Griffin Brendan J
Centre for Microscopy and Microanalysis, The University of Western Australia, Crawley, Western Australia, 6009, Australia.
Methods Mol Biol. 2007;369:467-95. doi: 10.1007/978-1-59745-294-6_23.
The use of elevated gas pressures in the sample chamber of a scanning electron microscope (i.e., variable pressure SEM, or VPSEM) together with specialized electron detectors create imaging conditions that allow biological samples to be examined without any preparation. Specific operating conditions of elevated pressures combined with sample cooling (usually restricted to the environmental SEM range) can allow hydrated samples to be maintained in a pristine state for long periods of time. Dynamic processes also can be easily observed. A wider range of detector options and imaging parameters introduce greater complexity to the VPSEM operation than is present in routine SEM. The current instrumentation with field emission electron sources has nanometer-scale beam resolution (approx 1 nm) and low-voltage beam capability (0.1 kV). However, under the more extreme variable pressure conditions, useful biological sample information can be achieved by skilled operators at image resolutions to 2 to 4 nm and with primary electron beam voltages down to 1.0 kV. Imaging relating to electron charge behavior in some biological samples, generally referred to as charge contrast imaging, provides information unique to this VPSEM and environmental SEM that closely relates to luminescence imaged by confocal microscopy.
在扫描电子显微镜的样品室中使用高压气体（即可变压力扫描电子显微镜，或VPSEM），并结合专门的电子探测器，可创造出无需任何制备就能检查生物样品的成像条件。高压与样品冷却相结合的特定操作条件（通常限于环境扫描电子显微镜范围），可使水合样品长时间保持原始状态。动态过程也能轻松观察到。与常规扫描电子显微镜相比，更多的探测器选项和成像参数给VPSEM操作带来了更大的复杂性。当前配备场发射电子源的仪器具有纳米级的束分辨率（约1纳米）和低电压束能力（0.1 kV）。然而，在更极端的可变压力条件下，熟练的操作人员在图像分辨率达到2至4纳米且一次电子束电压低至1.0 kV时，仍可获得有用的生物样品信息。一些生物样品中与电子电荷行为相关的成像，通常称为电荷对比成像，可提供这种VPSEM和环境扫描电子显微镜特有的信息，这些信息与共聚焦显微镜成像的发光密切相关。
