nn8b01396_si_002.mpg (32.64 MB)

Nanometer-Resolved Mapping of Cell–Substrate Distances of Contracting Cardiomyocytes Using Surface Plasmon Resonance Microscopy

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posted on 04.09.2018, 00:00 by Eva Kreysing, Hossein Hassani, Nico Hampe, Andreas Offenhäusser
It has been shown that quantitative measurements of the cell–substrate distance of steady cells are possible with scanning surface plasmon resonance microscopy setups in combination with an angle resolved analysis. However, the accuracy of the determined cell–substrate distances as well as the capabilities for the investigation of cell dynamics remained limited due to the assumption of a homogeneous refractive index of the cytosol. Strong spatial or temporal deviations between the local refractive index and the average value can result in errors in the calculated cell–substrate distance of around 100 nm, while the average accuracy was determined to 37 nm. Here, we present a combination of acquisition and analysis techniques that enables the measurement of the cell–substrate distance of contractile cells as well as the study of intracellular processes through changes in the refractive index at the diffraction limit. By decoupling the measurement of the cell–substrate distance and the refractive index of the cytoplasm, we could increase the accuracy of the distance measurement on average by a factor of 25 reaching 1.5 nm under ideal conditions. We show a temporal and spatial mapping of changes in the refractive index and the cell–substrate distance which strongly correlate with the action potentials and reconstruct the three-dimensional profile of the basal cell membrane and its dynamics, while we reached an actual measurement accuracy of 2.3 nm.