Another Diatoms

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Waltermicrofossil
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Joined: Sat Jan 03, 2015 3:56 pm

Another Diatoms

#1 Post by Waltermicrofossil » Mon Jan 05, 2015 7:39 pm

Hello everyone,
I show you another set of famous diatoms use for test optic.
I have acquire a movie of 600 frames, then with a software called autostakkert2 i have stacking the single frame.
Then i have use wavelet filter with Registax which is much more selective than unsharp mask use in Photoshop for details.
This technique is used in astronomy to lower the noise created from the ccd and is very good to use in microscopy imaging.

Warmest regards
Walter
Attachments
Surirella-Gemma-(Ehrenberg).jpg
Surirella-Gemma-(Ehrenberg).jpg (247.98 KiB) Viewed 2115 times
Stauroneis Phoenicenteron (Ehrenberg).jpg
Stauroneis Phoenicenteron (Ehrenberg).jpg (248.83 KiB) Viewed 2115 times
Nitzschia sigma (Kuetz).jpg
Nitzschia sigma (Kuetz).jpg (211.07 KiB) Viewed 2115 times
Pleurosigma angulatum (Wm Smith).jpg
Pleurosigma angulatum (Wm Smith).jpg (252.29 KiB) Viewed 2115 times

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75RR
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Location: Estepona, Spain

Re: Another Diatoms

#2 Post by 75RR » Mon Jan 05, 2015 8:34 pm

Hi Walter,
I have acquire a movie of 600 frames, then with a software called autostakkert2 i have stacking the single frame.
Just to make sure I understand. You convert a 600 frame video of the diatom into a 600 image stack to produce one final image. Is that correct?
Presumably while filming you step focus as if taking several (600!) individual shots.
Do you do this by hand in one smooth continuous motion?

First I have heard of this.
Zeiss Standard WL (somewhat fashion challenged) & Wild M8
Olympus E-P2 (Micro Four Thirds Camera)

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Waltermicrofossil
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Re: Another Diatoms

#3 Post by Waltermicrofossil » Mon Jan 05, 2015 9:22 pm

This is a tipical procedure used in astronomy to have the greatest number of possible details from an image with minor noise and also works perfectly in microscopy.
When you have saved an AVI format file in the personal computer, you can process the file to obtain a single image with a good signal-to-noise ratio. For this type of procedure you can use a typical astronomical freeware program called “Autostakkert 2”.
With this software, you can sum and stack a sequence of images taken from a movie and save it as a single image. The technique of summing and stacking images has the advantage of increasing the signal, and decreasing the noise typically introduced by an electronic CCD.
Another method to eliminate CCD noise is to capture a photo of the electronic noise only, then subtract that single photo from the final image to eliminate electronic noise.
Then for obtain the largest number of details without altering the picture, instead of using the unsharp mask i using the filters wavlet which are more selective.
In the past year i have write an article in "Modernmicroscopy"
"How to Obtain the Best Resolution With a Modest Light Microscope" in the section Howto Tutorial series
which explains this technique.

Warmest regards
Walter

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gekko
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Location: Durham, NC, USA.

Re: Another Diatoms

#4 Post by gekko » Mon Jan 05, 2015 11:01 pm

Amazing resolutions. Thank you for posting those and also for the explanation of your methods.

JimT
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Re: Another Diatoms

#5 Post by JimT » Tue Jan 06, 2015 3:46 pm

Excelent images. As an ex amateur astronomer I am familiar with stacking AVI files and dark subtraction to eliminate noise.
What camera did you use and have you considered eliminating the few sensor spots in post processing?

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Waltermicrofossil
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Re: Another Diatoms

#6 Post by Waltermicrofossil » Tue Jan 06, 2015 5:34 pm

Thank's for your compliments.
I have two camera ASI120MM (monocrome camera) and ASI 120MC (Color camera).
I prefer to use the monochrome CCD because inside there is not bayer filter, and i have the possibility of using selective filters in a particular area of the optical spectrum.
For the spots in the photo I believe that it is dirt inside the prism of the microscope, but do not know how to clean them and if you can do.
A system to remove the spots is to take a picture with only the spot and then subtract them from the picture with objects.

Warmest regards
Walter

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vasselle
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Location: France

Re: Another Diatoms

#7 Post by vasselle » Tue Jan 06, 2015 8:24 pm

Bonjour
Très bon résultats
Cordialement seb
Microscope Leitz Laborlux k
Boitier EOS 1200D + EOS 1100D

Peter
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Re: Another Diatoms

#8 Post by Peter » Wed Jan 07, 2015 5:13 pm

Hi Walter,
The resolution you have achieved with the Amphipleura pellucida in your modern microscopy article is amazing, is the original film available on the web so that we can view it to assess the improvement made by your technique?
Great work.
Peter.

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Waltermicrofossil
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Re: Another Diatoms

#9 Post by Waltermicrofossil » Wed Jan 07, 2015 5:28 pm

To improve the resolution it is important to use the light in the frequency of ultraviolet (360-400 nm). Then capture images with a monochrome CCD equipped with an ultraviolet filter as the Wratten 47, or even more selective filter but still are very expensive. During the image elaboration there are some filters called Wavelet and deconvolution that further increase the image detail. There is a great software for this but unfortunately not freeware that is Astra Image 3.0 Pro.

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Waltermicrofossil
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Re: Another Diatoms

#10 Post by Waltermicrofossil » Wed Jan 07, 2015 5:36 pm

Also as you know much better than me, is very important to use the oil between objective and slide and especially between the slide and the condenser.
I must say that the condenser of the Lomo: Aplanatic N.A. 1.4 and oblique illumination is one of the best ever.
With this type of condenser can not only adjust the oblique illumination but rotating the condenser find the perfect point of illumination to observe fine details.

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