Stomata in tulip leaf

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MicroBob
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Stomata in tulip leaf

#1 Post by MicroBob » Sat Apr 18, 2020 5:59 pm

Hi together,
I'm preparing a video conference meeting on stomata and did a couple of nail polish impressions and hand sections today. Here is a photo of an impression in UV hardening nail polish of the underside of a tulip leaf. I put a tiny droplet of the nail polish on the leaf and then press a 18mm rectangular cover slip with pincers on it. Through the gap between the pincer's legs I shine with an UV flashlight for 30 seconds. The impression can be removed together with the cover slip. Then it is put onto a slide and sealed with narrow strips of adhesive tape.

Here an image with the old Zeiss DIC, just the green channel.

Bob
Attachments
Tulpenblatt Topcoat Luft DIC 1024 .jpg
Tulpenblatt Topcoat Luft DIC 1024 .jpg (206.12 KiB) Viewed 919 times

MichaelG.
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Re: Stomata in tulip leaf

#2 Post by MichaelG. » Sat Apr 18, 2020 6:12 pm

Very impressive, Bob !!

MichaelG.

[ do please forgive the pun ]
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daruosha
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Re: Stomata in tulip leaf

#3 Post by daruosha » Sat Apr 18, 2020 6:13 pm

That's very cool Bob. Any reason why did you use UV curing nail polish? To make it dry faster, I assume. Am i right?
Daruosh.

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KD Arvidsson
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Re: Stomata in tulip leaf

#4 Post by KD Arvidsson » Sat Apr 18, 2020 7:42 pm

I agree very impressive Bob! //KD
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Re: Stomata in tulip leaf

#5 Post by Hobbyst46 » Sat Apr 18, 2020 7:44 pm

Very nice and inspiring to try it, Bob.
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MicroBob
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Re: Stomata in tulip leaf

#6 Post by MicroBob » Sat Apr 18, 2020 8:19 pm

Well, this is how I came to UV hardening nail polish: I used to teach commercial apprentices some technical stuff at work, each one a month. One morning I came into my office and my apprentice of the month held her hands in my direction and said "I did this myself!". My interest was awakened: "New DIY hands? "No", she said, "can't you see my new finger nails???" This was my introduction in UV hardening substances and I immediately bought an UV lamp, in pink: https://www.ebay.de/itm/36W-UV-Lampe-Li ... SwlxRaU4s8

The big advantage of using a UV glue is the quick curing. The leaf doesn't get much time to react and shrivel while the glue hardens. here I didn't use the pink lamp but an UV torch that allowed to apply the light in a very conentrated way.

I tried mounting with LOCA TP2500, an elastic UV hardening glue used for phone displays. By eye the imprint stays visible, und the microscope-invisible, even in DIC and phase.
Next was Pleurax: Contrast is better but overall not a great method.
Since the imprint is already attatched to the cover slip there is no glass-air-imprint combination, but just glass-imprint. I didn't try but high power work would work.

This chinese "Topcoat" nail polish is quite hard and brittle. It can be used to make small lenses for jndemanding purposes. It doesn't adhere well to a cover slip when applied in thick layers like a drop hanging from the slide.

I even made some comparisons of contrast techniques. Oblique and DIC were best, plain bright field and phase were not able to compete.

Bob

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75RR
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Re: Stomata in tulip leaf

#7 Post by 75RR » Sat Apr 18, 2020 8:45 pm

That is pretty nifty!
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MicroBob
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Re: Stomata in tulip leaf

#8 Post by MicroBob » Sun Apr 19, 2020 10:03 am

To show that no expensive DIC is necessary here a single shot with oblique lighting.
Attachments
Tulpenblatt Topcoat Luft schiefe Beleuchtung DSC_9922 1024.jpg
Tulpenblatt Topcoat Luft schiefe Beleuchtung DSC_9922 1024.jpg (244.09 KiB) Viewed 848 times

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daruosha
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Re: Stomata in tulip leaf

#9 Post by daruosha » Sun Apr 19, 2020 10:11 am

Nice job Bob. I have a few tubes of UV curable glue for repairing PCBs. I'll like to try your recipe and see how it works. The glue I have is not fully transparent, but I think it doesn't matter.
Daruosh.

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Re: Stomata in tulip leaf

#10 Post by MicroBob » Sun Apr 19, 2020 10:59 am

Hi Daroush,
I would just try what you have. A semi-transparent glue might be just perfect as you get an amplitude object from it. One useful property would be good adherance to hard surfaces, this should be given in a glue that is intended for mounting electronic components. The glue should have a suitable viscosity to fill the pores and there should be no particles in it like flakes, it should be stained right through.

What might also work is quick hardening 2k epoxy as it hardens by itself and leaves an imprint that is directly attatched to the cover slip. The classic method would be solvent based nail polish or UHU hart glue that dries on the leaf and is then peeled of and mounted. But this will probably not giv ethe same quality of slide.

To carry the UV nail polish attempt further would be to stain it a see how a light stain would influence the image quality.

The stomata are mostly on the lower sides of the leaves and they are arranged differently from plant to plant. It is possible open and close the stomata by means of light and shadow, sugar solution (e.g. 0,5 mol) or plain water. So with enough determination the functionality of the stomata can be shown nicely.

Bob

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Re: Stomata in tulip leaf

#11 Post by tgss » Sun Apr 19, 2020 11:22 am

Very interesting series of posts. Thanks for starting it off, MicroBob, with your example of a variation on an old method that appears to give excellent results.
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Re: Stomata in tulip leaf

#12 Post by Wes » Sun Apr 19, 2020 1:49 pm

Great idea Bob! Very interesting approach for studying surfaces of objects otherwise not suitable for transmitted light microscopy.

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Re: Stomata in tulip leaf

#13 Post by MicroBob » Sun Apr 19, 2020 4:08 pm

Here a rough comparison of images taken with different contrast techniques and little image editing, all single shots. On cause there is a lot of room for improvement in every shot, it just shows round about how much potential is there.

Bob
Attachments
Tulpenblatt Topcoat Luft schiefe Beleuchtung DSC_9922 1024.jpg
Tulpenblatt Topcoat Luft schiefe Beleuchtung DSC_9922 1024.jpg (244.09 KiB) Viewed 795 times
Vergleich Kontrastverfahren DIC 1024.jpg
Vergleich Kontrastverfahren DIC 1024.jpg (334.25 KiB) Viewed 795 times
Vergleich Kontrastverfahren Hellfeld 1024.jpg
Vergleich Kontrastverfahren Hellfeld 1024.jpg (141.66 KiB) Viewed 795 times
Vergleich Kontrastverfahren Phasenkontrast 1024.jpg
Vergleich Kontrastverfahren Phasenkontrast 1024.jpg (285.51 KiB) Viewed 795 times

MicroBob
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Re: Stomata in tulip leaf

#14 Post by MicroBob » Sun Apr 19, 2020 8:18 pm

Another simple way to observe stomata: Hand sections!
By omitting the microtome one gains flexibility and speed. Aim is not a planparallel section but a wedge that at least has the right thickness in an area.
For thin objects it is feasible to try to cut planparallel and advance in tiny increments. This can be done by using the finger that holds the specimen as a fulcrum and angle it back from cut to cut: https://vimeo.com/409525546

It is possible to do crosscuts as shown on the pine needle (Etzold FCA stain)
It is alos posible to do flat sections that remove a bit of epidermis, shown here on a Red Lucky Clover.
Attachments
Spaltöffnungen 40-1 1024.jpg
Spaltöffnungen 40-1 1024.jpg (125.34 KiB) Viewed 769 times
Fichtennadel FCA 1024 DSC_9998.jpg
Fichtennadel FCA 1024 DSC_9998.jpg (225.45 KiB) Viewed 769 times
Spaltöffnung fichtennadel FCA DSC_9999.jpg
Spaltöffnung fichtennadel FCA DSC_9999.jpg (176.8 KiB) Viewed 769 times

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daruosha
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Re: Stomata in tulip leaf

#15 Post by daruosha » Mon Apr 20, 2020 11:07 am

The first image is very eye catching to me. Nice tips/techniques too.
Daruosh.

sinabro
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Re: Stomata in tulip leaf

#16 Post by sinabro » Tue Apr 21, 2020 1:02 pm

I agree, daruosha.
I really like the first image.

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