基于高倍物镜的相位显微成像方法

Phase Microscopy Using High Magnification Objective Lens

当前基于干涉技术的相位显微成像方法已经被广泛研究,但基于非干涉方法(相位恢复)的相位显微成像研究还很少.本文将相位恢复与显微成像技术相结合,提出并论证了一种基于高倍物镜的相位显微成像方法,适用于微米级相位型物体的显微成像.该方法简单有效,针对高倍物镜显微系统的特性,采用同时移动CCD及显微物镜的拍摄方法,解决了传统拍摄模式下,高倍物镜的采样距离无法满足成像关系的问题.通过CCD采集物面在不同传播的距离衍射放大像,利用测得的强度信息重建物体相位信息.采用直径5μm的聚苯乙烯微球作为待重建样品,使用100×物镜进行显微成像实验.相位分布重建结果反映了样品真实高度的分布变化,证明了该方法的有效性.与基于干涉技术的相位显微方法相比,该方法所使用的实验装置更加简单,易于操作且成本更低.对于生物细胞或组织等相位型样品的厚度或折射率分布测量具有实际应用意义.

Phase microscopy based on interference technology has been widely studied,while phase microscopy based on non-interference method(phase retrieval)has rarely been studied.A phase micro-imaging technique based on phase retrieval method was proposed and demonstrated,which was applicable for micron-scale phase objects imaging with a high magnification objective lens.The proposed high magnification imaging system had a problem that the sampling distance cannot satisfy the imaging formula.The problem was solved by moving the CCD and objective lens simultaneously.The phase distribution was reconstructed by the magnified intensity information measured at different diffraction planes.Phase imaging experiment with a 100×magnification lens was presented using the polystyrene microspheres with 5μm diameter as the sample.The phase reconstruction results reflected the height distributions of the specimen,which proved the effectiveness of the proposed method.The proposed microscopy technique had advantages in less complexity,easier in operation and lower in cost,comparing with the experiment setup used in the interferometry method.This approach was applicable for refractive index or thickness measurements of biological cells or tissue.