Intensity measurements by Gaussian peak fitting

[WantobeImageAnalyst]

I have a spinning disk confocal image which has a lot of noise and
particle(my interest). I need the only total intensity of spots in 2D and 3D
images using ImageJ or Matlab in diffraction-limited images. My particles
are slowly moving.

I'm using Track mate to detect particle and get their coordinates.
I want to apply 2D Gaussian fitting to particles which have a different
offset of noise.  Then, I would measure the intensity of each particle.
What and How can I do for that?
<http://imagej.1557.x6.nabble.com/file/t382308/sample0505.jpg> 



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[Herbie]

Good day,

this is a cross-post from the Forum.

The problem with your image is that it is so noisy that I can't decide
by eye and without any previous knowledge of how your particles look
like and what is signal (particle) and what is noise.

Furthermore, the sample image is spatially badly resolved and I doubt
that they are really diffraction-limited.

Finally, be aware of the fact that by filtering such noisy images you
can get nearly every signal or shape you want, depending on the type of
filter you apply.

Careful Fourier-spectral analysis of the sample image shows that the
particles in question are suspected to show significant spectral
components up to about 27 percent of the Nyquist frequency. With this
limit I conclude that a Gauss-filter of sigma 1.4 pixel may serve as a
starting point for image filtering the proposed by you.

Please find the filtered image attached to this message.

Regardless what approach you use, you will not get significantly better
results.

Good luck

Herbie

:::::::::::::::::::::::::::::::::::::::::::::::::::::::::::
Am 05.05.19 um 14:04 schrieb WantobeImageAnalyst:

> I have a spinning disk confocal image which has a lot of noise and
> particle(my interest). I need the only total intensity of spots in 2D and 3D
> images using ImageJ or Matlab in diffraction-limited images. My particles
> are slowly moving.
>
> I want to apply 2D Gaussian fitting to particles which have a different
> offset of noise. Then, I would measure the intensity of each particle.
> What and How can I do for that?
>
> <http://imagej.1557.x6.nabble.com/file/t382308/sample0505.jpg>
>
>
>
> --
> Sent from: http://imagej.1557.x6.nabble.com/
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>

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[WantobeImageAnalyst]

Thank you, Herbie.

I used 'Track mate' to detect my particles like the attached image based on
blob size and intensity threshold. I have coordinates of their centroid. And
I want to apply 2D and 3D Gaussian fitting of different shapes according to
each particle to discard noise.(Then, it will provide total or mean
intensity of the region of interest(?))

P.S.I cannot upload the whole image(3D, timelapse) because it exceeded the
file size limit. If you let me share the image through Google drive, I'll do
that for a better understanding.

<http://imagej.1557.x6.nabble.com/file/t382308/sample.jpg> of these
particles.



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[Herbie]

As I've written,

I doubt that you will get reliable results from such images.

Please post the result from trackmate and explain exactly how you've
set-up this plugin.

You don't reply at all to my suggestions and results. Why?

What about your claim that the images are diffraction-limited. How do
you test this?

What about the poor spatial resolution of your images?

What about previous knowledge of size and shape of your particles?

"WantobeImageAnalyst"
Are you serious?

Regards

Herbie

:::::::::::::::::::::::::::::::::::::::::::::::::
Am 05.05.19 um 18:48 schrieb WantobeImageAnalyst:

> Thank you, Herbie.
>
> I used 'Track mate' to detect my particles like the attached image based on
> blob size and intensity threshold. I have coordinates of their centroid. And
> I want to apply 2D and 3D Gaussian fitting of different shapes according to
> each particle to discard noise.(Then, it will provide total or mean
> intensity of the region of interest(?))
>
> P.S.I cannot upload the whole image(3D, timelapse) because it exceeded the
> file size limit. If you let me share the image through Google drive, I'll do
> that for a better understanding.
>
> <http://imagej.1557.x6.nabble.com/file/t382308/sample.jpg> of these
> particles.
>
>
>
> --
> Sent from: http://imagej.1557.x6.nabble.com/
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>

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[Arne Seitz]

Dear Herbie,

I agree with the content of your replies but I think the tone is unappropriated.
Getting personally insulting shall to my understanding be avoided in such a forum.

Best regards
Arne


---------------------------------------------------------------
Dr. Arne Seitz
Head of Bioimaging and Optics Platform (BIOP)
Swiss Institute of Technology (EPFL)
Faculty of Life Sciences
Station 15
CH-1015 Lausanne

Phone: [phone]
Fax:      [phone]
http://biop.epfl.ch/
---------------------------------------------------------------



 

-----Original Message-----
From: ImageJ Interest Group <[hidden email]> On Behalf Of Herbie
Sent: Sonntag, 5. Mai 2019 19:04
To: [hidden email]
Subject: Re: Intensity measurements by Gaussian peak fitting

As I've written,

I doubt that you will get reliable results from such images.

Please post the result from trackmate and explain exactly how you've set-up this plugin.

You don't reply at all to my suggestions and results. Why?

What about your claim that the images are diffraction-limited. How do you test this?

What about the poor spatial resolution of your images?

What about previous knowledge of size and shape of your particles?

"WantobeImageAnalyst"
Are you serious?

Regards

Herbie

:::::::::::::::::::::::::::::::::::::::::::::::::
Am 05.05.19 um 18:48 schrieb WantobeImageAnalyst:

> Thank you, Herbie.
>
> I used 'Track mate' to detect my particles like the attached image
> based on blob size and intensity threshold. I have coordinates of
> their centroid. And I want to apply 2D and 3D Gaussian fitting of
> different shapes according to each particle to discard noise.(Then, it
> will provide total or mean intensity of the region of interest(?))
>
> P.S.I cannot upload the whole image(3D, timelapse) because it exceeded
> the file size limit. If you let me share the image through Google
> drive, I'll do that for a better understanding.
>
> <http://imagej.1557.x6.nabble.com/file/t382308/sample.jpg> of these
> particles.
>
>
>
> --
> Sent from: http://imagej.1557.x6.nabble.com/
>
> --
> ImageJ mailing list: http://imagej.nih.gov/ij/list.html
>

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