Posted by
daschneider9 on
URL: http://imagej.273.s1.nabble.com/Re-Counting-particle-fluorescence-labelled-using-ImageJ-tp3688447p3688454.html
Sindy,
Perhaps the use of a Gaussian blur filter might help. When I change your image to 16-bit, apply Gaussian blur filter with Sigma = 1.0, and apply auto default threshold for a dark background, then particle analyzer detects 615 particles. Using the generated ROIs to map these particles back to the original image looks like a good _first_ approximation. Perhaps further tweaking the Gaussian blur Sigma value, using watershed, and setting minimum size cutoffs might return values closer to your eyeball count.
In the end you might still need to "filter" the results in order to extract information only on those shape characteristics that you consider valid for actual clusters. For example, the results on the 615 include 350 with an "area" between 1-15 (I used the histogram function on the results window to get a feel for the particles detected). Perhaps you'd say these do not meet your threshold for inclusion in the counts (validated of course by your own manual counts). With this example of 615, you could re-run the particle analyzer and set "size" to exclude particles below 15. Now the analyzer detects 285 with minimum area of 15.
I have no idea if this is correct for your purposes but I hope it helps demonstrate some possibilities you can consider. I hope others might comment on this suggestion as I am also very new to this and eager to learn.
Good luck.
Dave
-----Original Message-----
From: ImageJ Interest Group [mailto:
[hidden email]] On Behalf Of Sindy Kueh
Sent: Monday, February 01, 2010 9:26 PM
To:
[hidden email]
Subject: Re: Counting particle (fluorescence labelled) using ImageJ
Hi all
I am a newbie with using imageJ and am trying to count the number of
clusters and also to measure the cluster size of GABAA receptor a1
subunit (fluorescence labelled). I have tried looking up on the ImageJ
homepage and could not find or do not know where to look for information
on how to correctly set threshold etc on counting these
clusters/particles. I have downloaded a 2 page pdf file which explains
how to count particle etc, but have difficulty locating the file-embryos
in which is needed to do the "tutorial".
I read that in order to perform the analysis, I have to change the file
type to either 16-bit or 8 bit-gray scale, which I did... however, I had
difficulty trying to set the threshold and have returned results in the
order of thousands of clusters where only a few hundreds are present. I
have attached an example of a file of what I need to measure... the
clusters are labelled green.
I hope someone can help.
Many thanks
Best regards
Sindy
<<Example1.JPG>>