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Thanks Kris
Just to re-emphasize why this issue is so important. To get accurate
intensities and segmentation of the nuclei images some type of image
restoration must be done (either deconvolution or AI based restoration).
The segmentations I've seen of the nuclei image, so far, looked good at
first glance (round well separated objects), however are merely
simplifications which fit a spheroid to the data, but do not capture the
true shape or intensities.
Anyway, I will spend some time this week or next looking over the docker
release and see if I can tell what happened with the PSF. Seems like it is
probably a resampling issue, perhaps even as simple as incorrect reading of
spacings from the meta-data, but hard to know for sure.
Brian
Brian
On Sun, Apr 17, 2022 at 8:08 PM Kubow, Kristopher E - kubowke <
[log in to unmask]> wrote:
> *****
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> *****
>
> Question was cross-posted on image.sc forum please see forum for response
> and continuation of discussion:
> https://forum.image.sc/t/3d-image-analysis-tools-and-reproducibility-event-april-27th-via-zoom/64671/5?u=kris_kubow
>
> ________________________________
> From: Brian Northan <[log in to unmask]>
> Sent: Friday, April 15, 2022 6:09 AM
> Subject: Re: 3D Image Analysis Tools and Reproducibility Event - April
> 27th via Zoom
>
> *****
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> *****
>
> Hi Kris
>
> Any update on this? Why not just release the forward model code publically
> and have the community look it over? I don't understand how ABRF can claim
> they are addressing 'reproducibility' when they are working in such a
> closed manner. Isn't openness and communication the number one principle
> of reproducibility?
>
> Brian
>
> On Thu, Mar 24, 2022 at 3:28 PM Kubow, Kristopher E - kubowke <
> [log in to unmask]> wrote:
>
> > *****
> > To join or leave the confocal microscopy listserv or to change your email
> > address, go to:
> >
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> and include the link in your posting.
> > *****
> >
> > Thanks Brian and David. We agree that there does seem to be a mismatch
> > between the spacing parameters that were given and what is actually used
> in
> > the nuclei images. This will impact deconvolution of the images, if
> people
> > elect to do that as part of their segmentation protocol for the study.
> > We’re currently looking into the problem and are in contact with the
> group
> > that developed the simulation software. We'll post updates (when we get
> > them) on the image.sc forum:
> >
> https://urldefense.proofpoint.com/v2/url?u=https-3A__forum.image.sc_t_3d-2Dimage-2Danalysis-2Dtools-2Dand-2Dreproducibility-2Devent-2Dapril-2D27th-2Dvia-2Dzoom_64671&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=hBsapqAzr5hAx-G9-qM3LSc9TEwTGgBoh0n4rbwkBks&s=SMse04bgkg546B_NFjehOds-hRMIW4bovk4mbCxMeqc&e=
> >
> > ________________________________
> > From: Brian Northan <[log in to unmask]>
> > Sent: Wednesday, March 23, 2022 9:26 AM
> > Subject: Re: 3D Image Analysis Tools and Reproducibility Event - April
> > 27th via Zoom
> >
> > *****
> > To join or leave the confocal microscopy listserv or to change your email
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> > Post images on
> >
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> > and include the link in your posting.
> > *****
> >
> > Interestingly I used similar spacings to those David estimated. After
> > first using the given spacings and getting terrible deconvolution
> results,
> > I simply used the Nyquist spacings and got a much better result.
> > From there I tweaked the spacings a bit and used 0.1 XY and 0.8 Z and
> got a
> > pretty good deconvolution.
> >
> > I mentioned this to Kris Kublow, one of the organizers several months ago
> > and he said the group would look into it, but I never heard if they did.
> >
> > Jessica, did your group ever run tests to validate the PSF, forward model
> > and spacings? If so, how did you convince yourself the simulation was
> > correct?
> >
> > Brian
> >
> >
> > On Mon, Mar 21, 2022 at 1:09 PM David Biggs <[log in to unmask]> wrote:
> >
> > > *****
> > > To join or leave the confocal microscopy listserv or to change your
> email
> > > address, go to:
> > >
> >
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> > > Post images on
> >
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> > and include the link in your posting.
> > > *****
> > >
> > > In reference to Brian's observations, I also concur that the parameters
> > > provided for the widefield nuclei dataset do not match the data
> provided.
> > > For a 0.75 NA air lens, I estimate the Z spacing is closer to 0.75um,
> and
> > > the XY spacing is closer to 0.1um.
> > > I roughly estimated these from the shape of the missing cone in the
> > Fourier
> > > domain.
> > >
> > > Any clarification would be appreciated.
> > >
> > > Thanks,
> > > David
> > >
> > >
> > >
> > > On Sat, Mar 19, 2022 at 1:44 PM Brian Northan <[log in to unmask]>
> > wrote:
> > >
> > > > *****
> > > > To join or leave the confocal microscopy listserv or to change your
> > email
> > > > address, go to:
> > > >
> >
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> > > > Post images on
> >
> https://urldefense.proofpoint.com/v2/url?u=http-3A__www.imgur.com&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=Bbo8EpRKzdBsReltSfbg8Udr6_-vbdBTWzo2_VmZUow&s=uQ0i7zYMybxCqH94JEtgiMLq5Tqf4RT45jB8kGVrshU&e=
> > and include the link in your
> > > posting.
> > > > *****
> > > >
> > > > Has anyone else tried to deconvolve the nuclei images used in the
> > > study? I
> > > > was only able to obtain a good deconvolution result after
> introducing a
> > > > fudge factor into the PSF generation. Is anybody else running into
> the
> > > > same problem? My concern is that there is a potential sampling issue
> > in
> > > > the PSF used in the simulation, and that in turn could (falsely) make
> > it
> > > > seem as if deconvolution doesn't work. That would be a shame,
> > > > deconvolution is an important tool for processing 3D Widefield
> > > fluorescence
> > > > images, and it would be nice if this study could be used, to
> > demonstrate
> > > > the benefits of deconvolution.
> > > >
> > > > Contact me off list if you would like. I'd like to verify that these
> > > > images can be deconvolved using standard tools.
> > > >
> > > > Brian
> > > >
> > > > On Fri, Mar 18, 2022 at 9:44 AM Jessica Hornick <
> > > > [log in to unmask]>
> > > > wrote:
> > > >
> > > > > *****
> > > > > To join or leave the confocal microscopy listserv or to change your
> > > email
> > > > > address, go to:
> > > > >
> >
> https://urldefense.proofpoint.com/v2/url?u=https-3A__lists.umn.edu_cgi-2Dbin_wa-3FSUBED1-3Dconfocalmicroscopy-26A-3D1&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=Bbo8EpRKzdBsReltSfbg8Udr6_-vbdBTWzo2_VmZUow&s=-owGyRKzOtmTtqOXx9hHPa81IsG5cCt5iCVjWV4ILTg&e=
> > > > > Post images on
> >
> https://urldefense.proofpoint.com/v2/url?u=http-3A__www.imgur.com&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=Bbo8EpRKzdBsReltSfbg8Udr6_-vbdBTWzo2_VmZUow&s=uQ0i7zYMybxCqH94JEtgiMLq5Tqf4RT45jB8kGVrshU&e=
> > and include the link in your
> > > > posting.
> > > > > *****
> > > > >
> > > > > Come learn more about 3D image analysis tools and how they can help
> > > > > improve reproducibility at this two-hour online workshop hosted by
> > the
> > > > > Light Microscopy Research Group of the ABRF!
> > > > >
> > > > > This event will be centered around the LMRG’s ongoing study of
> > > > > reproducibility in 3D image analysis in which we seek to
> characterize
> > > and
> > > > > identify sources of (ir)reproducibility in 3D segmentation. Try
> your
> > > hand
> > > > > at analyzing the study images and then attend the workshop to see
> how
> > > the
> > > > > pros would do it. Representatives from several major 3D analysis
> > > > platforms
> > > > > will walk attendees through how to segment and analyze the images
> > from
> > > > our
> > > > > study and will discuss how their platforms support reproducible
> image
> > > > > analysis. These demonstrations will occur simultaneously in
> breakout
> > > > rooms
> > > > > and attendees may choose up to two to attend. There will also be an
> > > open
> > > > > session at the conclusion of the program during which attendees may
> > > visit
> > > > > and talk with any representative they like. Note that the platforms
> > > will
> > > > > not be competing against each other–the purposes of the event are
> > > > > instructional/informational and to promote reproducibility in image
> > > > > analysis.
> > > > >
> > > > > We will host representatives from the following platforms:
> > ImageJ/FIJI,
> > > > > Napari, CellProfiler, Arivis, and Volocity. We hope to add more to
> > this
> > > > > list before the event.
> > > > >
> > > > > The event will take place on April 27th at 8:00 am Pacific Time /
> > 11:00
> > > > am
> > > > > Eastern Time via Zoom.
> > > > >
> > > > > ** Register for the event here:
> > > > >
> > > >
> > >
> >
> https://urldefense.proofpoint.com/v2/url?u=https-3A__us02web.zoom.us_meeting_register_tZEsceuvpzIvH9e0kFHEuWdxRMBAgrrpUwzj&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=Bbo8EpRKzdBsReltSfbg8Udr6_-vbdBTWzo2_VmZUow&s=df5s8P1S7R061gnHC6OXrqAOlhVJ3FaJP5Wt8KsC0Qo&e=
> > > > >
> > > > > This event is targeted to people who are interested in learning
> more
> > > > about
> > > > > a specific image analysis platform, learning how to use an image
> > > analysis
> > > > > tool better, and/or learning about how reproducibility is addressed
> > by
> > > > > these platforms.
> > > > >
> > > > > We strongly encourage attendees to participate in the study before
> > the
> > > > > event so that you are familiar with the image analysis problem.
> > > Attendees
> > > > > who participate in the study before the event will be entered in a
> > $50
> > > > > Amazon gift card drawing. (If you’ve already participated in the
> > study
> > > > and
> > > > > didn’t win in our last drawing, you’re automatically entered.)
> Study
> > > info
> > > > > can be found here:
> > > >
> >
> https://urldefense.proofpoint.com/v2/url?u=https-3A__sites.google.com_view_lmrg-2Dimage-2Danalysis-2Dstudy&d=DwIFaQ&c=eLbWYnpnzycBCgmb7vCI4uqNEB9RSjOdn_5nBEmmeq0&r=wgGHXpNkPeE6PJJsl0IvoA&m=Bbo8EpRKzdBsReltSfbg8Udr6_-vbdBTWzo2_VmZUow&s=AvCI1kJRHZzgNZUP-7MfZJdOZZaCKUJOvPZAamulY-0&e=
> > > > >
> > > > > If you have questions about the event, please contact Jessica
> > Hornick (
> > > > > [log in to unmask]).
> > > > >
> > > >
> > >
> >
>
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