> Leila,
>
> Try something like this, and remove all the other 'setThreshold',
> runThreshold, and setAutothreshold lines.
> setAutoThreshold("Huang"); // or other method
> run("Convert to Mask"); // display white 0 on black 255
> //run("Threshold...");
> //setAutoThreshold("Default");
> //setThreshold(u0, 65535);
> You may need to try setAutoThreshold("Huang dark"); or add run("Invert");
> depending on your images.
>
> At the end of your macro, do you get what you need with just two lines?
> Areas of each particle.
> // id = getImageID();
> // selectImage(id+1);
> sourceTitle = getTitle();
> // selectImage(id);
> run("Set Measurements...", "area mean min display decimal=3");
> // run("Set Measurements...", "area mean min display
> redirect=["+sourceTitle+ "] decimal=3");
> run("Analyze Particles...", "size=90-350 pixel circularity=0.00-1.00
> show=Outlines display");
> // selectImage(id+2);
> // sourceTitle = getTitle();
> // selectImage(id);
> // run("Set Measurements...", "area mean min display
> redirect=["+sourceTitle+ "] decimal=3");
> // run("Analyze Particles...", "size=90-350 pixel circularity=0.00-1.00
> show=Outlines display include");
>
> Or if you want total area of all the particle masks
> run("Analyze Particles...", "size=90-350 circularity=0.00-1.00 show=Masks
> display");
> run("Measure");
>
> Charles
>
> -----Original Message-----
> From: ImageJ Interest Group [mailto:
[hidden email]] On Behalf Of
> Leila Alieh
> Sent: Saturday, October 12, 2013 4:28 AM
> To:
[hidden email]
> Subject: Re: Threshold problems
>
> Hello Curtis,
>
> thank you very much for your help!
> I studied the link you suggested me but I still have some problem. When I
> run the Auto Threshold I get an image with colour satured particles (the
> cells I'm interested in) on a black background (or on the original image in
> the montage when I run the "Try all" option), I don't get a black on white
> mask as I was used to.
> But the main thing is that when I select a specific method and then I
> select "Analyze particles" and I choose and appropriate size range I get an
> error message:
>
> "No particles were detected. The assumed thredhold (0-0) may not be
> correct"
>
> If I don't choose the size range it counts something near the image
> borders but not he selected particles (I checked with the outlines).
> I tried also to select the algorithm in normal Threshold. In this case it
> works, I get the mask and it measures the selected particles, but the
> thresold value is different than the one obtained with the same algorithm
> in the Auto Threshold method, is it normal?
> Once I have chosen the algorithm can I include it in the macro and run it
> automatically? How should I do?
> Then I will also need to measure the entire area of my cut images. Does
> someone already developed a macro for this? if not do you have any
> suggestion?
>
> Thank you very much!!!
>
>
>
> 2013/10/12 Leila Alieh <
[hidden email]>
>
> > Hello Curtis,
> >
> > thank you very much for your help!
> > I studied the link you suggested me but I still have some problem.
> > When I run the Auto Threshold I get an image with colour satured
> > particles (the cells I'm interested in) on a black background (or on
> > the original image in the montage when I run the "Try all" option), I
> > don't get a black on white mask as I was used to.
> > But the main thing is that when I select a specific method and then I
> > select "Analyze particles" and I choose and appropriate size range I
> > get an error message:
> >
> > "No particles were detected. The assumed thredhold (0-0) may not be
> > correct"
> >
> > If I don't choose the size range it counts something near the image
> > borders but not he selected particles (I checked with the outlines).
> > I tried also to select the algorithm in normal Threshold. In this case
> > it works, I get the mask and it measures the selected particles, but
> > the thresold value is different than the one obtained with the same
> > algorithm in the Auto Threshold method, is it normal?
> > Once I have chosen the algorithm can I include it in the macro and run
> > it automatically? How should I do?
> > Here the macro I'm using:
> >
> > origen = getDirectory("Images to process"); lista =
> > getFileList(origen);
> > u0 = getNumber("threshold", u0);
> > setBatchMode(true);
> > for (i=0; i<lista.length; i++) {
> > showProgress(i+1, lista.length);
> > open(origen+lista[i]);
> > nombre = lista[i];
> > run("Split Channels");
> > run("Duplicate...", "title=[]");
> > run("Subtract Background...", "rolling=50");
> > run("Gaussian Blur...", "sigma=1");
> > setAutoThreshold("Default");
> > //run("Threshold...");
> > setAutoThreshold("Default");
> > setThreshold(u0, 65535);
> > run("Convert to Mask");
> > run("Close-");
> > run("Fill Holes");
> > id = getImageID();
> > selectImage(id+1);
> > sourceTitle = getTitle();
> > selectImage(id);
> > run("Set Measurements...", "area mean min display redirect=["+
> > sourceTitle+ "] decimal=3");
> > run("Analyze Particles...", "size=90-350 pixel
> > circularity=0.00-1.00 show=Outlines display");
> > selectImage(id+2);
> > sourceTitle = getTitle();
> > selectImage(id);
> > run("Set Measurements...", "area mean min display redirect=["+
> > sourceTitle+ "] decimal=3");
> > run("Analyze Particles...", "size=90-350 pixel
> > circularity=0.00-1.00 show=Outlines display include");
> > }
> >
> >
> > Then I will also need to measure the entire area of my modified images.
> > Does someone already developed a macro for this? if not do you have
> > any suggestion?
> >
> > Thank you very much!!!
> >
> >
> > 2013/10/9 Curtis Rueden <
[hidden email]>
> >
> >> Hi Leila,
> >>
> >> > What do you mean with the auto-threshold algorithm?
> >>
> >> An auto-thresholding algorithm chooses a (hopefully) appropriate
> >> threshold by analyzing the histogram of your data and guessing at a
> >> good cutoff point.
> >>
> >> See the docs at:
> >>
http://imagej.net/docs/guide/146-28.html#toc-Subsubsection-28.2.4> >>
> >> You can also use Fiji's "Image > Adjust > Auto Threshold" plugin with
> >> method of "Try all" to create a mosaic of the mask produced by each
> >> auto-thresholding algorithm, which makes the results very easy to
> >> compare quickly.
> >>
> >> Regards,
> >> Curtis
> >>
> >>
> >> On Wed, Oct 9, 2013 at 5:02 AM, Leila Alieh <
[hidden email]>
> >> wrote:
> >>
> >> > Hi Curtis!!!
> >> > Thank you for the reply!
> >> > I tried to set the threshold without the macro, to see how reliable
> >> > it
> >> is.
> >> > There is a huge difference in intensity across the images...I
> >> > really
> >> can't
> >> > choose a good threshold for all of them. What do you mean with the
> >> > auto-threshold algorithm?
> >> >
> >> >
> >> > 2013/10/8 Curtis Rueden <
[hidden email]>
> >> >
> >> > > Hi Leila,
> >> > >
> >> > > Are you sure ImageJ isn't autoscaling your display differently
> >> > > across
> >> > your
> >> > > images?
> >> > >
> >> > > What does the pixel probe say? Mouse over your data and look at
> >> > > the
> >> raw
> >> > > numbers spit out in the main ImageJ window's status bar.
> >> > >
> >> > > Also, can you use an auto-threshold algorithm across all your
> >> > > images
> >> to
> >> > > make your analysis reproducible without needing to hardcode a
> >> > > specific threshold value?
> >> > >
> >> > > Regards,
> >> > > Curtis
> >> > >
> >> > >
> >> > > On Tue, Oct 8, 2013 at 1:56 PM, Leila Alieh
> >> > > <
[hidden email]>
> >> > wrote:
> >> > >
> >> > > > Hello!
> >> > > >
> >> > > > I developed a macro to count the number of cells positives for
> >> > > > a
> >> given
> >> > > > antibody. The cells are selected depending on size, circularity
> >> > > > and intensity. I found an intensity value good for some images
> >> > > > and I run
> >> > the
> >> > > > macro. After that I found that the choosen threshold is too
> >> > > > high for
> >> > one
> >> > > > image, so that to have a reliable valuation for that image I
> >> > > > should
> >> use
> >> > > > half of the threshold chosen for the other images, but this
> >> > > > latter threshold is really too low for the other images and it
> >> > > > gives a
> >> wrong
> >> > > > result. I can't understand why this image is so different
> >> > > > because
> >> the
> >> > > > intensity of the cells is not so different by eyes (it's a good
> >> quality
> >> > > > image, the ab signal is strong and there are no shadows). I was
> >> > wondering
> >> > > > if I did some mistake during the "editing": I had to cut the
> >> > > > images
> >> > (I'm
> >> > > > interested only in a small area of them) and save them in a
> >> > > > tiff
> >> > format.
> >> > > Is
> >> > > > it possible that this editing chenged the intensity values?
> >> > > > Thank you for your help!!!!
> >> > > >
> >> > > > Leila
> >> > > >
> >> > > > --
> >> > > > 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> >> >
> >>
> >> --
> >> ImageJ mailing list:
http://imagej.nih.gov/ij/list.html> >>
> >
> >
>
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