A mysterious ball

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Myriophyllum
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Location: Schaumburg, North of Germany

A mysterious ball

Post by Myriophyllum »

Hi,

here some pictures of a mysteriously looking ball in a cover with funnellike craters.
Has anyone an idea what that can be?
A cyst of amoebia (I hope that's english ;-) )?

Image


Like many of my favourite "micromodels" it was found on sphagnum-moss at a pond in the woods of Schaumburg (Germany) this november.

Zeiss Standard; Planpo 40/0,95; S-Kpl 10/20; slide film; one image each;
the picture above was made using asmmetric col-lightning with additional darkfield component (I know, it's some kind of crazy light but it makes the ball looking even more mysteriously).
The images below are darkfield.


Best wishes

Jens

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Ken Ramos
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Post by Ken Ramos »

I would almost venture going out on a limb and saying that it could be a cyst but I really don't think so, however I could be wrong. I am far from an expert, even the least bit knowledgeble, in microorganisms. More like an egg of some kind maybe. Cysts are more compact and hardened in appearance. At least most of the ones I have seen. :-k

Some very good photograph you have posted here though. :D What is asymmetric col-lighting? I am a little familar with COL. :-k
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Charles Krebs
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Post by Charles Krebs »

Jens... I'm not a good one for IDing these things... but my first guess was some sort of encysted creature. (Based mostly on the lower right image). You have, once again, given us very interesting, detailed images, so hopefully someone more knowledgeable will be of more help with an ID.

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Ken Ramos
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Post by Ken Ramos »

See I was wrong. :lol: I believe this link will answer your question Jens :D http://protist.i.hosei.ac.jp/PDB/Images ... ata_2.html
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Kenneth Ramos
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Charles Krebs
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Post by Charles Krebs »

Hey Ken... you got it!
(Does that mean you are more knowledgeable than me? :wink: :roll: )

Myriophyllum
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Let's talk about studio photography...

Post by Myriophyllum »

Wow, fantastic, Ken! :-)
Bursaria ovata! Even the size is almost the same.

Thanks a lot.


About your question - you wrote: " What is asymmetric col-lighting? I am a little familar with COL. "

How you can make a illumination like I used it can be described in three steps:

First let's have a typical col stop as described by Paul James:

Image

The stop is optimized for a objective 40/0.95 to be placed above the filter tray (diameter of the outer ring 31 mm, the whole thing can be printed easily on a 33x75 mm piece of inkjet film).

We have four areas:
A) useless zone, can stay dark
B) the col-part of it
C) zone for an additional darkfield compound
D) central col-stop
Dashed or gray lines are just to show where darkfield ends.


Now some modifications leading to stop two:

Image

E and F) the darkfield zone and the col zone were made asmmetric. This idea came as I read an article about studio photography. Here in most cases a strong light beside the object is combined by a week light (or a white thing) on the other side.
G) smaller central stop for a more natural look.


The studio photograph certainly has many possibilities to optimize the background on his photo - so have we (usually called Reinberg lightning):

Image

H) the col area is filled with gray to get the background darker

That's it :-)

Jens

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Ken Ramos
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Post by Ken Ramos »

Charlie said:
Hey Ken... you got it!
(Does that mean you are more knowledgeable than me? )
No. Just means I was lucky enough to beat you to it Charlie :lol:

Jens wrote:
Wow, fantastic, Ken!
Bursaria ovata! Even the size is almost the same.

Thanks a lot.
Hey no problem Jens. :D I just went over to Protist Information Server and clicked on cysts and there it was on the front page.

Now about this asymmetric col. Looks somewhat similar to Zeiss's VAREL Contrasting. I had the privilage of using or testing a prototype here a few months back. Although it did not fit my Axiostar exactly and was a bit hard to use. It is the last diagram that looks like the VAREL annulus.

Thanks Jens. :D
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Myriophyllum
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Post by Myriophyllum »

Hello Ken,

I just took a look at the Zeiss websites to learn about Varel.
Why was Varel a bit hard to use? Theoretically it sounds quite simple (if once installed).

As I understoud it means to have an annulus in the condenser where light can only pass in a small area (like about a sixth part of a condenser Ph-ring but wider).
This area can be moved from the border to the middle producing various kinds of light.
If a Ph-objective is used we have darkfield or oblique light or a Ph plus oblique light combination, depending upon the offset of the Varel stop.

The difference about the stops I used is that here each part of the light can be combined freely, including how oblique the light is and how much Col type or darkfield (see picture) you wish. The full aperture of both condenser and objective is used all the time.

It's important to find a good balance between darkfield and col.
One example is this picture of Tetraspora sp. : left photos with too much darkfield, right a good balance (for my taste).

Image

(Planapo 40/0.95; S-Kpl 10x/20)


For some kind of flat objects you results like this:

Image

(Plan-Neofluar 25/0.80; S-Kpl 16x/12.5)


Greetings
Jens

PS: The snow starts falling right now :-), merry christmas to all of you.

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Ken Ramos
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Post by Ken Ramos »

Jens asked:
Why was Varel a bit hard to use? Theoretically it sounds quite simple (if once installed).

It is not that it was hard to use but that the system I had was a prototype for the Axiostar Plus. The VAREL system was initially made for the Axioskop 40 and the other higher end Zeiss models and was ill fitting on the Axiostar. Soon it may be available for the Axiostar, at least I hope so. :D
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Kens Microscopy
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Myriophyllum
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Post by Myriophyllum »

Hi,

I just read my col describtion again...

B) and C) were mixed up, sorry.

Jens

Charles Krebs
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Post by Charles Krebs »

Jens,
Thanks for the details of your specialized condensor stop. Your results are excellent!

It really is too bad so many "modern" microscope condensors have no easy way to place "stops" in a filter holder near the aperture. It is an easy an effective way to get very useful lighting effects.

Harry Yosh
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Post by Harry Yosh »

Based on the first image taken by Myriophyllum, I synthesized a stereograph as shown below (view: parallel eyes / with permission by Myriophyllum).
Image
(software: Stereographer)

rjlittlefield
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Post by rjlittlefield »

Jens,

Looking at the stereograph has made me realize that I am confused by your images.

If the cyst is really a ball, then when you are focused on its front surface, the ball should get fuzzier and fuzzier toward its edges.

But your images seem to be uniformly sharp across the whole width of the cyst.

Why? What is the shape of the detail that I am seeing? Is this thing pressed against the coverslip so that its front surface is really planar?

--Rik

Myriophyllum
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Post by Myriophyllum »

Hi Rik,

well, thats a good question...

First of all I want to say that the larger image was made from one single shot and it isn't sharpened.

If we take a closer look at the image with more contrast below, the middle (A) is less sharp then the other surrounding parts (B, C, D):

Image

So I think the front surface isn't planar.
As it was also possible to bring details above the surface (of the same kind as we see them sharp beside the ball) to focus, the ball can't be flattened very much.

If anybody thinks I'm going wrong, just tell me.

Greetings
Jens

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Post by rjlittlefield »

Jens, here is the reason for my question.

You're using a 40X NA 0.95 objective to image something that's roughly 100 microns wide. But DOF at that magnification & NA is very small -- for example Table 1 at http://www.microscopyu.com/articles/for ... depth.html lists only 1 micron for 40X NA 0.65. I'm seeing well over half the diameter of the smooth inner sphere in good focus, as well as almost the entire diameter of that apparently folded outer pellicle. That's telling me that all those things have to be within only a few microns of planar, way too flat for being on the surface of a 100 micron sphere. That's why I thought maybe it was pressed up against the cover slip.

I'm having trouble finding anything like an image of a 100 micron sphere to compare directly against. There are of course the cysts at the link that Ken gave, but those are advertised as being "slightly flattened by coverglass". The example I know best is at http://www.janrik.net/insects/ExtendedD ... tereo.html. The third figure shows a single frame of a 500 micron butterfly egg through 10X NA 0.25. This is a much easier problem, but still only a small part of the slightly flattened sphere is in sharp focus at once. (The microscopyu.com table says DOF = 8.5 microns in this case, which is consistent with the image.)

Am I missing something here?

--Rik

PS. I'm sure you don't need me to tell you, but you have made a beautiful image -- it would look good printed and framed!

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