Mild detergent treatment of thin carbon substrate films

Abstract

The preparation of biological specimens for the STEM by the wet film technique and subsequent freeze-drying, has been shown to preserve biological structures reasonably well and give good mass measurements on them. However, the thin (2-3 nm) carbon substrate film, prepared by ultra-high vacuum evaporation onto a freshly cleaved crystal of rock salt, is never perfect for the attachment of fragile biological structures. The film is floated on a dish of clean water and grids covered with holey film are placed face down on the floating thin carbon film. The grids are picked up from above one at a time such that the carbon film retains a droplet of water. This water is exchanged by washing and wicking and the specimen is injected into the drop followed by further washing and wicking. After the final wash, the grid is blotted between two pieces of filter paper (retaining solution less than 1 μm thick), plunged into liquid nitrogen slush, and freeze-dried under vacuum overnight.