四维飞秒CARS共焦显微成像方法研究

Coherent anti-Stokes Raman scattering (CARS) is a spectral analysis tool with high spectral resolution, but the problems, such as monochromatic testing, low imaging contrast factor due to nonresonant disturbance,poor real-time performance due to point-by-point imaging , etc, have influenced the applications of CARS microscopy to the study of human tissues and cells in vivo. The research project proposes a four dimensional femto-second CARS confocal microscopy, which combines a femto-second laser parallel confocal microscopy with a four-dimensional reconstruction theory and algorithms including three dimensional space and one dimensional spectrum: accomplishes a fast broad spectrum microscopy based on a multiple CARS and acousto-optic tunable method; utilizes polarization interference with phase adjusting to enhance contrast ratio; adopts a microlens and micro-pin array to accomplish the laser parallel confocal microscopy to acquire wide-field CARS signals. The project researches on the theory, evaluation of image quality and corresponding experiments, and fusion and reconstruction algorithms of the four-dimensional femto-second CARS confocal microscopy, to solve the key problems existing in conventional CARS microscopies, such as poor real-time performance, low spatial and spectral resolution, etc. The achievements of this project will supply an approach of multiple dimensional imaging for human cells, chromosomes, etc, in vivo and have a broad application perspective in biology and iatrology.

相干反斯托克斯拉曼散射(CARS)是一种具有高光谱分辨率的分析工具, 但探测波长单一、非共振背景噪声强、逐点成像实时性差等问题影响了CARS显微成像在人体活体组织及细胞研究中的应用.项目提出一种四维飞秒CARS共焦显微成像方法, 将飞秒激光并行共焦显微成像方法与包括空间及光谱维的四维重建理论及算法结合: 拟基于飞秒多元CARS与声光可调滤波方法实现快速宽光谱显微成像;采用偏振干涉相角调节方法提高CARS显微图像对比度;采用微透镜及微针孔阵列方法实现飞秒激光并行共焦显微, 以获取宽场CARS信号.项目进行四维飞秒CARS共焦显微成像的理论、像质影响与实验研究, 并进行四维CARS显微图像融合及重建算法研究, 以解决目前CARS显微成像中的实时性、空间及光谱分辨等关键问题. 项目研究成果可提供一种人体细胞、染色体等微小活体组织结构的多维成像方法,在医学领域有较大应用前景.

相干反斯托克斯拉曼散射(CARS)是一种具有高光谱分辨率的分析工具, 但探测波长单一、非共振背景噪声强、逐点成像实时性差等问题影响了CARS显微成像在人体活体组织及细胞研究中的应用.项目提出一种四维飞秒CARS共焦显微成像方法, 将飞秒激光并行共焦显微成像方法与包括空间及光谱维的四维重建理论及算法结合: 拟基于飞秒多元CARS与声光可调滤波方法实现快速宽光谱显微成像;采用偏振干涉相角调节方法提高CARS显微图像对比度;采用微透镜及微针孔阵列方法实现飞秒激光并行共焦显微, 以获取宽场CARS信号.项目进行四维飞秒CARS共焦显微成像的理论、像质影响与实验研究, 并进行四维CARS显微图像融合及重建算法研究, 以解决目前CARS显微成像中的实时性、空间及光谱分辨等关键问题. 项目研究成果可提供一种人体细胞、染色体等微小活体组织结构的多维成像方法,在医学领域有较大应用前景.. 项目取得的成果包括：. 构建了超连续谱激发的时间分辨CARS光谱探测系统，探测到分子振动频率范围在477cm-1-3622cm-1的CARS信号光谱。利用CARS光谱探测系统对二甲基亚砜样品进行了在分子振动模式的光谱探测，获得了波数在500cm-1-3500cm-1范围内CARS光谱信号，由飞秒脉冲泵浦光子晶体光纤产生的超连续谱激光作为泵浦光和斯托克斯光，能够引起此范围内分子的不同分子振动模式的共振加强。实验结果可知利用探测光与超连续谱激光之间适当的时间延迟，可以有效的抑制非共振背景噪声。此外对不同温度下对CARS光谱信号的影响进行了实验研究，结果表明，温度对典型的分子振动模式对应的CARS光谱频移没有影响，只是在温度升高时，CARS信号会有所加强。构建了飞秒激光泵浦双光子共焦荧光扫描成像系统，获取了空间分辨率为10微米的双光子荧光共焦扫描显微图像。. 项目共计完成资助论文15篇，其他论文1篇，申请(授权)发明专利9项。