Nanoscale characterization of vesicle adhesion by normalized total internal reflection fluorescence microscopy

Area of Science:

• Biophysics
• Surface Science
• Materials Science

Background:

• Studying vesicle adhesion is crucial for understanding biological processes and developing new materials.
• Existing methods for measuring vesicle-surface interactions have limitations in resolution or range.

Purpose of the Study:

• To demonstrate a straightforward fluorescence microscopy technique for quantifying vesicle adhesion.
• To validate the technique's ability to measure a wide range of membrane-surface interactions, from ultra-weak to strong.

Main Methods:

• Utilized a dual-observation fluorescence microscopy approach combining epi-fluorescence and total internal reflection fluorescence (TIRF).
• Normalized TIRF images with epi-fluorescence images to map membrane/substrate separation distance.
• Achieved nanometric resolution (~20 nm) with a working range of 300-400 nm.

Main Results:

• Successfully quantified both unspecific and specific adhesion of Giant Unilamellar Vesicles (GUVs).
• Investigated unspecific adhesion by controlling electrostatic forces between charged GUVs and functionalized surfaces.
• Demonstrated specific adhesion using biotin/streptavidin recognition for lock-and-key interactions.

Conclusions:

• The proposed fluorescence microscopy technique is effective for quantifying vesicle adhesion across a broad spectrum of interaction strengths.
• This method provides a valuable tool for detailed studies of membrane-surface interactions in various contexts.
• The technique offers high resolution and a practical working range for vesicle adhesion analysis.