In this issue we will discuss a very, very special topic: what is the best way to take a photograph of an egg? As we are within the Water Bear web base area, we will not discuss this for everyday hen eggs, of course, but for water bear eggs only.

As a rule sphaeroidic or egg-shaped objects are difficult to master under a light microsope - in particular when they are not fully transparent. The limited focus depth will not be sufficient to render a crisp image of the overall object. Instead, we will get optical "slices" only. This has been an important argument for teachers not to forget drawing techniques as the only means to collect all the information of the various focus slices into one combined representation covering all details of an object.

So one might end up in thinking that light microscopic photography of those "thick" objects would always end up in an imaging catastrophy.
Of course this would be a pity. Some of the water bear eggs are real marvels of 3D construction symmetry similar to those fascinating, must-see  diatom shells  and   radiolaria .

Now the microscopic amateur living somewhere in our really incredible microscopic & networking world might take out his "Gerlach" book (in Germany) or another comprehensive textbook covering microscopy and might have a look at the curves representing the relations between objective properties, overall magnification and focus depth.
For a 40x objective with a numerical aperture of 0.65 the theoretical value of focus depth is about 2 microns only. This sounds not very good when keeping in mind that the typical water bear egg is about 0.1 mm in diameter which mean als that it is 0.1 mm (100 microns) in depth as well.

We might photograph the different focus slices and study them in series, one by one:

But this is a little bit frustrating and will never satisfy our wish for a good overall documentation.

This is the point where normally the drawing artist comes in: he adjusts focus, uses the pencil a little bit, adjusts focus, draws a little bit again ...

Well, in case you should be a really rich amateur you might simply go out and buy a scanning electron microscope (SEM) which works perfectly well in 3D space as can be seen from the literature, e.g. in the Italian monography about tardigrades by Walter Maucci or in Ian M. Kinchins tardigrade book (see literature below). We cannot show this for copyright reasons here. But even the SEM has its drawbacks: the image information is in black and white only and furthermore we can study only eggs which have been chemically prepared to death.

So, for the wise amateur, is it time to give up?

Not yet! Have a look at the image below:

water bear egg,
overall diameter about 85 µm.
not pressed and living.

How is this possible, so many details, a great focus depth and color? The ingenious solution comes from computing: AstroStack© by Robert J. Stekelenburg (copyright protected internet freeware) does the job. It combines a series of partially crisp photographs into one overall crisp image. For the image shown above only three photographs were used.

Below you will find a screenshot of the program at work. I think it is a fascinating perspective and you will not have to sell all your property in exchange for a SEM ;-)

Literature

Dieter Gerlach: Das Lichtmikroskop. p. 118. 2nd edition. Stuttgart 1985.

REM-photographs of water bear eggs e.g. can be found within:
Ian M. Kinchin: The Biology of Tardigrades. p. 63. London 1994.
Walter Maucci: Tardigrada. Fauna d'Italia. 24. p. 19. Bologna 1986.

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© Text, images, and video clips by  Martin Mach  (webmaster@baertierchen.de).
Water Bear web base is a licensed and revised version of the German language monthly magazine  Bärtierchen-Journal . Style and grammar amendments by native speakers are warmly welcomed.

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