Optical artefact removal: reply

[Jacqueline Ross]

Dear All,

 

Thanks very much to all of those who made suggestions both microscope-related and ImageJ solutions. Thanks especially to those who confirmed that this is a known issue with some Coolpix cameras and gave some suggestions about the cause and I might try to improve the imaging.

 

A few people asked questions which I will briefly answer below.

 

The artifact is seen with all objectives and I have cleaned them myself to make sure it is not due to any film on the lens. It is not visible down the eyepieces but only apparent on the images from the camera.

 

As Joel also commented, the size of the artifact is consistent through all objectives and is visible at different focal planes.

 

The effect is not a moiré effect but like a diffraction (circular rings) pattern. If you look at the images on the website (http://www.health.auckland.ac.nz/biru/exptal_images/index.html) , you should be able to see it particularly in the right hand images which don't have tissue/cells. You may need to download one if you have difficulty seeing it.

 

There doesn't seem to have any light leak and this microscope only has one light source. However, the research group sometimes use this particular camera on a fluorescence microscope where they also experience problems with the images although I haven't seen these as yet.

 

Before writing to the listserver, I tried out the Image Calculator with the images with/without tissue or cells but the resulting image is very dark. Andy suggested using a multiplication factor to restore brightness. Does anyone know what I should try?

 

I also have NIH Image on another computer so I will try that as well as suggested by John. Maybe it works differently.

 

Joel, I've tried out your suggestion but am having trouble to balance the colours back again. I also still seemed to see the artifact in one channel (blue). Perhaps I'm doing something wrong so I will persevere.

 

Thanks also for the other more complex suggestions (Gabriel, Jonathan) which I will also try.

 

Unfortunately I think this research group is purchasing another Coolpix camera. Does anyone know whether the newer ones are OK or do they still have problems with this artifact?

 

Cheers,

 

Jacqui.

 

Jacqueline Ross
Biomedical Imaging Research Unit
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/ <http://www.health.auckland.ac.nz/biru/>  

 

[Aryeh Weiss]

Jacqui Ross wrote:

>
> The artifact is seen with all objectives and I have cleaned them myself to
> make sure it is not due to any film on the lens. It is not visible down the
> eyepieces but only apparent on the images from the camera.
>
>
>
> As Joel also commented, the size of the artifact is consistent through all
> objectives and is visible at different focal planes.
>
>
>

You should check whether the ring pattern changes (or disappears) when you
change the condenser diaphragm. If the condenser diaphragm is too small, then
you are providing partially coherent illumination, and this allows a speck of
dust in a non-image plane along the light path to produce that kind of pattern.
If you increase the NA of the condenser, this may remove the artifact, but then
be sure to cut down the camera gain (exposure) to avoid saturation.

A corollary of this is that the entire light path may need to be cleaned, but
the dust generally returns as fast as you can clean it off.

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

Ph:  972-3-5317638
FAX: [phone]

[Monique Vasseur]

Do you have an infrared filter after your lamp (heat absorbing filter)?
Some cameras have an infrared filter in front of their lens, others not, do you know if the Coolpix has one?

Monique Vasseur
Microscopie et imagerie
Département de biochimie
Université de Montréal
tél. [phone] poste 5148
-----Message d'origine-----
De : ImageJ Interest Group [mailto:[hidden email]] De la part de Jacqui Ross
Envoyé : 19 juillet 2006 02:35
À : [hidden email]
Objet : Optical artefact removal: reply

Dear All,

 

Thanks very much to all of those who made suggestions both microscope-related and ImageJ solutions. Thanks especially to those who confirmed that this is a known issue with some Coolpix cameras and gave some suggestions about the cause and I might try to improve the imaging.

 

A few people asked questions which I will briefly answer below.

 

The artifact is seen with all objectives and I have cleaned them myself to make sure it is not due to any film on the lens. It is not visible down the eyepieces but only apparent on the images from the camera.

 

As Joel also commented, the size of the artifact is consistent through all objectives and is visible at different focal planes.

 

The effect is not a moiré effect but like a diffraction (circular rings) pattern. If you look at the images on the website (http://www.health.auckland.ac.nz/biru/exptal_images/index.html) , you should be able to see it particularly in the right hand images which don't have tissue/cells. You may need to download one if you have difficulty seeing it.

 

There doesn't seem to have any light leak and this microscope only has one light source. However, the research group sometimes use this particular camera on a fluorescence microscope where they also experience problems with the images although I haven't seen these as yet.

 

Before writing to the listserver, I tried out the Image Calculator with the images with/without tissue or cells but the resulting image is very dark. Andy suggested using a multiplication factor to restore brightness. Does anyone know what I should try?

 

I also have NIH Image on another computer so I will try that as well as suggested by John. Maybe it works differently.

 

Joel, I've tried out your suggestion but am having trouble to balance the colours back again. I also still seemed to see the artifact in one channel (blue). Perhaps I'm doing something wrong so I will persevere.

 

Thanks also for the other more complex suggestions (Gabriel, Jonathan) which I will also try.

 

Unfortunately I think this research group is purchasing another Coolpix camera. Does anyone know whether the newer ones are OK or do they still have problems with this artifact?

 

Cheers,

 

Jacqui.

 

Jacqueline Ross
Biomedical Imaging Research Unit
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/ <http://www.health.auckland.ac.nz/biru/>  

 

[Henry Barwood]

I know that this has been gone over quite a bit, but looking at the images,
I wonder if it could be diffraction off of the microscope tube. One way to
test would be to remove the cameras and eyepiece and simply look down the
illuminated tube. If you see concentric rings and/or bright spots it is
sometimes caused by too-small or improperly aligned light stops. I have also
had components diffract off the edges of an insert or component module. Just
a thought.

Henry Barwood
Associate Professor of Science, Earth Sciences
Department of Math and Physics
MSCX 312G
Troy University
Troy, Alabama
[hidden email]

[Jacqueline Ross]

Hi Monique,

No, I don't know if the camera has an infrared filter. I know one of our cameras does. I'll see if I can find out. Or someone on the list may know?

Thanks for the idea.

Cheers,

Jacqui.

Jacqueline Ross
Biomedical Imaging Research Unit
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/ 
-----Original Message-----
From: ImageJ Interest Group [mailto:[hidden email]] On Behalf Of Vasseur Monique
Sent: 20 July 2006 02:28
To: [hidden email]
Subject: Re: Optical artefact removal: reply

Do you have an infrared filter after your lamp (heat absorbing filter)?
Some cameras have an infrared filter in front of their lens, others not, do you know if the Coolpix has one?

Monique Vasseur
Microscopie et imagerie
Département de biochimie
Université de Montréal
tél. [phone] poste 5148
-----Message d'origine-----
De : ImageJ Interest Group [mailto:[hidden email]] De la part de Jacqui Ross
Envoyé : 19 juillet 2006 02:35
À : [hidden email]
Objet : Optical artefact removal: reply

Dear All,

 

Thanks very much to all of those who made suggestions both microscope-related and ImageJ solutions. Thanks especially to those who confirmed that this is a known issue with some Coolpix cameras and gave some suggestions about the cause and I might try to improve the imaging.

 

A few people asked questions which I will briefly answer below.

 

The artifact is seen with all objectives and I have cleaned them myself to make sure it is not due to any film on the lens. It is not visible down the eyepieces but only apparent on the images from the camera.

 

As Joel also commented, the size of the artifact is consistent through all objectives and is visible at different focal planes.

 

The effect is not a moiré effect but like a diffraction (circular rings) pattern. If you look at the images on the website (http://www.health.auckland.ac.nz/biru/exptal_images/index.html) , you should be able to see it particularly in the right hand images which don't have tissue/cells. You may need to download one if you have difficulty seeing it.

 

There doesn't seem to have any light leak and this microscope only has one light source. However, the research group sometimes use this particular camera on a fluorescence microscope where they also experience problems with the images although I haven't seen these as yet.

 

Before writing to the listserver, I tried out the Image Calculator with the images with/without tissue or cells but the resulting image is very dark. Andy suggested using a multiplication factor to restore brightness. Does anyone know what I should try?

 

I also have NIH Image on another computer so I will try that as well as suggested by John. Maybe it works differently.

 

Joel, I've tried out your suggestion but am having trouble to balance the colours back again. I also still seemed to see the artifact in one channel (blue). Perhaps I'm doing something wrong so I will persevere.

 

Thanks also for the other more complex suggestions (Gabriel, Jonathan) which I will also try.

 

Unfortunately I think this research group is purchasing another Coolpix camera. Does anyone know whether the newer ones are OK or do they still have problems with this artifact?

 

Cheers,

 

Jacqui.

 

Jacqueline Ross
Biomedical Imaging Research Unit
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/ <http://www.health.auckland.ac.nz/biru/>  

 

[Jacqueline Ross]

Hi Aryeh,

Thanks for your comments. The field diaphragm and condenser aperture are
set up for Koehler so as to match the NA of the objective lens. The
consistency of the pattern and its central location do seem to indicate
a systemic fault.

Cheers,

Jacqui.

Jacqueline Ross
Biomedical Imaging Research Unit
School of Medical Sciences
Faculty of Medical & Health Sciences
The University of Auckland
Private Bag 92019
Auckland, NEW ZEALAND

Tel: [phone] Ext 87438
Fax: [phone]

http://www.health.auckland.ac.nz/biru/ 
-----Original Message-----
From: ImageJ Interest Group [mailto:[hidden email]] On Behalf Of
Aryeh Weiss
Sent: 19 July 2006 19:13
To: [hidden email]
Subject: Re: Optical artefact removal: reply

Jacqui Ross wrote:


>
> The artifact is seen with all objectives and I have cleaned them
myself to
> make sure it is not due to any film on the lens. It is not visible
down the
> eyepieces but only apparent on the images from the camera.
>
>
>
> As Joel also commented, the size of the artifact is consistent through
all
> objectives and is visible at different focal planes.
>
>
>

You should check whether the ring pattern changes (or disappears) when
you
change the condenser diaphragm. If the condenser diaphragm is too small,
then
you are providing partially coherent illumination, and this allows a
speck of
dust in a non-image plane along the light path to produce that kind of
pattern.
If you increase the NA of the condenser, this may remove the artifact,
but then
be sure to cut down the camera gain (exposure) to avoid saturation.

A corollary of this is that the entire light path may need to be
cleaned, but
the dust generally returns as fast as you can clean it off.

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

Ph:  972-3-5317638
FAX: [phone]