基于多参量共模测量的多维光声显微成像技术的研究

Adopted photoacoustic microscopy with high compatibility and high sensitivity, the common mode imaging of the same cell that is used to get the structural and functional information of the cells in the same time and realize the multi-parameter imging of the cells. In this imaging method, two-photon absorption coefficient, optical absorption coeffcient, optical scattering coefficient and viscoelasticity coefficient are used to be the imaging parameters can monitor the dynamic biological behaviour that the cells’ responses to the medication in real time is realized. Aimming at solving the problem of the poor specific imaging capability, we proposed an imaging modality that use two-photon photoacoustic spectrum which has molecular recognition ability. This method can greatly improve the specific imaging capability and be used in functional imaging of the cells. The imaging modality that use optical scattering coefficient as the imaging parameters could be used for photoacoustic microscopy of transparent cells without cell staining. It means the interference of the stain can be avoied. The imaging modality that use viscoelasticity coefficient as the imaging parameters could be used for monitering the cells' activity. Then the four imaging modalities of photoacousitic imaging were integrated to the confocal microscope, and develope the “Photoacoustic-Confocal ”microscopy. This common mode imaging with multi-parameter microscopy has not only high resolution which is the characteristic of nonlinear optical microscopy, but also specific imaging ability. It can be used in researching the biological behaviour that the cells’ responses to the medication at the molecular level. That has important significance in cell morphology,molecular cell biology，targeted pharmacology.

为了同时获得细胞的结构和功能信息，选择能表征细胞不同生物学行为的多个不同物理量如双光子吸收系数、光吸收系数、光散射系数及粘弹系数等作为成像物理量对同一细胞进行共模测量光声显微成像。采用具有分子识别能力的双光子光谱进行光声成像，可以极大提高光声显微术的特异性成像能力，实现分子成像和细胞的功能成像；利用光散射系数进行光声显微成像，可以对透明细胞进行光声显微成像，无需对细胞进行染色，从而避免了染色剂对药物的干扰；采用粘弹系数进行光声显微成像，可以监测细胞的活性及其对药物的反应机制。进而采用系统集成方法，将多种光声成像模式集成到共焦显微镜上，研制成功多参量共模测量的多维光声显微镜。这种多参量共模显微成像技术，既具有非线性显微镜的高分辨率特点，又具有特异性成像的能力，可以在分子水平上实时监测细胞的各种生物学行为，在细胞形态学、分子细胞生物学、靶向药物学和药理学等研究领域具有重要意义。

传统光声显微术是以物质的线性光谱吸收系数做为成像物理量进行成像，由于线性吸收光谱系数的光谱分辨率低，导致传统光声显微术的特异性成像能力差，很难实现生物组织的功能成像。针对这一技术难题，本项目提出利用生物组织的弱吸收系数、双光子吸收系数、光散射系数以及双折射系数作为成像物理量，对生物组织进行多维成像，从而获得生物组织更多的有用信息，以实现生物组织的功能成像。为此，本项目首先设计研制成功了适应多参量共模测量的微腔光声探测器和共振型微腔光声探测器（亥姆霍兹腔），实现了弱吸收-光声测量、双光子-光声测量、散射-光声测量。在此基础上，把微腔光声探测器集成到共焦显微成像系统上，以弱吸收系数、双光子吸收系数、光散射系数作为成像物理量，实现弱吸收系数、双光子吸收系数、光散射系数的共模光声显微成像--称为多维光声显微术。在此基础上，进而把多维光声显微术与光热层析显微成像技术、偏振态显微成像集成在一起，对同一目标进行弱吸收系数、双光子吸收系数、光散射系数和双折射系数的共模成像，从而实现多参量共模显微成像，研制成功多参量共模测量的多维光声显微镜。双光子光声显微成像模式，可以提高光声显微术的特异性成像能力，实现分子成像；光散-光声显微成像模式，可以对透明细胞进行光声显微成像，无需对细胞进行染色，从而避免了染色剂对药物的干扰；偏振态成像模式，由于采用双折射系数进行成像，可以获得细胞膜的厚度和结构信息。这种多参量共模显微成像技术，可以对同一目标同时进行多参量测量和多维成像，可以在分子水平上实时监测细胞的各种生物学行为，在细胞形态学、分子细胞生物学、靶向药物学和药理学等研究领域具有重要意义。