Super-Resolution Microscopy of Trypanosomes

Trypanosomes expressing the mNG::IFT81 fusion protein were spread on glass coverslips in medium, and SIM was performed on an LSM780 Elyra PS1 microscope (ZEISS) using 100×/1.46 NA oil Plan Apochromat objective and an electron-multiplying charge-coupled device (EMCCD) Andor IXon 887 1-K camera for the detection at Institut Pasteur. 15 images per plane per channel (five phases; three angles) were acquired to perform the SIM imaging. SIM imaging was processed with ZEN software (ZEISS). The SIMcheck plugin (Ball et al., 2015 (link)) in ImageJ (Schneider et al., 2012 (link)) was used to evaluate the acquisition and the processing parameters.

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Bertiaux E., Mallet A., Fort C., Blisnick T., Bonnefoy S., Jung J., Lemos M., Marco S., Vaughan S., Trépout S., Tinevez J.Y, & Bastin P. (2018). Bidirectional intraflagellar transport is restricted to two sets of microtubule doublets in the trypanosome flagellum. The Journal of Cell Biology, 217(12), 4284-4297.

Publication 2018

LSM780 Elyra PS1 microscope IXon 887 1-K camera ZEN software

Device Electron Microscope Protein Trypanosomes

Corresponding Organization : Inserm

Other organizations : Sorbonne Université, Université Paris-Saclay, Université Paris-Sud, Centre National de la Recherche Scientifique, Institut Curie, Oxford Brookes University, Science Oxford

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Variable analysis

independent variables

Expression of mNG::IFT81 fusion protein in trypanosomes

dependent variables

Structural features of trypanosomes expressing mNG::IFT81 fusion protein observed through SIM imaging

control variables

Glass coverslips
Medium
LSM780 Elyra PS1 microscope (ZEISS) with 100×/1.46 NA oil Plan Apochromat objective
Electron-multiplying charge-coupled device (EMCCD) Andor IXon 887 1-K camera for detection
Acquisition parameters (15 images per plane per channel, five phases, three angles)
SIM imaging processing with ZEN software (ZEISS)
Evaluation of acquisition and processing parameters using SIMcheck plugin in ImageJ

controls

No positive or negative controls were explicitly mentioned in the provided information.

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