Ms. Kristýna Holanová Profile

Kristýna Holanová

at Institute of Photonics and Electronics CAS

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Proceedings Article | 11 April 2019 Paper

Interferometric scattering (iSCAT) microscopy with optimized reference wave

Dominika Jurdova, Kristyna Holanova, Milan Vala, Marek Piliarik

Proceedings Volume 11028, 110282P (2019) https://doi.org/10.1117/12.2522360

KEYWORDS: Optical microscopy, Scattering, Microscopes, Interferometry, Light scattering, Microscopy, Nanoparticles, Reflection, Signal detection

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The understanding of nanoscale biological processes is limited by the level of details we can achieve when observing their dynamics. Addressing molecules of interest using fluorescent labels is the most common contrast mechanism in biological nano-imaging. However, the complex photophysics of fluorescent labels limits the localization precision as well as observation times in practical experiments. As an alternative to fluorescence-based microscopy interferometric scattering microscopy (iSCAT) was recently introduced. It is an optical microscopy technique allowing to detect and track nanoscale objects with sub-nanometre localization precision. The basic concept of this technique is the interference of light scattered on the particle with a reference wave light partially reflected at the microscopic slide. Recent advancements pushed the sensitivity and high-speed tracking down to a level of a single unlabelled protein by balancing the amplitudes of scattering and reference waves. This is often achieved by optimizing the reference wave, e.g. via placing a partially transparent mask near the back focal plane of a high numerical aperture microscope. In this contribution we introduce and demonstrate an innovative layout of the iSCAT microscope with optimized reference wave and minimized interferometric artefacts. We benchmark the detection capabilities of the new layout using series of extremely small spherical gold nanoparticles and demonstrate possible applications of the novel detection scheme.

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