High-Density Super-Resolution Localization Imaging with Blinking Carbon Dots

Molecular
fluorescence blinking provides a simple and attractive
way to achieve super-resolution localization via conventional fluorescence
microscopy. However, success in super-resolution imaging relies heavily
on their blinking characteristics. We here report easily prepared
and photostable nanoparticles, carbon dots (CDs), with desirable fluorescence
blinking for high-density super-resolution imaging. The CDs exhibit
a low duty cycle (∼0.003) and high photon output (∼8000)
per switching event, as well as show much higher resistance to photobleaching
than Alexa 647 or Cy5 typically used in single molecule localization
microscopy. The stable blinking of CDs allows to perform high-density
localization imaging at a resolution of 25 nm by sequentially recording
the particle positions. The CD-based super-resolution imaging is further
demonstrated by rendering CD-stained tubular peptide self-assemblies,
CD-packed clusters with well-defined patterns, and CD-stained microtubules
in a cell. Furthermore, this method has been validated as a valuable
tool to detect the clustering and distribution of protein receptors
in the plasma membrane that are not discerned with normal fluorescence
imaging.