Immunohistochemistry Experiments
1. Provide the raw images obtained by light or fluorescence microscopy
Not applicable.
2. Provide the raw images obtained by confocal microscopy
Images were acquired using a Yokogawa W1 Spinning Disk confocal microscope (Zeiss) with a 63x oil objective (1.4 numerical aperture).
3. Provide the name and version of the software employed to obtain images
MetaMorph® (version 7.10, Molecular Devices, LLC. San Jose, California, USA).
4. Provide a spreadsheet of the raw quantifications and a description of analysis parameters and program
Deconvolution. Except for DAPI, all channels were deconvoluted with Huygens Essential software (version 19.04, Scientific Volume Imaging, The Netherlands; http://svi.nl), using the classic maximum likehood estimation algorithm and a signal-to-noise ratio of 50 (for the immunofluorescence channel) or 20 (for the FISH channel), a quality change threshold of 0.01, and 150 iterations at most.
The experimental PSF was obtained using carboxylate microsphere beads (diameter: 170 nm; Invitrogen/thermoFisher Corp.).
Imaging analysis. AstroDot (as described in (Oudart et al., 2020)).
Statistical analysis. AstroStat (as described in (Oudart et al., 2020)).
Oudart, M., Tortuyaux, R., Mailly, P., Mazare, N., Boulay, A. C., & Cohen-Salmon, M. (2020). AstroDot - a new method for studying the spatial distribution of mRNA in astrocytes. J Cell Sci, 133(7).
