Department of Chemical Engineering and Biotechnology, University of Cambridge, CB3 0AS, Cambridge, UK.
Department of Physics and Technology, UiT-The Arctic University of Norway, 9037, Tromsø, Norway.
Sci Rep. 2019 Apr 23;9(1):6425. doi: 10.1038/s41598-019-42743-4.
The deconvolution of widefield fluorescence images provides only guesses of spatial frequency information along the optical axis due to the so called missing cone in the optical transfer function. Retaining the single-shot imaging speed of deconvolution microscopy while gaining access to missing cone information is thus highly desirable for microscopy of volumetric samples. Here, we present a concept that superimposes two orthogonally polarized excitation lattices with a phase-shift of p between them. In conjunction with a non-iterative image reconstruction algorithm this permits the restoration of missing cone information. We show how fluorescence anisotropy could be used as a method to encode and decode the patterns simultaneously and develop a rigorous theoretical framework for the method. Through in-silico experiments and imaging of fixed biological cells on a structured illumination microscope that emulates the proposed setup we validate the feasibility of the method.
宽场荧光图像的反卷积仅能提供沿光轴的空间频率信息的猜测，这是由于在光学传递函数中存在所谓的“缺失锥体”。因此，在保持反卷积显微镜单次成像速度的同时，获取缺失锥体信息对于体积样品的显微镜观察非常重要。在这里，我们提出了一种概念，即将两个正交偏振的激发晶格叠加在一起，并在它们之间进行 p 相移。结合非迭代图像重建算法，这允许恢复缺失锥体信息。我们展示了如何将荧光各向异性用作同时编码和解码图案的方法，并为该方法开发了严格的理论框架。通过在模拟所提出的设置的结构光照明显微镜上对固定生物细胞进行的计算机模拟实验和成像，验证了该方法的可行性。
