Re: processing and Z cropping tiled Leica images ** Commercial Response **

Posted by Barlow, Andrew on
URL: http://confocal-microscopy-list.275.s1.nabble.com/processing-and-Z-cropping-tiled-Leica-images-tp7578835p7578849.html

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Hi Dave,

We've recently implemented stitching of 3D, time resolved datasets in Volocity.
The relative positions of tiles is read from Leica lif files, which greatly speeds the process up (compared to blind stitching).
We've had some great feedback about this feature, please contact me offline if you'd like a demonstration.
Regards,
Andrew


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> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:[hidden email]] On Behalf Of David Johnston
> Sent: 16 August 2012 08:30
> To: [hidden email]
> Subject: processing and Z cropping tiled Leica images
>
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> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Hi all,
>
> I have a user who is assessing bacterial biofilm clearance by ciliated
> airway epithelial cells grown on air liquid interface cultures. He then
> fixes the cultures, cuts out and divides the membrane and does FISH for
> the bacteria, immuno for the cilia plus nuclear counterstain. The
> filter slices are mounted in mountant between 2 coverslips and imaged
> on SP5 in tile scan mode, imaging a long, thin radial strip of approx
> 45x1 fields at 512x512 resolution and multiple Z levels (up to 50ish)
> It is impossible to get the filters perfectly flat - they tilt and
> buckle so we need to set a large Z range to encompass absolute highest
> and lowest points and many Z slices. Ultimately he wants to calculate
> an stimate of biofilm volume
>
> My 2 problems, seeking advice and solutions:
> (1) because of the wide Z range, some slices on some fields cut into
> the filter (autofluorescence and non specific binding at holes) or
> close to coverslip (flash). Because of the undulations of the membrane,
> this varies from field of view to field of view. Therefore I need a way
> to crop or blank Z slices off the data set in a manner which is field
> of view specific, rather than global, but still retains the relative Z
> position in the overall stack.
>
> (2) Despite the fields forming an absolute linear strip, Leica's
> autostitcher sometimes makes a complete hash of it. Is there any easy
> way to get a stitch of Z stacks based simply on relative location.
>
> We are talking reasonably large datafiles here (several GB)
>
> Thanks in Advance,
> Dave Johnston,
> Biomedical Imaging Unit, Southampton, UK.
>
> PS, learned the hard way, if doing tile sets, make sure scan head
> rotation is set to 0 :-)