Non Radiative Excitation Fluorescence Microscopy: A New Method for Studying Membrane Adhesion at the Nanoscale
Résumé
Non-radiative Excitation Fluorescence Microscopy (NEFM) is a promising technique allowing the observation of biological samples beyond the diffraction limit. By coating a substrate with an homogenous monolayer of quantum dots (QDs), Förster Resonance Energy Transfer (FRET) could be achieved from QDs (donors) to dye molecules located in the sample (acceptors). Therefore, the excitation depth of the sample is then given by the Förster radius, which corresponds to few nanometers above the surface. Here, we present this original method to probe the adhesion of Giant Unilamellar Vesicles (GUVs), negatively charged, in strong interaction with a positively charged surface ( QDs layer is coated with Poly-L-Lysine ). Distances between the surface and GUVs are lower than 5 nm. We used the QD-quenching level to calculate and map the absolute distance between the membrane and the surface with a nanometer resolution. By tuning the electrostatic interactions between the surface and the membrane, we have been able to measure a displacement of about 1 nm of the lipid membrane height.