Multi-colour direct STORM with red emitting carbocyanines.

Area of Science:

• Cell Biology
• Microscopy
• Nanotechnology

Background:

• Single molecule super-resolution microscopy is crucial for studying nanoscale cellular structures.
• Multi-colour applications in super-resolution have been limited by complexity.
• Direct stochastic optical reconstruction microscopy (dSTORM) offers simpler single-molecule imaging but faces challenges with optimal dye pairs for dual-colour applications due to differing buffer requirements.

Purpose of the Study:

• To develop a user-friendly and efficient multi-colour super-resolution microscopy method.
• To overcome limitations of existing dSTORM techniques regarding dye compatibility and buffer conditions.
• To enable reliable co-localization studies in cell biology.

Main Methods:

• Developed a novel variant of dSTORM termed spectral demixing dSTORM (SD-dSTORM).
• Integrated spectral demixing with buffer-compatible blinking properties of red-emitting carbocyanine dyes.
• Utilized commercially available probes and avoided cross-talk correction.

Main Results:

• SD-dSTORM requires reduced laser power and fewer imaging frames for high-resolution reconstruction.
• Achieved faithful reconstruction of super-resolved biological nanostructures.
• Enabled reliable co-localization without error-prone cross-talk correction.

Conclusions:

• SD-dSTORM represents a significant advancement in single molecule localization microscopy.
• The method offers a user-friendly approach to fast, reliable, and efficient multi-colour dSTORM.
• Combines advantages of spectral demixing and optimized dye blinking for advanced biological imaging.