RT Journal Article
SR Electronic
T1 Photon-free (s)CMOS camera characterization for artifact reduction in high- and super-resolution microscopy
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.04.16.440125
DO 10.1101/2021.04.16.440125
A1 Diekmann, Robin
A1 Deschamps, Joran
A1 Li, Yiming
A1 Tschanz, Aline
A1 Kahnwald, Maurice
A1 Matti, Ulf
A1 Ries, Jonas
YR 2021
UL http://biorxiv.org/content/early/2021/04/16/2021.04.16.440125.abstract
AB Modern implementations of widefield fluorescence microscopy often rely on sCMOS cameras, but this camera architecture inherently features pixel-to-pixel variations. Such variations lead to image artifacts and render quantitative image interpretation difficult. Although a variety of algorithmic corrections exists, they require a thorough characterization of the camera, which typically is not easy to access or perform. Here, we developed a fully automated pipeline for camera characterization based solely on thermally generated signal, and implemented it in the popular open-source software Micro-Manager and ImageJ/Fiji. Besides supplying the conventional camera maps of noise, offset and gain, our pipeline also gives access to dark current and thermal noise as functions of the exposure time. This allowed us to avoid structural bias in single-molecule localization microscopy (SMLM), which without correction is substantial even for scientific-grade, cooled cameras. In addition, our approach enables high-quality 3D super-resolution as well as live-cell time-lapse microscopy with cheap, industry-grade cameras. As our approach for camera characterization does not require any user interventions or additional hardware implementations, numerous correction algorithms demanding camera characterization become easily applicable.Competing Interest StatementThe authors have declared no competing interest.