I studied the link you suggested me but I still have some problem. When I
"No particles were detected. The assumed thredhold (0-0) may not be correct"
I tried also to select the algorithm in normal Threshold. In this case it
Then I will also need to measure the entire area of my cut images. Does
> 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>>
>
>