Print Email Facebook Twitter Retarding Field Integrated Fluorescence and Electron Microscope Title Retarding Field Integrated Fluorescence and Electron Microscope Author Vos, Y. (TU Delft ImPhys/Microscopy Instrumentation & Techniques) Lane, R. (TU Delft ImPhys/Microscopy Instrumentation & Techniques) Peddie, Chris J. (Francis Crick Institute) Wolters, A.E. (University Medical Center Groningen) Hoogenboom, J.P. (TU Delft ImPhys/Microscopy Instrumentation & Techniques) Date 2020 Abstract The authors present the application of a retarding field between the electron objective lens and sample in an integrated fluorescence and electron microscope. The retarding field enhances signal collection and signal strength in the electron microscope. This is beneficial for samples prepared for integrated fluorescence and electron microscopy as the amount of staining material added to enhance electron microscopy signal is typically lower compared to conventional samples in order to preserve fluorescence. We demonstrate signal enhancement through the applied retarding field for both 80-nm post-embedding immunolabeled sections and 100-nm in-resin preserved fluorescence sections. Moreover, we show that tuning the electron landing energy particularly improves imaging conditions for ultra-thin (50 nm) sections, where optimization of both retarding field and interaction volume contribute to the signal improvement. Finally, we show that our integrated retarding field setup allows landing energies down to a few electron volts with 0.3 eV dispersion, which opens new prospects for assessing electron beam induced damage by in situ quantification of the observed bleaching of the fluorescence following irradiation. Subject backscattered electron detectioncorrelative light and electron microscopyelectron beam induced damageretarding fieldSEM To reference this document use: http://resolver.tudelft.nl/uuid:f46169f1-2773-4b68-afcd-0c71732c9cfa DOI https://doi.org/10.1017/S1431927620024745 ISSN 1431-9276 Source Microscopy and Microanalysis, 27 (1), 109-120 Part of collection Institutional Repository Document type journal article Rights © 2020 Y. Vos, R. Lane, Chris J. Peddie, A.E. Wolters, J.P. Hoogenboom Files PDF retarding_field_integrate ... oscope.pdf 1.05 MB Close viewer /islandora/object/uuid:f46169f1-2773-4b68-afcd-0c71732c9cfa/datastream/OBJ/view