Dahlberg, P. D. et al. J. Am. Chem. Soc. 140, 12310–12313 (2018).
The resolution that can be achieved in single-molecule localization microscopy measurements is dependent on the number of photons emitted by individual fluorophores. Dahlberg et al. show that carrying out localization microscopy at cryogenic temperatures can improve the localization precision by a factor of four, which they find is largely due to increased photon counts from reduced photobleaching. To enable these studies, they first had to identify a probe that was suitable for localization microscopy at cryogenic temperatures. They screened several fluorescent proteins and found that the photoactivatable fluorescent protein PAmKate could be activated at temperatures as low as 77 K. Using this probe, the researchers carried out cryogenic super-resolution microscopy to study the localization of a bacterial protein to microdomains at the cell poles in Caulobacter crescentus. This finding should help enable super-resolution correlated light and electron microscopy.