Combining confocal and atomic force microscopy to quantify single-virus binding to mammalian cell surfaces

Area of Science:

• Biophysics
• Cell Biology
• Microscopy

Background:

• Atomic force microscopy (AFM) has advanced to multiparametric imaging of biological samples.
• Force-distance curve-based AFM (FD-based AFM) enables high-resolution imaging and characterization of ligand-receptor interactions on living cells.

Purpose of the Study:

• To develop and detail an FD-based AFM method for quantifying single enveloped virus binding to living animal cells.
• To gain insights into the early stages of virus-cell interactions and measure binding affinity.

Main Methods:

• Functionalized AFM tips with ligands to probe specific virus-receptor interactions.
• Simultaneous AFM imaging and fluorescence microscopy of living cells.
• Analysis of force-distance curves to quantify binding kinetics and thermodynamics.
• Detailed protocols for tip/cell preparation, imaging, and data analysis.

Main Results:

• Quantified binding events of single enveloped viruses to cognate receptors on living cells.
• Measured the affinity of virus-cell interactions.
• Observed rapid establishment of multivalent interactions, with sequential bond formation strengthening viral attachment.
• Provided insights into the initial stages of virus-cell adhesion.

Conclusions:

• FD-based AFM is a powerful tool for studying virus-cell interactions at the single-molecule level.
• The developed protocol allows for detailed characterization of virus binding dynamics.
• This method offers a comprehensive understanding of early virus-host cell engagement.