ABSTRACT
Since the early 1960s, much has been written regarding the preparation of thin plastic, plastic-carbon and carbon films, their varying properties and the treatments given to them before use as specimen supports for negative staining and other purposes (Hayat and Miller, 1990; Sommerville and Scheer, 1987). Often, one has the impression that electron microscopists, similar to all other scientists, revel in the reinvention or improvement of the wheel. Sometimes there are technical improvements and something really new, but often there are only minor innovations, with apparent oblivion to, or careless neglect of, earlier technical publications! To quote Thomas F. Anderson (1966) “Unfortunately, the original literature on techniques is becoming so voluminous that it is impossible for one to keep himself informed of all current advances”, so it is hardly surprising that 30 years later the situation is even worse. In reality, the various negative staining procedures given below tend to be variants of a very limited number of ‘original themes’. This inherent limitation apart, it is my intention to present some of the principal approaches, when possible giving a number of alternatives, with emphasis upon the overall simplicity of the techniques. Details of the three main approaches for the production of negatively stained specimens will be given below (Sections 3.3, 3.4, 3.5); namely, the droplet, floating and mica–carbon transfer techniques. Whilst at first glance it might appear that these techniques are totally independent of one another, this is not the case. There is considerable interplay between the techniques and indeed a close relationship to the techniques used for the preparation of support films. Again, this indicates the overall sparsity of technical possibilities, which has led investigators to seek out the most useful combinations. It will be noted by some that the spraying techniques are not included. In a safety-conscious scientific world, I think that these techniques should not be routinely employed unless absolutely necessary, and then only performed within an appropriate negative pressure extraction cabinet (see also Section 3.3). In general, suitable precautions should always be observed when handling viruses and potentially toxic or dangerous samples, with the inclusion of UV or chemically produced inactivation when necessary.
