Spatial fluorescence cross correlation spectroscopy by means of a spatial light modulator

TL;DR

This paper introduces a novel spatial fluorescence cross correlation spectroscopy technique using a spatial light modulator to measure molecular flows and membrane permeability with high spatial resolution.

Contribution

It demonstrates the feasibility of using a spatial light modulator for spatial FCS, enabling new measurements of molecular transport and membrane permeability.

Findings

Successfully measured molecular flows of 0.2-1.5 μm/ms.

Investigated membrane permeability of GUVs to different molecules.

Proposed theoretical models and discussed alternative methods.

Abstract

Spatial Fluorescence Cross Correlation Spectroscopy is a rarely investigated version of Fluorescence Correlation Spectroscopy, in which the fluorescence signals from different observation volumes are cross-correlated. In the reported experiments, two observation volumes, typically shifted by a few $\mu$m, are produced, with a Spatial Light Modulator and two adjustable pinholes. We illustrated the feasibility and potentiality of this technique by: i) measuring molecular flows, in the range 0.2 - 1.5 $\mu$m/ms, of so-lutions seeded with fluorescent nanobeads or rhodamine molecules (simulating active transport phenomenons); ii) investigating the permeability of the phospholipidic membrane of Giant Unilamellar Vesicles versus hydrophilic or hydrophobic molecules (in that case the laser spots were set on both sides of the membrane). Theoretical descriptions are proposed together with a discus-sion about Fluorescence Correlation Spectroscopy based, alternative methods.