Kühn Jonas, Niraula Bimochan, Liewer Kurt, Kent Wallace J, Serabyn Eugene, Graff Emilio, Lindensmith Christian, Nadeau Jay L
Jet Propulsion Laboratory, California Institute of Technology, 4800 Oak Grove Dr., Pasadena, California 91009, USA.
Department of Biomedical Engineering, McGill University, 3775 University St., Montreal, Quebec H3A 2B4, Canada.
Rev Sci Instrum. 2014 Dec;85(12):123113. doi: 10.1063/1.4904449.
Digital holographic microscopy is an ideal tool for investigation of microbial motility. However, most designs do not exhibit sufficient spatial resolution for imaging bacteria. In this study we present an off-axis Mach-Zehnder design of a holographic microscope with spatial resolution of better than 800 nm and the ability to resolve bacterial samples at varying densities over a 380 μm × 380 μm × 600 μm three-dimensional field of view. Larger organisms, such as protozoa, can be resolved in detail, including cilia and flagella. The instrument design and performance are presented, including images and tracks of bacterial and protozoal mixed samples and pure cultures of six selected species. Organisms as small as 1 μm (bacterial spores) and as large as 60 μm (Paramecium bursaria) may be resolved and tracked without changes in the instrument configuration. Finally, we present a dilution series investigating the maximum cell density that can be imaged, a type of analysis that has not been presented in previous holographic microscopy studies.
数字全息显微镜是研究微生物运动性的理想工具。然而，大多数设计在对细菌成像时没有表现出足够的空间分辨率。在本研究中，我们展示了一种离轴马赫-曾德全息显微镜设计，其空间分辨率优于800纳米，能够在380μm×380μm×600μm的三维视野内分辨不同密度的细菌样本。更大的生物体，如原生动物，可以被详细分辨，包括纤毛和鞭毛。本文介绍了该仪器的设计和性能，包括细菌和原生动物混合样本以及六种选定物种的纯培养物的图像和轨迹。小至1μm（细菌孢子）、大至60μm（绿草履虫）的生物体都可以在不改变仪器配置的情况下被分辨和追踪。最后，我们展示了一个稀释系列，研究了能够成像的最大细胞密度，这种分析类型在以前的全息显微镜研究中尚未出现。
