Electrochemical Dye Switching Assisted Spectral Demixing and 3D STORM Imaging

Area of Science:

• Cellular Imaging
• Super-resolution Microscopy
• Biophysics

Background:

• Super-resolution microscopy offers nanoscale resolution for cellular imaging.
• Reliable multicolor imaging is essential for analyzing multiple targets in complex biological samples.

Purpose of the Study:

• To develop a robust two-color imaging strategy for super-resolution microscopy.
• To precisely control the switching behavior of cyanine dyes for improved multicolor STORM imaging.

Main Methods:

• Demonstrated a two-color imaging strategy using electrochemically controlled stochastic optical reconstruction microscopy (EC-STORM).
• Utilized oscillating electrochemical potential to control emitter density and molecule ON time.
• Combined spectral demixing with a biplane detection configuration for 3D localization.

Main Results:

• Achieved precise control over the switching behavior of Alexa 647 and CF680 dyes.
• Minimized spatial and temporal overlap of emitters for effective spectral demixing.
• Enabled 3D localization of Alexa 647 using photon-intensity ratios in a biplane setup.

Conclusions:

• EC-STORM provides precise control for multicolor super-resolution imaging.
• The integrated approach offers a practical solution for high-density multicolor 3D STORM imaging.
• This method enhances the ability to resolve multiple targets in complex cellular environments.