Screen thresholded objects for in/out of focus

[TimFeinstein]

Hello,

I have a field of non-touching fluorescent objects that are segmented by thresholding, and I want to discard objects which are out of focus.  In-focus objects (cilia) should have a clear and straight edge that an algorithm like Modulation Transfer Function should be able to detect.  However, MTF plugins are overkill for my needs and they don't seem like they'd be easy to incorporate into a macro script.  I need a single number that represents the sharp-ness or focused-ness of each cilium, so that a macro can automatically discard fuzzy cilia before measuring them.  I like, for example, the Gaussian gradient function in Matlab, but I'm not sure something similar is available in ImageJ.  

Any suggestions would be appreciated.  Thanks!


TF

Timothy Feinstein, Ph.D.
Research Scientist
Department of Developmental Biology
University of Pittsburgh

 



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[Aryeh Weiss]

On 06/08/2018 22:54, Feinstein, Timothy N wrote:
> Hello,
>
> I have a field of non-touching fluorescent objects that are segmented by thresholding, and I want to discard objects which are out of focus.  In-focus objects (cilia) should have a clear and straight edge that an algorithm like Modulation Transfer Function should be able to detect.  However, MTF plugins are overkill for my needs and they don't seem like they'd be easy to incorporate into a macro script.  I need a single number that represents the sharp-ness or focused-ness of each cilium, so that a macro can automatically discard fuzzy cilia before measuring them.  I like, for example, the Gaussian gradient function in Matlab, but I'm not sure something similar is available in ImageJ.
>
> Any suggestions would be appreciated.  Thanks!

Try any of the variance o other edge detection operators (eg,
Processing>Filteres... or Processing> Find Edges) and tehn .threshold on
the result.

--aryeh

>
> TF
>
> Timothy Feinstein, Ph.D.
> Research Scientist
> Department of Developmental Biology
> University of Pittsburgh
>
>  
>
>
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html

--
Aryeh Weiss
Faculty of Engineering
Bar Ilan University
Ramat Gan 52900 Israel

Ph:  972-3-5317638
FAX: [phone]

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[Herbie]

Good day Timothy,

absolute focus judgments are difficult, relative judgments* are rather
easy to accomplish.

*If you have a focus series you can use LaPlace filtering followed by
power/energy evaluation.

In your case, the problem is that relative judgments appear limited to
in-image comparisons but the cells will most likely have different shape
etc.

I doubt that there is a simple *and* reliable way to do what you want,
except the un-sharp cells are strongly out of focus.

Regards

Herbie

::::::::::::::::::::::::::::::::::::::::::::::::::
Am 06.08.18 um 21:54 schrieb Feinstein, Timothy N:

> Hello,
>
> I have a field of non-touching fluorescent objects that are segmented by thresholding, and I want to discard objects which are out of focus.  In-focus objects (cilia) should have a clear and straight edge that an algorithm like Modulation Transfer Function should be able to detect.  However, MTF plugins are overkill for my needs and they don't seem like they'd be easy to incorporate into a macro script.  I need a single number that represents the sharp-ness or focused-ness of each cilium, so that a macro can automatically discard fuzzy cilia before measuring them.  I like, for example, the Gaussian gradient function in Matlab, but I'm not sure something similar is available in ImageJ.
>
> Any suggestions would be appreciated.  Thanks!
>
>
> TF
>
> Timothy Feinstein, Ph.D.
> Research Scientist
> Department of Developmental Biology
> University of Pittsburgh

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ImageJ mailing list: http://imagej.nih.gov/ij/list.html