Small-molecule labeling of live cell surfaces for three-dimensional super-resolution microscopy

Area of Science:

• Microscopy and Imaging
• Chemical Biology
• Bacterial Cell Biology

Background:

• Bacterial cell surface imaging requires super-resolution methods due to size limitations.
• Conventional rhodamine spirolactams use damaging UV light for photoswitching.

Purpose of the Study:

• To develop a photocontrollable rhodamine spirolactam for non-toxic, 3D super-resolution (SR) imaging.
• To enable specific labeling of live bacterial cell surfaces.

Main Methods:

• Iterative synthesis and spectroscopic characterization of rhodamine spirolactams.
• Functionalization with N-hydroxysuccinimide for covalent amine labeling.
• 3D super-resolution (SR) microscopy of labeled Caulobacter crescentus.

Main Results:

• Optimized rhodamine spirolactam photoswitches with visible light (>400 nm).
• Covalent labeling achieved uniform and specific sampling of bacterial cell surfaces.
• High-resolution (10-20 nm) 3D SR reconstructions revealed cell stalk length distribution and growth sites.

Conclusions:

• Photocontrollable rhodamine spirolactams offer a non-toxic, specific labeling strategy for live-cell SR imaging.
• This method allows detailed study of bacterial surface dynamics and structures.
• The optimized dye is a versatile tool for nanoscale biological imaging.