More phase contrast photos of cleaned diatoms

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Hobbyst46
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More phase contrast photos of cleaned diatoms

#1 Post by Hobbyst46 » Sun Apr 19, 2020 8:51 pm

This is a continuation of
viewtopic.php?f=10&t=9079

Further slide surfing revealed a Cyclotella meneghiniana Kutzing (tentative idn.). I was very happy since the Centrales are a rare find. So here are images, one with a 40X0.75 Neofluar (dry), the other with 100X1.3 Planapo (oil). Both phase contrast. Images were resized and individually cropped to yield about the same apparent size of the diatom on the screen. Stacks, slightly tweaked (brightness and contrast, untouched color/hue).
In principle, best resolution is expected under green light; here, the purpose was just aesthetics.
Hope it was achieved.
Attachments
Cyclotella stack of 2 40X Neofluar.jpg
Cyclotella stack of 2 40X Neofluar.jpg (61.54 KiB) Viewed 1690 times
Cyclotella stack of 3 100X.jpg
Cyclotella stack of 3 100X.jpg (74.59 KiB) Viewed 1690 times
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DonSchaeffer
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Re: More phase contrast photos of cleaned diatoms

#2 Post by DonSchaeffer » Sun Apr 19, 2020 8:53 pm

These are amazing!

Hobbyst46
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Re: More phase contrast photos of cleaned diatoms

#3 Post by Hobbyst46 » Sun Apr 19, 2020 9:59 pm

Thanks DonSchaeffer.
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MichaelG.
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Re: More phase contrast photos of cleaned diatoms

#4 Post by MichaelG. » Sun Apr 19, 2020 10:26 pm

Hobbyst46 wrote:
Sun Apr 19, 2020 8:51 pm
In principle, best resolution is expected under green light; here, the purpose was just aesthetics.
Hope it was achieved.
It certainly was achieved !

I’m a little puzzled by your ‘green light’ comment though ... would blue light not give better resolution?

MichaelG.
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Re: More phase contrast photos of cleaned diatoms

#5 Post by Hobbyst46 » Mon Apr 20, 2020 9:11 am

MichaelG. wrote:
Sun Apr 19, 2020 10:26 pm
Hobbyst46 wrote:
Sun Apr 19, 2020 8:51 pm
In principle, best resolution is expected under green light; here, the purpose was just aesthetics.
Hope it was achieved.
It certainly was achieved !

I’m a little puzzled by your ‘green light’ comment though ... would blue light not give better resolution?

MichaelG.
Thanks, Michael!
In theory, blue light is supposed to give better resolution due to the shorter wavelength, but I have read that in practice, the optical components of the phase contrast are optimized for green light (546nm); and also in practice, all common-use phase contrast scopes that I have met, like those used to inspect tissue cultures, are configured with a green filter. Once upon a time I recorded diatoms with green light and with blue light and to my eye, there was no difference...
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Re: More phase contrast photos of cleaned diatoms

#6 Post by MichaelG. » Mon Apr 20, 2020 11:24 am

Thanks for the reply, Doron

I can think of three plausible reasons why Green might be considered optimum [as distinct from maximum]
  • The human eye is most sensitive in the Green region
  • many monochrome cameras are also tuned to that range
  • most colour cameras use the Bayer mosaic, which has twice the number of Green sub-pixels
These are also good reasons for optimising the optics to suit.

One day [!] we should probably try some controlled experiments; but meanwhile, I respect your judgement.

MichaelG.
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Re: More phase contrast photos of cleaned diatoms

#7 Post by Hobbyst46 » Mon Apr 20, 2020 12:46 pm

MichaelG. wrote:
Mon Apr 20, 2020 11:24 am
Thanks for the reply, Doron

I can think of three plausible reasons why Green might be considered optimum [as distinct from maximum]
  • The human eye is most sensitive in the Green region
  • many monochrome cameras are also tuned to that range
  • most colour cameras use the Bayer mosaic, which has twice the number of Green sub-pixels
These are also good reasons for optimising the optics to suit.

One day [!] we should probably try some controlled experiments;
Willingly so. When I find a "standard", amphipleura pellucida (very rare here) or pleurosigma (supposedly common but missed it so far), and manage to mount them well.
Last edited by Hobbyst46 on Mon Apr 20, 2020 1:02 pm, edited 1 time in total.
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Re: More phase contrast photos of cleaned diatoms

#8 Post by MichaelG. » Mon Apr 20, 2020 12:58 pm

Hobbyst46 wrote:
Mon Apr 20, 2020 12:46 pm
Willingly so. When I find a "standard", amphipleura pellucida (very rare here) or pleurosigma (supposedly common but missed it so far), and manage to mount them well.
:D :D :D
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Re: More phase contrast photos of cleaned diatoms

#9 Post by MicroBob » Mon Apr 20, 2020 2:09 pm

Hi together,

a nice set of images again, Doron!
Green: I frequently divide the image apart ito th RGB channels when I only need a b/w image. I usually end up with the green channel. Blue is usually quite similar and also quite good, red is quite different in image ans usually a lot worse. But this is the result after photography with white LED-light. It might no be the same with tungsten light or with light that has already passed a colour filter.

Bob

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Re: More phase contrast photos of cleaned diatoms

#10 Post by daruosha » Mon Apr 20, 2020 3:07 pm

Hobbyst46, they are beautiful and stunning.

May i ask how do you clean your diatoms? (Sorry for an irrelevant question).
Daruosh.

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Re: More phase contrast photos of cleaned diatoms

#11 Post by Hobbyst46 » Mon Apr 20, 2020 3:58 pm

Thanks, Bob and darousha.

@darousha:
Diatoms were cleaned with APS - see this post:
viewtopic.php?f=10&t=8101&p=72044&hilit=APS#p72044
One step of boiling in APS, then, instead of repeating it, I rinsed with distilled water, then applied house-grade hypochlorite (clear liquid, not a gel or cream or mixture with anything else). For a couple of hours without warming. Then rinsed.
The whole process is not as efficient and comprehensive as some well-known laboratory procedures that rely on hazardous chemicals. On the other hand, it is much safer for the amateur.
Still, some knowledge and experience of handling materials are essential, and appropriate safety measures must be taken.
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Re: More phase contrast photos of cleaned diatoms

#12 Post by daruosha » Mon Apr 20, 2020 6:11 pm

Hobbyst46 wrote:
Mon Apr 20, 2020 3:58 pm
Thanks, Bob and darousha.

@darousha:
Diatoms were cleaned with APS - see this post:
viewtopic.php?f=10&t=8101&p=72044&hilit=APS#p72044
One step of boiling in APS, then, instead of repeating it, I rinsed with distilled water, then applied house-grade hypochlorite (clear liquid, not a gel or cream or mixture with anything else). For a couple of hours without warming. Then rinsed.
The whole process is not as efficient and comprehensive as some well-known laboratory procedures that rely on hazardous chemicals. On the other hand, it is much safer for the amateur.
Still, some knowledge and experience of handling materials are essential, and appropriate safety measures must be taken.
Huge thanks for the explanation. I've watched a video on YouTube showing how to clean and mount the diatoms, but I cannot reproduce the results shown in the same video.



I'll try sodium hypochlorite. Thanks again.
Daruosh.

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Re: More phase contrast photos of cleaned diatoms

#13 Post by Hobbyst46 » Mon Apr 20, 2020 6:18 pm

@daruosha:

Here is an article about cleaning diatoms with hypochlorite:

A Simple, Inexpensive Method for Cleaning Diatoms

Author(s): Jennifer M. Carr, Gary L. Hergenrader, Nels H. Troelstrup and Jr.
Source: Transactions of the American Microscopical Society, Vol. 105, No. 2 (Apr., 1986),pp. 152-157
Published by: Wiley on behalf of American Microscopical Society
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Re: More phase contrast photos of cleaned diatoms

#14 Post by daruosha » Mon Apr 20, 2020 6:29 pm

Hobbyst46 wrote:
Mon Apr 20, 2020 6:18 pm
@daruosha:

Here is an article about cleaning diatoms with hypochlorite:

A Simple, Inexpensive Method for Cleaning Diatoms

Author(s): Jennifer M. Carr, Gary L. Hergenrader, Nels H. Troelstrup and Jr.
Source: Transactions of the American Microscopical Society, Vol. 105, No. 2 (Apr., 1986),pp. 152-157
Published by: Wiley on behalf of American Microscopical Society
Found the paper. Thanks a lot. ;)
Daruosh.

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Re: More phase contrast photos of cleaned diatoms

#15 Post by MicroBob » Mon Apr 20, 2020 7:07 pm

daruosha wrote:
Mon Apr 20, 2020 6:11 pm
I've watched a video on YouTube showing how to clean and mount the diatoms, but I cannot reproduce the results shown in the same video.
Hi Daruosh,
I have tried his method and was quite happy with the results. Here in Germany "bleach" is sold under the name "Clorix" and contains 2,6% sodiumhypochlorite.

This is a caustic substance and will destroy the diatoms if applied for too long.

Generally diatom cleaning consists of a range of methods and rarely just one step will be enough. Sand grains e.g. would practically unremovable chemically.

A while ago I presented diatom cleaning as a group topic: http://www.mikrohamburg.de/Programm/Pro ... 170917.pdf
Today I would present different methods: Heat, but only up to 450°C, sodium hypoclorite and Dorons APS method. From my perspective today these are the most promising methods for the home lab when effect, speed and safety are regarded. But the combination with physical methods as described in the (german) document in indispensable.

Here is an other simple method to clean diatoms:

1. Place mud with diatoms in a flat dish
2. cover with a fine net (e.g. 50 to 200µ hole size)
3. cover with water from where the mud is
4. place in a bright place for a day
5. Remove net with diatoms who have crept towards the light

Bob

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Re: More phase contrast photos of cleaned diatoms

#16 Post by Wes » Mon Apr 20, 2020 7:48 pm

Really nice comparison. The second one looks stunning.

Btw what mounting media are you guys using?

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Re: More phase contrast photos of cleaned diatoms

#17 Post by Hobbyst46 » Mon Apr 20, 2020 7:52 pm

Wes wrote:
Mon Apr 20, 2020 7:48 pm
Really nice comparison. The second one looks stunning.

Btw what mounting media are you guys using?
Thanks Wes !
The mounting medium is Pleurax.
Some time ago I tried Norland's NOA 61, it is a lot easier to apply than Pleurax but less satisfactory because it does not always penetrate into the frustule. The mounted frustule then contains an ugly air bubble. Pleurax is better in this respect.
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Re: More phase contrast photos of cleaned diatoms

#18 Post by KD Arvidsson » Mon Apr 20, 2020 9:18 pm

Very beatiful! Hobbyst46. Second picture is fantastic, sharp and very very nice! //KD
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Re: More phase contrast photos of cleaned diatoms

#19 Post by Hobbyst46 » Mon Apr 20, 2020 10:02 pm

Thanks, KD Arvidsson.
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