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Updated: May 6, 2026

Nanotopology of Cell Adhesion upon Variable-Angle Total Internal Reflection Fluorescence Microscopy VA-TIRFM
Published on: October 2, 2012
Nanoscale contact line visualization based on Total Internal Reflection Fluorescence Microscopy
Abstract:
We describe a novel measurement method to study the contact line of a droplet at nanoscale level. The method is based on Total Internal Reflection Fluorescence Microscopy (TIRFM), which uses an evanescent excitation field produced by total internal reflection of light. The evanescent field depends on the angle of the incident light and has an exponential intensity decay, characterized by the penetration depth. The penetration depth is determined by imaging a fluorescent particle probe that is traversed using an Atomic Force Microscopy (AFM) setup. The result confirms the exponential behavior of the evanescent field intensity, and the value of the penetration depth also corresponds with the value predicted based on the optical configuration. By using the intensity distribution of a fluorescent dye and the value for the penetration depth of the evanescent wave, it is possible to reconstruct the interface of a partial wetting droplet. The reconstructed interface based on TIRFM is in good agreement with the interface obtained from two reference measurements: non-disturbing AFM-imaging and conventional contact angle measurement. The latter lacks spatial resolution, while the former is limited to particular droplets. This new non-contact measurement does not suffer from these drawbacks, making it a very useful tool to study the fundamental wetting behavior of both stationary and dynamic interfaces.
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