Projector plugin does not attach phototargetting sequence to MDA

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Projector plugin does not attach phototargetting sequence to MDA

tamara7
Hi,

I want to attach photo-activation with Andor Mosaic Duet device to
multi-dimensional acquisition (MDA) using Projector plugin. Projector plugin
controls correctly the Mosaic in Live mode, however it does not seem to
attach phototargeting ROI sequence to MDA. I use different channel presets
for phototargeting and stimulation (different cube, light source, shutter).
In Projector I set the phototargeting channel and shutter. I create the
sequence of FRAP ROI, which works correctly when press run in Sequence
window (Projector plugin). Then I press ‘Attach to acquisition’ and start
MDA. Microscope switches correctly to phototargeting channel in between
imaging frames, however almost instantaneously moves back to imaging cube
(even if phototargeting ROI exposure time is many seconds), waiting to snap
another picturein MDA imaging sequence.

I would be grateful for any help on how to attach ptototargeting sequence to
multidimensional aquisition using Projector plugin.

Tamara




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Re: -|EXT|- Projector plugin does not attach phototargetting sequence to MDA

AndorThirdParty
Hi Tamara,

I don't have much experience with the Projector Plugin, but I had a look at this.

It looks like the sequence is attached to the MDA acquisition, rather than to a specific channel. The sequence begins exposing on the Mosaic3 when the MDA is started, and continues until the sequence is completed.

I think you want to take an image, switch to another channel to shine light on your specimen, then switch back and acquire another image. Is this correct? It doesn't seem like there is principled way to achieve this in the plug-in.

By adding custom time intervals in the MDA window you can cause the acquisition to wait in your preset channel for the duration of the interval. Maybe you could use this to expose your specimen to light between frames, by causing the acquisition to pause in your phototargeting channel while the Mosaic is running its sequence. You could use the Off Duration of the Mosaic masks to avoid continuing to run the sequence during the acquisition step.

If synchronisation is important for your experiment this approach may not be any use. If the synchronisation is important, you could set up the Mosaic3 to use external start triggering and trigger the start of the sequence when your other hardware is prepared. I'm not sure if your microscope has an appropriate trigger output. I think you would still need to add intervals to your MDA in order to keep your microscope in the correct state for the duration of the sequence.

In the projector plug-in guide it mentions:
"Pressing "Run ROIs now!" is a good way to phototarget the specimen during a Multi-Dimensional Acquisition. Just start the MDA, wait for the right moment, and then hit "Run Rois now!" to cause phototargeting to start, while acquisition continues simultaneously."

The controls on the ROI tab don't support the Mosaic, but you could do this by clicking "Run" in the Andor Mosaic 3 ROI Sequencing window.

It looks like the "During imaging" and "Between images" tabs in the ROI tab were designed to do something like your use case, but unfortunately they do not support the Mosaic3. The Mosaic3 device adapter was developed outside of Andor, so I haven't managed to get access to the code. I'll see if I can get a copy.

Thanks,
Jack

-----Original Message-----
From: tamara7 [mailto:[hidden email]]
Sent: 19 March 2018 15:07
To: [hidden email]
Subject: -|EXT|- [micro-manager-general] Projector plugin does not attach phototargetting sequence to MDA

Hi,

I want to attach photo-activation with Andor Mosaic Duet device to multi-dimensional acquisition (MDA) using Projector plugin. Projector plugin controls correctly the Mosaic in Live mode, however it does not seem to attach phototargeting ROI sequence to MDA. I use different channel presets for phototargeting and stimulation (different cube, light source, shutter).
In Projector I set the phototargeting channel and shutter. I create the sequence of FRAP ROI, which works correctly when press run in Sequence window (Projector plugin). Then I press ‘Attach to acquisition’ and start MDA. Microscope switches correctly to phototargeting channel in between imaging frames, however almost instantaneously moves back to imaging cube (even if phototargeting ROI exposure time is many seconds), waiting to snap another picturein MDA imaging sequence.

I would be grateful for any help on how to attach ptototargeting sequence to multidimensional aquisition using Projector plugin.

Tamara




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Re: -|EXT|- Projector plugin does not attach phototargetting sequence to MDA

tamara7
Dear Jack,

thank you very much for your detailed reply. I am a bit supprised to know
that the 'run during MDA' does not work for mosaic 3, since thre is even a
wiki: https://micro-manager.org/wiki/Mosaic3.

I will follow your advices, adding pauses in MDA, creating dummy imaging
chanel with my phototargetting cube and using external trigger to run
phototargetting sequence (I use Andor BOBD control box with PCI DIO24 card).
If you have any news on this subject, please let me know,

best wishes

Tamara





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Re: -|EXT|- Projector plugin does not attach phototargetting sequence to MDA

tamara7
In reply to this post by AndorThirdParty
Hi Jack,

thanks, your advice worked very well. Unfortunatelly Mosaic 3 cannot be
triggered when operating in sequencing mode, but is fine in single shot and
referencing modes.

cheers

Tamara



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