Biology subjects

Roos, J.

Publications and source records attributed to Roos, J..

3 recordsLinked to original sources

bioRxiv2024

FASER: A TOOL TO SIMULATE PSF DISTORTIONS IN STEDMICROSCOPY

We introduce Faser, a software package developed in Python as a plugin for the open-source napari platform, designed to simulate the excitation point spread functions (PSFs) of microscopes. Using a full-vectorial computational approach to simulate the electromagnetic fields within the focal region, it makes precise predictions and allows detailed analyses of excitation PSFs. Faser is intended as a pedagogical tool enabling users to explore the impact of various geometrical and optical parameters of practical importance on imaging performance. It supports the modeling of complex beam profiles, including donut and bottle-shaped beams, which are instrumental in advanced microscopy techniques such as Stimulated Emission Depletion (STED) microscopy. Through specific simulations and accessible illustrations, we showcase Fasers capabilities in replicating the distinctive properties of STED beams, making it a valuable resource for researchers and students in optical microscopy to explore and optimize high-resolution imaging techniques.

neuroscience↗

bioRxiv2023

Impact of a tilted coverslip on two-photon and STED microscopy

The advent of super-resolution microscopy has opened up new avenues to unveil brain structures with unprecedented spatial resolution in the living state. Yet, its application to live animals remains a genuine challenge. Getting optical access to the brain in vivo requires the use of a cranial window, whose mounting greatly influences image quality. Indeed, the coverslip used for the cranial window should lie as orthogonal as possible to the optical axis of the objective, or else significant optical aberrations occur. In this work, we assess the effect of the tilt angle of the coverslip on STED and two-photon microscopy, in particular image brightness and spatial resolution. We then propose an approach to measure and reduce the tilt using a simple device added to the microscope, which can ensure orthogonality with a precision of 0.07{degrees}.

neuroscience↗

bioRxiv2023

Imaging dendritic spines in the hippocampus of a living mouse by 3D-STED microscopy

STED microscopy has been used to address a wide range of neurobiological questions in optically well-accessible samples like cell culture or brain slices. However, the application of STED to deeply embedded structures in the brain of living animals remains technically challenging. In previous work, we established chronic STED imaging in the hippocampus in vivo but the gain in spatial resolution was restricted to the lateral plane. In this study, we report on extending the gain in STED resolution into the optical axis to visualize dendritic spines in the hippocampus in vivo. The approach is based on a spatial light modulator to shape the focal STED light intensity in all three dimensions and a conically shaped window that is compatible with an objective that has a long working distance and a high numerical aperture. Moreover, we corrected distortions of the laser wavefront to optimize the shape of the bottle beam of the STED laser, which is required for 3D-STED microscopy. In summary, we present a methodology to improve the axial resolution for STED microscopy in the deeply embedded hippocampus in vivo, facilitating longitudinal studies of neuroanatomical plasticity at the nanoscale in a wide range of (patho-)physiological contexts.

neuroscience↗