It has become clear that the standard cartoon, in which macromolecular particles prepared for electron cryo-microscopy are shown to be surrounded completely by vitreous ice, often is not accurate. In particular, the standard picture does not include the fact that diffusion to the air-water interface, followed by adsorption and possibly denaturation, can occur on the time scale that normally is required to make thin specimens. The extensive literature on interaction of proteins with the air-water interface suggests that many proteins can bind to the interface, either directly or indirectly via a sacrificial layer of already-denatured protein. In the process, the particles of interest can, in some cases, become preferentially oriented, and in other cases they can be damaged and/or aggregated at the surface. Thus, although a number of methods and recipes have evolved for dealing with protein complexes that prove to be difficult, making good cryo-grids can still be a major challenge for each new type of specimen. Recognition that the air-water interface is a very dangerous place to be has inspired work on some novel approaches for preparing cryo-grids. At the moment, two of the most promising ones appear to be: (1) thin and vitrify the specimen much faster than is done currently or (2) immobilize the particles onto a structure-friendly support film so that they cannot diffuse to the air-water interface.

很明显，标准动画通常是不准确的，其中显示为电子冷冻显微镜制备的大分子颗粒完全被玻璃冰包围。特别是，标准图片不包括这样的事实：扩散到空气-水界面，然后是吸附和可能的变性，可以在制作薄样品通常所需的时间尺度上发生。关于蛋白质与空气-水界面相互作用的大量文献表明，许多蛋白质可以直接或通过已变性蛋白质的牺牲层间接结合到界面上。在此过程中，在某些情况下，感兴趣的颗粒可能会优先定向，而在其他情况下，它们可能会被损坏和/或聚集在表面。因此，尽管已经发展出许多处理蛋白质复合物的方法和配方，但事实证明这些方法和配方很困难，但制造良好的冷冻网格仍然是每种新型标本的主要挑战。认识到空气-水界面是一个非常危险的地方，激发了人们对一些制备低温网格的新方法的研究。目前，最有希望的两种方法似乎是：（1）比目前更快地薄化和玻璃化样品，或（2）将颗粒固定在结构友好的支撑膜上，使它们无法扩散到空气中-水界面。