Peters K R
Scan Electron Microsc. 1979(2):133-48.
In this report, conditions for attaining high resolution in scanning electron microscopy with soft biological specimens are described using the currently available high resolution scanning electron microscopes in emission mode of low energy electrons (secondary and charging electrons). Retinal rod outer segments, red blood cells, intestinal mucosa, and ferritin molecules were all used as biological test specimens. From uncoated specimens a new source of signal, referred to as a discharge signal, can provide a high yield of low energy electrons from an excitation area approximately the size of the beam's cross section. Additionally, under these conditions sufficient topographic contrast can be achieved by applying ultra thin metal coatins. A 0.5 nm thick gold film is found sufficient for generating the total signal, whereas increased coating thickness causes additional topographic background signal. However, a 2.0 nm film is needed for imaging surface details with the present instrument. Ultra thin, even, and grainless tantalum films have been found effective in eliminating the charging artifacts caused by external fields, and the decoration artifacts caused by crystal growth as seen in gold films. To improve, in high magnification work on ultra thin coated specimen, signal-to-noise ratio, methods for obtaining saturation of the signal with discharge electrons are shown. The necessity of confirming the information obtained in SEM by independent techniques (TEM of stereo-replicas or ultra thin sections) is discussed.
在本报告中，我们描述了使用当前可用的低能电子发射模式（二次电子和充电电子）的高分辨率扫描电子显微镜，对软生物标本进行高分辨率扫描电子显微镜检查的条件。视网膜杆状外段、红细胞、肠黏膜和铁蛋白分子均用作生物测试标本。对于未镀膜的标本，一种新的信号源，称为放电信号，可以从大约与电子束横截面大小相同的激发区域提供高产率的低能电子。此外，在这些条件下，通过应用超薄金属涂层可以实现足够的形貌对比度。发现0.5nm厚的金膜足以产生总信号，而增加涂层厚度会导致额外的形貌背景信号。然而，用目前的仪器成像表面细节需要2.0nm的膜。已经发现超薄、均匀且无颗粒的钽膜可有效消除外部场引起的充电伪像以及金膜中出现的晶体生长引起的装饰伪像。为了在超薄涂层标本的高倍工作中提高信噪比，展示了用放电电子使信号饱和的方法。讨论了通过独立技术（立体复制品或超薄切片的透射电子显微镜）确认扫描电子显微镜中获得的信息的必要性。
