A new view on polarization microscopy
A new view on polarization microscopy
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DOI：
10.1038/381811a0
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发表时间：
1996-06-27
期刊：
影响因子：
64.8
通讯作者：
Oldenbourg, R
中科院分区：
文献类型：
--
作者：
Oldenbourg, R
图1从受精海胆卵中分离并用Pol-Scope成像的有丝分裂纺锤体（Strongylocentrotus purpartus，由美国费城宾夕法尼亚大学John Murray制备）。延迟量图像，白色：5 nm延迟;黑色：0 nm。微管束从两个黑色的中心体放射出来;染色体在两个星星结构之间可见。动态和非破坏性地测量样品中的亚微观分子有序度，通常可以保持在自然环境条件下。此外，由于其固有的光学特性，特定的结构，如细丝和膜，被突出显示，而不需要染色或标记它们。然而，与传统的偏振光显微镜，单一的图像显示只有那些各向异性的结构，相对于显微镜的偏振轴的取向范围有限。此外，快速测量被限制在一个单一的图像点或单一的区域，表现出均匀的双折射显微镜与新颖的电光设备，算法来计算标本双折射和今天的数字图像捕捉和处理能力。图1和图2展示了这种新型显微镜拍摄的图像的精致分辨率、高对比度和分析强度。延迟的大小是双折射率乘以通过样品的路径长度。慢轴方向或延迟方位是指折射率最大的主轴的方向）。在图像的所有点处测量延迟，并且与双折射轴的取向无关。原始图像数据以640 × 480图像点的分辨率在不到半秒的时间内被记录，并且使用台式计算机在不到一秒的时间内被转换成延迟幅度和取向值。根据所使用的透镜，可以测量低至0.02 nm和高至几个波长的延迟值，空间分辨率为0.2 µm或更大。测量结果以前所未有的清晰度和细节水平显示为代表亚微观分子顺序的图像。
FIG. 1 Mitotic spindle isolated from fertilized sea urchin egg and imaged with the Pol-Scope (Strongylocentrotus purpartus, preparation by John Murray, University of Pennsylvania, Philadelphia, USA). Retardance magnitude image, white: 5 nm retardance; black: 0 nm. Microtubule bundles radiate out from two black centrosomes; chromosomes visible between the two star formations (asters). tial to measure submicroscopic molecular order dynamically and nondestructively in samples that, in general, can be kept in native environmental conditions. Furthermore, specific structures, such as filaments and membranes, are highlighted owing to their intrinsic optical properties, without the need to stain or label them. With the traditional polarized light microscope, however, single images display only those anisotropic structures that have a limited range of orientations with respect to the polarization axes of the microscope. In addition, rapid measurements are restricted to a single image point or single area that exhibits uniform birefrinmicroscope with novel electro-optical devices, algorithms to compute specimen birefringences and today's digital image capture and processing capabilities. Figures 1 and 2 exhibit the exquisite resolution, high contrast and analytical strength of images taken with this new microscope. gence or other forms of optical anisotropy', while measurements comparing several image points take an inordinately long time7•Expressed in distance, the retardance magnitude is the birefringence multiplied with the pathlength through the specimen. The slow axis orientation or retardance azimuth refers to the orientation of the principal axis with the largest refractive index). The retardance is measured at all points of the image and irrespective of orientation of the birefringence axes. Raw image data are recorded in less than half a second at a resolution of 640 by 480 image points and converted to retardance magnitude and orientation values in less than one second using a desktop computer. Retardance values as low as 0.02 nm and as high as several wavelengths can be measured, at a spatial resolution of 0.2 µm or larger, depending on the lenses used. Measurements are displayed as images representing submicroscopic molecular order at an unprecedented level of clarity and detail.