Novel perspectives for the application of total internal reflection microscopy

TL;DR

This paper introduces a new method for Total Internal Reflection Microscopy (TIRM) that determines the intensity-distance relationship experimentally, broadening its applicability to complex and reflective surfaces.

Contribution

The authors develop a novel approach to measure I(z) directly from hydrodynamic interactions, enabling TIRM use in previously inaccessible conditions.

Findings

Method extends TIRM applicability to reflective surfaces.

Allows measurements on gold surfaces with complex reflections.

Enhances the versatility of TIRM in various experimental setups.

Abstract

Total Internal Reflection Microscopy (TIRM) is a sensitive non-invasive technique to measure the interaction potentials between a colloidal particle and a wall with femtonewton resolution. The equilibrium distribution of the particle-wall separation distance z is sampled monitoring the intensity I scattered by the Brownian particle under evanescent illumination. Central to the data analysis is the knowledge of the relation between I and the corresponding z, which typically must be known a priori. This poses considerable constraints to the experimental conditions where TIRM can be applied (short penetration depth of the evanescent wave, transparent surfaces). Here, we introduce a method to experimentally determine I(z) by relying only on the distance-dependent particle-wall hydrodynamic interactions. We demonstrate that this method largely extends the range of conditions accessible with TIRM, and even allows measurements on highly reflecting gold surfaces where multiple reflections lead to a complex I(z).