Laser speckle contrast imaging (LSCI) is a powerful tool for monitoring blood flow changes in tissue or vessels in vivo, but its applications are limited by shallow penetration depth under reflective imaging configuration. The traditional LSCI setup has been used in transmissive imaging for depth extension up to 2lt–3lt (lt is the transport mean free path), but the blood flow estimation is biased due to the depth uncertainty in large depth of field (DOF) images. In this study, we propose a transmissive multifocal LSCI for depth-resolved blood flow in thick tissue, further extending the transmissive LSCI for tissue thickness up to 12lt. The limited-DOF imaging system is applied to the multifocal acquisition, and the depth of the vessel is estimated using a robust visibility parameter Vr in the coherent domain. The accuracy and linearity of depth estimation are tested by Monte Carlo simulations. Based on the proposed method, the model of contrast analysis resolving the depth information is established and verified in a phantom experiment. We demonstrated its effectiveness in acquiring depth-resolved vessel structures and flow dynamics in in vivo imaging of chick embryos.

为进一步提高双光子多焦点结构光照明显微技术（2P-MSIM）的空间分辨率，笔者提出并发展了一种双光子亚衍射多焦点结构光照明显微成像方法（2P-sMSIM）。首先，通过改进的Gerchberg-Saxton（GS）相位恢复算法设计亚衍射聚焦点阵，生成相位图，利用高速相位型空间光调制器产生亚衍射聚焦点阵。通过计算机模拟的仿真实验，探究算法的可行性，并通过对荧光染料溶液的激发成像，证明了每个亚衍射聚焦点阵的平均尺寸为正常衍射受限点阵聚焦点尺寸的80%。其次，将该点阵引入2P-MSIM系统，对固定在BS-C-1细胞内的微管和商用线粒体切片分别进行了超分辨成像实验，证明了在亚衍射聚焦点阵激发下，2P-MSIM的分辨率和成像质量得到了进一步提高，这对于2P-MSIM的发展具有重要意义。

为了进一步提高成像速度和分辨率，提出了基于双螺旋点扩展函数（DH-PSF）工程的多焦点双光子激光扫描显微成像方法和系统（DH-MTPLSM）。在激发光路中，通过高速相位型空间光调制器（SLM）同时实现了三维多焦点阵列的产生和在样品面上的高精度并行数字寻址扫描；在探测光路中，通过双螺旋相位片将系统探测PSF调制为DH-PSF，从而提供样品的轴向信息，减少轴向扫描层数，进而提高三维成像速度；结合基于DH-PSF的数字重聚焦算法，恢复出不同深度样本的宽场图像，通过单次二维扫描获得样品的三维光切片信息。在此基础上，利用搭建的DH-MTPLSM系统开展了小鼠肾组织切片的双光子成像实验，验证了该方法的快速三维高分辨成像能力，这对于MTPLSM的发展具有重要的意义。

为了解决传统立体显示器成像不满足人眼正常成像规律的问题,同时考虑到穿戴设备兼具质量小、体积小的特点,在计算分析光学系统参数的基础上,结合数字微镜元件(DMD)和压电可变形反射镜(PDM),利用Zemax软件设计出了具有多焦平面投影功能的光学系统。该光学系统由7片透镜组成,总长为200 mm,视场角为40°,采用双远心光路结构。对光学系统的整体分析结果表明,改变PDM的曲率半径,可实现多焦平面的成像。人眼根据自身的调节作用,在特定位置处可观察到由各个焦面位置处(屈光度范围为0~3 m ^-1)的二维图像重叠所带来的整体三维效果。最后对系统的成像质量进行分析,结果表明该系统在极限分辨率为37 lp/mm时,各视场处的调制传递函数(MTF)均高于0.4,性能良好,满足设计要求。

为了实现人工晶状体植入人眼后在远、中、近三个视距范围内均清晰成像，帮助白内障患者获得更好的视觉体验，设计了多焦点菲涅耳透镜，其环带面采用自由曲面的形式。通过每个环带交替控制焦距的方法，避免了随着瞳孔收缩而出现的焦点丢失的问题。基于折射定律的矢量形式，采取迭代计算的方法，设计了自由曲面形式的菲涅耳透镜，并模拟了不同焦点处的能量分布。基于ISO 11979-2标准眼模型模拟了自由曲面菲涅耳三焦点人工晶状体植入眼成像系统，附加光焦度分别为+1.66 D和+3.32 D。通过光线追迹计算了不同光焦度下的调制传递函数；计算结果表明：在空间频率为50 lp/mm时，远焦点、中焦点及近焦点处调制传递函数均在0.2以上，满足患者的使用需求。

随着人口老龄化,年龄相关性白内障患者日益增多,通过植入人工晶状体可以使白内障患者的视力基本得到恢复。由于白内障患者对术后的视觉质量要求越来越高,个性化设计的人工晶状体已逐渐在临床上推广使用,而多焦点人工晶状体的出现,则成功解决了单焦点人工晶状体术后难以解决的近视力问题。对人工晶状体的发展历史做了简要的阐述,详细地说明了单焦点和多焦点人工晶状体的设计思想及特点,介绍了多焦点人工晶状体的发展现状,最后对人工晶状体未来的发展进行了展望。

Multifocal multiphoton microscopy (MMM) has greatly improved the utilization of excitation light and imaging speed due to parallel multiphoton excitation of the samples and simultaneous detection of the signals, which allows it to perform three-dimensional fast fluorescence imaging. Stochastic scanning can provide continuous, uniform and high-speed excitation of the sample, which makes it a suitable scanning scheme for MMM. In this paper, the graphical programming language — LabVIEW is used to achieve stochastic scanning of the two-dimensional galvo scanners by using white noise signals to control the x and y mirrors independently. Moreover, the stochastic scanning process is simulated by using Monte Carlo method. Our results show that MMM can avoid oversampling or subsampling in the scanning area and meet the requirements of uniform sampling by stochastically scanning the individual units of the N × N foci array. Therefore, continuous and uniform scanning in the whole field of view is implemented.

多焦点多光子显微（MMM）和单点扫描多光子显微相比，可显著提高光能利用率和成像速度，在生命科学研究领域具有广泛的应用前景。但现有MMM 技术成像质量和成像速度之间存在矛盾，增加单位面积内的焦点数目可提高成像速度，但过小的焦点间距离会产生子光束串扰，从而影响纵向空间分辨率。利用Zemax 软件模拟产生了高密度激发点阵，将传统的不小于6 mm 的相邻焦点间距缩小到约3 mm；分析了高密度点阵激发成像产生噪声的原因；根据时间复用技术的原理设计正方形时间延板，对相邻子光束脉冲进行不同时间的延迟，从而可以消除MMM 系统中高密度激发点阵子光束间的串扰。

Multifocal multiphoton microscopy (MMM) has recently become an important tool in biomedicine for performing three-dimensional fast fluorescence imaging. Using various beamsplitting techniques, MMM splits the near-infrared laser beam into multiple beamlets and produces a multifocal array on the sample for parallel multiphoton excitation and then records fluorescence signal from all foci simultaneously with an area array detector, which significantly improves the imaging speed of multiphoton microscopy and allows for high efficiency in use of the excitation light. In this paper, we discuss the features of several MMM setups using different beamsplitting devices, including a Nipkow spinning disk, a microlens array, a set of beamsplitting mirrors, or a diffractive optical element (DOE). In particular, we present our recent work on the development of an MMM using a spatial light modulator (SLM).