Re: Spinning disk or (new) point scanning confocals for live imaging of 3D engineered tissues?

Posted by Sylvie Le Guyader on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Spinning-disk-or-new-point-scanning-confocals-for-live-imaging-of-3D-engineered-tissues-tp7590297p7590347.html

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Hi Francesco

Are there any single point scanning microscopes (i.e. not spinning disk confocals) that scan 2048x2048 pixels at Nyquist pixel size for the 60x objective at 100 fps?
I do not think so, not even with a resonance scanner cannot. You can go very fast with a point scanning confocal but you need to compromise the FOV or pixel size (resolution). And that would be even worse for lower magnification objectives. You need a camera based system.

For your applications, I would definitely go for a spinning disk with bypass to go widefield which you might need for your calcium imaging on monolayers.

Med vänlig hälsning / Best regards
Sylvie
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Sylvie Le Guyader, PhD
Live Cell Imaging Facility Manager
Karolinska Institutet- Bionut Dpt
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-----Original Message-----
From: Confocal Microscopy List <[hidden email]> On Behalf Of Francesco Pasqualini
Sent: 13 December 2019 17:18
To: [hidden email]
Subject: Spinning disk or (new) point scanning confocals for live imaging of 3D engineered tissues?

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Hi,
I will start my ERC-funded lab in Italy in 2020, which means I am fortunate to be shopping for a confocal instrument right about now.

Since I do mostly live experiments (more details below) I was going to get a spinning disk system. But, I realized that the new scanning confocals are also relatively fast and gentle. The question is how much (if at all) slower and harsher are they with respect to spinning disks?

More details:
- I do a lot of live experiments (traction force microscopy and voltage/calcium-sensitive dyes) on engineered cells (300x300 um FOV) and tissues (600x600 um FOV) in 2D or 3D (<300 um thick) that I complement with immunostainings after fixation. Needed acquisition rates go from <1 fps to 100s depending on the application.

- Originally, I was oriented towards a spinning disk confocal (Yokogawa/SORA, Crest) and was looking at ways to deal with the issues of confocality on the 3D tissue (600x600x300 um volumes) case. But, I realized all vendors have new point scanning confocal (980+Airyscan2, FV3000, A1R) with resonant scanners that acquire full frames/ROIs at 10s/100s of fps

- Of course, structured illumination (Elyra-7, N-SIM) and light-sheet microscopes (QuVi) are also appealing but relatively untested in my applications of interests....

The application specialists from all vendors in my area have been great to work with but since my lab is not up and running, yet, I can't demo these systems directly. Therefore, I could use help and feedback, especially from people who have had related experiences in the recent past.

Thanks,
Francesco

--
Francesco S. Pasqualini
Visiting Professor University of Pavia
Associate Harvard University

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