Imaging live cell membranes via surface plasmon-enhanced fluorescence and phase microscopy

Area of Science:

• Biophysics
• Microscopy
• Cell Biology

Background:

• Observing living cells and their interactions with substrates requires advanced imaging techniques.
• Existing methods often lack the sensitivity or simultaneous multi-modal capabilities for comprehensive analysis.
• Surface plasmon (SP) phenomena offer unique optical properties for enhanced imaging.

Purpose of the Study:

• To develop and demonstrate a novel wide-field microscopy technique combining surface plasmon (SP) phase microscopy and SP-enhanced fluorescence microscopy.
• To achieve simultaneous, high-sensitivity imaging of living cells and their contacts with a bio-substrate.
• To enable real-time observation of cellular behavior at the cell-substrate interface.

Main Methods:

• Integration of phase-shift interferometry for high-sensitivity phase imaging.
• Utilizing SP-enhanced fluorescence microscopy for bright cellular visualization.
• Employing a high numerical aperture objective and a 30 nm silver film for SP excitation.
• Developing a common-path optical setup for long-term stability.

Main Results:

• Successfully combined wide-field SP phase and SP-enhanced fluorescence microscopy.
• Achieved simultaneous real-time imaging of living cell fluorescence and cell-substrate contact phase.
• Demonstrated high-sensitivity phase information and bright fluorescence imaging of transfected cells near the membrane.
• Validated the technique for observing cellular interactions on bio-substrates.

Conclusions:

• The developed combined SP microscope offers a powerful new tool for live-cell imaging.
• This technique provides simultaneous, high-sensitivity visualization of cellular morphology and substrate interactions.
• It opens new avenues for studying cell dynamics, adhesion, and signaling at the cell-material interface.