WO2011097851A1 - 细胞光声显微成像方法及其装置 - Google Patents

细胞光声显微成像方法及其装置 Download PDF

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WO2011097851A1
WO2011097851A1 PCT/CN2010/073274 CN2010073274W WO2011097851A1 WO 2011097851 A1 WO2011097851 A1 WO 2011097851A1 CN 2010073274 W CN2010073274 W CN 2010073274W WO 2011097851 A1 WO2011097851 A1 WO 2011097851A1
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photoacoustic
photo
cell
acoustic
laser
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PCT/CN2010/073274
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French (fr)
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唐志列
谭治良
吴泳波
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华南师范大学
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N29/00—Investigating or analysing materials by the use of ultrasonic, sonic or infrasonic waves; Visualisation of the interior of objects by transmitting ultrasonic or sonic waves through the object
    • G01N29/04—Analysing solids
    • G01N29/06—Visualisation of the interior, e.g. acoustic microscopy
    • G01N29/0654—Imaging
    • G01N29/0681—Imaging by acoustic microscopy, e.g. scanning acoustic microscopy
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N29/00—Investigating or analysing materials by the use of ultrasonic, sonic or infrasonic waves; Visualisation of the interior of objects by transmitting ultrasonic or sonic waves through the object
    • G01N29/22—Details, e.g. general constructional or apparatus details
    • G01N29/24—Probes
    • G01N29/2418—Probes using optoacoustic interaction with the material, e.g. laser radiation, photoacoustics

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  • the present invention relates to microscopic imaging techniques, in particular to a cellular photoacoustic microscopy imaging method, and to a cellular photoacoustic microscopy imaging device.
  • Photoacoustic imaging is performed by a method of 'light excitation-induced photoacoustic signal-photoacoustic detection-image reconstruction'.
  • Photoacoustic imaging technology has significant advantages over many traditional imaging technologies in image resolution, contrast and information volume.
  • photoacoustic imaging methods mainly include the following three imaging methods: acoustic lens imaging, phase-controlled focusing imaging and inversion. Imaging method.
  • the acoustic lens imaging method can only achieve a resolution of 1 mm due to the acoustic diffraction effect and the aberration of the acoustic lens.
  • the phase-controlled focusing imaging method uses the multi-line array ultrasonic transducer combined with the phase-controlled focusing algorithm for image reconstruction.
  • the resolution reaches 100 micrometers, which can image blood vessels.
  • the inversion imaging method uses Radon inversion algorithm and filtered back projection algorithm for image reconstruction. Because it avoids the limitation of sonic diffraction effect, it can achieve higher resolution.
  • the rate of photoacoustic imaging with a resolution of 15 microns, allows imaging of capillaries. However, this resolution still does not distinguish cells, and it is impossible to image cells.
  • photoacoustic imaging technology can obtain characteristic information such as biological behavior (such as growth, apoptosis, metabolism, pathology, mutation) of the tissue, thereby achieving biological function imaging.
  • biological behavior such as growth, apoptosis, metabolism, pathology, mutation
  • the resolution of 15 micrometers can only be achieved, and structural images and functional images at the cellular level cannot be observed.
  • the cell photoacoustic microscopy imaging method of the present invention comprises:
  • the laser is irradiated onto the cells through the coverslip, and the cells absorb the light to produce a photoacoustic effect, and the sound pressure change generated by the photoacoustic effect is transmitted to the photoacoustic sensor, and then The photoacoustic sensor detects and outputs a photoacoustic signal to realize photoacoustic detection on a single cell;
  • the cell photoacoustic microscopic imaging device of the present invention comprises a laser scanning imaging mechanism, a photoacoustic sensor and a signal processor.
  • the laser scanning imaging mechanism is composed of a laser, a scanning galvanometer and a microscope objective lens;
  • the signal processor is composed of a chopper, a lock-in amplifier, a capture card and a computer in turn.
  • the laser light emitted by the laser 1 is modulated by the chopper 2, enters the scanning galvanometer 3 for two-dimensional scanning, and then focused by the microscope objective 4 onto the surface of the sample 5, and the photoacoustic sound generated by the sample 5
  • the signal is detected by the photoacoustic sensor 6, and the output signal of the photoacoustic sensor 6 is amplified and processed by the lock-in amplifier 7, and then sent to the acquisition card 8 for data acquisition, and then stored and reconstructed by the computer 9.
  • the invention has the following advantages:
  • the signal is then photoacoustic sensor for photoacoustic detection, and the photoacoustic microscopy of the cells is performed with a resolution of less than 1 micron.
  • Figure 1 is a block diagram showing the structure of a photoacoustic microscopic imaging device of the present invention
  • 1 is a laser
  • 2 is a chopper
  • 3 is a laser scanning galvanometer
  • 4 is a microscope objective.
  • 5 is the sample
  • 6 is the photoacoustic sensor
  • 7 is the lock-in amplifier
  • 8 is the acquisition card
  • 9 is the computer;
  • Figure 2 is an optical microscopic image of onion cells
  • Figure 3 is a photoacoustic microscopic image of onion cells.
  • the apparatus of the present invention comprises a laser scanning imaging mechanism, a photoacoustic sensor and a signal processor.
  • the laser scanning imaging mechanism is composed of a laser 1, a scanning galvanometer 3, and a microscope objective lens 4; during operation, the scanning galvanometer 3, the microscope objective lens 4 and the photoacoustic sensor 6 can be mounted on the microscope bracket;
  • the signal processor consists of a chopper 2, a lock-in amplifier 7, a capture card 8 and a computer 9, which are in turn electrically connected.
  • the working process of the present invention is as follows: the laser light emitted by the laser 1 is modulated by the chopper 2, then enters the laser scanning galvanometer 3 for two-dimensional scanning, and then focused by the microscope objective 4 onto the surface of the sample 5, and the sample 5 is produced.
  • the photoacoustic signal is detected by the photoacoustic sensor 6, and the output signal of the photoacoustic sensor 6 is amplified and processed by the lock-in amplifier 7, and then sent to the acquisition card 8 for data acquisition, and then stored and reconstructed by the computer 9.
  • the experimental results are shown in FIG. 2 and FIG. 3.
  • the photoacoustic microscopic imaging of the present invention has the advantages of large amount of information and high contrast, and can obtain not only structural information of cells but also function of cells. information.
  • the cells in the figure are clearly contoured and can show a single cell image.

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  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Pathology (AREA)
  • General Health & Medical Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Acoustics & Sound (AREA)
  • Optics & Photonics (AREA)
  • Investigating Or Analyzing Materials By The Use Of Ultrasonic Waves (AREA)
  • Investigating Or Analysing Materials By Optical Means (AREA)
  • Microscoopes, Condenser (AREA)

Description

细胞光声显微成像方法及其装置 技术领域
本发明涉及显微成像技术，具体是一种细胞光声显微成像方法，本发明还涉及细胞光声显微成像装置。
背景技术
光声成像是采用'光激发-诱导光声信号-光声探测-图像重建'的方法进行成像。光声成像技术在图像分辨率、对比度和信息量等方面比许多传统影像技术具有显著优势，目前光声成像方法主要包括以下三种成像方法：声透镜成像法、相控聚焦成像法和反演成像法。声透镜成像法由于受声波衍射效应和声透镜像差的限制，分辨率只能达到1毫米的水平；相控聚焦成像法是利用多元线阵超声换能器结合相控聚焦算法进行图像重构，其分辨率达到了100微米，可以对血管进行成像；反演成像法是利用Radon反演算法和滤波反投影算法进行图像重构，由于避免了声波衍射效应的限制，从而可以实现较高分辨率的光声成像，其分辨率达到了15微米，可以对毛细血管进行成像。但这种分辨率仍无法分辨细胞，更无法对细胞进行成像。
由于光声成像技术可以获得组织的生物学行为（如生长、凋亡、代谢、病变、突变）等特征信息，从而实现生物功能成像。然而，由于目前光声成像技术的分辨率还比较低，只能达到15微米的分辨率，无法观察细胞水平的结构图像和功能图像。
发明内容
本发明的目的是提供一种能观察到细胞水平的高分辨率细胞光声显微成像方法，实现高分辨率光声显微成像，分辨率小于1微米。
本发明的目的还在于提供一种细胞光声显微成像装置。
本发明的细胞光声显微成像方法包括：
--将细胞置于盖玻片的凹面上，激光透过盖玻片照射在细胞上，细胞吸收光产生光声效应，由该光声效应所产生的声压变化传递到光声传感器，然后由光声传感器进行探测并输出光声信号，对单细胞实现光声探测；
--通过光束扫描和高分辨率的显微物镜，对细胞进行二维扫描，实现单细胞的光声显微成像。
本发明的细胞光声显微成像装置包括激光扫描成像机构、光声传感器和信号处理器三部分。
其中，激光扫描成像机构由激光器、扫描振镜、显微物镜构成；信号处理器由斩波器、锁相放大器、采集卡和计算机依次电气连接构成。
如图1所示，激光器1发出的激光，通过斩波器2进行调制，进入扫描振镜3进行二维扫描，再由显微物镜4聚焦到样品5的表面，样品5所产生的光声信号，由光声传感器6进行探测，光声传感器6的输出信号经过锁相放大器7放大和处理后，输送到采集卡8进行数据采集，再由计算机9进行存储和图像重建。
本发明与现有技术相比，具有如下优点：
1 、采用无机械噪声的光束扫描技术结合高分辨率的显微物镜，产生具有高空间分辨率的光声
信号，再采用光声传感器进行光声探测，对细胞进行光声显微成像，分辨率小于1微米。
2 、采用连续激光进行激发，无须采用脉冲激光来进行光声成像。
3 、不仅获得细胞的结构信息，还可对细胞的物质成份进行成像，获得细胞的功能信息。
附图说明
图1是本发明细胞光声显微成像装置结构框图；
图中：1是激光器， 2是斩波器， 3是激光扫描振镜， 4是显微物镜， 5是样品，6是光声传感器，7是锁相放大器，8是采集卡，9是计算机；
图2是洋葱细胞的光学显微图像；
图3是洋葱细胞的光声显微图像。
具体实施方式
如图1所示，本发明装置包括激光扫描成像机构、光声传感器和信号处理器三部分。其 中，激光扫描成像机构由激光器1、扫描振镜3、显微物镜4构成；工作时，扫描振镜3、显微物镜4和光声传感器6可以安装在显微镜支架上；
信号处理器由斩波器2、锁相放大器7、采集卡8和计算机9组成，它们依次电气连接。
本发明的工作过程如下：激光器1发出的激光，通过斩波器2进行调制，然后进入激光扫描振镜3进行二维扫描，再由显微物镜4聚焦到样品5的表面，样品5所产生的光声信号，由光声传感器6进行探测，光声传感器6的输出信号经过锁相放大器7放大和处理后，输送到采集卡8进行数据采集，再由计算机9进行存储和图像重建。实验结果如图2、图3所示，与光学显微成像相比，本发明的光声显微成像具有信息量大、对比度高等优点，不仅可以获得细胞的结构信息，还可以获得细胞的功能信息。图中细胞轮廓清晰，可以显示出单个细胞图像。

Claims (3)

1 、一种细胞光声显微成像方法，其特征在于：
--将细胞置于盖玻片的凹面上，激光透过盖玻片照射在细胞上，细胞吸收光产生光声效应，由该光声效应所产生的声压变化传递到光声传感器，然后由光声传感器进行探测并输出光声信号，对单细胞实现光声探测；
--通过光束扫描和高分辨率的显微物镜，对细胞进行二维扫描，实现单细胞的光声显微成像。
2 、一种细胞光声显微成像装置，其特征在于包括激光扫描成像机构、光声传感器和信号处理
器三部分；其中，激光扫描成像机构由激光器、扫描振镜、显微物镜构成；信号处理器由斩波器、锁相放大器、采集卡和计算机依次电气连接构成。
3 、根据权利要求2所示的装置，其特征在于激光器（1）发出的激光，通过斩波器（2进行调制，进入扫描振镜（3）进行二维扫描，再由显微物镜（4）聚焦到样品（5）的表面，样品（5）所产生的光声信号，由光声传感器（6）进行探测，光声传感器（6）的输出信号经过锁相放大器（7）放大和处理后，输送到采集卡（8）进行数据采集，再由计算机（9）进行存储和图像重建。
PCT/CN2010/073274 2010-02-11 2010-05-26 细胞光声显微成像方法及其装置 WO2011097851A1 (zh)

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CN111948147B (zh) * 2020-09-25 2023-07-25 广东工业大学 一种无盲区的全场域超声显微镜成像系统及其方法
CN112444492A (zh) * 2020-10-09 2021-03-05 中国科学院深圳先进技术研究院 一种光声成像方法、计算机设备、系统以及存储介质
CN112444492B (zh) * 2020-10-09 2023-05-12 中国科学院深圳先进技术研究院 一种光声成像方法、计算机设备、系统以及存储介质
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