taking multi-channel images repeatedly with same settings

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taking multi-channel images repeatedly with same settings

David Knecht
We would like to be able to image a series of slides in 3 fluorescence channels with the same settings for the channels for each slide.  Normally, our channels Group does not include things like laser power, exposure, camera gain etc. so we set that individually and then snap an image.  But it is time consuming to acquire three channels with a slide and then go back and repeat that for a new slide using the same channel settings each time because the settings are different for each channel.  Perhaps a solution is to build everything into the channel group, but I am not sure how to do that since exposure (for instance) is a separate element of the interface.  Can a parameter be part of multiple configuation groups and does updating in one place update it in all?  Another solution is to use the multiD to set the channel and exposure settings, but then we get a separate folder/file for each slide and I don’t know of a way to get the images into a single Album to use the viewer (we want to see the color overlay of the three channels in the end).  I am hoping others have faced this issue and can suggest a good solution.  Thanks- Dave

David Knecht, Ph.D.    
Professor and Head of Core Microscopy Facility
Department of Molecular and Cell Biology
U-3125
91 N. Eagleville Rd.
University of Connecticut
Storrs, CT 06269
860-486-2200
860-486-4331 (fax)






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Re: taking multi-channel images repeatedly with same settings

Brian Teague
On 12/12/2013 10:10 AM, David Knecht wrote:

> We would like to be able to image a series of slides in 3 fluorescence
> channels with the same settings for the channels for each slide.
>   Normally, our channels Group does not include things like laser power,
> exposure, camera gain etc. so we set that individually and then snap an
> image.  But it is time consuming to acquire three channels with a slide
> and then go back and repeat that for a new slide using the same channel
> settings each time because the settings are different for each channel.
>   Perhaps a solution is to build everything into the channel group, but
> I am not sure how to do that since exposure (for instance) is a separate
> element of the interface.  Can a parameter be part of multiple
> configuation groups and does updating in one place update it in all?
>   Another solution is to use the multiD to set the channel and exposure
> settings, but then we get a separate folder/file for each slide and I
> don’t know of a way to get the images into a single Album to use the
> viewer (we want to see the color overlay of the three channels in the
> end).  I am hoping others have faced this issue and can suggest a good
> solution.  Thanks- Dave
Hi David -- let me see if I understand what you're asking.  You want to
keep the exposure, laser power, gain etc. the same across a set of
slides, "snap" a multi-channel image for each slide, overlay the
channels in color, then view them in all in a single album (multi-TIFF
file.)

I can help you with all but the last.  A configuration "group" is just a
bunch of "preset"s with the same properties.  So, you can have a group
with just laser power, and another group with both laser power and
exposure time.  You can assign the same property to multiple groups.  As
far as I know, you can only take a Multi-D multi-channel acquisition
from the presets in a single group.

Also, the fact that a property is present in the user interface
(sometimes several places) doesn't mean you can't make a preset out of
it.  Any property in the device / property browser can be added as a
member of a configuration -- that includes exposure, shutter on/off, etc.

Here's what I think I would do in your situation.  I would make a new
Group with all the parameters you want to keep the same, including the
usual channel settings as well as laser power, exposure, etc.  Then, I
would open Multi-D Acquisition.  Set up a multi-channel acquisition
using that group, and setting the colors appropriately. (Note - if
you've set the exposure as part of the config group, I would set the
same value in the multi-channel setup.  If you have different settings
in the preset and the Multi-D dialog, I don't know which one gets
chosen.)  Then, place each slide on the stage, find/focus your specimin,
and "Acquire" the multi-dimensional image.

As far as combining the images in a single "Album", I'm not sure what
the best thing to do here is.  As far as I know, a Micro-Manager image
stack file (what the Multi-D acquisition produces) is just an ImageJ
hyperstack with some extra meta-data; maybe the best thing to do is to
use ImageJ combine the individual Multi-D acquisitions (perhaps as
you're acquiring them) into a multi-channel, multi-position hyperstack.

Hope this helps,
Brian

--
Brian Teague
[hidden email]
Weiss Group, Synthetic Biology Center @ MIT


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Re: taking multi-channel images repeatedly with same settings

David Knecht

On Dec 12, 2013, at 10:57 AM, Brian Teague <[hidden email]> wrote:

On 12/12/2013 10:10 AM, David Knecht wrote:
We would like to be able to image a series of slides in 3 fluorescence
channels with the same settings for the channels for each slide.
 Normally, our channels Group does not include things like laser power,
exposure, camera gain etc. so we set that individually and then snap an
image.  But it is time consuming to acquire three channels with a slide
and then go back and repeat that for a new slide using the same channel
settings each time because the settings are different for each channel.
 Perhaps a solution is to build everything into the channel group, but
I am not sure how to do that since exposure (for instance) is a separate
element of the interface.  Can a parameter be part of multiple
configuation groups and does updating in one place update it in all?
 Another solution is to use the multiD to set the channel and exposure
settings, but then we get a separate folder/file for each slide and I
don’t know of a way to get the images into a single Album to use the
viewer (we want to see the color overlay of the three channels in the
end).  I am hoping others have faced this issue and can suggest a good
solution.  Thanks- Dave

Hi David -- let me see if I understand what you're asking.  You want to keep the exposure, laser power, gain etc. the same across a set of slides, "snap" a multi-channel image for each slide, overlay the channels in color, then view them in all in a single album (multi-TIFF file.)
yes- same for each slide for a channel, different for different channels.  I have now solved this problem using a new Enable command that was recently added to the Andor laser control software.  It allowed me to keep only one laser on at a time in the individual channels while controlling each laser power from the laser power configuration group what is not part of the channel.  Now I can use the multiD to set up each slide capture as a sequential series of channels. and repeat for each slide.  Thanks to Pariksheet for making that work

As far as combining the images in a single "Album", I'm not sure what the best thing to do here is.  As far as I know, a Micro-Manager image stack file (what the Multi-D acquisition produces) is just an ImageJ hyperstack with some extra meta-data; maybe the best thing to do is to use ImageJ combine the individual Multi-D acquisitions (perhaps as you're acquiring them) into a multi-channel, multi-position hyperstack.
I thought there was a way to do this with Bioformats importer, but I can’t find a way to make it open all the files in multiple folders.

Hope this helps,
Brian

--
Brian Teague
[hidden email]
Weiss Group, Synthetic Biology Center @ MIT

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David Knecht, Ph.D.    
Professor and Head of Core Microscopy Facility
Department of Molecular and Cell Biology
U-3125
91 N. Eagleville Rd.
University of Connecticut
Storrs, CT 06269
860-486-2200
860-486-4331 (fax)






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