture and also has CW lasers do not have the property that trapping can be achieved at lower average power. Mao Fanglin and others theoretically proposed and calculated the transverse optical force of femtosecond optical tweezers for the first time in 2004. In the same year, B.Agate et al. used femtosecond optical tweezers to realize two-photon fluorescence excitation and in-situ control, and found that the capture required The average light intensity of the laser is significantly lower than that of the continuous wave laser; Rain Qiang and others first discovered the phenomenon of optical trap splitting due to the nonlinear capture of strongly focused femtosecond optical tweezers in 2010; Goswami et al. based on the new femtosecond optical tweezers The microrheological method directly measures the temperature at the solid-liquid interface in a small volume in situ. Compared with the traditional optical tweezers, the complex characteristics of the femtosecond laser itself make the relevant experimental research of the femtosecond optical tweezers very limited. It is very necessary to establish a high-sensitivity and high-precision imaging measurement method for the measurement of the kinetic information of colloidal particles.

发明内容Contents of the invention

针对上述问题,本发明的目的是提供一种可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统,能够对胶体体系、细胞进行精确主动操控的同时还能够进行高分辨率、高灵敏度的单分子测量与成像,进而得到胶体粒子或细胞在溶液中的动力学信息。In view of the above problems, the purpose of the present invention is to provide a femtosecond optical tweezers system that can obtain the dynamics and imaging information of colloidal particles in solution, which can perform precise and active manipulation of colloidal systems and cells, and can also perform high-resolution, high-resolution imaging. Sensitive single-molecule measurement and imaging, and then obtain kinetic information of colloidal particles or cells in solution.

为实现上述目的,本发明采取以下技术方案:一种可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统,该系统包括:用于作为飞秒捕获激光为待测样品提供稳定捕获光阱的飞秒光镊捕获激光单元;用于探测被捕获待测样品在相对于光阱中心做受限布朗运动时位置分布的位置探测单元;用于作为激发光照射待测样品进行荧光激发的激发光源单元;用于将飞秒捕获激光收集到所述位置探测单元,且将所述激发光源单元出射的激发光引入到位于捕获光阱中的待测样品,使得待测样品的荧光染料分子受激发产生荧光信号的光学显微单元;用于收集待测样品产生的荧光信号,完成对荧光染料包被的单个胶体粒子的实时荧光成像,获得待测样品在不同粘弹性特征体系中的单个胶体粒子的动力学信息的单分子荧光成像单元。In order to achieve the above object, the present invention adopts the following technical solutions: a femtosecond optical tweezers system that can obtain colloidal particle dynamics and imaging information in solution, the system includes: used as a femtosecond capture laser to provide stable capture for the sample to be tested The femtosecond optical tweezers capture laser unit of the optical trap; the position detection unit used to detect the position distribution of the trapped sample to be measured when it is doing limited Brownian motion relative to the center of the optical trap; used to irradiate the sample to be measured with excitation light for fluorescence excitation An excitation light source unit; used to collect the femtosecond capture laser light to the position detection unit, and introduce the excitation light emitted by the excitation light source unit into the sample to be measured in the capture optical trap, so that the fluorescent dye of the sample to be measured An optical microscopic unit that generates fluorescent signals when molecules are excited; it is used to collect the fluorescent signals generated by the samples to be tested, to complete real-time fluorescence imaging of single colloidal particles coated with fluorescent dyes, and to obtain the characteristics of the samples to be tested in different viscoelastic characteristic systems. Single-molecule fluorescence imaging unit for kinetic information of individual colloidal particles.

进一步地,所述光学显微单元采用倒置荧光显微镜结构,包括显微物镜、第一~第三二向色镜和聚光镜;激发光经所述显微物镜聚焦到位于捕获光阱中的待测样品,经激发光激发待测样品产生的荧光经所述显微物镜收集后发射到第一二向色镜,经所述第一二向色镜出射的荧光信号发射到所述单分子荧光成像测量单元。Further, the optical microscope unit adopts an inverted fluorescence microscope structure, including a microscope objective lens, first to third dichroic mirrors, and a condenser lens; For the sample, the fluorescence generated by the sample to be tested is collected by the microscope objective lens and emitted to the first dichroic mirror, and the fluorescence signal emitted by the first dichroic mirror is emitted to the single-molecule fluorescence imaging unit of measurement.

进一步地,所述飞秒光镊捕获激光单元包括飞秒脉冲激光器、扩束和准直镜以及偏转镜,所述飞秒脉冲激光器出射的飞秒脉冲激光经所述扩束和准直镜进行扩束和准直,扩束和准直后的飞秒脉冲激光经所述偏转镜进行偏转后发射到第二二向色镜,经所述第二二向色镜出射的光发射到所述显微物镜。Further, the femtosecond optical tweezers capture laser unit includes a femtosecond pulse laser, a beam expander and collimator mirror, and a deflection mirror, and the femtosecond pulse laser emitted by the femtosecond pulse laser passes through the beam expander and collimator mirror. Beam expansion and collimation, the femtosecond pulse laser after beam expansion and collimation is deflected by the deflection mirror and then sent to the second dichroic mirror, and the light emitted by the second dichroic mirror is sent to the microscope objective.

进一步地,所述位置探测单元包括会聚透镜、位置探测装置、数据采集卡和计算机;经所述显微物镜出射的前向散射光形成的干涉图样经所述聚光镜收集后发射到第三二向色镜,经所述第三二向色镜出射的光经所述会聚透镜聚焦所述位置探测装置,所述位置探测装置通过所述数据采集卡连接所述计算机获得待测样品所处光阱的刚度系数以及待测样品在不同粘弹性的体系中的粘度。Further, the position detection unit includes a converging lens, a position detection device, a data acquisition card and a computer; the interference pattern formed by the forward scattered light emitted by the microscope objective lens is collected by the condenser lens and then emitted to the third dichotomous A color mirror, the light emitted by the third dichroic mirror is focused on the position detection device through the converging lens, and the position detection device is connected to the computer through the data acquisition card to obtain the optical trap where the sample to be measured is located The coefficient of stiffness and the viscosity of the sample to be tested in different viscoelastic systems.

进一步地,所述位置探测装置采用位置敏感探测器或数字相机。Further, the position detecting device adopts a position sensitive detector or a digital camera.

进一步地,所述激发光源单元包括固体激光器、格兰泰勒棱镜、四分之一波片、扩束和准直镜、透镜以及滤光片;所述固体激光器发出的连续波依次经所述格兰-泰勒棱镜和四分之一波片发射到所述扩束和准直镜,经所述扩束和准直镜出射的圆偏振光经依次经上所述透镜和滤光片聚焦所述显微物镜的后焦面上。Further, the excitation light source unit includes a solid-state laser, a Glan-Taylor prism, a quarter-wave plate, a beam expander and collimator mirror, a lens, and an optical filter; the continuous wave emitted by the solid-state laser passes through the lattice The blue-Taylor prism and the quarter-wave plate emit to the beam expander and collimator mirror, and the circularly polarized light emitted by the beam expander and collimator mirror is focused by the above lens and filter in turn. back focal plane of the microscope objective.

进一步地,所述固体激光器出口处设置中性密度滤波片。Further, a neutral density filter is arranged at the exit of the solid-state laser.

进一步地,所述单分子荧光成像单元采用EMCCD相机和PCI数据采集卡,所述EMCCD相机经所述PCI数据采集卡连接所述计算机。Further, the single-molecule fluorescence imaging unit adopts an EMCCD camera and a PCI data acquisition card, and the EMCCD camera is connected to the computer via the PCI data acquisition card.

进一步地,所述待测样品为聚苯乙烯荧光小球、金纳米颗粒、无荧光的聚苯乙烯小球或二氧化硅小球、人体活性细胞。Further, the sample to be tested is polystyrene fluorescent beads, gold nanoparticles, non-fluorescent polystyrene beads or silicon dioxide beads, human active cells.

本发明由于采取以上技术方案,其具有以下优点:1、本发明将高重复频率的飞秒脉冲光镊与单分子全内反射荧光成像相结合,能够获取溶液中被测量体系的动力学信息。2、本发明在能够实现捕获操控的同时,采用高灵敏度的EMCCD相机对待测样品进行激发光激发以及荧光信号的收集,进而实现包括双光子荧光信号以及普通多荧光信号的探测。3、本发明的待测样品为不同粘弹性体系中的胶体小球或金纳米颗粒等,因此可以实现不同粘弹性体系、不同尺寸的胶体粒子进行测量。本发明可以广泛地应用于纳米颗粒、胶体粒子、凝胶、细胞等多种体系的相关动力学研究,探索其各自特殊性质的基础物理根源。Due to the adoption of the above technical solutions, the present invention has the following advantages: 1. The present invention combines femtosecond pulsed optical tweezers with high repetition rate and single-molecule total internal reflection fluorescence imaging to obtain dynamic information of the measured system in solution. 2. While the present invention can achieve capture control, it uses a high-sensitivity EMCCD camera to excite the sample to be tested with excitation light and collect fluorescence signals, thereby realizing detection including two-photon fluorescence signals and ordinary multi-fluorescence signals. 3. The samples to be tested in the present invention are colloid balls or gold nanoparticles in different viscoelastic systems, so colloid particles of different viscoelastic systems and different sizes can be measured. The present invention can be widely applied to related dynamic research of various systems such as nanoparticles, colloidal particles, gels, cells, etc., and explores the basic physical roots of their respective special properties.

附图说明Description of drawings

图1是本发明的飞秒光镊系统测量原理示意图;Fig. 1 is a schematic diagram of the measurement principle of the femtosecond optical tweezers system of the present invention;

图2是本发明的飞秒激光捕获光源单元与激发光源单元光路示意图;Fig. 2 is a schematic diagram of the light path of the femtosecond laser capture light source unit and the excitation light source unit of the present invention;

图3是本发明实施例的胶体粒子在光阱中运动的位移分布示意图;3 is a schematic diagram of the displacement distribution of colloidal particles moving in the optical trap according to an embodiment of the present invention;

图4是本发明实施例中被捕获的半径100nm的荧光PS小球的双光子激发成像图。Fig. 4 is a two-photon excitation imaging diagram of trapped fluorescent PS beads with a radius of 100 nm in an embodiment of the present invention.

具体实施方式Detailed ways

以下结合附图来对本发明进行详细的描绘。然而应当理解,附图的提供仅为了更好地理解本发明,它们不应该理解成对本发明的限制。在本发明的描述中,需要理解的是,术语“第一”、“第二”等仅仅是用于描述的目的,而不能理解为指示或暗示相对重要性。The present invention will be described in detail below in conjunction with the accompanying drawings. However, it should be understood that the accompanying drawings are provided only for better understanding of the present invention, and they should not be construed as limiting the present invention. In the description of the present invention, it should be understood that the terms "first", "second" and so on are only used for the purpose of description, and should not be understood as indicating or implying relative importance.

如图1所示,本发明提供的可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统,包括飞秒光镊捕获激光单元1、位置探测单元2、激发光源单元3、光学显微单元4和单分子荧光成像单元5,其中:As shown in Figure 1, the femtosecond optical tweezers system provided by the present invention, which can obtain the dynamics and imaging information of colloidal particles in the solution, includes a femtosecond optical tweezers capture laser unit 1, a position detection unit 2, an excitation light source unit 3, an optical display Micro unit 4 and single molecule fluorescence imaging unit 5, wherein:

飞秒光镊捕获激光单元1用于作为飞秒捕获激光为待测样品提供稳定的捕获光阱;The femtosecond optical tweezers capture laser unit 1 is used as a femtosecond capture laser to provide a stable capture optical trap for the sample to be tested;

位置探测单元2用于探测被捕获待测样品在相对于光阱中心做受限布朗运动时的位置分布;The position detection unit 2 is used to detect the position distribution of the captured sample to be measured when performing limited Brownian motion relative to the center of the optical trap;

激发光源单元3用于作为激发光照射待测样品进行荧光激发;The excitation light source unit 3 is used as excitation light to irradiate the sample to be tested for fluorescence excitation;

光学显微单元4用于将飞秒捕获激光收集到位置探测单元2,以及将激发光源单元3出射的激发光引入到位于捕获光阱中的待测样品,使得待测样品的荧光染料分子受激发产生荧光信号;The optical microscope unit 4 is used to collect the femtosecond capture laser light into the position detection unit 2, and introduce the excitation light emitted by the excitation light source unit 3 into the sample to be measured in the capture light trap, so that the fluorescent dye molecules of the sample to be measured are subjected to Excited to generate a fluorescent signal;

单分子荧光成像单元5用于收集待测样品产生的荧光信号,完成对荧光染料包被的单个胶体粒子的实时荧光成像,获得待测样品在不同粘弹性特征体系中的单个胶体粒子的动力学信息;另外,单分子荧光成像单元5也可用于满足柯勒照明条件的普通的明场成像。The single-molecule fluorescence imaging unit 5 is used to collect the fluorescent signal generated by the sample to be tested, complete the real-time fluorescence imaging of the single colloidal particle coated with the fluorescent dye, and obtain the dynamics of the single colloidal particle of the sample to be tested in different viscoelastic characteristic systems Information; In addition, the single-molecule fluorescence imaging unit 5 can also be used for ordinary bright-field imaging that meets Koehler illumination conditions.

在一个优选的实施例中,如图1所示,光学显微单元4可以采用倒置荧光显微镜结构,包括显微物镜40、三个二向色镜41、聚光镜42、反射镜43和光阑44;激发光经显微物镜40聚焦到位于捕获光阱中的待测样品,经激发光激发待测样品产生的荧光经显微物镜40收集后发射到二向色镜41c,经二向色镜41c出射的荧光信号经发射光高通滤波片发射到反射镜43,经反射镜43反射的荧光经光阑44发射到单分子荧光成像测量单元5。In a preferred embodiment, as shown in FIG. 1 , the optical microscope unit 4 can adopt an inverted fluorescence microscope structure, including a microscope objective lens 40, three dichroic mirrors 41, a condenser lens 42, a mirror 43 and an aperture 44; The excitation light is focused by the microscope objective lens 40 to the sample to be measured in the capture light trap, and the fluorescence generated by the excitation light to excite the sample to be measured is collected by the microscope objective lens 40 and then emitted to the dichroic mirror 41c, and passed through the dichroic mirror 41c The emitted fluorescence signal is transmitted to the reflector 43 through the high-pass filter of the emitted light, and the fluorescence reflected by the reflector 43 is transmitted to the single-molecule fluorescence imaging measurement unit 5 through the aperture 44 .

在一个优选的实施例中,如图2所示,飞秒光镊捕获激光单元1包括提供飞秒捕获激光的飞秒脉冲激光器10、若干反射镜11、扩束和准直镜12、光阑13、偏转镜14和两带通滤光片15。飞秒脉冲激光器10可以采用TEM00模式的钛蓝宝石光纤激光器,飞秒脉冲激光器10出射的飞秒脉冲激光经反射镜11a反射到扩束和准直镜12进行扩束和准直,扩束和准直后的飞秒脉冲激光依次经光阑13和反射镜11b和11c发射到偏转镜14,经偏转镜14偏转出射的光经带通滤光片15a发射到二向色镜41a,经二向色镜41a出射的光经带通滤光片15b发射到显微物镜40,扩束后束径大小应等于或略大于显微物镜40后瞳大小,不同数值孔径的显微物镜40会略有差别,约5~6.3mm,这样能够保证获得尽可能大的梯度力,从而有利于实现并保持稳定的捕获,其中,偏转镜14可以采用压电驱动的高精度偏转镜,可以通过计算机控制驱动偏转镜14发生微弧度偏转,实现飞秒光镊的主动操控。In a preferred embodiment, as shown in Figure 2, the femtosecond optical tweezers capture laser unit 1 includes a femtosecond pulsed laser 10 that provides femtosecond capture laser light, several mirrors 11, a beam expander and collimator mirror 12, an aperture 13. A deflection mirror 14 and two bandpass filters 15. Femtosecond pulsed laser 10 can adopt the titanium sapphire fiber laser of TEM00 mode, the femtosecond pulsed laser light that femtosecond pulsed laser 10 emerges is reflected to beam expander and collimating mirror 12 by reflector 11a and carries out beam expansion and collimation, beam expander and The collimated femtosecond pulsed laser light is transmitted to the deflection mirror 14 through the diaphragm 13 and the mirrors 11b and 11c in turn, and the light deflected by the deflection mirror 14 is transmitted to the dichroic mirror 41a through the band-pass filter 15a, and passed through the dichroic mirror 41a. The light emitted by the chromatic mirror 41a is emitted to the microscopic objective lens 40 through the band-pass filter 15b. There is a difference, about 5-6.3mm, which can ensure the gradient force as large as possible, which is conducive to realizing and maintaining stable capture. Among them, the deflection mirror 14 can be a high-precision deflection mirror driven by piezoelectricity, which can be controlled by a computer The deflection mirror 14 is driven to deflect in micro-radians, so as to realize the active control of the femtosecond optical tweezers.

在一个优选的实施例中,如图1所示,位置探测单元2包括会聚透镜20、高通低阻滤光片21、位置敏感探测器22、NI数据采集卡23和计算机24;经显微物镜40出射的前向散射光形成的干涉图样经聚光镜42(condenser,使光束更好地照到样品上的部件,目的是尽可能的收集前向散射光)收集后发射到二向色镜41b,经二向色镜41b出射的光依次经会聚透镜20和高通低阻滤光片21聚焦位置敏感探测器22,位置敏感探测器22通过NI数据采集卡23连接计算机24。需要说明的是,位置敏感探测器22可以替换为数字相机,数字相机将探测接收的光强信号转换为电信号经NI数据采集卡23发送到计算机24也可以进行分析获得位置与电压的关系,进而获得待测样品所处光阱的刚度系数以及获得待测样品在不同粘弹性的体系中的粘度,其中,高通低阻滤光片21可以采用可见光和红外干涉滤光片。In a preferred embodiment, as shown in Figure 1, the position detection unit 2 includes a converging lens 20, a high-pass low-resistance filter 21, a position-sensitive detector 22, a NI data acquisition card 23 and a computer 24; The interference pattern formed by the forward scattered light emitted by 40 is collected by the condenser lens 42 (condenser, the part that makes the light beam better shine on the sample, the purpose is to collect the forward scattered light as much as possible) and emits to the dichroic mirror 41b, The light emitted by the dichroic mirror 41b is focused on the position-sensitive detector 22 through the converging lens 20 and the high-pass low-resistance filter 21 in turn, and the position-sensitive detector 22 is connected to the computer 24 through the NI data acquisition card 23 . It should be noted that the position-sensitive detector 22 can be replaced by a digital camera, and the digital camera converts the detected light intensity signal into an electrical signal and sends it to the computer 24 through the NI data acquisition card 23, which can also be analyzed to obtain the relationship between position and voltage. Further, the stiffness coefficient of the optical trap where the sample to be measured is located and the viscosity of the sample to be measured in different viscoelastic systems are obtained, wherein, the high-pass low-resistance filter 21 can use visible light and infrared interference filters.

在一个优选的实施例中,高重复频率的飞秒脉冲激光由于自身的非线性效应可以实现双光子激发,在捕获的同时又可以获得双光子激发,因此所选择荧光染料的激发波长要适当,本发明的激发光源单元3采用高稳定性TEM00模式连续波激光,连续波激光的波长需要与所激发的染料分子相匹配,激发光源单元3包括固体激光器30(本实施例采用532nm波长,以此为例,不限于此)、中性密度滤波片31(中性密度滤波片31可以采用激发光带通滤波片)、若干反射镜32、格兰泰勒棱镜33、四分之一波片34、扩束和准直镜35、透镜36以及两滤光片37;固体激光器30发出的连续波激光经中性密度滤波片31发射到反射镜32a,经反射镜32a出射的激光依次经格兰-泰勒棱镜33、四分之一波片34和反射镜32b发射到扩束和准直镜35,经扩束和准直镜出射的圆偏振光经依次经透镜36、滤光片37a、反射镜32c和滤光片37b聚焦到显微物镜40的后焦面上,由于高倍显微物镜40具有很高的数值孔径可以实现全内反射,能够实现比宽场荧光成像信噪比更高的全内反射荧光成像。其中,此处扩束的目的是用于将激发光束直径扩大以确保其尺寸大于显微物镜40的进光孔以确保激发光是高质量的圆偏振光。In a preferred embodiment, the high repetition rate femtosecond pulsed laser can realize two-photon excitation due to its own nonlinear effect, and can obtain two-photon excitation while capturing, so the excitation wavelength of the selected fluorescent dye should be appropriate, The excitation light source unit 3 of the present invention adopts a high-stability TEM 00 mode continuous wave laser, the wavelength of the continuous wave laser needs to match the excited dye molecules, and the excitation light source unit 3 includes a solid-state laser 30 (the present embodiment adopts a 532nm wavelength, with This is an example, not limited thereto), neutral density filter 31 (neutral density filter 31 can adopt excitation light band-pass filter), some reflection mirrors 32, Glan Taylor prism 33, quarter wave plate 34 , beam expander and collimating mirror 35, lens 36 and two optical filters 37; The continuous wave laser that solid-state laser 30 sends is transmitted to reflecting mirror 32a through neutral density filter 31, and the laser light that exits through reflecting mirror 32a passes successively through gran -Taylor prism 33, quarter-wave plate 34 and reflector 32b emit to beam expander and collimator mirror 35, and the circularly polarized light through beam expander and collimator mirror exits through lens 36, optical filter 37a, reflection successively The mirror 32c and the optical filter 37b are focused on the back focal plane of the microscopic objective lens 40. Since the high-magnification microscopic objective lens 40 has a very high numerical aperture, total internal reflection can be realized, and a higher signal-to-noise ratio than wide-field fluorescence imaging can be achieved. Total internal reflection fluorescence imaging. Wherein, the purpose of beam expansion here is to expand the diameter of the excitation beam to ensure that its size is larger than the light entrance hole of the microscope objective lens 40 to ensure that the excitation light is high-quality circularly polarized light.

在一个优选的实施例中,单分子荧光成像单元5可以采用EMCCD相机和PCI数据采集卡,EMCCD相机通过PCI数据卡连接计算机24,EMCCD相机具备单光子的极高灵敏度,可以获取单个聚合物分子的散焦图像。In a preferred embodiment, single-molecule fluorescence imaging unit 5 can adopt EMCCD camera and PCI data acquisition card, and EMCCD camera is connected computer 24 by PCI data card, and EMCCD camera possesses the extremely high sensitivity of single photon, can obtain single polymer molecule defocused image.

在一个优选的实施例中,待测样品可以是聚苯乙烯荧光小球、金纳米颗粒、量子点、无荧光的聚苯乙烯小球或二氧化硅小球、人体活性细胞;其中,聚苯乙烯荧光小球半径约为100nm,金纳米颗粒和量子点的半径为25nm、30nm等,远小于飞秒脉冲激光器10的束腰半径,无荧光聚苯乙烯小球或二氧化硅小球半径为0.80μm、1.08μm、2.02μm等,接近或大于飞秒脉冲激光器10的束腰半径,人体活性细胞的半径为5μm,远大于飞秒脉冲激光器10的束腰半径,因此,本发明可以捕获不同尺寸的米氏粒子,获得直观的图像。In a preferred embodiment, the sample to be tested can be polystyrene fluorescent beads, gold nanoparticles, quantum dots, non-fluorescent polystyrene beads or silica beads, human active cells; wherein, polystyrene The radius of the ethylene fluorescent ball is about 100nm, the radius of the gold nanoparticles and quantum dots is 25nm, 30nm, etc., which are far smaller than the beam waist radius of the femtosecond pulsed laser 10, and the radius of the non-fluorescent polystyrene ball or silicon dioxide ball is 0.80 μm, 1.08 μm, 2.02 μm, etc., close to or greater than the beam waist radius of femtosecond pulsed laser 10, the radius of human active cells is 5 μm, much larger than the beam waist radius of femtosecond pulsed laser 10, therefore, the present invention can capture different Mie particles of different sizes to obtain intuitive images.

下面通过具体实施例详细说明采用本发明的可获取溶液中胶体粒子动力学及成像信息的飞秒光镊系统的使用过程。The process of using the femtosecond optical tweezers system of the present invention capable of obtaining dynamics and imaging information of colloidal particles in solution will be described in detail below through specific examples.

实施例1:采用本发明获取不同粘度下溶液中单个胶体粒子的动力学信息,具体过程为:Embodiment 1: adopt the present invention to obtain the kinetic information of a single colloidal particle in the solution under different viscosities, the specific process is:

1、将本发明的飞秒光镊系统放置在光学平台上,调整飞秒光镊系统中的各光学器件的位置使得符合本发明飞秒光镊系统的光路传播条件;1. Place the femtosecond optical tweezers system of the present invention on an optical platform, adjust the position of each optical device in the femtosecond optical tweezers system so as to meet the optical path propagation conditions of the femtosecond optical tweezers system of the present invention;

2、将半径为1.08μm聚苯乙烯微球作为待测样品放置于样品池中,样品池底部为0.17mm的盖玻片;2. Place polystyrene microspheres with a radius of 1.08 μm as the sample to be tested in the sample pool, and the bottom of the sample pool is a 0.17 mm cover glass;

3、打开飞秒捕获激光光源单元1的飞秒脉冲激光器10,使得飞秒脉冲激光器10发出的激光经扩束和准直镜12扩大到束径满足显微物镜40的孔径要求,扩束后的激光经准直使之成为平行光;3. Turn on the femtosecond pulse laser 10 of the femtosecond capture laser light source unit 1, so that the laser beam emitted by the femtosecond pulse laser 10 is expanded to a beam diameter that meets the aperture requirements of the microscopic objective lens 40 through beam expansion and collimator mirror 12. After beam expansion The laser beam is collimated to make it parallel light;

4、将显微物镜40切换到油浸显微物镜,在显微物镜40上加入20~100μL镜油,并对其进行调节使之达到合适的聚焦位置;4. Switch the microscopic objective lens 40 to an oil-immersed microscopic objective lens, add 20-100 μL of lens oil to the microscopic objective lens 40, and adjust it so that it reaches a suitable focus position;

5、打开显微镜40的卤素灯,并调节到合适的亮度,通过数字相机或EMCCD相机,初步找到在溶液中悬浮的胶体小球;5. Turn on the halogen lamp of the microscope 40 and adjust it to a suitable brightness, and use a digital camera or EMCCD camera to initially find the colloidal beads suspended in the solution;

6、打开NI数据采集卡23采集数据;6. Open the NI data acquisition card 23 to collect data;

7、结束数据集后将数据导出,做出位置的统计概率分布图(如图3所示),通过Boltzmann统计法校准得到光阱刚度为0.800pN/μm,粘度为0.0013泊(室内温度22℃)。7. After finishing the data set, export the data, make a statistical probability distribution map of the position (as shown in Figure 3), and calibrate through the Boltzmann statistical method to obtain an optical trap with a stiffness of 0.800pN/μm and a viscosity of 0.0013 poise (indoor temperature 22°C ).

8、根据斯托克斯-爱因斯坦公式计算得到待测样品的扩散系数D为0.016μm2/s。8. According to the Stokes-Einstein formula, the diffusion coefficient D of the sample to be tested is calculated to be 0.016 μm 2 /s.

实施例2:采用本发明完成光镊捕获下纳米荧光小球荧光成像,具体过程为:Embodiment 2: Using the present invention to complete fluorescence imaging of nano-fluorescence beads captured by optical tweezers, the specific process is as follows:

1、将本发明的飞秒光镊系统放置在光学平台上,将荧光染料包被的聚苯乙烯小球置于显微物镜4正上方,调整各光学器件使得符合本发明的光路传播条件,1. Place the femtosecond optical tweezers system of the present invention on an optical platform, place the polystyrene pellets coated with fluorescent dye directly above the microscope objective lens 4, adjust each optical device so as to meet the optical path propagation conditions of the present invention,

2、将待测样品放置于样品池中,样品池底部使用0.17mm的盖玻片;2. Place the sample to be tested in the sample cell, and use a 0.17mm cover glass at the bottom of the sample cell;

3、打开激发光源单元3的固体激光器30,使固体激光器30出射的连续波激光经扩束调整后使激发光在显微物镜40的后焦平面处聚焦,从而使得通过显微物镜40后出射为平行光;3. Turn on the solid-state laser 30 of the excitation light source unit 3, so that the continuous-wave laser emitted by the solid-state laser 30 is adjusted by beam expansion to focus the excitation light at the back focal plane of the microscopic objective lens 40, so that it is emitted after passing through the microscopic objective lens 40 for parallel light;

4、将显微物镜40切换到油浸显微物镜,在显微物镜40上加入20-100μL镜油,并对其进行调节使之达到最佳聚焦位置;4. Switch the microscopic objective 40 to an oil-immersed microscopic objective, add 20-100 μL of lens oil to the microscopic objective 40, and adjust it to achieve the best focus position;

5、打开飞秒捕获激光光源单元中1的飞秒脉冲激光器10并对其光强进行调节;5. Turn on the femtosecond pulse laser 10 in the femtosecond capture laser light source unit 1 and adjust its light intensity;

6、打开单分子荧光成像单元5对单个荧光小球进行实时荧光成像(如图4所示),通过荧光信号强度随时间阶梯式的变化从而获取被捕获胶体粒子变化个数的信息。6. Turn on the single-molecule fluorescence imaging unit 5 to perform real-time fluorescence imaging of a single fluorescent bead (as shown in FIG. 4 ), and obtain information on the number of captured colloidal particles through the stepwise change of fluorescence signal intensity with time.

上述各实施例仅用于说明本发明,其中各部件的结构、连接方式和制作工艺等都是可以有所变化的,凡是在本发明技术方案的基础上进行的等同变换和改进,均不应排除在本发明的保护范围之外。The above-mentioned embodiments are only used to illustrate the present invention, wherein the structure, connection mode and manufacturing process of each component can be changed to some extent, and any equivalent transformation and improvement carried out on the basis of the technical solution of the present invention should not excluded from the protection scope of the present invention.

Claims (9)

1. a kind of obtaining the femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution, it is characterised in that the system Including:
Stablize the femtosecond optical tweezer capture laser cell for capturing ligh trap for capturing laser as femtosecond and being provided for sample to be tested;
Position sensing for detecting captured sample to be tested position distribution when doing limited Brownian movement relative to ligh trap center Unit;
Excitation light source unit for carrying out fluorescence excitation as exciting light irradiation sample to be tested;
For femtosecond capture laser to be collected into the position detection unit, and the exciting light that the excitation light source unit is emitted The sample to be tested being introduced into capture ligh trap so that the luminescent dye molecule of sample to be tested, which is stimulated, generates fluorescence signal Optical microphotograph unit;
Fluorescence signal for collecting sample to be tested generation completes the real-time fluorescence of single colloidal particle coated to fluorescent dye Imaging obtains the single molecular fluorescence of the dynamic information of single colloidal particle of the sample to be tested in different viscoelasticity characteristic systems Imaging unit.
2. femtosecond optical optical tweezers system according to claim 1, which is characterized in that the optical microphotograph unit is using inversion fluorescence Microscopic structure, including microcobjective, first~third dichroscope and condenser;Exciting light is focused on through the microcobjective Sample to be tested in capture ligh trap, the fluorescence generated through excitation sample to be tested are sent out after microcobjective collection It is mapped to the first dichroscope, the fluorescence signal being emitted through first dichroscope is emitted to the single molecular fluorescence imaging measurement Unit.
3. femtosecond optical optical tweezers system according to claim 2, which is characterized in that the femtosecond optical tweezer captures laser cell and includes Femtosecond pulse laser expands and collimating mirror and deflecting mirror, the femtosecond pulse warp of the femtosecond pulse laser outgoing Described expand is expanded and is collimated with collimating mirror, and the femtosecond pulse after expanding and collimating is through the deflecting mirror into horizontal deflection After be emitted to the second dichroscope, the light emitting through second dichroscope outgoing is to the microcobjective.
4. femtosecond optical optical tweezers system according to claim 2, which is characterized in that the position detection unit includes assembling thoroughly Mirror, positional detecting device, data collecting card and computer;The interference pattern that forward scattering light through microcobjective outgoing is formed Sample is emitted to third dichroscope after condenser collection, and the light through third dichroscope outgoing is assembled thoroughly through described Mirror focuses the positional detecting device, and the positional detecting device connects the computer by the data collecting card and waited for Viscosity of the stiffness coefficient and sample to be tested of ligh trap residing for sample in different viscoelastic systems.
5. femtosecond optical optical tweezers system according to claim 4, which is characterized in that the positional detecting device uses position sensing Detector or digital camera.
6. femtosecond optical optical tweezers system according to claim 2, which is characterized in that the excitation light source unit includes Solid State Laser Device, quarter-wave plate, expands and collimating mirror, lens and optical filter Glan-Taylor prism;What the solid state laser was sent out Continuous wave successively through the Glan-Taylor prism and quarter-wave plate be emitted to it is described expand and collimating mirror, expanded through described Circularly polarized light with collimating mirror outgoing through the upper lens and optical filter through focusing on the back focal plane of the microcobjective successively.
7. femtosecond optical optical tweezers system according to claim 6, which is characterized in that the solid state laser exit setting is neutral Density filtering piece.
8. femtosecond optical optical tweezers system according to claim 4, which is characterized in that the single molecular fluorescence imaging unit uses EMCCD cameras and pci data capture card, the EMCCD cameras connect the computer through the pci data capture card.
9. femtosecond optical optical tweezers system according to claim 1, which is characterized in that the sample to be tested is that polystyrene fluorescence is small Ball, gold nano grain, non-blooming polystyrene sphere or silicon oxide pellets, Viable human cell.
CN201810353723.8A 2018-04-19 2018-04-19 The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained Pending CN108801863A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201810353723.8A CN108801863A (en) 2018-04-19 2018-04-19 The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201810353723.8A CN108801863A (en) 2018-04-19 2018-04-19 The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained

Publications (1)

Publication Number Publication Date
CN108801863A true CN108801863A (en) 2018-11-13

Family

ID=64092921

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201810353723.8A Pending CN108801863A (en) 2018-04-19 2018-04-19 The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained

Country Status (1)

Country Link
CN (1) CN108801863A (en)

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110631992A (en) * 2019-10-09 2019-12-31 南京理工大学 A feedback device and method for longitudinal positioning of optical tweezers based on fluorescence coupling output
CN110664369A (en) * 2019-09-19 2020-01-10 哈尔滨工业大学 An adaptive confocal line scan harmonic microscopy imaging method and device
CN111366510A (en) * 2020-03-02 2020-07-03 清华大学深圳国际研究生院 Suspended particulate matter flux measuring device utilizing synchronous polarization and fluorescence
CN111965050A (en) * 2020-08-19 2020-11-20 天津大学 A kind of testing method of colloidal particle microfluidic shear force
CN113295888A (en) * 2021-05-25 2021-08-24 中国人民解放军国防科技大学 Method and device for measuring van der waals force by utilizing photon force microscope
CN114324339A (en) * 2022-01-12 2022-04-12 山西大学 A kind of spherical object mass measurement system and measurement method
CN114755200A (en) * 2022-03-21 2022-07-15 北京大学长三角光电科学研究院 Visual monitoring system and method based on photodynamic therapy
CN115112632A (en) * 2022-07-11 2022-09-27 浙江大学 An aerosol chemical reaction device based on dual-camera multi-trap optical tweezers
DE102023123483B3 (en) * 2023-08-31 2025-01-23 Histolution GmbH Laser microscope for histological examination of a tissue sample and histological examination procedure

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090122402A1 (en) * 2007-11-14 2009-05-14 Jds Uniphase Corporation Achromatic Converter Of A Spatial Distribution Of Polarization Of Light
CN102181361A (en) * 2011-03-25 2011-09-14 哈尔滨工业大学(威海) Device and method for sorting cells
CN102743159A (en) * 2012-07-26 2012-10-24 中国科学院自动化研究所 Optical projection tomographic imaging system
CN103604787A (en) * 2013-11-14 2014-02-26 北京大学 Laser scanning phase microimaging method and system

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090122402A1 (en) * 2007-11-14 2009-05-14 Jds Uniphase Corporation Achromatic Converter Of A Spatial Distribution Of Polarization Of Light
CN101470226A (en) * 2007-11-14 2009-07-01 Jds尤尼弗思公司 Achromatic converter of a spatial distribution of polarization of light
CN102181361A (en) * 2011-03-25 2011-09-14 哈尔滨工业大学(威海) Device and method for sorting cells
CN102743159A (en) * 2012-07-26 2012-10-24 中国科学院自动化研究所 Optical projection tomographic imaging system
CN103604787A (en) * 2013-11-14 2014-02-26 北京大学 Laser scanning phase microimaging method and system

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
YUQIANG JIANG等: "Nonlinear optical effects in trapping nanoparticles with femtosecond pulses", 《NATURE PHYSICS》 *
ZHENG ZHONGLI 等: "Segmental dynamics near the chain end of polystyrene in its ultrathin films: a study by single-molecule fluorescence de-focus microscopy", 《SCIENCE CHINA CHEMISTRY》 *
任洪亮: "光镊测量胶体粒子间及界面与粒子间相互作用", 《中国科学技术大学2008年博士学位论文》 *
王锴等: "利用飞秒激光光镊捕获生物细胞", 《光电子·激光》 *

Cited By (14)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110664369A (en) * 2019-09-19 2020-01-10 哈尔滨工业大学 An adaptive confocal line scan harmonic microscopy imaging method and device
CN110631992A (en) * 2019-10-09 2019-12-31 南京理工大学 A feedback device and method for longitudinal positioning of optical tweezers based on fluorescence coupling output
CN111366510B (en) * 2020-03-02 2022-06-03 清华大学深圳国际研究生院 Suspended particulate matter flux measuring device utilizing synchronous polarization and fluorescence
CN111366510A (en) * 2020-03-02 2020-07-03 清华大学深圳国际研究生院 Suspended particulate matter flux measuring device utilizing synchronous polarization and fluorescence
CN111965050A (en) * 2020-08-19 2020-11-20 天津大学 A kind of testing method of colloidal particle microfluidic shear force
CN111965050B (en) * 2020-08-19 2022-06-07 天津大学 A kind of testing method of colloidal particle microfluidic shear force
CN113295888A (en) * 2021-05-25 2021-08-24 中国人民解放军国防科技大学 Method and device for measuring van der waals force by utilizing photon force microscope
CN114324339A (en) * 2022-01-12 2022-04-12 山西大学 A kind of spherical object mass measurement system and measurement method
CN114324339B (en) * 2022-01-12 2024-08-27 山西大学 Spherical object quality measurement system and measurement method
CN114755200A (en) * 2022-03-21 2022-07-15 北京大学长三角光电科学研究院 Visual monitoring system and method based on photodynamic therapy
CN114755200B (en) * 2022-03-21 2022-11-08 北京大学长三角光电科学研究院 Visual monitoring system and method based on photodynamic therapy
CN115112632A (en) * 2022-07-11 2022-09-27 浙江大学 An aerosol chemical reaction device based on dual-camera multi-trap optical tweezers
CN115112632B (en) * 2022-07-11 2023-06-02 浙江大学 Aerosol chemical reaction device based on double-camera multi-optical-trap optical tweezers
DE102023123483B3 (en) * 2023-08-31 2025-01-23 Histolution GmbH Laser microscope for histological examination of a tissue sample and histological examination procedure

Similar Documents

Publication Publication Date Title
CN108801863A (en) The femtosecond optical optical tweezers system of colloidal particle dynamics and image-forming information in solution can be obtained
CN109211847B (en) A method for analyzing the chemical composition of a single suspended particle using an analytical device
CN104568886B (en) A kind of dark field illumination method based on total internal reflection
CN103743720B (en) A kind of confocal Raman microscopy with angle resoluting ability
CN102116929B (en) High-speed wide-field coherent anti-Stokes Raman scattering microscopic system and method
Arpali et al. High-throughput screening of large volumes of whole blood using structured illumination and fluorescent on-chip imaging
CN105699344A (en) Measuring system for obtaining single molecular spectrum and imaging under shear field
CN1912587A (en) Time resolution fluorescence spectral measuring and image forming method and its device
CN111458312B (en) Detection optical system for fluorescent defects of micro-regions on processing surface layer of soft and brittle optical crystal
CN104568857B (en) A kind of two-dimentional light scattering quiescence cells instrument method and device
CN106896095B (en) Composite Surface Plasmon Resonance and Surface Enhanced Raman Microscopic Imaging Technology
CN107861230B (en) Zoom optical tweezers confocal microscopy imaging device and method
CN104793329B (en) A kind of device and method of femtosecond laser rotation manipulation optical tweezer
CN104677830A (en) Spectrophotometric pupil confocal-photoacoustic microimaging device and method
CN107167455A (en) Light splitting pupil laser differential confocal CARS micro-spectrometer method and devices
US20110068279A1 (en) Ultra dark field microscope
CN107167457A (en) The confocal CARS micro-spectrometers method and device of transmission-type
JP2011191199A (en) Microscope system
CN105043988A (en) Single-point deconvolution microscopic system and imaging method based on scanning galvanometers
CN107229133A (en) Based on SiO2Super-resolution imaging method of dielectric microsphere
CN110646872A (en) Raman spectrum scanning imaging system based on SPP thermoelectric optical tweezers
CN105044066B (en) A kind of nanometer OCT image method and system based on broadband stimulated radiation
CN110082330A (en) Combined system is imaged in a kind of laser optical tweezer Raman spectrum and multi-photon
CN104568710A (en) High time-space resolution optical detection and microscopic imaging method and device
JP2004361087A (en) Biomolecule analyzer

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
RJ01 Rejection of invention patent application after publication

Application publication date: 20181113

RJ01 Rejection of invention patent application after publication