[ome-users] Mirror before stitching?

[Björn Quast]

Hi Sam,

have you tried to give the order of the tiles by hand, i. e. choose Type: row 
by row, Order: Left & Up ? Or other configurations, ignoring your metadata?


Björn


> Attached is a screenshot of the issue I'm seeing, even when I deselect
> "compute overlap" and set the slider to 0. On the right is the LOCI
> stitching, which seems reasonable except the flipped y axis. The left is
> the output of the plugin, in which images overlap erroneously (?). When I
> "ignore calibration," there is no change.
> 
> Is my metadata wrong? Does the plugin handle composite (multi-channel)
> images well?
> 
> -Sam
> 
> On Jun 19, 2013, at 11:29 AM, Curtis Rueden <ctrueden at wisc.edu> wrote:
> > Hi Sam,
> > 
> > > Also, what does the "ignore calibration" option do?
> > 
> > I would test with this option both on and off. It controls whether the
> > stage position coordinates are interpreted as pixels, or in calibrated
> > physical units (e.g., microns).
> > 
> > Regards,
> > Curtis
> > 
> > 
> > On Wed, Jun 19, 2013 at 1:27 PM, Sam Lord <sjlord at berkeley.edu> wrote:
> > Thanks for the response, Curtis.
> > 
> > I tried setting the overlap slider to 0 and deselecting all other options.
> > I thought this would reproduce the results that the LOCI importer
> > stitching gave me. Instead, the final fused image was about half the
> > dimensions, and several tiles were fused on top of each other. Maybe
> > there's an issue reading the positions from the metadata?
> > 
> > I'll keep playing around with it.
> > 
> > Also, what does the "ignore calibration" option do?
> > 
> > Thanks,
> > 
> > -Sam
> > 
> > On Jun 19, 2013, at 9:39 AM, Curtis Rueden <ctrueden at wisc.edu> wrote:
> >> Hi Sam,
> >> 
> >> > I've been having a lot of trouble getting the Fiji Grid/Collection
> >> > Stitching plugin to operate how I want.
> >> 
> >> Thanks for the report. I am forwarding this thread to fiji-devel, which
> >> is a better place to report issues with Fiji plugins such as
> >> Grid/Collection Stitching.>> 
> >> > -If I deselect "compute overlap," I would expect the plugin to simply
> >> > place the tiles where the stage position in the metadata claims it is.
> >> > Instead, the output seems to not include most of the images.
> >> 
> >> Try reducing the overlap slider to 0. IIRC, it defaults to 10, and that
> >> factor still applies even if you uncheck "compute overlap.">> 
> >> > -When I do select "compute overlap," the images often aren't placed
> >> > anywhere near the right locations. For instance, it will output an
> >> > irregular shape, when the fused image should be a rectangle.
> >> 
> >> There are various ways of tuning how it approaches the computation, but
> >> if your stage coordinates are already very accurate, you probably don't
> >> need the "compute overlap" feature anyway.>> 
> >> > I'm sure some of these issues result from a non-perfect pixel
> >> > calibration and the fact that I did not include any overlap in my
> >> > tiles when acquiring them.
> >> 
> >> Ah, yes. If you do not include any overlap, then the "Compute overlap"
> >> feature will always fail. It relies on overlap between tiles to detect
> >> how things should be laid out. Your only option when collecting fully
> >> non-overlapping tiles is to uncheck "Compute overlap" and reduce the
> >> overlap slider to 0. It should then lay them out exactly how you
> >> collected them.>> 
> >> > However, the LOCI importer seems to not worry about that. Instead, it
> >> > happy tiles my images. The only problem is that I can tell the images
> >> > are flipped in the y direction.
> >> 
> >> Right, the LOCI importer will stitch your image *exactly* how the
> >> metadata says, and cannot do anything else such as flip tiles inverted
> >> in Y. So I suggest you try the Grid/Collection Stitching plugin again as
> >> I described above. If you still cannot get it work, and are willing to
> >> post a sample non-working dataset, we can investigate further.
> >> 
> >> Regards,
> >> Curtis
> >> 
> >> 
> >> On Wed, Jun 19, 2013 at 11:29 AM, Sam Lord <sjlord at berkeley.edu> wrote:
> >> Hi Curtis,
> >> 
> >> I've been having a lot of trouble getting the Fiji Grid/Collection
> >> Stitching plugin to operate how I want. I'm asking it to select
> >> positions from file and instructing it to use the image metadata. Here
> >> are some issues I've found:
> >> 
> >> -The color scheme does not import. Neither does some other metadata (e.g.
> >> pixel size). -If I deselect "compute overlap," I would expect the plugin
> >> to simply place the tiles where the stage position in the metadata
> >> claims it is. Instead, the output seems to not include most of the
> >> images. -When I do select "compute overlap," the images often aren't
> >> placed anywhere near the right locations. For instance, it will output
> >> an irregular shape, when the fused image should be a rectangle.
> >> 
> >> I'm sure some of these issues result from a non-perfect pixel calibration
> >> and the fact that I did not include any overlap in my tiles when
> >> acquiring them. However, the LOCI importer seems to not worry about
> >> that. Instead, it happy tiles my images. The only problem is that I can
> >> tell the images are flipped in the y direction.
> >> 
> >> Is there a way to "brainlessly" tile my images from metadata location AND
> >> flip y?
> >> 
> >> -Sam
> >> 
> >> On Jun 16, 2013, at 11:49 AM, Curtis Rueden <ctrueden at wisc.edu> wrote:
> >>> Hi Sam,
> >>> 
> >>> We recently added an "invert Y axis coordinates" option (and one for X
> >>> as well) to the Stitching plugins. Did you try that?
> >>> 
> >>> The Fiji Grid/Collection Stitching plugin, in case you haven't tried it,
> >>> uses Bio-Formats and is much more powerful than the BF Importer's basic
> >>> stitching feature.
> >>> 
> >>> Regards,
> >>> Curtis
> >>> On Jun 16, 2013 12:01 PM, "Sam Lord" <sjlord at berkeley.edu> wrote:
> >>> Importing stacks from Micro-Manager works well using Bio-Formats
> >>> Importer, and the stacks option seems to read location directly from
> >>> metadata. The only issue I have is that my camera must be rotated 180
> >>> degrees, because the columns stitch together well, but each row is
> >>> upside-down.
> >>> 
> >>> I tried importing the stack, flipping, exporting, then reimorting, but
> >>> something must have gotten lost in the metadata in that maneuver.
> >>> 
> >>> Any chance there is a simple way to tell the stitching that the images
> >>> are flipped?
> >>> 
> >>> -Sam
> >>> 
> >>> _______________________________________________
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