Abstracts
Fast, long-time STED imaging of endoplasmic reticulum dynamics living cells
Ashwin Balakrishnan
Goethe - Universität Frankfurt, Institute of Physical and Theoretical Chemistry, Max-von-Laue str. 7, 60438 Frankfurt am Main [DE], balakrishnan@chemie.uni-frankfurt.de
Author(s):
Ashwin Balakrishnan, Johanna Rahm, Alexandra Kaminer, Mike Heilemann
Super-resolution microscopy techniques have been crucial in recent years in realizing structural cell biology a reality.However, biomolecules in a cell are highly dynamic and in order to assess these dynamics, two challenges need to be adressed: first, photobleaching limits the information that can be extracted from a cell. Second, phototoxicity leads to cell damage. Here, we demonstrate long-time STED microscopy with fast temporal resolution in living cells. We achieve this by employing very low irradiation intensities, and restore information by using convolutional neural network-based image denoising. We demonstrate live-cell imaging for hours with a temporal resolution of seconds. We use this approach to study the dynamics of the endoplasmic reticulum (ER) in healthy and autophagy-induced cells. By extending this further with a neural network-based segmentation, we quantify morphological rearrangements. We find that the ER is highly dynamic and visualize morphological changes under stress-induced conditions that have proved elusive before. In summary, we present an experimental approach that can be transferred to other live-cell super-resolution organelle dynamics.