Negative-stain TEM: Preparation, acquisition, and data processing
Sequence-programmable covalent bonding of designed DNA assemblies
Sci Adv, Aug 17, 2018; DOI: 10.1126/sciadv.aau1157

Samples were adsorbed on glow-discharged, collodion-supported, carbon-coated (10 nm) Cu400 TEM grids (in-house production) and stained using a 2% aqueous uranyl formate solution containing 25 mM sodium hydroxide. Samples were incubated for 15 to 300 s depending on the buffer/solvent used. For samples dissolved in solvents including low concentrations of positively charged ions, we used higher monomer concentrations (50 nM) and longer incubation times. We used magnifications between ×10,000 and ×30,000 to acquire the data.

Imaging was performed on different microscopes (see Table 2). TEM micrographs used in the figures were high-pass–filtered to remove long-range staining gradients, and the contrast was autoleveled (Adobe Photoshop CS6).

For 2D image processing, libraries of individual particle micrographs were created by particle picking using the RELION-2 picking routine (65). Generation of average 2D particle micrographs was performed using RELION-2 (65). Typically, around 2000 individual particles were averaged.