Northwestern University, Department of Biomedical Engineering, Evanston, Illinois, United States.
Northwestern University, Department of Electrical Engineering, Evanston, Illinois, United States.
J Biomed Opt. 2017 Mar 1;22(3):30901. doi: 10.1117/1.JBO.22.3.030901.
Optical microscopy is the staple technique in the examination of microscale material structure in basic science and applied research. Of particular importance to biology and medical research is the visualization and analysis of the weakly scattering biological cells and tissues. However, the resolution of optical microscopy is limited to ? 200 ?? nm due to the fundamental diffraction limit of light. We review one distinct form of the spectroscopic microscopy (SM) method, which is founded in the analysis of the second-order spectral statistic of a wavelength-dependent bright-field far-zone reflected-light microscope image. This technique offers clear advantages for biomedical research by alleviating two notorious challenges of the optical evaluation of biomaterials: the diffraction limit of light and the lack of sensitivity to biological, optically transparent structures. Addressing the first issue, it has been shown that the spectroscopic content of a bright-field microscope image quantifies structural composition of samples at arbitrarily small length scales, limited by the signal-to-noise ratio of the detector, without necessarily resolving them. Addressing the second issue, SM utilizes a reference arm, sample arm interference scheme, which allows us to elevate the weak scattering signal from biomaterials above the instrument noise floor.
光学显微镜是基础科学和应用研究中检查微观材料结构的主要技术。对生物学和医学研究特别重要的是对弱散射生物细胞和组织的可视化和分析。然而,由于光的基本衍射极限,光学显微镜的分辨率限制在? 200??nm。我们回顾了光谱显微镜 (SM) 方法的一种独特形式,该方法基于对波长相关明场远场反射光显微镜图像的二阶光谱统计的分析。该技术通过缓解生物材料光学评估的两个臭名昭著的挑战,为生物医学研究提供了明显的优势:光的衍射极限和对生物、光学透明结构的缺乏敏感性。为了解决第一个问题,已经表明,明场显微镜图像的光谱内容可以定量地描述样品的结构组成,其长度尺度任意小,受限于探测器的信噪比,而不一定需要解析它们。为了解决第二个问题,SM 利用参考臂和样品臂干涉方案,使我们能够将生物材料的弱散射信号提升到仪器噪声基底之上。