swimming spores

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iconoclastica
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swimming spores

#1 Post by iconoclastica » Tue Oct 25, 2022 6:21 pm

Fern spores are much like pollen. They vary in size, but typically they're about 50 µm long. Often I need to take a quick look at them to see whether they're spiny, wartly, smooth or whatever. I stress the word 'quick' in the previous sentence because at such moments I am not busy with microscopy but with ferns and just need some more magnification. To do so, I move some spores onto a slide, add a small drop of water and add a cover glass. After viewing the samples are disposed of.

At times, though, it turns out the spores are interesting enough to make a photo of them. Then I use the 100x objective and immersion. And several problems pop up. Either the water evaporates and the slide dries from the sides. In itself that is nothing too bad and can even be remedied by adding a droplet near the edge of the cover glass. But excessive evaporation causes currents that move the spores I am photographing. Since I make stacks of a few tens of exposures, I'd rather they would stay where they are during the session.
Or there's too much water (apparently) and the spores move again, by the presure of the objective front or even jump around for the light vibration of the camera shutter.

I can't use glycerin in stead of water for it makes the spores more transparent and the surface features too hard to see.

Sometimes the samples behave very well, but it is unpredictable. What are to options to improve the stability of the slides?

Chas
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Re: swimming spores

#2 Post by Chas » Wed Oct 26, 2022 1:52 pm

I had a box of 24x60mm coverslips... they seemed to float less with a given amount of water (a 'drop')

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Wes
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Re: swimming spores

#3 Post by Wes » Wed Oct 26, 2022 7:09 pm

You can deposit a thin strip of petroleum jelly on the underside edge of the coverslip. Alternatively, after you've assembled your slide coat the boarder between coverslip and glass slide with nail polish. Once the nail polish dries out (takes a few minutes) water evaporation happens much slower.
Zeiss Photomicroscope III BF/DF/Pol/Ph/DIC/FL/Jamin-Lebedeff
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Chas
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Re: swimming spores

#4 Post by Chas » Thu Oct 27, 2022 1:38 pm

Rolf Vossen uses a light cooking oil when examining pollen:
https://microscopyofnature.com/pollen

I used some in the summer and the biggest surprise/pleasure was the lack of evaporation.... the ability to look at 'quickies' the next day or a week later is really nice.

I dug out the 24 x 60mm coverslips, mentioned above, and tried a water mount of some fine diatom powder. Using an oil immersion lens & A-type oil, the long coverslip didnt budge at all ...I couldnt see any drift, just lots of brownian motion.
Also had a stab with cooking oil and the long slips, it seemed to take 10 minutes or so to suck itself along the length of the coverslip and in the process trapped quite a lot of small bubbles, however whilst the coverslip wasnt as glued to the slide as tightly as it was with the water there didnt seem to be a problem with items moving except adajacent to the bubbles.
Maybe there is room for some experimentation with different oils and these long slips?

(I notice that Zeiss make an immersion oil with nearly the same refractive index as water ..Immersol W )

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iconoclastica
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Re: swimming spores

#5 Post by iconoclastica » Sun Oct 30, 2022 11:48 am

Thanks Chas, those are useful remarks. I definitely will try oily media, it's easy enough to obtain them ;)
But I am afraid it will make the spores to transparent.
Rolf wrote:In an oil, pollen grains become more clear and transparent and the structures can be better observed
That is exactly the problem I get with glycerin: the characteristic surface details disappear and the internal structure that is not interesting comes in place.
I expect that an oil-ethanol mixture will flow more easily and possibly even flattens out more when the alcohol starts evaporating.
Interesting note by Rolf that in water "some pollen grains will swell and eventually burst". Spores do the opposite: glycerin and more so, alcohol, tend to make them eject their internals. After that there's an ugly blob of cell content next to the shell and the latter again looks too transparent.

I have been reading through the basics of microscopy the past days to see if there's anything I may be overlooking. One thing that was brought to mind again is that with the 100x objective only the top of the sample can be focused. Which means I have to zoom in to the nearest spores. This may be part of the issue: pressure excerted by the objective that dislodges previously stable spores and introduces stress on the medium so that they after being shaken by the camera shutter don't resume their exact position again. I think it's possible that this happens before the objective visibly hits the cover glass and evering beneath it starts racing around.

Chas
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Re: swimming spores

#6 Post by Chas » Sun Oct 30, 2022 2:09 pm

iconoclastica... there might be one thing; are you using the thinner type of immersion oil? ["Type A"] it is much more watery and so ought to flow in and out from the underneath of the objective more easily.
I find it easier to use.
e.g. https://amscope.com/products/ml-a

The 24x60 coverslips are a bit extreme (and maybe a bit too big to be perfectly convenient) I see that there are reasonably priced 24x50 and 24 x40 slips for sale on ebay.

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iconoclastica
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Re: swimming spores

#7 Post by iconoclastica » Sun Oct 30, 2022 3:37 pm

The label on my oil excells in being non-informative...
"Labtek immersion oil R.I. 1.518"


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First attempt with olive oil moderately unsuccesful: not too transparent, that's good, but very few spores in the sample, so none within focus reach. Wll try again.

Chas
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Re: swimming spores

#8 Post by Chas » Sun Oct 30, 2022 4:56 pm

Well I guess, on the basis of probabilty, that it will be the thicker stuff.
As a rough check; if you put a drop of it on a slide and a drop of glycerine and then tilt the slide ...does it start to run off before the glycerine?
( The viscosities given on the Cargille website are Type-A 150 cSt , Type-B 1250 cSt and a figure pulled of the web for pure glycerine is ~650 ).
The thin stuff seems to be less available and is more expensive, but as you only use a drop, it ought to last a long time [well I am hoping that is true ] :-)

EYE C U
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Re: swimming spores

#9 Post by EYE C U » Sun Oct 30, 2022 9:11 pm

you talking about these?
FERNs.jpg
FERNs.jpg (113.67 KiB) Viewed 2433 times

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iconoclastica
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Re: swimming spores

#10 Post by iconoclastica » Mon Oct 31, 2022 9:16 am

EYE C U wrote:
Sun Oct 30, 2022 9:11 pm
you talking about these? `
Nope. Those are the sporangia or spore capsules.
The spores are inside them:


HF19113 spore 100x.jpg
HF19113 spore 100x.jpg
HF19113 spore 100x.jpg (29.75 KiB) Viewed 2403 times

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iconoclastica
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Re: swimming spores

#11 Post by iconoclastica » Mon Oct 31, 2022 9:30 am

Chas wrote:
Thu Oct 27, 2022 1:38 pm
Also had a stab with cooking oil

Code: Select all

olive oil:		1.46-1.48		(dependent on wavelength)
coconut oil:		1.45-1.46
Peanut oil:		1.460 – 1.465
Corn oil:		1.465 – 1.468
Sesame oil:		1.465 – 1.469
Sunflower oil:		1.461 – 1.468
Palm oil:		1.449 – 1.455
Sabah oil:		1.4633 ± 0.0021

ref:

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iconoclastica
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Re: swimming spores

#12 Post by iconoclastica » Wed Nov 02, 2022 9:10 pm

It seems that there's nothing to it than making a new slide for observation at the highest magnification. So be it then, comaperd with all the hassle of viewing 100x making an addional slide isn't that much work after all. I am having some succes now with pre-made water-gelatin as mounting medium. I cut off a tiny blob of it and smear it through the spores. Then I apply some pressure to flatten the sample. Additional advantage is that it doesn't dry overnight. In fact, it may be that loosing some water - which it does nonetheless - makes the sample even thinner.


todays work: Dryopteris villarii spore 100x
todays work: Dryopteris villarii spore 100x
HF15025 spores 100x.jpg (26.8 KiB) Viewed 2300 times

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