Re: IMAGEJ Digest - 3 Jan 2011 to 4 Jan 2011 (#2011-4)

[Daniel James White]

Hi Scott,


On Jan 5, 2011, at 6:00 AM, IMAGEJ automatic digest system wrote:

>
> Date:    Tue, 4 Jan 2011 10:41:01 -0800
> From:    schamber789 <[hidden email]>
> Subject: Counting empty circles
>
> Hello,
>
> I have images (see below) that have many glandular trichomes (the round
> shiny objects) which I would like to quantify. I simply want to know how
> many of them there are.  It seems like I should be able to ask ImageJ to
> count the number of these glandular trichomes, but I don't know how to do
> this. I have tried many things already, but have hit a wall. The smaller
> circles within each gland seem to be very consistent in size. I wonder if
> this consistency in size can be used somehow. Thanks so much!
>

Our eyes and brains are very good at recognizing patterns,
even in images like this where contrast is low,
the edges of the interesting blobs are not sharp but fuzzy
and there is lots of other interfering stuff in the image.... that makes it hard for a simple method to work.  

Since it would be easy to manually count these blobs in this image,
I assume you want to count the blobs in many many images,
automagically...

We face here, the classic problem of segmentation,
and its a whole branch of science in itself,
and the method that works is nearly always a customized solution for a particular case.

Looking at this image, I can not tell if it is transmitted light,
or a multi channel color merge image from fluorescence....

splitting the image into its RGB components doe not make the image any clearer really...
but suggests its a transmitted or reflected white light light colour image?

Can you tell us some details of the sample, staining if any
and the microscopy setup (camera, objective lens, etc. )

The best approach here is to use a specific label for the structures of interest
either a fluorescent dye attached to an antibody, or even a GFP genetic label,
or a visible dye

(You might also try dark field illumination to get a cleaner image).

Using specific fluorescence labeling only the structure you want is lit up,
and its them much easier for the computer to count them using a very simple method
as seen in the segmentation part of this teaching material - blobs segmentation and analysis by threshold and analyse particles imageJ functions:

http://ifn.mpi-cbg.de/wiki/ifn/images/8/8d/2010-12-Basics_of_Imaging_Processing_Course.pdf

It is possible to get the structure you want from this image using a clever method.... but one would have to take time to figure that out.

....much better is to start with an image where only the interesting structures give a signal.

cheers

Dan



> Sincerely,
>
> Scott Chamberlain
> Rice University, EEB Dept.
>
>
> http://imagej.588099.n2.nabble.com/file/n5889708/2309-1.jpg 

Dr. Daniel James White BSc. (Hons.) PhD
Senior Microscopist / Image Visualisation, Processing and Analysis
Light Microscopy and Image Processing Facilities
Max Planck Institute of Molecular Cell Biology and Genetics
Pfotenhauerstrasse 108
01307 DRESDEN
Germany

[phone] (German Mobile)
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