Posted by
Stoyan Pavlov on
Mar 13, 2013; 6:57am
URL: http://imagej.273.s1.nabble.com/quantification-of-fluorescent-signal-per-a-cell-tp5002137p5002140.html
Hi Lee,
I had similar problem several years ago, so i think i may be able to give
you some advice.
I had also to run measurements inside multiple cells (RoIs) so i had to
develop my own macro. It is crude but it did the job for me so if you can
you might try and modify it to your needs. This macro is run after your
images are segmented, the RoIs are generated and loaded in the RoIManager.
// This Macro counts particles and measures particle covered area inside
specific Roi and otputs the numbers //to TAB separated Log window which
after saving can be imported to excel.
print ("RoiNumber"," ","SliceLabel"," ","RoIName"," ","Count","
","RoIArea"," ","RoIPerimeter"," "," ParticlesCoveredArea","
","AreaFraction") ;
setAutoThreshold("Default dark");
n = roiManager("count");
for (i=0; i<n; i++) {
roiManager("select", i);
run("Set Measurements...", "area perimeter redirect=None
decimal=4");
run("Measure");
r=getResult("Area");
u=getResult("Perim.");
run("Set Measurements...", "area area_fraction limit display redirect=None
decimal=4");
run("Measure");
s=getResult("Area");
t=getResult("%Area");
run("Analyze Particles...", "size=0-20 circularity=0.00-1.00 show=Nothing
display clear ");
m=nResults;
q=getInfo("slice.label");
print (i," ",q," ",call("ij.plugin.frame.RoiManager.getName",
i)," ",m," ",r," ",u," ",s," ",t) ;
}
As to how to segment your images there are many plugins and explanations,
depending on your needs and goals. One of the easiest and quickest is to
simply threshold them. As it wan't work perfectly for your images you need
to try different segmentation strategies and see which one is the best for
you. For example there is a plugin for FIJI that does something similar to
what you were asking for - Trainable Segmentation. It can be found here
http://fiji.sc/Trainable_Segmentation_PluginDon't worry once you choose your method it can easily be automated using
the macro recording functions of ImageJ (or Fiji). For example there is one
very nice macro that lets you process entire folders and it can be found
here (
http://rsb.info.nih.gov/ij/macros/BatchProcessFolders.txt).
Hope this can help you a little bit. If you have further questions ask them
as there are many people here that can help you much more than me.
2013/3/13 Junsung LEE <
[hidden email]>
> I want to quantify fluorescent signal (punctiform, like cellular vesicles)
> in one cell, but I don't know how. I have tried a few things as follows
> (but all of them were not successful) :
>
>
> 1. first, I selected one cell with 'polygon selections tool', and tried to
> analyse fluorescent signal with 'RGB profile (Plugins>Graphics>RGB
> profile)'. But I failed because 'RGB profile' is working only with 'line or
> rectangular selections tool'. I can apply 'rectangular selections tool' in
> some cases, but when cells are overlapped partially, it is not applicable.
>
> > is there any ways to make 'RGB profile' work with 'polygon selections
> tool' or 'freehand selections tool'?
>
>
> 2. second, I marked fluorescent signals (they are dot-shaped, as I said
> above) with 'multi-point selections tool' one by one, and analysed them
> with 'measure (analyse>measure (ctrl+M)'. This method is working but
> marking many fluorescent dots one by one is really hard.
>
> > I really wonder how to select fluorescent signals (dots) automatically.
> Once I designate one dot as signal, it is possible to select dots
> automatically, I think, but I don't know how.
>
>
> I will be appreciated if you tell me how. If appropriate method is exist
> except what I said above, please don't hesitate to let me know. Please
> refer to attached image file.
>
> Thanks in advance.
>
>
> --
> ImageJ mailing list:
http://imagej.nih.gov/ij/list.html>
--
Dr. Stoyan P. Pavlov, MD, PhD
Departament of Anatomy, Histology and Embryology
Medical University "Prof. Dr. Paraskev Stoyanov", Varna
Prof. Marin Drinov Str.55
9002 Varna
Bulgaria
Tel: +359 (0) 52 - 677 - 086
e-mail:
[hidden email]
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