Posted by
Ed Simmons on
May 25, 2014; 6:12pm
URL: http://imagej.273.s1.nabble.com/How-can-we-Distinguish-In-Out-of-focus-cells-in-confocal-microscope-tp5007911p5007916.html
On 25/05/14 18:00, Mohamed Tleis wrote:
> Dear Joel,
>
> Thank you for your reply, My Cells are simple yeast cells. I do Image
> Analysis for some images acquired by Biologists. they try to change
> the focal level to the most optimal one then they acquire their
> images. However, In the analysis part we need to be aware of out of
> focus cells, because they can wrongly affect the final measurement
> results. So my question is: is there any means to deal with such issues?
>
> Best Regards,
>
> M. Tleis
> Phd. Candidate
> LIACS, Leiden University
> The Netherlands.
>
>
>
Hi Mohammed,
Typically to deal with this sort of go/no-go situation, if your images
have good contrast, I would try to score the focus of each cell based on
it's rate of intensity change in an area with good contrast (eg edges or
details in the cell).
To get an idea for what I mean, find examples of well focused and poorly
focused cells and draw a line selection through the cell, beginning and
ending off the cell either side. Then plot a line profile (Ctrl+K I
think) and study the 'shoulders' of the curve, the well focused cell
should show much steeper rises at the edges than the poorly focused cell.
It is possible to make use of this method automatically by writing a
plugin/macro to suit your needs.
Choosing the largest difference between two adjacent pixels on the edges
of the curve as your sharpness score is often sufficient.
Setting some threshold for this value gives an easy indication of
go/no-go for focus dependent measurements.
I hope that helps with the issue, please let me know if you need further
assistance...
Best regards,
Ed
--
[hidden email]
www.esimagingsolutions.com
--
ImageJ mailing list:
http://imagej.nih.gov/ij/list.html