Nijsse J, van Aelst A C
Department of Plant Sciences, Wageningen University, The Netherlands.
Scanning. 1999 Nov-Dec;21(6):372-8. doi: 10.1002/sca.4950210603.
In the past decade, investigators of cryo-planing for low-temperature scanning electron microscopy (cryo-SEM) have developed techniques that enable observations of flat sample surfaces. This study reviews these sample preparation techniques, compares and contrasts their results, and introduces modifications that improve results from cryo-planing. A prerequisite for all successful cryo-planing required a stable attachment of the specimen to a holder. In most cases, clamping with a screw mechanism and using indium as space-filler sufficed. Once this problem was solved, any of three existing cryo-planing methods could be used to provide successful results: cryo-milling, microtomy in a cold room, and cryo-ultramicrotomy. This study introduces modifications to the cryo-planing technique that produces flat surfaces of any desired plane through a specimen. These flat surfaces of frozen, fully hydrated samples can be used to improve observations from cryo-SEM as well as to enhance results from x-ray microanalysis and (digital) image analysis. Cryo-planing results of chrysanthemum (Dendranthema x grandiflorum Tzvelev) stems, hazel (Corylus avelane L.) stems, and repeseed (Brassica napus L.) pistils are presented to illustrate the use of the planing method on fibrous, hard, and delicate materials, respectively.
在过去十年中，用于低温扫描电子显微镜（cryo-SEM）的冷冻刨光技术研究人员开发了能够观察平坦样品表面的技术。本研究回顾了这些样品制备技术，比较并对比了它们的结果，并介绍了改进冷冻刨光结果的改良方法。所有成功的冷冻刨光的一个先决条件是将标本稳定地附着在支架上。在大多数情况下，用螺丝机构夹紧并使用铟作为填充材料就足够了。一旦这个问题得到解决，现有的三种冷冻刨光方法中的任何一种都可以用来获得成功的结果：冷冻研磨、在冷室中进行切片以及冷冻超薄切片。本研究介绍了对冷冻刨光技术的改良，该技术可通过标本产生任何所需平面的平坦表面。这些冷冻的、完全水合的样品的平坦表面可用于改善冷冻扫描电子显微镜的观察结果，以及增强X射线微分析和（数字）图像分析的结果。展示了菊花（Dendranthema x grandiflorum Tzvelev）茎、榛子（Corylus avelane L.）茎和油菜籽（Brassica napus L.）雌蕊的冷冻刨光结果，分别说明刨光方法在纤维状、坚硬和易碎材料上的应用。
