nn8b06047_si_002.pdf (22.92 MB)
Combined Use of Unnatural Amino Acids Enables Dual-Color Super-Resolution Imaging of Proteins via Click Chemistry
journal contribution
posted on 2018-12-10, 00:00 authored by Kim-A. Saal, Frank Richter, Peter Rehling, Silvio O. RizzoliRecent
advances in optical nanoscopy have brought the imaging resolution
to the size of the individual macromolecules, thereby setting stringent
requirements for the fluorescent labels. Such requirements are optimally
fulfilled by the incorporation of unnatural amino acids (UAAs) in
the proteins of interest (POIs), followed by fluorophore conjugation via click chemistry. However, this approach has been limited
to single POIs in mammalian cells. Here we solve this problem by incorporating
different UAAs in different POIs, which are expressed in independent
cell sets. The cells are then fused, thereby combining the different
proteins and organelles, and are easily imaged by dual-color super-resolution
microscopy. This procedure, which we termed Fuse2Click, is simple,
requires only the well-established Amber codon, and allows the use
of all previously optimized UAAs and tRNA/RS pairs. This should render
it a tool of choice for multicolor click-based imaging.