Re: IMAGEJ Digest - 30 Apr 2015 to 1 May 2015 (#2015-141)

[chaitanya athale]

Hi jacqui,
For rod shaped bacterial cells we found a simple gradient threshold filter
worked quite well. While you can look it up here:

Athale and Chaidhari (2011) bioinformatics

we have an experimental imageJ plugin we are still testing. If you send an
example image we can share the code if it works. In essence it works on
maximum filtering of previously smoothed images.

Cheers,
Chaitanya

On Saturday 2 May 2015, IMAGEJ automatic digest system <
[hidden email]> wrote:

> There are 11 messages totaling 3077 lines in this issue.
>
> Topics of the day:
>
>   1. Segmentation of DIC or Hoffman Modulation Contrast images - help
> please
>      (6)
>   2. FW: Segmentation of DIC or Hoffman Modulation Contrast images - help
>      please
>   3. Problem using Image Sequence command from a macro (2)
>   4. Labels and default values for @parameter in scripts
>   5. Saving large files in ome.tif or ids/ics
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ----------------------------------------------------------------------
>
> Date:    Fri, 1 May 2015 04:37:08 +0000
> From:    Jacqui Ross <[hidden email] <javascript:;>>
> Subject: Segmentation of DIC or Hoffman Modulation Contrast images - help
> please
>
> Hi All,
>
> I'm helping a PhD student with analysing some Hoffman modulation contrast
> images of cells. She's primarily interested in changes in diameter. The
> cells are embedded in a 3D matrix and compression is being applied.
>
> In the images, there are nice cells in focus with clear boundaries, plus
> others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> These kind of images are really tricky to segment as anyone who has tried,
> already knows. I've tried lots of different filters (edge, etc.) , FFT
> filtering and the Trainable Weka Segmentation but have been unable to
> achieve good enough results to be able to then threshold the cells
> automatically.
>
> I've come to the end of the line for now so am asking for your expert help
> in case anyone has some suggestions:). I note that in 2006 Monique Vasseur
> offered some DIC images to a PhD student called Daniel Mauch in Germany but
> I'm not sure if anything came of that project. There are a few papers out
> there (some mention Hilbert Transform, FFT) but I haven't been successful
> in implementing anything from those papers as yet.
>
> In the meantime, my solution is to use the Pseudo flat field correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very small
> radius (5) to flatten the background and out of focus cells while
> preserving the in focus cell outlines. We can then use the Cell Magic Wand
> (Thanks Theo!) to create selections that can be loaded into the ROI Manager
> and then measured. This works really well but requires that the cells be
> selected manually.
>
> The Cell Magic Wand Tool works on the colour or grayscale so we can also
> split the channels from the colour image if needed and use the channel
> image with the most contrast.
>
> I've attached an image in case anyone has any ideas. The image has been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
>
> Look forward to hearing any suggestions!
>
> Kind regards,
>
> Jacqui
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 08:05:23 +0300
> From:    Aryeh Weiss <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Try converting to 8-bit and running a variance filter
> (Process>Filters>Variance...) followed by thresholding. This will enhance
> the cells in focus due to their texture.
>
> --aryeh
>
> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is being
> applied.
> >
> > In the images, there are nice cells in focus with clear boundaries, plus
> others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge, etc.) ,
> FFT filtering and the Trainable Weka Segmentation but have been unable to
> achieve good enough results to be able to then threshold the cells
> automatically.
> >
> > I've come to the end of the line for now so am asking for your expert
> help in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur offered some DIC images to a PhD student called Daniel Mauch in
> Germany but I'm not sure if anything came of that project. There are a few
> papers out there (some mention Hilbert Transform, FFT) but I haven't been
> successful in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very small
> radius (5) to flatten the background and out of focus cells while
> preserving the in focus cell outlines. We can then use the Cell Magic Wand
> (Thanks Theo!) to create selections that can be loaded into the ROI Manager
> and then measured. This works really well but requires that the cells be
> selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can also
> split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences
> > The University of Auckland
> > Private Bag 92019
> > Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 06:00:05 +0000
> From:    Jacqui Ross <[hidden email] <javascript:;>>
> Subject: FW: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Hi All,
>
> Just realised my reply to Aryeh didn't get to the listserv. Please see
> below.
>
> Cheers,
>
> Jacqui
>
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> -----Original Message-----
> From: Jacqui Ross
> Sent: Friday, 1 May 2015 5:48 p.m.
> To: '[hidden email] <javascript:;>'
> Subject: RE: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Hi Aryeh,
>
> Thanks for your reply. I did try using a variance filter (the built-in one
> under Process - Filters - Variance) with different radii but I was unable
> to achieve a good result. The resultant circles were often incomplete so
> that when I then converted to binary, I had to do a lot of additional
> processing (Closing, filling holes, etc.) and then the outlines weren't
> very accurate.
>
> I know that you presented on the Trainable Weka Segmentation at the ImageJ
> conference that I attended a couple of years ago. My notes on that weren't
> fantastic (I pulled them out!) but in your case you also had some
> fluorescence labelling to help inform the segmentation.
>
> Kind regards,
>
> Jacqui
>
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> -----Original Message-----
> From: Aryeh Weiss [mailto:[hidden email] <javascript:;>] On
> Behalf Of Aryeh Weiss
> Sent: Friday, 1 May 2015 5:07 p.m.
> To: Jacqui Ross
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
>
> Try converting to 8-bit and running a variance filter
> (Process>Filters>Variance...) followed by thresholding. This will enhance
> the cells in focus due to their texture.
>
> --aryeh
>
> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is being
> applied.
> >
> > In the images, there are nice cells in focus with clear boundaries, plus
> others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge, etc.) ,
> FFT filtering and the Trainable Weka Segmentation but have been unable to
> achieve good enough results to be able to then threshold the cells
> automatically.
> >
> > I've come to the end of the line for now so am asking for your expert
> help in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur offered some DIC images to a PhD student called Daniel Mauch in
> Germany but I'm not sure if anything came of that project. There are a few
> papers out there (some mention Hilbert Transform, FFT) but I haven't been
> successful in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very small
> radius (5) to flatten the background and out of focus cells while
> preserving the in focus cell outlines. We can then use the Cell Magic Wand
> (Thanks Theo!) to create selections that can be loaded into the ROI Manager
> and then measured. This works really well but requires that the cells be
> selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can also
> split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences
> > The University of Auckland
> > Private Bag 92019
> > Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 16:02:08 +0300
> From:    Aryeh Weiss <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> On 5/1/15 8:48 AM, Jacqui Ross wrote:
> > Hi Aryeh,
> >
> > Thanks for your reply. I did try using a variance filter (the built-in
> one under Process - Filters - Variance) with different radii but I was
> unable to achieve a good result. The resultant circles were often
> incomplete so that when I then converted to binary, I had to do a lot of
> additional processing (Closing, filling holes, etc.) and then the outlines
> weren't very accurate.
> Yes -- these methods are better at marking objects than getting accurate
> boundaries. You might be able to use the inaccurate segmentation that
> produces as a mask against the original variance image, which produces
> reasonable arcs around your in-focus cells.
> > I know that you presented on the Trainable Weka Segmentation at the
> ImageJ conference that I attended a couple of years ago. My notes on that
> weren't fantastic (I pulled them out!) but in your case you also had some
> fluorescence labelling to help inform the segmentation.
> That problem was easier than yours (isn't it always that way?). The
> texture was much better defined, and I did not have a continuum of
> out-of-focus or partially out-of-focus cells. There was a DAPI channel
> which I used to exclude artitfacts which were not cells (since they did
> not contain nuclei). However, that will not help you, since your out of
> focus cells are still cells.
>
> You might have an easier time here if you can acquire a z-stack (even
> though it is wide-field) and use the EDF plugin or similar to have all
> of your cells in-focus. That would be easier to segment. Alternatively,
> if you choose to be very strict in your segmentation (ie, take only the
> cells that are really sharp and textured), then I think you will be able
> to variance and it relatives to get an accurate segmentation and boundary.
>
> Best regards
> --aryeh
>
> >
> > -----Original Message-----
> > From: Aryeh Weiss [mailto:[hidden email] <javascript:;>] On
> Behalf Of Aryeh Weiss
> > Sent: Friday, 1 May 2015 5:07 p.m.
> > To: Jacqui Ross
> > Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
> >
> >
> > Try converting to 8-bit and running a variance filter
> > (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
> >
> > --aryeh
> >
> > On 5/1/15 7:37 AM, Jacqui Ross wrote:
> >> Hi All,
> >>
> >> I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is being
> applied.
> >>
> >> In the images, there are nice cells in focus with clear boundaries,
> plus others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> >> These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge, etc.) ,
> FFT filtering and the Trainable Weka Segmentation but have been unable to
> achieve good enough results to be able to then threshold the cells
> automatically.
> >>
> >> I've come to the end of the line for now so am asking for your expert
> help in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur offered some DIC images to a PhD student called Daniel Mauch in
> Germany but I'm not sure if anything came of that project. There are a few
> papers out there (some mention Hilbert Transform, FFT) but I haven't been
> successful in implementing anything from those papers as yet.
> >>
> >> In the meantime, my solution is to use the Pseudo flat field correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very small
> radius (5) to flatten the background and out of focus cells while
> preserving the in focus cell outlines. We can then use the Cell Magic Wand
> (Thanks Theo!) to create selections that can be loaded into the ROI Manager
> and then measured. This works really well but requires that the cells be
> selected manually.
> >>
> >> The Cell Magic Wand Tool works on the colour or grayscale so we can
> also split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >>
> >> I've attached an image in case anyone has any ideas. The image has been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >>
> >> Look forward to hearing any suggestions!
> >>
> >> Kind regards,
> >>
> >> Jacqui
> >> Jacqueline Ross
> >> Biomedical Imaging Microscopist
> >> Biomedical Imaging Research Unit
> >> School of Medical Sciences
> >> Faculty of Medical & Health Sciences
> >> The University of Auckland
> >> Private Bag 92019
> >> Auckland 1142, NEW ZEALAND
> >>
> >> Tel: [phone]
> >> Fax: [phone]
> >>
> >> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >
> > --
> > Aryeh Weiss
> > Faculty of Engineering
> > Bar Ilan University
> > Ramat Gan 52900 Israel
> >
> > Ph:  972-3-5317638
> > FAX: [phone]
> >
> >
> > .
> >
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 09:07:27 -0500
> From:    Kenneth Sloan <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> I would consider an approach that is one level higher than raw image
> processing.  DIC images produce distinctive doublets (a bright region
> adjacent to a darker region - always (for the same setup) at the same angle.
>
> Once you can identify a cell from it’s doublet, and have a reasonable
> guess as to its size, finding the boundary and estimating area should
> become easier.
>
> Just a thought.
>
> My first try might be to design a custom convolution kernel that responds
> preferentially to doublets at a fixed angle.  There is still the problem of
> estimating the angle (if it’s not stable)  but that should be doable based
> either on meta-information or user input.
>
>
> Are all the cells as well-behaved as these?
>
> I second the idea of using a texture measurement to eliminate regions of
> the image that are out of focus (variance is one - I would perhaps go a
> step further and use coefficient of variation - stdDev/mean - but there are
> others)
>
> --
> Kenneth Sloan
> [hidden email] <javascript:;>
> Vision is the art of seeing what is invisible to others.
>
>
>
>
> > On May 1, 2015, at 08:02 , Aryeh Weiss <[hidden email]
> <javascript:;>> wrote:
> >
> > On 5/1/15 8:48 AM, Jacqui Ross wrote:
> >> Hi Aryeh,
> >>
> >> Thanks for your reply. I did try using a variance filter (the built-in
> one under Process - Filters - Variance) with different radii but I was
> unable to achieve a good result. The resultant circles were often
> incomplete so that when I then converted to binary, I had to do a lot of
> additional processing (Closing, filling holes, etc.) and then the outlines
> weren't very accurate.
> > Yes -- these methods are better at marking objects than getting accurate
> boundaries. You might be able to use the inaccurate segmentation that
> produces as a mask against the original variance image, which produces
> reasonable arcs around your in-focus cells.
> >> I know that you presented on the Trainable Weka Segmentation at the
> ImageJ conference that I attended a couple of years ago. My notes on that
> weren't fantastic (I pulled them out!) but in your case you also had some
> fluorescence labelling to help inform the segmentation.
> > That problem was easier than yours (isn't it always that way?). The
> texture was much better defined, and I did not have a continuum of
> out-of-focus or partially out-of-focus cells. There was a DAPI channel
> which I used to exclude artitfacts which were not cells (since they did not
> contain nuclei). However, that will not help you, since your out of focus
> cells are still cells.
> >
> > You might have an easier time here if you can acquire a z-stack (even
> though it is wide-field) and use the EDF plugin or similar to have all of
> your cells in-focus. That would be easier to segment. Alternatively, if you
> choose to be very strict in your segmentation (ie, take only the cells that
> are really sharp and textured), then I think you will be able to variance
> and it relatives to get an accurate segmentation and boundary.
> >
> > Best regards
> > --aryeh
> >
> >>
> >> -----Original Message-----
> >> From: Aryeh Weiss [mailto:[hidden email] <javascript:;>] On
> Behalf Of Aryeh Weiss
> >> Sent: Friday, 1 May 2015 5:07 p.m.
> >> To: Jacqui Ross
> >> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
> >>
> >>
> >> Try converting to 8-bit and running a variance filter
> >> (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
> >>
> >> --aryeh
> >>
> >> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> >>> Hi All,
> >>>
> >>> I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is being
> applied.
> >>>
> >>> In the images, there are nice cells in focus with clear boundaries,
> plus others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> >>> These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge, etc.) ,
> FFT filtering and the Trainable Weka Segmentation but have been unable to
> achieve good enough results to be able to then threshold the cells
> automatically.
> >>>
> >>> I've come to the end of the line for now so am asking for your expert
> help in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur offered some DIC images to a PhD student called Daniel Mauch in
> Germany but I'm not sure if anything came of that project. There are a few
> papers out there (some mention Hilbert Transform, FFT) but I haven't been
> successful in implementing anything from those papers as yet.
> >>>
> >>> In the meantime, my solution is to use the Pseudo flat field
> correction plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a
> very small radius (5) to flatten the background and out of focus cells
> while preserving the in focus cell outlines. We can then use the Cell Magic
> Wand (Thanks Theo!) to create selections that can be loaded into the ROI
> Manager and then measured. This works really well but requires that the
> cells be selected manually.
> >>>
> >>> The Cell Magic Wand Tool works on the colour or grayscale so we can
> also split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >>>
> >>> I've attached an image in case anyone has any ideas. The image has
> been cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >>>
> >>> Look forward to hearing any suggestions!
> >>>
> >>> Kind regards,
> >>>
> >>> Jacqui
> >>> Jacqueline Ross
> >>> Biomedical Imaging Microscopist
> >>> Biomedical Imaging Research Unit
> >>> School of Medical Sciences
> >>> Faculty of Medical & Health Sciences
> >>> The University of Auckland
> >>> Private Bag 92019
> >>> Auckland 1142, NEW ZEALAND
> >>>
> >>> Tel: [phone]
> >>> Fax: [phone]
> >>>
> >>> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>>
> >>>
> >>> --
> >>> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >> --
> >> Aryeh Weiss
> >> Faculty of Engineering
> >> Bar Ilan University
> >> Ramat Gan 52900 Israel
> >>
> >> Ph:  972-3-5317638
> >> FAX: [phone]
> >>
> >>
> >> .
> >>
> >
> >
> > --
> > Aryeh Weiss
> > Faculty of Engineering
> > Bar Ilan University
> > Ramat Gan 52900 Israel
> >
> > Ph:  972-3-5317638
> > FAX: [phone]
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 10:10:20 -0400
> From:    "JOEL B. SHEFFIELD" <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Hi Jacqui,
>
> These images are particularly difficult to segment because the edges are
> assymetric --dark on one side, and light on the other.  I was able to get
> some enhancement by using an unsharp mask (on your image, pixel radius of
> 100, weight .60), followed by the "find edges" convolution.  It wasn't
> perfect, but might help.
>
> Joel
>
>
>
> Joel B. Sheffield, Ph.D
> Department of Biology
> Temple University
> Philadelphia, PA 19122
> Voice: 215 204 8839
> e-mail: [hidden email] <javascript:;>
> URL:  *http://tinyurl.com/khbouft <http://tinyurl.com/khbouft>*
>
> On Fri, May 1, 2015 at 12:37 AM, Jacqui Ross <[hidden email]
> <javascript:;>>
> wrote:
>
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation contrast
> > images of cells. She's primarily interested in changes in diameter. The
> > cells are embedded in a 3D matrix and compression is being applied.
> >
> > In the images, there are nice cells in focus with clear boundaries, plus
> > others which are out of focus which we don't want to measure as any
> > measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried,
> > already knows. I've tried lots of different filters (edge, etc.) , FFT
> > filtering and the Trainable Weka Segmentation but have been unable to
> > achieve good enough results to be able to then threshold the cells
> > automatically.
> >
> > I've come to the end of the line for now so am asking for your expert
> help
> > in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur
> > offered some DIC images to a PhD student called Daniel Mauch in Germany
> but
> > I'm not sure if anything came of that project. There are a few papers out
> > there (some mention Hilbert Transform, FFT) but I haven't been successful
> > in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field correction
> > plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very
> small
> > radius (5) to flatten the background and out of focus cells while
> > preserving the in focus cell outlines. We can then use the Cell Magic
> Wand
> > (Thanks Theo!) to create selections that can be loaded into the ROI
> Manager
> > and then measured. This works really well but requires that the cells be
> > selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can also
> > split the channels from the colour image if needed and use the channel
> > image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has been
> > cropped out of a larger image so that it's not too big and it's pink
> > because there's cell culture medium there (in case anyone was
> wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences
> > The University of Auckland
> > Private Bag 92019
> > Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 14:36:03 +0000
> From:    Kenneth R Sloan <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Again - I’ll promote taking a higher level approach.  Simple SEGMENTATION
> should be able to pull out “Bright blobs” and “Dark blobs” - segmenting the
> CELLS requires combining that evidence (and then perhaps going back to the
> image data in “verification vision” mode - making predictions about what
> the cell boundary will look like, and localizing it.  But, this is probably
> beyond the scope of a script combining standard image processing operators.
>
> --
> Kenneth Sloan
> [hidden email] <javascript:;>
> Vision is the art of seeing what is invisible to others.
>
>
>
>
> > On May 1, 2015, at 09:10 , JOEL B. SHEFFIELD <[hidden email]
> <javascript:;>> wrote:
> >
> > Hi Jacqui,
> >
> > These images are particularly difficult to segment because the edges are
> > assymetric --dark on one side, and light on the other.  I was able to get
> > some enhancement by using an unsharp mask (on your image, pixel radius of
> > 100, weight .60), followed by the "find edges" convolution.  It wasn't
> > perfect, but might help.
> >
> > Joel
> >
> >
> >
> > Joel B. Sheffield, Ph.D
> > Department of Biology
> > Temple University
> > Philadelphia, PA 19122
> > Voice: 215 204 8839
> > e-mail: [hidden email] <javascript:;>
> > URL:  *http://tinyurl.com/khbouft <http://tinyurl.com/khbouft>*
> >
> > On Fri, May 1, 2015 at 12:37 AM, Jacqui Ross <[hidden email]
> <javascript:;>>
> > wrote:
> >
> >> Hi All,
> >>
> >> I'm helping a PhD student with analysing some Hoffman modulation
> contrast
> >> images of cells. She's primarily interested in changes in diameter. The
> >> cells are embedded in a 3D matrix and compression is being applied.
> >>
> >> In the images, there are nice cells in focus with clear boundaries, plus
> >> others which are out of focus which we don't want to measure as any
> >> measurements won't be accurate.
> >> These kind of images are really tricky to segment as anyone who has
> tried,
> >> already knows. I've tried lots of different filters (edge, etc.) , FFT
> >> filtering and the Trainable Weka Segmentation but have been unable to
> >> achieve good enough results to be able to then threshold the cells
> >> automatically.
> >>
> >> I've come to the end of the line for now so am asking for your expert
> help
> >> in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur
> >> offered some DIC images to a PhD student called Daniel Mauch in Germany
> but
> >> I'm not sure if anything came of that project. There are a few papers
> out
> >> there (some mention Hilbert Transform, FFT) but I haven't been
> successful
> >> in implementing anything from those papers as yet.
> >>
> >> In the meantime, my solution is to use the Pseudo flat field correction
> >> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very
> small
> >> radius (5) to flatten the background and out of focus cells while
> >> preserving the in focus cell outlines. We can then use the Cell Magic
> Wand
> >> (Thanks Theo!) to create selections that can be loaded into the ROI
> Manager
> >> and then measured. This works really well but requires that the cells be
> >> selected manually.
> >>
> >> The Cell Magic Wand Tool works on the colour or grayscale so we can also
> >> split the channels from the colour image if needed and use the channel
> >> image with the most contrast.
> >>
> >> I've attached an image in case anyone has any ideas. The image has been
> >> cropped out of a larger image so that it's not too big and it's pink
> >> because there's cell culture medium there (in case anyone was
> wondering..).
> >>
> >> Look forward to hearing any suggestions!
> >>
> >> Kind regards,
> >>
> >> Jacqui
> >> Jacqueline Ross
> >> Biomedical Imaging Microscopist
> >> Biomedical Imaging Research Unit
> >> School of Medical Sciences
> >> Faculty of Medical & Health Sciences
> >> The University of Auckland
> >> Private Bag 92019
> >> Auckland 1142, NEW ZEALAND
> >>
> >> Tel: [phone]
> >> Fax: [phone]
> >>
> >> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 19:15:40 +0000
> From:    "Paletzki, Ron (NIH/NIMH) [C]" <[hidden email]
> <javascript:;>>
> Subject: Problem using Image Sequence command from a macro
>
> I have a macro that creates a stack and then tries to save it to a folder
> using the Image Sequence command.   Problem is it works fine if the folder
> it’s saving to has no spaces in the name but fails if it does.   I’m
> wondering if this is a bug since it has never been an issue before in other
> macros using other save and open commands.  This is the first macro using
> the Image Sequence command.
>
> Here is an example macro that demonstrates the bug.  With a stack open if
> I run this macro and select a folder with a name that contains a space in
> it I get an error
>  “File Saving error (IOException):
>
> If I select a folder that does not contain a space in the name it works
> fine.
>
> macro "Save Stack  [f2]"{
>     dirOutput = getDirectory("Choose Output Directory for stack");
>     Dialog.create(" Macro Options ");
>     Dialog.addString("Enter Filename for stack ","fileName");
>     Dialog.show();
>     fileNameStack = Dialog.getString();
>     run("Image Sequence... ", "format=TIFF name="+fileNameStack+" digits=3
> save="+dirOutput);
> }
>
>
> By the way this in running on a Mac using Image 149s8
>
> Thanks
> Ron
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 13:03:53 -0700
> From:    Glen MacDonald <[hidden email] <javascript:;>>
> Subject: Re: Problem using Image Sequence command from a macro
>
> Ron,
> Use brackets to denote string variables with spaces.
>
> macro "Save Stack  [f2]"{
>    dirOutput = getDirectory("Choose Output Directory for stack");
>    Dialog.create(" Macro Options ");
>    Dialog.addString("Enter Filename for stack ","fileName");
>    Dialog.show();
>    fileNameStack = Dialog.getString();
>    run("Image Sequence... ", "format=TIFF name=["+fileNameStack+"]
> digits=3 save=[dirOutput]");
> }
>
>
> Glen MacDonald
>         Core for Communication Research
> Virginia Merrill Bloedel Hearing Research Center
>         Cellular Morphology Core
> Center on Human Development and Disability
> Box 357923
> University of Washington
> Seattle, WA 98195-7923  USA
> [phone]
> [hidden email] <javascript:;>
>
>
>
>
>
>
>
> On May 1, 2015, at 12:15 PM, Paletzki, Ron (NIH/NIMH) [C] <
> [hidden email] <javascript:;>> wrote:
>
> > I have a macro that creates a stack and then tries to save it to a
> folder using the Image Sequence command.   Problem is it works fine if the
> folder it’s saving to has no spaces in the name but fails if it does.   I’m
> wondering if this is a bug since it has never been an issue before in other
> macros using other save and open commands.  This is the first macro using
> the Image Sequence command.
> >
> > Here is an example macro that demonstrates the bug.  With a stack open
> if I run this macro and select a folder with a name that contains a space
> in it I get an error
> > “File Saving error (IOException):
> >
> > If I select a folder that does not contain a space in the name it works
> fine.
> >
> > macro "Save Stack  [f2]"{
> >    dirOutput = getDirectory("Choose Output Directory for stack");
> >    Dialog.create(" Macro Options ");
> >    Dialog.addString("Enter Filename for stack ","fileName");
> >    Dialog.show();
> >    fileNameStack = Dialog.getString();
> >    run("Image Sequence... ", "format=TIFF name="+fileNameStack+"
> digits=3 save="+dirOutput);
> > }
> >
> >
> > By the way this in running on a Mac using Image 149s8
> >
> > Thanks
> > Ron
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 15:10:07 -0500
> From:    Mark Hiner <[hidden email] <javascript:;>>
> Subject: Re: Labels and default values for @parameter in scripts
>
> Hello,
>
> Just wanted you to know that if you update to the latest release[1] many of
> the issues you brought up have been resolved and improved.
>
> >You can't select the full input field by using a double click
>
> In my original reply I forgot to mention that I believe this is a Java-ism.
> The most reliable way to select the full text of an input field with the
> mouse is to triple click.
>
> Thanks again.
>
> Best,
> Mark
>
> [1] http://imagej.net/2015-05-01_-_ImageJ_2.0.0-rc-30
>
> On Thu, Apr 30, 2015 at 9:32 AM, Mark Hiner <[hidden email] <javascript:;>>
> wrote:
>
> > Hi Michael,
> >
> > It's great to see people using script parameters. These are a new feature
> > introduced with ImageJ2[1], and thus are still relatively new - so
> feedback
> > like this is both helpful and necessary in making these features robust.
> > For ImageJ2-specific features like this, you can also use the
> imagej-devel
> > mailing list[2] or file issues directly on GitHub[3].
> >
> > > Except this page I can't find any documentation on using script
> > parameters in ImageJ. Is there some more documentation, I was not able to
> > find?
> >
> > This is the intended documentation page. It's definitely sparse right
> now.
> > I am updating it in response to your feedback, but if you have any more
> > suggestions/requests for what would be helpful here, please let us know.
> >
> > > In plugins you can define labels and default values for parameters. In
> > scripts this is not possible.
> >
> > I had to dig around for this a bit, but it actually is possible! It uses
> > the same Java syntax as plugins, but requires the properties (like labels
> > and default values) to be in a parenthetical expression. Examples are now
> > on the wiki[4].
> >
> > >The default values are strange.
> >
> > Agreed. I added an issue[5] for this.
> >
> > > You can't select the full input field by using a double click.
> > > Using [Tab] to switch between fields works bad
> >
> > These are UI-specific[6] issues and so shouldn't be problems with
> > scripting itself. I was able to confirm this behavior on a mac, so I'll
> > look into it.
> >
> > Thank you for the comments!
> >
> > Best,
> > Mark
> >
> > [1] http://imagej.net/ImageJ2
> > [2] http://imagej.net/Mailing_lists
> > [3] http://imagej.net/Source_code#Source_code
> > [4] http://imagej.net/Script_parameters#Parameter_properties
> > [5] https://github.com/scijava/scijava-common/issues/160
> > [6] https://github.com/scijava/scijava-ui-swing
> >
> > On Thu, Apr 30, 2015 at 6:28 AM, Michael Entrup <
> > [hidden email] <javascript:;>> wrote:
> >
> >> Hi,
> >>
> >> using script parameters might be a nice solution to write small scripts
> >> that need user inputs. But there are some drawbacks that keep me away
> from
> >> using script parameters.
> >>
> >> In plugins you can define labels and default values for parameters. In
> >> scripts this is not possible.
> >>
> >> The default values are strange. When using @Integer the default value is
> >> '-2.147.483.648'.
> >>
> >> You can't select the full input field by using a double click. The minus
> >> sign is not selected, when double clicking the number.
> >>
> >> Using [Tab] to switch between fields works bad. Normally I expect that
> >> the content of the next input field is selected, but only the cursor is
> >> placed at the next field. [Ctrl]+[A] is necessary to replace the default
> >> value.
> >>
> >> The attached screen shot shows the dialog that is shown, when using 4
> >> different script parameters.
> >>
> >> Is all this intended, or is still work in progress?
> >> Except this page [1] I can't find any documentation on using script
> >> parameters in ImageJ. Is there some more documentation, I was not able
> to
> >> find?
> >>
> >> Best regards
> >> Michael
> >>
> >>
> >> [1] http://imagej.net/Script_parameters
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >
> >
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 19:37:59 -0400
> From:    Jason Miller <[hidden email] <javascript:;>>
> Subject: Re: Saving large files in ome.tif or ids/ics
>
> Hi all-
>
> I received an answer from the OME folks about my question and figured I'd
> post here to keep this in the same thread. This was helpful info for me.
>
> -------------------------------
> Dear Jason,
>
> > Similar to recent past posts on this list, I'm having trouble saving
> > files that are > 4GB into ome.tif or IDS/ICS formats. I have a 5GB file,
> > open as a virtual file, and when I go to export via Bioformats Exporter
> > to IDS/ICS format, I get the following error:
> [stacktrace]
>
> For IDS/ICS, this was fixed in this PR last week:
>    https://github.com/openmicroscopy/bioformats/pull/1738
> I have tested this with >5GB images and it works correctly.
>
> > When I go to export to ome.tif format, I get the same error previously
> > reported on this list:
> > loci.formats.FormatException: File is too large; call setBigTiff(true)
> >
> > Is it possible to retain the ome.tif format for >4GB? In my case, this
> > is for importing into Huygens for deconvolution.
>
> For plain (not OME) TIFF:
>    https://github.com/openmicroscopy/bioformats/pull/1744
>
> Both of the above fixes are in the newly-released Bio-Formats 5.1.1
> release.  If using Fiji, you should automatically get this update; for
> ImageJ you can download the updated plugin from
>    http://downloads.openmicroscopy.org/bio-formats/5.1.1/
>
> For OME-TIFF, it's not currently possible to enable BigTIFF when
> exporting from ImageJ using the exporter dialogue.  For the plain TIFF
> case, rather than relying on an API call to enable BigTIFF, we now
> enable it if the image data is >4GiB or a "big" TIFF file extension such
> as .btf, .tf8 or .tf2 is used.  We would like to do the same for
> OME-TIFF, but this first requires updating the OME-TIFF specification
> which currently only allows a .ome.tiff or .ome.tif file extension.
> Once that's done, you would be able to use e.g. ".ome.btf" and get a
> "big" OME-TIFF file.  However, this does require some consideration
> since (for example) these files would not be readable with older
> versions of Bio-Formats unless you renamed them to ".ome.tiff" after
> export.
>
> To summarise, it isn't currently possible to export >4GB ome.tiff files
> using the Bio-Formats Exporter.  It is possible via the Java API by
> calling writer.setBigTiff(true) if this is an option for you.  This is a
> known limitation which we are working on; see the discussion in the PR
> #1744 above.
>
> There is a ticket open here for this:
>    https://trac.openmicroscopy.org/ome/ticket/12858
> I can add you as a CC on it if you like, so you'll be kept up to date
> with its progress?
>
>
> Kind regards,
> Roger
>
> --
> Dr Roger Leigh -- Open Microscopy Environment
> Wellcome Trust Centre for Gene Regulation and Expression,
> College of Life Sciences, University of Dundee, Dow Street,
> Dundee DD1 5EH Scotland UK   Tel: [phone]
>
> The University of Dundee is a registered Scottish Charity, No: SC015096
>
> On Thu, Apr 30, 2015 at 5:28 PM, Jason Miller <[hidden email]
> <javascript:;>>
> wrote:
>
> > Hi all-
> >
> > This may be a Bioformats issue as well, so I will separately e-mail the
> > OME folks, but similar to recent past posts on this list, I'm having
> > trouble saving files that are > 4GB into ome.tif or IDS/ICS formats. I
> have
> > a 5GB file, open as a virtual file, and when I go to export via
> Bioformats
> > Exporter to IDS/ICS format, I get the following error:
> >
> > (Fiji Is Just) ImageJ 2.0.0-rc-15/1.49p; Java 1.6.0_24 [64-bit]; Windows
> 7
> > 6.1; 98MB of 12209MB (<1%)
> >
> > java.lang.IllegalArgumentException: Negative position
> > at sun.nio.ch.FileChannelImpl.read(FileChannelImpl.java:600)
> > at
> >
> loci.common.NIOByteBufferProvider.allocateDirect(NIOByteBufferProvider.java:131)
> > at
> >
> loci.common.NIOByteBufferProvider.allocate(NIOByteBufferProvider.java:116)
> > at loci.common.NIOFileHandle.buffer(NIOFileHandle.java:551)
> > at loci.common.NIOFileHandle.seek(NIOFileHandle.java:273)
> > at
> >
> loci.common.RandomAccessOutputStream.seek(RandomAccessOutputStream.java:79)
> > at loci.formats.out.ICSWriter.saveBytes(ICSWriter.java:139)
> > at loci.formats.FormatWriter.saveBytes(FormatWriter.java:126)
> > at loci.plugins.out.Exporter.run(Exporter.java:629)
> > at loci.plugins.LociExporter.run(LociExporter.java:77)
> > at
> >
> ij.plugin.filter.PlugInFilterRunner.processOneImage(PlugInFilterRunner.java:263)
> > at
> ij.plugin.filter.PlugInFilterRunner.<init>(PlugInFilterRunner.java:112)
> > at ij.IJ.runUserPlugIn(IJ.java:201)
> > at ij.IJ.runPlugIn(IJ.java:163)
> > at ij.Executer.runCommand(Executer.java:131)
> > at ij.Executer.run(Executer.java:64)
> > at java.lang.Thread.run(Thread.java:662)
> >
> >
> > When I go to export to ome.tif format, I get the same error previously
> > reported on this list:
> > loci.formats.FormatException: File is too large; call setBigTiff(true)
> >
> > Is it possible to retain the ome.tif format for >4GB? In my case, this is
> > for importing into Huygens for deconvolution.
> >
> > Thanks so very much
> > -Jason
> > --
> >
> > Jason Miller, MD, PhD
> >
> > University of Michigan Kellogg Eye Center
> >
> > --
> >
> > Home address:
> >
> > 117 Worden Ave
> >
> > Ann Arbor, MI 48103
> >
> > Cell: [phone]
> >
> > E-mail: *[hidden email] <javascript:;>
> > <[hidden email] <javascript:;>>*
> >
> >
> > "When I was 5 years old, my mother always told me that happiness was the
> > key to life. When I went to school, they asked me what I wanted to be
> when
> > I grew up. I wrote down, 'Happy.' They told me I didn't understand the
> > assignment. I told them they didn't understand life." - John Lennon
> >
>
>
>
> --
>
> Jason Miller, MD, PhD
>
> University of Michigan Kellogg Eye Center
>
> --
>
> Home address:
>
> 117 Worden Ave
>
> Ann Arbor, MI 48103
>
> Cell: [phone]
>
> E-mail: *[hidden email] <javascript:;> <
> [hidden email] <javascript:;>>*
>
>
> "When I was 5 years old, my mother always told me that happiness was the
> key to life. When I went to school, they asked me what I wanted to be when
> I grew up. I wrote down, 'Happy.' They told me I didn't understand the
> assignment. I told them they didn't understand life." - John Lennon
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> End of IMAGEJ Digest - 30 Apr 2015 to 1 May 2015 (#2015-141)
> ************************************************************
>

--
-------------------------
Chaitanya Athale,
Pune, India
18° 31' N, 73° 55' E, 560m.
-------------------------

--
ImageJ mailing list: http://imagej.nih.gov/ij/list.html

[Jacqueline Ross]

Hi Chaitanya,

I've had a look at your paper and the images do look similar to ours, other than being rod-shaped.

It would be fantastic if you could try out your plugin on one of Sophia's images.

The original images are about 14Mb so too large to send via email. However, I will upload one to our Web Dropoff box for you.

You will receive an email from The University of Auckland Dropoff box with a link for download.

I'll be very interested to hear how you get on.

Kind regards,

Jacqui

Jacqueline Ross
Biomedical Imaging Microscopist
Biomedical Imaging Research Unit 
School of Medical Sciences 
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland 1142, NEW ZEALAND

Tel: [phone]
Fax: [phone]

http://www.fmhs.auckland.ac.nz/sms/biru/


-----Original Message-----
From: ImageJ Interest Group [mailto:[hidden email]] On Behalf Of chaitanya athale
Sent: Sunday, 3 May 2015 2:21 a.m.
To: [hidden email]
Subject: Re: IMAGEJ Digest - 30 Apr 2015 to 1 May 2015 (#2015-141)

Hi jacqui,
For rod shaped bacterial cells we found a simple gradient threshold filter worked quite well. While you can look it up here:

Athale and Chaidhari (2011) bioinformatics

we have an experimental imageJ plugin we are still testing. If you send an example image we can share the code if it works. In essence it works on maximum filtering of previously smoothed images.

Cheers,
Chaitanya

On Saturday 2 May 2015, IMAGEJ automatic digest system < [hidden email]> wrote:

> There are 11 messages totaling 3077 lines in this issue.
>
> Topics of the day:
>
>   1. Segmentation of DIC or Hoffman Modulation Contrast images - help
> please
>      (6)
>   2. FW: Segmentation of DIC or Hoffman Modulation Contrast images - help
>      please
>   3. Problem using Image Sequence command from a macro (2)
>   4. Labels and default values for @parameter in scripts
>   5. Saving large files in ome.tif or ids/ics
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ----------------------------------------------------------------------
>
> Date:    Fri, 1 May 2015 04:37:08 +0000
> From:    Jacqui Ross <[hidden email] <javascript:;>>
> Subject: Segmentation of DIC or Hoffman Modulation Contrast images -
> help please
>
> Hi All,
>
> I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is being applied.
>
> In the images, there are nice cells in focus with clear boundaries,
> plus others which are out of focus which we don't want to measure as
> any measurements won't be accurate.
> These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge,
> etc.) , FFT filtering and the Trainable Weka Segmentation but have
> been unable to achieve good enough results to be able to then
> threshold the cells automatically.
>
> I've come to the end of the line for now so am asking for your expert
> help in case anyone has some suggestions:). I note that in 2006
> Monique Vasseur offered some DIC images to a PhD student called Daniel
> Mauch in Germany but I'm not sure if anything came of that project.
> There are a few papers out there (some mention Hilbert Transform, FFT)
> but I haven't been successful in implementing anything from those papers as yet.
>
> In the meantime, my solution is to use the Pseudo flat field
> correction plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!)
> with a very small radius (5) to flatten the background and out of
> focus cells while preserving the in focus cell outlines. We can then
> use the Cell Magic Wand (Thanks Theo!) to create selections that can
> be loaded into the ROI Manager and then measured. This works really
> well but requires that the cells be selected manually.
>
> The Cell Magic Wand Tool works on the colour or grayscale so we can
> also split the channels from the colour image if needed and use the
> channel image with the most contrast.
>
> I've attached an image in case anyone has any ideas. The image has
> been cropped out of a larger image so that it's not too big and it's
> pink because there's cell culture medium there (in case anyone was wondering..).
>
> Look forward to hearing any suggestions!
>
> Kind regards,
>
> Jacqui
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 08:05:23 +0300
> From:    Aryeh Weiss <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> Try converting to 8-bit and running a variance filter
> (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
>
> --aryeh
>
> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is
> being applied.
> >
> > In the images, there are nice cells in focus with clear boundaries,
> > plus
> others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge,
> etc.) , FFT filtering and the Trainable Weka Segmentation but have
> been unable to achieve good enough results to be able to then
> threshold the cells automatically.
> >
> > I've come to the end of the line for now so am asking for your
> > expert
> help in case anyone has some suggestions:). I note that in 2006
> Monique Vasseur offered some DIC images to a PhD student called Daniel
> Mauch in Germany but I'm not sure if anything came of that project.
> There are a few papers out there (some mention Hilbert Transform, FFT)
> but I haven't been successful in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field
> > correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very
> small radius (5) to flatten the background and out of focus cells
> while preserving the in focus cell outlines. We can then use the Cell
> Magic Wand (Thanks Theo!) to create selections that can be loaded into
> the ROI Manager and then measured. This works really well but requires
> that the cells be selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can
> > also
> split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has
> > been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences The University of Auckland
> > Private Bag 92019 Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 06:00:05 +0000
> From:    Jacqui Ross <[hidden email] <javascript:;>>
> Subject: FW: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> Hi All,
>
> Just realised my reply to Aryeh didn't get to the listserv. Please see
> below.
>
> Cheers,
>
> Jacqui
>
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> -----Original Message-----
> From: Jacqui Ross
> Sent: Friday, 1 May 2015 5:48 p.m.
> To: '[hidden email] <javascript:;>'
> Subject: RE: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> Hi Aryeh,
>
> Thanks for your reply. I did try using a variance filter (the built-in
> one under Process - Filters - Variance) with different radii but I was
> unable to achieve a good result. The resultant circles were often
> incomplete so that when I then converted to binary, I had to do a lot
> of additional processing (Closing, filling holes, etc.) and then the
> outlines weren't very accurate.
>
> I know that you presented on the Trainable Weka Segmentation at the
> ImageJ conference that I attended a couple of years ago. My notes on
> that weren't fantastic (I pulled them out!) but in your case you also
> had some fluorescence labelling to help inform the segmentation.
>
> Kind regards,
>
> Jacqui
>
> Jacqueline Ross
> Biomedical Imaging Microscopist
> Biomedical Imaging Research Unit
> School of Medical Sciences
> Faculty of Medical & Health Sciences
> The University of Auckland
> Private Bag 92019
> Auckland 1142, NEW ZEALAND
>
> Tel: [phone]
> Fax: [phone]
>
> http://www.fmhs.auckland.ac.nz/sms/biru/
>
>
> -----Original Message-----
> From: Aryeh Weiss [mailto:[hidden email] <javascript:;>] On
> Behalf Of Aryeh Weiss
> Sent: Friday, 1 May 2015 5:07 p.m.
> To: Jacqui Ross
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
>
> Try converting to 8-bit and running a variance filter
> (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
>
> --aryeh
>
> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is
> being applied.
> >
> > In the images, there are nice cells in focus with clear boundaries,
> > plus
> others which are out of focus which we don't want to measure as any
> measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge,
> etc.) , FFT filtering and the Trainable Weka Segmentation but have
> been unable to achieve good enough results to be able to then
> threshold the cells automatically.
> >
> > I've come to the end of the line for now so am asking for your
> > expert
> help in case anyone has some suggestions:). I note that in 2006
> Monique Vasseur offered some DIC images to a PhD student called Daniel
> Mauch in Germany but I'm not sure if anything came of that project.
> There are a few papers out there (some mention Hilbert Transform, FFT)
> but I haven't been successful in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field
> > correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very
> small radius (5) to flatten the background and out of focus cells
> while preserving the in focus cell outlines. We can then use the Cell
> Magic Wand (Thanks Theo!) to create selections that can be loaded into
> the ROI Manager and then measured. This works really well but requires
> that the cells be selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can
> > also
> split the channels from the colour image if needed and use the channel
> image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has
> > been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences The University of Auckland
> > Private Bag 92019 Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 16:02:08 +0300
> From:    Aryeh Weiss <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> On 5/1/15 8:48 AM, Jacqui Ross wrote:
> > Hi Aryeh,
> >
> > Thanks for your reply. I did try using a variance filter (the
> > built-in
> one under Process - Filters - Variance) with different radii but I was
> unable to achieve a good result. The resultant circles were often
> incomplete so that when I then converted to binary, I had to do a lot
> of additional processing (Closing, filling holes, etc.) and then the
> outlines weren't very accurate.
> Yes -- these methods are better at marking objects than getting
> accurate boundaries. You might be able to use the inaccurate
> segmentation that produces as a mask against the original variance
> image, which produces reasonable arcs around your in-focus cells.
> > I know that you presented on the Trainable Weka Segmentation at the
> ImageJ conference that I attended a couple of years ago. My notes on
> that weren't fantastic (I pulled them out!) but in your case you also
> had some fluorescence labelling to help inform the segmentation.
> That problem was easier than yours (isn't it always that way?). The
> texture was much better defined, and I did not have a continuum of
> out-of-focus or partially out-of-focus cells. There was a DAPI channel
> which I used to exclude artitfacts which were not cells (since they
> did not contain nuclei). However, that will not help you, since your
> out of focus cells are still cells.
>
> You might have an easier time here if you can acquire a z-stack (even
> though it is wide-field) and use the EDF plugin or similar to have all
> of your cells in-focus. That would be easier to segment.
> Alternatively, if you choose to be very strict in your segmentation
> (ie, take only the cells that are really sharp and textured), then I
> think you will be able to variance and it relatives to get an accurate segmentation and boundary.
>
> Best regards
> --aryeh
>
> >
> > -----Original Message-----
> > From: Aryeh Weiss [mailto:[hidden email] <javascript:;>] On
> Behalf Of Aryeh Weiss
> > Sent: Friday, 1 May 2015 5:07 p.m.
> > To: Jacqui Ross
> > Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast
> > images -
> help please
> >
> >
> > Try converting to 8-bit and running a variance filter
> > (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
> >
> > --aryeh
> >
> > On 5/1/15 7:37 AM, Jacqui Ross wrote:
> >> Hi All,
> >>
> >> I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is
> being applied.
> >>
> >> In the images, there are nice cells in focus with clear boundaries,
> plus others which are out of focus which we don't want to measure as
> any measurements won't be accurate.
> >> These kind of images are really tricky to segment as anyone who has
> tried, already knows. I've tried lots of different filters (edge,
> etc.) , FFT filtering and the Trainable Weka Segmentation but have
> been unable to achieve good enough results to be able to then
> threshold the cells automatically.
> >>
> >> I've come to the end of the line for now so am asking for your
> >> expert
> help in case anyone has some suggestions:). I note that in 2006
> Monique Vasseur offered some DIC images to a PhD student called Daniel
> Mauch in Germany but I'm not sure if anything came of that project.
> There are a few papers out there (some mention Hilbert Transform, FFT)
> but I haven't been successful in implementing anything from those papers as yet.
> >>
> >> In the meantime, my solution is to use the Pseudo flat field
> >> correction
> plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!) with a very
> small radius (5) to flatten the background and out of focus cells
> while preserving the in focus cell outlines. We can then use the Cell
> Magic Wand (Thanks Theo!) to create selections that can be loaded into
> the ROI Manager and then measured. This works really well but requires
> that the cells be selected manually.
> >>
> >> The Cell Magic Wand Tool works on the colour or grayscale so we can
> also split the channels from the colour image if needed and use the
> channel image with the most contrast.
> >>
> >> I've attached an image in case anyone has any ideas. The image has
> >> been
> cropped out of a larger image so that it's not too big and it's pink
> because there's cell culture medium there (in case anyone was wondering..).
> >>
> >> Look forward to hearing any suggestions!
> >>
> >> Kind regards,
> >>
> >> Jacqui
> >> Jacqueline Ross
> >> Biomedical Imaging Microscopist
> >> Biomedical Imaging Research Unit
> >> School of Medical Sciences
> >> Faculty of Medical & Health Sciences The University of Auckland
> >> Private Bag 92019 Auckland 1142, NEW ZEALAND
> >>
> >> Tel: [phone]
> >> Fax: [phone]
> >>
> >> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >
> > --
> > Aryeh Weiss
> > Faculty of Engineering
> > Bar Ilan University
> > Ramat Gan 52900 Israel
> >
> > Ph:  972-3-5317638
> > FAX: [phone]
> >
> >
> > .
> >
>
>
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph:  972-3-5317638
> FAX: [phone]
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 09:07:27 -0500
> From:    Kenneth Sloan <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> I would consider an approach that is one level higher than raw image
> processing.  DIC images produce distinctive doublets (a bright region
> adjacent to a darker region - always (for the same setup) at the same angle.
>
> Once you can identify a cell from it’s doublet, and have a reasonable
> guess as to its size, finding the boundary and estimating area should
> become easier.
>
> Just a thought.
>
> My first try might be to design a custom convolution kernel that
> responds preferentially to doublets at a fixed angle.  There is still
> the problem of estimating the angle (if it’s not stable)  but that
> should be doable based either on meta-information or user input.
>
>
> Are all the cells as well-behaved as these?
>
> I second the idea of using a texture measurement to eliminate regions
> of the image that are out of focus (variance is one - I would perhaps
> go a step further and use coefficient of variation - stdDev/mean - but
> there are
> others)
>
> --
> Kenneth Sloan
> [hidden email] <javascript:;> Vision is the art of seeing
> what is invisible to others.
>
>
>
>
> > On May 1, 2015, at 08:02 , Aryeh Weiss <[hidden email]
> <javascript:;>> wrote:
> >
> > On 5/1/15 8:48 AM, Jacqui Ross wrote:
> >> Hi Aryeh,
> >>
> >> Thanks for your reply. I did try using a variance filter (the
> >> built-in
> one under Process - Filters - Variance) with different radii but I was
> unable to achieve a good result. The resultant circles were often
> incomplete so that when I then converted to binary, I had to do a lot
> of additional processing (Closing, filling holes, etc.) and then the
> outlines weren't very accurate.
> > Yes -- these methods are better at marking objects than getting
> > accurate
> boundaries. You might be able to use the inaccurate segmentation that
> produces as a mask against the original variance image, which produces
> reasonable arcs around your in-focus cells.
> >> I know that you presented on the Trainable Weka Segmentation at the
> ImageJ conference that I attended a couple of years ago. My notes on
> that weren't fantastic (I pulled them out!) but in your case you also
> had some fluorescence labelling to help inform the segmentation.
> > That problem was easier than yours (isn't it always that way?). The
> texture was much better defined, and I did not have a continuum of
> out-of-focus or partially out-of-focus cells. There was a DAPI channel
> which I used to exclude artitfacts which were not cells (since they
> did not contain nuclei). However, that will not help you, since your
> out of focus cells are still cells.
> >
> > You might have an easier time here if you can acquire a z-stack
> > (even
> though it is wide-field) and use the EDF plugin or similar to have all
> of your cells in-focus. That would be easier to segment.
> Alternatively, if you choose to be very strict in your segmentation
> (ie, take only the cells that are really sharp and textured), then I
> think you will be able to variance and it relatives to get an accurate segmentation and boundary.
> >
> > Best regards
> > --aryeh
> >
> >>
> >> -----Original Message-----
> >> From: Aryeh Weiss [mailto:[hidden email] <javascript:;>]
> >> On
> Behalf Of Aryeh Weiss
> >> Sent: Friday, 1 May 2015 5:07 p.m.
> >> To: Jacqui Ross
> >> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast
> >> images
> - help please
> >>
> >>
> >> Try converting to 8-bit and running a variance filter
> >> (Process>Filters>Variance...) followed by thresholding. This will
> enhance the cells in focus due to their texture.
> >>
> >> --aryeh
> >>
> >> On 5/1/15 7:37 AM, Jacqui Ross wrote:
> >>> Hi All,
> >>>
> >>> I'm helping a PhD student with analysing some Hoffman modulation
> contrast images of cells. She's primarily interested in changes in
> diameter. The cells are embedded in a 3D matrix and compression is
> being applied.
> >>>
> >>> In the images, there are nice cells in focus with clear
> >>> boundaries,
> plus others which are out of focus which we don't want to measure as
> any measurements won't be accurate.
> >>> These kind of images are really tricky to segment as anyone who
> >>> has
> tried, already knows. I've tried lots of different filters (edge,
> etc.) , FFT filtering and the Trainable Weka Segmentation but have
> been unable to achieve good enough results to be able to then
> threshold the cells automatically.
> >>>
> >>> I've come to the end of the line for now so am asking for your
> >>> expert
> help in case anyone has some suggestions:). I note that in 2006
> Monique Vasseur offered some DIC images to a PhD student called Daniel
> Mauch in Germany but I'm not sure if anything came of that project.
> There are a few papers out there (some mention Hilbert Transform, FFT)
> but I haven't been successful in implementing anything from those papers as yet.
> >>>
> >>> In the meantime, my solution is to use the Pseudo flat field
> correction plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!)
> with a very small radius (5) to flatten the background and out of
> focus cells while preserving the in focus cell outlines. We can then
> use the Cell Magic Wand (Thanks Theo!) to create selections that can
> be loaded into the ROI Manager and then measured. This works really
> well but requires that the cells be selected manually.
> >>>
> >>> The Cell Magic Wand Tool works on the colour or grayscale so we
> >>> can
> also split the channels from the colour image if needed and use the
> channel image with the most contrast.
> >>>
> >>> I've attached an image in case anyone has any ideas. The image has
> been cropped out of a larger image so that it's not too big and it's
> pink because there's cell culture medium there (in case anyone was wondering..).
> >>>
> >>> Look forward to hearing any suggestions!
> >>>
> >>> Kind regards,
> >>>
> >>> Jacqui
> >>> Jacqueline Ross
> >>> Biomedical Imaging Microscopist
> >>> Biomedical Imaging Research Unit
> >>> School of Medical Sciences
> >>> Faculty of Medical & Health Sciences The University of Auckland
> >>> Private Bag 92019 Auckland 1142, NEW ZEALAND
> >>>
> >>> Tel: [phone]
> >>> Fax: [phone]
> >>>
> >>> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>>
> >>>
> >>> --
> >>> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >> --
> >> Aryeh Weiss
> >> Faculty of Engineering
> >> Bar Ilan University
> >> Ramat Gan 52900 Israel
> >>
> >> Ph:  972-3-5317638
> >> FAX: [phone]
> >>
> >>
> >> .
> >>
> >
> >
> > --
> > Aryeh Weiss
> > Faculty of Engineering
> > Bar Ilan University
> > Ramat Gan 52900 Israel
> >
> > Ph:  972-3-5317638
> > FAX: [phone]
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 10:10:20 -0400
> From:    "JOEL B. SHEFFIELD" <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> Hi Jacqui,
>
> These images are particularly difficult to segment because the edges
> are assymetric --dark on one side, and light on the other.  I was able
> to get some enhancement by using an unsharp mask (on your image, pixel
> radius of 100, weight .60), followed by the "find edges" convolution.  
> It wasn't perfect, but might help.
>
> Joel
>
>
>
> Joel B. Sheffield, Ph.D
> Department of Biology
> Temple University
> Philadelphia, PA 19122
> Voice: 215 204 8839
> e-mail: [hidden email] <javascript:;>
> URL:  *http://tinyurl.com/khbouft <http://tinyurl.com/khbouft>*
>
> On Fri, May 1, 2015 at 12:37 AM, Jacqui Ross
> <[hidden email] <javascript:;>>
> wrote:
>
> > Hi All,
> >
> > I'm helping a PhD student with analysing some Hoffman modulation
> > contrast images of cells. She's primarily interested in changes in
> > diameter. The cells are embedded in a 3D matrix and compression is being applied.
> >
> > In the images, there are nice cells in focus with clear boundaries,
> > plus others which are out of focus which we don't want to measure as
> > any measurements won't be accurate.
> > These kind of images are really tricky to segment as anyone who has
> tried,
> > already knows. I've tried lots of different filters (edge, etc.) ,
> > FFT filtering and the Trainable Weka Segmentation but have been
> > unable to achieve good enough results to be able to then threshold
> > the cells automatically.
> >
> > I've come to the end of the line for now so am asking for your
> > expert
> help
> > in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur
> > offered some DIC images to a PhD student called Daniel Mauch in
> > Germany
> but
> > I'm not sure if anything came of that project. There are a few
> > papers out there (some mention Hilbert Transform, FFT) but I haven't
> > been successful in implementing anything from those papers as yet.
> >
> > In the meantime, my solution is to use the Pseudo flat field
> > correction plugin from Jan Brocher's BioVoxxel Toolbox (Thanks Jan!)
> > with a very
> small
> > radius (5) to flatten the background and out of focus cells while
> > preserving the in focus cell outlines. We can then use the Cell
> > Magic
> Wand
> > (Thanks Theo!) to create selections that can be loaded into the ROI
> Manager
> > and then measured. This works really well but requires that the
> > cells be selected manually.
> >
> > The Cell Magic Wand Tool works on the colour or grayscale so we can
> > also split the channels from the colour image if needed and use the
> > channel image with the most contrast.
> >
> > I've attached an image in case anyone has any ideas. The image has
> > been cropped out of a larger image so that it's not too big and it's
> > pink because there's cell culture medium there (in case anyone was
> wondering..).
> >
> > Look forward to hearing any suggestions!
> >
> > Kind regards,
> >
> > Jacqui
> > Jacqueline Ross
> > Biomedical Imaging Microscopist
> > Biomedical Imaging Research Unit
> > School of Medical Sciences
> > Faculty of Medical & Health Sciences The University of Auckland
> > Private Bag 92019 Auckland 1142, NEW ZEALAND
> >
> > Tel: [phone]
> > Fax: [phone]
> >
> > http://www.fmhs.auckland.ac.nz/sms/biru/
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 14:36:03 +0000
> From:    Kenneth R Sloan <[hidden email] <javascript:;>>
> Subject: Re: Segmentation of DIC or Hoffman Modulation Contrast images
> - help please
>
> Again - I’ll promote taking a higher level approach.  Simple
> SEGMENTATION should be able to pull out “Bright blobs” and “Dark
> blobs” - segmenting the CELLS requires combining that evidence (and
> then perhaps going back to the image data in “verification vision”
> mode - making predictions about what the cell boundary will look like,
> and localizing it.  But, this is probably beyond the scope of a script combining standard image processing operators.
>
> --
> Kenneth Sloan
> [hidden email] <javascript:;>
> Vision is the art of seeing what is invisible to others.
>
>
>
>
> > On May 1, 2015, at 09:10 , JOEL B. SHEFFIELD <[hidden email]
> <javascript:;>> wrote:
> >
> > Hi Jacqui,
> >
> > These images are particularly difficult to segment because the edges
> > are assymetric --dark on one side, and light on the other.  I was
> > able to get some enhancement by using an unsharp mask (on your
> > image, pixel radius of 100, weight .60), followed by the "find
> > edges" convolution.  It wasn't perfect, but might help.
> >
> > Joel
> >
> >
> >
> > Joel B. Sheffield, Ph.D
> > Department of Biology
> > Temple University
> > Philadelphia, PA 19122
> > Voice: 215 204 8839
> > e-mail: [hidden email] <javascript:;>
> > URL:  *http://tinyurl.com/khbouft <http://tinyurl.com/khbouft>*
> >
> > On Fri, May 1, 2015 at 12:37 AM, Jacqui Ross
> > <[hidden email]
> <javascript:;>>
> > wrote:
> >
> >> Hi All,
> >>
> >> I'm helping a PhD student with analysing some Hoffman modulation
> contrast
> >> images of cells. She's primarily interested in changes in diameter.
> >> The cells are embedded in a 3D matrix and compression is being applied.
> >>
> >> In the images, there are nice cells in focus with clear boundaries,
> >> plus others which are out of focus which we don't want to measure
> >> as any measurements won't be accurate.
> >> These kind of images are really tricky to segment as anyone who has
> tried,
> >> already knows. I've tried lots of different filters (edge, etc.) ,
> >> FFT filtering and the Trainable Weka Segmentation but have been
> >> unable to achieve good enough results to be able to then threshold
> >> the cells automatically.
> >>
> >> I've come to the end of the line for now so am asking for your
> >> expert
> help
> >> in case anyone has some suggestions:). I note that in 2006 Monique
> Vasseur
> >> offered some DIC images to a PhD student called Daniel Mauch in
> >> Germany
> but
> >> I'm not sure if anything came of that project. There are a few
> >> papers
> out
> >> there (some mention Hilbert Transform, FFT) but I haven't been
> successful
> >> in implementing anything from those papers as yet.
> >>
> >> In the meantime, my solution is to use the Pseudo flat field
> >> correction plugin from Jan Brocher's BioVoxxel Toolbox (Thanks
> >> Jan!) with a very
> small
> >> radius (5) to flatten the background and out of focus cells while
> >> preserving the in focus cell outlines. We can then use the Cell
> >> Magic
> Wand
> >> (Thanks Theo!) to create selections that can be loaded into the ROI
> Manager
> >> and then measured. This works really well but requires that the
> >> cells be selected manually.
> >>
> >> The Cell Magic Wand Tool works on the colour or grayscale so we can
> >> also split the channels from the colour image if needed and use the
> >> channel image with the most contrast.
> >>
> >> I've attached an image in case anyone has any ideas. The image has
> >> been cropped out of a larger image so that it's not too big and
> >> it's pink because there's cell culture medium there (in case anyone
> >> was
> wondering..).
> >>
> >> Look forward to hearing any suggestions!
> >>
> >> Kind regards,
> >>
> >> Jacqui
> >> Jacqueline Ross
> >> Biomedical Imaging Microscopist
> >> Biomedical Imaging Research Unit
> >> School of Medical Sciences
> >> Faculty of Medical & Health Sciences The University of Auckland
> >> Private Bag 92019 Auckland 1142, NEW ZEALAND
> >>
> >> Tel: [phone]
> >> Fax: [phone]
> >>
> >> http://www.fmhs.auckland.ac.nz/sms/biru/
> >>
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 19:15:40 +0000
> From:    "Paletzki, Ron (NIH/NIMH) [C]" <[hidden email]
> <javascript:;>>
> Subject: Problem using Image Sequence command from a macro
>
> I have a macro that creates a stack and then tries to save it to a folder
> using the Image Sequence command.   Problem is it works fine if the folder
> it’s saving to has no spaces in the name but fails if it does.   I’m
> wondering if this is a bug since it has never been an issue before in
> other macros using other save and open commands.  This is the first
> macro using the Image Sequence command.
>
> Here is an example macro that demonstrates the bug.  With a stack open
> if I run this macro and select a folder with a name that contains a
> space in it I get an error  “File Saving error (IOException):
>
> If I select a folder that does not contain a space in the name it
> works fine.
>
> macro "Save Stack  [f2]"{
>     dirOutput = getDirectory("Choose Output Directory for stack");
>     Dialog.create(" Macro Options ");
>     Dialog.addString("Enter Filename for stack ","fileName");
>     Dialog.show();
>     fileNameStack = Dialog.getString();
>     run("Image Sequence... ", "format=TIFF name="+fileNameStack+"
> digits=3 save="+dirOutput); }
>
>
> By the way this in running on a Mac using Image 149s8
>
> Thanks
> Ron
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 13:03:53 -0700
> From:    Glen MacDonald <[hidden email] <javascript:;>>
> Subject: Re: Problem using Image Sequence command from a macro
>
> Ron,
> Use brackets to denote string variables with spaces.
>
> macro "Save Stack  [f2]"{
>    dirOutput = getDirectory("Choose Output Directory for stack");
>    Dialog.create(" Macro Options ");
>    Dialog.addString("Enter Filename for stack ","fileName");
>    Dialog.show();
>    fileNameStack = Dialog.getString();
>    run("Image Sequence... ", "format=TIFF name=["+fileNameStack+"]
> digits=3 save=[dirOutput]");
> }
>
>
> Glen MacDonald
>         Core for Communication Research Virginia Merrill Bloedel
> Hearing Research Center
>         Cellular Morphology Core
> Center on Human Development and Disability Box 357923 University of
> Washington Seattle, WA 98195-7923  USA
> [phone]
> [hidden email] <javascript:;>
>
>
>
>
>
>
>
> On May 1, 2015, at 12:15 PM, Paletzki, Ron (NIH/NIMH) [C] <
> [hidden email] <javascript:;>> wrote:
>
> > I have a macro that creates a stack and then tries to save it to a
> folder using the Image Sequence command.   Problem is it works fine if the
> folder it’s saving to has no spaces in the name but fails if it does.   I’m
> wondering if this is a bug since it has never been an issue before in other
> macros using other save and open commands.  This is the first macro using
> the Image Sequence command.
> >
> > Here is an example macro that demonstrates the bug.  With a stack open
> if I run this macro and select a folder with a name that contains a space
> in it I get an error
> > “File Saving error (IOException):
> >
> > If I select a folder that does not contain a space in the name it works
> fine.
> >
> > macro "Save Stack  [f2]"{
> >    dirOutput = getDirectory("Choose Output Directory for stack");
> >    Dialog.create(" Macro Options ");
> >    Dialog.addString("Enter Filename for stack ","fileName");
> >    Dialog.show();
> >    fileNameStack = Dialog.getString();
> >    run("Image Sequence... ", "format=TIFF name="+fileNameStack+"
> digits=3 save="+dirOutput);
> > }
> >
> >
> > By the way this in running on a Mac using Image 149s8
> >
> > Thanks
> > Ron
> >
> >
> > --
> > ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 15:10:07 -0500
> From:    Mark Hiner <[hidden email] <javascript:;>>
> Subject: Re: Labels and default values for @parameter in scripts
>
> Hello,
>
> Just wanted you to know that if you update to the latest release[1] many of
> the issues you brought up have been resolved and improved.
>
> >You can't select the full input field by using a double click
>
> In my original reply I forgot to mention that I believe this is a Java-ism.
> The most reliable way to select the full text of an input field with the
> mouse is to triple click.
>
> Thanks again.
>
> Best,
> Mark
>
> [1] http://imagej.net/2015-05-01_-_ImageJ_2.0.0-rc-30
>
> On Thu, Apr 30, 2015 at 9:32 AM, Mark Hiner <[hidden email] <javascript:;>>
> wrote:
>
> > Hi Michael,
> >
> > It's great to see people using script parameters. These are a new feature
> > introduced with ImageJ2[1], and thus are still relatively new - so
> feedback
> > like this is both helpful and necessary in making these features robust.
> > For ImageJ2-specific features like this, you can also use the
> imagej-devel
> > mailing list[2] or file issues directly on GitHub[3].
> >
> > > Except this page I can't find any documentation on using script
> > parameters in ImageJ. Is there some more documentation, I was not able to
> > find?
> >
> > This is the intended documentation page. It's definitely sparse right
> now.
> > I am updating it in response to your feedback, but if you have any more
> > suggestions/requests for what would be helpful here, please let us know.
> >
> > > In plugins you can define labels and default values for parameters. In
> > scripts this is not possible.
> >
> > I had to dig around for this a bit, but it actually is possible! It uses
> > the same Java syntax as plugins, but requires the properties (like labels
> > and default values) to be in a parenthetical expression. Examples are now
> > on the wiki[4].
> >
> > >The default values are strange.
> >
> > Agreed. I added an issue[5] for this.
> >
> > > You can't select the full input field by using a double click.
> > > Using [Tab] to switch between fields works bad
> >
> > These are UI-specific[6] issues and so shouldn't be problems with
> > scripting itself. I was able to confirm this behavior on a mac, so I'll
> > look into it.
> >
> > Thank you for the comments!
> >
> > Best,
> > Mark
> >
> > [1] http://imagej.net/ImageJ2
> > [2] http://imagej.net/Mailing_lists
> > [3] http://imagej.net/Source_code#Source_code
> > [4] http://imagej.net/Script_parameters#Parameter_properties
> > [5] https://github.com/scijava/scijava-common/issues/160
> > [6] https://github.com/scijava/scijava-ui-swing
> >
> > On Thu, Apr 30, 2015 at 6:28 AM, Michael Entrup <
> > [hidden email] <javascript:;>> wrote:
> >
> >> Hi,
> >>
> >> using script parameters might be a nice solution to write small scripts
> >> that need user inputs. But there are some drawbacks that keep me away
> from
> >> using script parameters.
> >>
> >> In plugins you can define labels and default values for parameters. In
> >> scripts this is not possible.
> >>
> >> The default values are strange. When using @Integer the default value is
> >> '-2.147.483.648'.
> >>
> >> You can't select the full input field by using a double click. The minus
> >> sign is not selected, when double clicking the number.
> >>
> >> Using [Tab] to switch between fields works bad. Normally I expect that
> >> the content of the next input field is selected, but only the cursor is
> >> placed at the next field. [Ctrl]+[A] is necessary to replace the default
> >> value.
> >>
> >> The attached screen shot shows the dialog that is shown, when using 4
> >> different script parameters.
> >>
> >> Is all this intended, or is still work in progress?
> >> Except this page [1] I can't find any documentation on using script
> >> parameters in ImageJ. Is there some more documentation, I was not able
> to
> >> find?
> >>
> >> Best regards
> >> Michael
> >>
> >>
> >> [1] http://imagej.net/Script_parameters
> >>
> >> --
> >> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
> >>
> >
> >
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> Date:    Fri, 1 May 2015 19:37:59 -0400
> From:    Jason Miller <[hidden email] <javascript:;>>
> Subject: Re: Saving large files in ome.tif or ids/ics
>
> Hi all-
>
> I received an answer from the OME folks about my question and figured I'd
> post here to keep this in the same thread. This was helpful info for me.
>
> -------------------------------
> Dear Jason,
>
> > Similar to recent past posts on this list, I'm having trouble saving
> > files that are > 4GB into ome.tif or IDS/ICS formats. I have a 5GB file,
> > open as a virtual file, and when I go to export via Bioformats Exporter
> > to IDS/ICS format, I get the following error:
> [stacktrace]
>
> For IDS/ICS, this was fixed in this PR last week:
>    https://github.com/openmicroscopy/bioformats/pull/1738
> I have tested this with >5GB images and it works correctly.
>
> > When I go to export to ome.tif format, I get the same error previously
> > reported on this list:
> > loci.formats.FormatException: File is too large; call setBigTiff(true)
> >
> > Is it possible to retain the ome.tif format for >4GB? In my case, this
> > is for importing into Huygens for deconvolution.
>
> For plain (not OME) TIFF:
>    https://github.com/openmicroscopy/bioformats/pull/1744
>
> Both of the above fixes are in the newly-released Bio-Formats 5.1.1
> release.  If using Fiji, you should automatically get this update; for
> ImageJ you can download the updated plugin from
>    http://downloads.openmicroscopy.org/bio-formats/5.1.1/
>
> For OME-TIFF, it's not currently possible to enable BigTIFF when
> exporting from ImageJ using the exporter dialogue.  For the plain TIFF
> case, rather than relying on an API call to enable BigTIFF, we now
> enable it if the image data is >4GiB or a "big" TIFF file extension such
> as .btf, .tf8 or .tf2 is used.  We would like to do the same for
> OME-TIFF, but this first requires updating the OME-TIFF specification
> which currently only allows a .ome.tiff or .ome.tif file extension.
> Once that's done, you would be able to use e.g. ".ome.btf" and get a
> "big" OME-TIFF file.  However, this does require some consideration
> since (for example) these files would not be readable with older
> versions of Bio-Formats unless you renamed them to ".ome.tiff" after
> export.
>
> To summarise, it isn't currently possible to export >4GB ome.tiff files
> using the Bio-Formats Exporter.  It is possible via the Java API by
> calling writer.setBigTiff(true) if this is an option for you.  This is a
> known limitation which we are working on; see the discussion in the PR
> #1744 above.
>
> There is a ticket open here for this:
>    https://trac.openmicroscopy.org/ome/ticket/12858
> I can add you as a CC on it if you like, so you'll be kept up to date
> with its progress?
>
>
> Kind regards,
> Roger
>
> --
> Dr Roger Leigh -- Open Microscopy Environment
> Wellcome Trust Centre for Gene Regulation and Expression,
> College of Life Sciences, University of Dundee, Dow Street,
> Dundee DD1 5EH Scotland UK   Tel: [phone]
>
> The University of Dundee is a registered Scottish Charity, No: SC015096
>
> On Thu, Apr 30, 2015 at 5:28 PM, Jason Miller <[hidden email]
> <javascript:;>>
> wrote:
>
> > Hi all-
> >
> > This may be a Bioformats issue as well, so I will separately e-mail the
> > OME folks, but similar to recent past posts on this list, I'm having
> > trouble saving files that are > 4GB into ome.tif or IDS/ICS formats. I
> have
> > a 5GB file, open as a virtual file, and when I go to export via
> Bioformats
> > Exporter to IDS/ICS format, I get the following error:
> >
> > (Fiji Is Just) ImageJ 2.0.0-rc-15/1.49p; Java 1.6.0_24 [64-bit]; Windows
> 7
> > 6.1; 98MB of 12209MB (<1%)
> >
> > java.lang.IllegalArgumentException: Negative position
> > at sun.nio.ch.FileChannelImpl.read(FileChannelImpl.java:600)
> > at
> >
> loci.common.NIOByteBufferProvider.allocateDirect(NIOByteBufferProvider.java:131)
> > at
> >
> loci.common.NIOByteBufferProvider.allocate(NIOByteBufferProvider.java:116)
> > at loci.common.NIOFileHandle.buffer(NIOFileHandle.java:551)
> > at loci.common.NIOFileHandle.seek(NIOFileHandle.java:273)
> > at
> >
> loci.common.RandomAccessOutputStream.seek(RandomAccessOutputStream.java:79)
> > at loci.formats.out.ICSWriter.saveBytes(ICSWriter.java:139)
> > at loci.formats.FormatWriter.saveBytes(FormatWriter.java:126)
> > at loci.plugins.out.Exporter.run(Exporter.java:629)
> > at loci.plugins.LociExporter.run(LociExporter.java:77)
> > at
> >
> ij.plugin.filter.PlugInFilterRunner.processOneImage(PlugInFilterRunner.java:263)
> > at
> ij.plugin.filter.PlugInFilterRunner.<init>(PlugInFilterRunner.java:112)
> > at ij.IJ.runUserPlugIn(IJ.java:201)
> > at ij.IJ.runPlugIn(IJ.java:163)
> > at ij.Executer.runCommand(Executer.java:131)
> > at ij.Executer.run(Executer.java:64)
> > at java.lang.Thread.run(Thread.java:662)
> >
> >
> > When I go to export to ome.tif format, I get the same error previously
> > reported on this list:
> > loci.formats.FormatException: File is too large; call setBigTiff(true)
> >
> > Is it possible to retain the ome.tif format for >4GB? In my case, this is
> > for importing into Huygens for deconvolution.
> >
> > Thanks so very much
> > -Jason
> > --
> >
> > Jason Miller, MD, PhD
> >
> > University of Michigan Kellogg Eye Center
> >
> > --
> >
> > Home address:
> >
> > 117 Worden Ave
> >
> > Ann Arbor, MI 48103
> >
> > Cell: [phone]
> >
> > E-mail: *[hidden email] <javascript:;>
> > <[hidden email] <javascript:;>>*
> >
> >
> > "When I was 5 years old, my mother always told me that happiness was the
> > key to life. When I went to school, they asked me what I wanted to be
> when
> > I grew up. I wrote down, 'Happy.' They told me I didn't understand the
> > assignment. I told them they didn't understand life." - John Lennon
> >
>
>
>
> --
>
> Jason Miller, MD, PhD
>
> University of Michigan Kellogg Eye Center
>
> --
>
> Home address:
>
> 117 Worden Ave
>
> Ann Arbor, MI 48103
>
> Cell: [phone]
>
> E-mail: *[hidden email] <javascript:;> <
> [hidden email] <javascript:;>>*
>
>
> "When I was 5 years old, my mother always told me that happiness was the
> key to life. When I went to school, they asked me what I wanted to be when
> I grew up. I wrote down, 'Happy.' They told me I didn't understand the
> assignment. I told them they didn't understand life." - John Lennon
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>
> ------------------------------
>
> End of IMAGEJ Digest - 30 Apr 2015 to 1 May 2015 (#2015-141)
> ************************************************************
>

--
-------------------------
Chaitanya Athale,
Pune, India
18° 31' N, 73° 55' E, 560m.
-------------------------

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