Moss leaves in glycerin: how to prevent cells from collapsing?

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JayAvery
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Moss leaves in glycerin: how to prevent cells from collapsing?

#1 Post by JayAvery » Wed Mar 04, 2020 12:37 pm

I want to make semi-permanent slides of moss leaves in glycerin. I've read that glycerin can be used with samples straight from water, but I had trouble the first time trying it. When I placed the moss leaf into a drop of pure glycerin, it immediately started to curl up. I managed to flatten it out under a cover slip, but when I examined it under the microscope the cells were shrivelled and collapsed (as if dried out). Why did this happen, and what can I do to prevent it?

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75RR
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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#2 Post by 75RR » Wed Mar 04, 2020 3:31 pm

glycerin is hygroscopic, so it absorbs water from the moss leaves.
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mrsonchus
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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#3 Post by mrsonchus » Wed Mar 04, 2020 4:49 pm

You may try a weak glycerin/water mix - perhaps 1:10 (v/v) - but with a glycerin content that is enough to cover the sample if used on it's own. The idea being to put the leaves into the mixture then allow the water content to slowly evaporate - thus gradually increasing the concentration of glycerin until the sample/leaf is in pure glycerin. At that stage perhaps a glycerin-gel mount may be OK. I may give this a try myself to see what happens if I get the chance.

I also wonder if at the stage of total infiltration (with glycerin) a glycerin gel may be employed - giving a solid mount that may even be sealed for permanence/longevity?

Let us know how you get along....
John B

JayAvery
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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#4 Post by JayAvery » Wed Mar 04, 2020 5:24 pm

mrsonchus wrote:
Wed Mar 04, 2020 4:49 pm
You may try a weak glycerin/water mix - perhaps 1:10 (v/v) - but with a glycerin content that is enough to cover the sample if used on it's own. The idea being to put the leaves into the mixture then allow the water content to slowly evaporate - thus gradually increasing the concentration of glycerin until the sample/leaf is in pure glycerin. At that stage perhaps a glycerin-gel mount may be OK. I may give this a try myself to see what happens if I get the chance.

I also wonder if at the stage of total infiltration (with glycerin) a glycerin gel may be employed - giving a solid mount that may even be sealed for permanence/longevity?

Let us know how you get along....
Thanks for the suggestion! I tried another attempt right after I posted this. I put the leaf in a drop of water on a slide, then added a drop of glycerin, and then heated it to boil away some of the water. So far the cells look in good condition, but that may just be because a lot of the water is still there, I have no idea how to tell whether it was all boiled off. :D I'll see how much it dries out over the next couple of days, and whether the leaf loses condition in the process.

I'm very much a beginner in making slides of my own, so I wanted to start off at the very simple end with pure glycerin rather than the extra steps of gelatin. But it's a good idea, so I may try it at some point - keep us filled in if you try it too! :)

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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#5 Post by JayAvery » Wed Mar 04, 2020 6:05 pm

Now I'm very confused because I just tried another attempt with pure glycerin and.. it went fine? The only difference is that with the first leaf I put a drop of glycerin down then placed the leaf into it, and with this attempt I spread the leaves on the slide and then dropped glycerin on top. Surely that can't have had that much of an effect?! But the leaves didn't curl or dehydrate, and the cells look perfectly normal. Maybe the first one was a fluke somehow, maybe the leaf was already a bit dry because it hadn't been in water for a few moments and so the glycerin just fixed it as it was rather than soaking in.

(And yes, I was using leaves from the same species, from the same sample, for every attempt!)

Anyway, I'll check back tomorrow for another update. :D

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75RR
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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#6 Post by 75RR » Wed Mar 04, 2020 7:52 pm

JayAvery wrote:
Wed Mar 04, 2020 6:05 pm
Now I'm very confused because I just tried another attempt with pure glycerin and.. it went fine? The only difference is that with the first leaf I put a drop of glycerin down then placed the leaf into it, and with this attempt I spread the leaves on the slide and then dropped glycerin on top. Surely that can't have had that much of an effect?! But the leaves didn't curl or dehydrate, and the cells look perfectly normal. Maybe the first one was a fluke somehow, maybe the leaf was already a bit dry because it hadn't been in water for a few moments and so the glycerin just fixed it as it was rather than soaking in.

(And yes, I was using leaves from the same species, from the same sample, for every attempt!)

Anyway, I'll check back tomorrow for another update. :D
Perhaps the variable is the glycerin? If it was left out or open it might have absorbed humidity and reached saturation point. Just a thought.
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JayAvery
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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#7 Post by JayAvery » Wed Mar 04, 2020 7:59 pm

Perhaps the variable is the glycerin? If it was left out or open it might have absorbed humidity and reached saturation point. Just a thought.
Interesting thought. It seems unlikely though because it was only a couple of hours apart, and the glycerin came straight out of the bottle both times and was sealed in between. 🤔

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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#8 Post by MicroBob » Wed Mar 04, 2020 8:38 pm

Hi Jay,
a different method to make moss slides is to let the moss dry out fully, place a coverslip on top and fix it with two drops of glue or nail polish. For observation a drip of dem. water is placed so is is sucked under the cover slip. I haven't tested this myself.

Bob

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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#9 Post by JayAvery » Thu Mar 05, 2020 6:53 pm

Update: the pure glycerin slide from yesterday hasn't dried out at all and the cells are still in perfect condition! I don't know what happened the first time but the subsequent attempt was fine. I'll definitely be doing more tests with different parts and different species to see if the cell-shrinking problem reappears.

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Re: Moss leaves in glycerin: how to prevent cells from collapsing?

#10 Post by GrbovicMicroscopy » Sun Mar 08, 2020 10:55 pm

mrsonchus wrote:
Wed Mar 04, 2020 4:49 pm
I also wonder if at the stage of total infiltration (with glycerin) a glycerin gel may be employed - giving a solid mount that may even be sealed for permanence/longevity?

Let us know how you get along....
I made a lot of permanent slides of plant stems with glycerin gel so far they have lasted about 6-8 months. I seem to have found out when i make slides with lower concentration of gel the specimen tends to get separated from the gel even tho i have nail polished the coverslip. But if mounted in gel with higher concentration of gel it does not happen... We will see how long they last. I wanted to post pictures here but the site wont let me for some reason..

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