Multiplane HiLo microscopy with speckle illumination and non-local means denoising.
Multiplane HiLo microscopy with speckle illumination and non-local means denoising.
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DOI：
10.1117/1.jbo.28.11.116502
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发表时间：
2023-11
影响因子：
3.5
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中科院分区：
文献类型：
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HiLo显微术将两幅图像合成为光学切片图像，其中一幅图像由均匀照明获得，另一幅图像由图案照明获得，如激光散斑。基于散斑的HiLo具有对像差的鲁棒性，但容易受到难以控制的残余散斑噪声的影响。我们提出了一种不影响分辨率的计算方法来减少这种残余噪声。此外，我们提高了HiLo显微镜的通用性，使同时多平面成像（这里有九个平面）。我们的目标是使用传统的基于相机的荧光显微镜进行快速、高对比度、多平面成像。多平面HiLo成像是通过使用单相机和z-分光棱镜实现的。散斑降噪是基于应用非局部均值（NLM）去噪方法对散斑颗粒进行集合平均。我们通过合成数据和在40 Hz帧率下对斑马鱼幼体的心脏和大脑活动进行成像，证明了具有NLM去噪的多平面HiLo的能力。借助NLM去噪的多平面HiLo显微镜为快速光学切片体成像提供了一种简单的工具，可用于荧光成像应用。
HiLo microscopy synthesizes an optically sectioned image from two images, one obtained with uniform and another with patterned illumination, such as laser speckle. Speckle-based HiLo has the advantage of being robust to aberrations but is susceptible to residual speckle noise that is difficult to control. We present a computational method to reduce this residual noise without undermining resolution. In addition, we improve the versatility of HiLo microscopy by enabling simultaneous multiplane imaging (here nine planes). Our goal is to perform fast, high-contrast, multiplane imaging with a conventional camera-based fluorescence microscope. Multiplane HiLo imaging is achieved with the use of a single camera and z-splitter prism. Speckle noise reduction is based on the application of a non-local means (NLM) denoising method to perform ensemble averaging of speckle grains. We demonstrate the capabilities of multiplane HiLo with NLM denoising both with synthesized data and by imaging cardiac and brain activity in zebrafish larvae at 40 Hz frame rates. Multiplane HiLo microscopy aided by NLM denoising provides a simple tool for fast optically sectioned volumetric imaging that can be of general utility for fluorescence imaging applications.