Negative-stain transmission electron microscopy (TEM) is a single-particle imaging method for DNA nanostructures1. In negative-stain TEM, DNA nanostructures are deposited onto a glow-discharged carbon-coated TEM grid and then a stain with heavy metal salts (typically uranyl acetate) is added. This stain surrounds the sample, however it is excluded from space occupied by the DNA nanostructures. This heavy-metal stain is electron-dense and strongly scatters electrons, while DNA is electron-sparse and is a weak scatterer. Therefore, DNA appears light against the dark, stained background.
TEM can achieve lateral resolution around 1–3 nm for single molecules, and averaging of thousands of particles can push resolution below 1 nm. TEM is performed on dried samples which also distorts the structure of DNA compared to when it is hydrated, and additionally interaction with the grid can bias certain conformations of the nanostructure. Compared to AFM, TEM enables higher-throughput imaging and more images of individual nanostructures can be obtained in less time.
References
- Douglas, S. M. et al. Self-assembly of DNA into nanoscale three-dimensional shapes. Nature 459, 414–418 (2009).