Slide mounting media for a beginner - observations

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Sauerkraut
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Slide mounting media for a beginner - observations

#1 Post by Sauerkraut » Fri Jun 07, 2019 5:36 pm

Inspired by one of Oliver's videos, I just started dabbling in slide making of simple subjects, such as small gnats, insect wings, and mosquito antennae, etc. and tried out a few different slide mounting media:

Hoyer's Slide Mounting Medium
Synthetic Canada Balsam in xylene
L'Oreal Shockproof base coat 905 nail polish

Perhaps a few observations could help another beginner. All three media seem to do a nice job of making slides. I didn't have any real issues with air bubbles in any of the 3 media. Both the Hoyer's and Canada Balsam have some warning labels/MSDS info that could give a person pause. They issue warnings about the fumes and keeping it out of your eyes and off of your skin. The Canada Balsam is also flammable. The nail polish doesn't have any specific hazard info but probably should not be inhaled and is possibly flammable.

As far as ease of use, imo the nail polish has the huge advantage of having an applicator brush and is drying quickly, while the Hoyer's seems to take the longest to cure - perhaps over a week or even more. Since fumes are an issue, I was storing the uncured slides outside until they dried. The Canada Balsam has the tendency to skim over quickly, so if a drop was applied to a slide and then a specimen was placed onto the drop, some struggling with sinking it in through the skim coat occurred. Perhaps it would be better to just put the specimen down first and then place a drop on top of it.

I can't speak on the slide longevity of any of these media as of yet and they were only used on dry specimens, no watery items.

So my conclusion for someone just curious about slide making and not sure where to start with mounting media is that starting out with inexpensive, widely available clear nail polish is a viable way to go to make semi-permanent slides.

Just my opinion, and I'm not saying nail polish is the best, just that it is useful enough for someone to start experimenting without buying more expensive and likely more toxic mounting media.

Heather

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Re: Slide mounting media for a beginner - observations

#2 Post by mrsonchus » Fri Jun 07, 2019 6:00 pm

Hi Heather and thanks for the interesting post!

It'd be really interesting and helpful to see some images if you can?

Please keep us updated re your adventures with mounting, especially the methods of the late-great Walter Dioni - a great inspiration for beginners, now's a really good time I think for some demos of the simple 'starter methods' with for example nail polish.
Keep up the good work!
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Re: Slide mounting media for a beginner - observations

#3 Post by Hobbyst46 » Fri Jun 07, 2019 6:19 pm

Hi Sauerkraut,

Such comparison is very useful, thanks for posting!
One difficulty that I encountered with nail polish (in fact, with all viscous media) is inclusion of tiny air bubbles. Microscopists on this forum have devised vacuum chambers for degassing in such cases.
For mounting plant parts (moss) I prefer PVA glue.
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Re: Slide mounting media for a beginner - observations

#4 Post by Sauerkraut » Fri Jun 07, 2019 6:51 pm

Hobbyst46 wrote: One difficulty that I encountered with nail polish (in fact, with all viscous media) is inclusion of tiny air bubbles. Microscopists on this forum have devised vacuum chambers for degassing in such cases.
For mounting plant parts (moss) I prefer PVA glue.
I am not seeing any air bubbles in the nail polish slides. Is it possible that a base coat nail polish is less viscous than regular nail polish?

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Re: Slide mounting media for a beginner - observations

#5 Post by Sauerkraut » Fri Jun 07, 2019 7:01 pm

mrsonchus wrote: It'd be really interesting and helpful to see some images if you can?
I'm not set up to capture decent images yet. But as far as how the specimens looked, all three media gave good results - a nice clear background that works well for bright field.

Thank you for the tip on Walter Dioni. I'll look into that.

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Re: Slide mounting media for a beginner - observations

#6 Post by MicroBob » Fri Jun 07, 2019 7:18 pm

Hi Heather,
thank you for sharing your experiences with mounting media! There is much room for new solutions, especially widely available ones.
Some thoughts about the media you use:
-Hoyers medium: It lightens some specimens over time, e.g. whole insects. One component makes it dangerous to swallow it so make shure to keep it out of reach of children. Experiences with the longevity vary: Many slides crystalize after half a year or so, some last a long time. I have no information what is the difference between these preparations.

-Canada Balsam: The specimen has to be water free so it has to be treated with a row of fluids, ending with xylene or toluene. The synthetic stuff could be everything, so it has to be seen how good it wourks. Sildes in classic Canada Balsam have often stayed somewhat fluid over decades, but have also lasted extremely long.

-Nail polish: It might shrink a lot because of the evaporation of whatever keeps it fluid. This might lead to cracks in the specimen or lifting of cover glasses.
Perhaps some of the solvent can be let evaporate before putting the cover slip on the slide.


What I can suggest for further experiments is LOCA, "liquid optical clear adhesive", sold cheaply to repair smartphone screens. It cures with UV, no shrinking and very good penetration. What I don't know is how specimens like a plant section can be prepared to include in LOCA. I have tried it with radiolaria and got great results.

Very easy to use for plant sections and some other specimens is Euparal if it is available to you.

One general hint: It is usually best to have the specimen close to the cover slip for best observation. So it is of advantage to prepare the specimen on the cover slip and keep the slide upside down as long as it is fluid.

Bob

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Re: Slide mounting media for a beginner - observations

#7 Post by Hobbyst46 » Fri Jun 07, 2019 8:05 pm

Sauerkraut wrote:
Hobbyst46 wrote: One difficulty that I encountered with nail polish (in fact, with all viscous media) is inclusion of tiny air bubbles. Microscopists on this forum have devised vacuum chambers for degassing in such cases.
For mounting plant parts (moss) I prefer PVA glue.
I am not seeing any air bubbles in the nail polish slides. Is it possible that a base coat nail polish is less viscous than regular nail polish?
Regretfully, regarding nail polish, I only know there is "ordinary" NP and "gel" (UV curable) NP. I used the former for mounting and the latter for sealing only, since it is too viscous.
However, warm NP is more fluid and less likely to hold stubborn air bubbles than cold NP. Also, shaking and swirling NP promote the formation of bubbles. Good to know that you obtained clear mounting, so success depends on proper application of the NP.
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Re: Slide mounting media for a beginner - observations

#8 Post by Sauerkraut » Sat Jun 08, 2019 3:05 pm

MicroBob wrote: One general hint: It is usually best to have the specimen close to the cover slip for best observation. So it is of advantage to prepare the specimen on the cover slip and keep the slide upside down as long as it is fluid.
Good to know. Thank you. It'll be interesting to see how long my slides with the different media last before they start to deteriorate.

Making my own slides has given me a definite appreciation for my set of Amscope prepared slides. Not all of them are presentable but a lot of them are decent, and to think they can mass produce those slides with acceptable results is impressive. It's not easy to slice samples thin enough and to stain them well enough to get decent slides from my failed attempts.

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Re: Slide mounting media for a beginner - observations

#9 Post by Sauerkraut » Sat Jun 08, 2019 3:11 pm

Hobbyst46 wrote:
Sauerkraut wrote:
Hobbyst46 wrote: One difficulty that I encountered with nail polish (in fact, with all viscous media) is inclusion of tiny air bubbles. Microscopists on this forum have devised vacuum chambers for degassing in such cases.
For mounting plant parts (moss) I prefer PVA glue.
I am not seeing any air bubbles in the nail polish slides. Is it possible that a base coat nail polish is less viscous than regular nail polish?
Regretfully, regarding nail polish, I only know there is "ordinary" NP and "gel" (UV curable) NP. I used the former for mounting and the latter for sealing only, since it is too viscous.
However, warm NP is more fluid and less likely to hold stubborn air bubbles than cold NP. Also, shaking and swirling NP promote the formation of bubbles. Good to know that you obtained clear mounting, so success depends on proper application of the NP.
I'm not really up on the finer points of nail polish but the L'Oreal base coat is not very viscous. That particular one seems to be discontinued but there must be other similar options available. I am careful not to shake the bottle and also remove any large air bubbles with tweezers under the stereoscope before landing the specimen in the medium.

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Re: Slide mounting media for a beginner - observations

#10 Post by mrsonchus » Sat Jun 08, 2019 7:00 pm

I think it may be easier and possibly even cheaper just to buy a small pot of 'Numount' resinous mountant from Brunel Micros?
An excellent and (relatively) fast-drying (to the 'able-to-use-slide' level) mountant with which I've made hundreds of permanently-mounted slides. Requires tissue dehydration via alcohol, to Xylene or Limone (e.g. 'Histoclear') of course.
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Re: Slide mounting media for a beginner - observations

#11 Post by MicroBob » Sat Jun 08, 2019 8:47 pm

For me it is important to learn to prepare slides, be it temporary or permanent. When microscopy consists of observing a set of ready made slides interest will faint soon. (Unless you have a really big slide collection of cause). It is like 3D-printing without the ability to design ones own 3D-models. I was told that the availability of prepared slides of good quality has dwindeled in the past decades in general.

To practise hand sectioning plant stems are nice, not too soft and not too hard ones, diameter 2-3mm. Then use a sharp rasor blade (the classic ones) and try to start thin and cut a wedge that get even thinner. Try to play microtome and deliver a firmly guided cut. Such a wedge will have an area where the thickness is just fine. It is fairly easy to pick the best cuts for further preparation so cut many sections. Remember that one section is always the result of two cuts , the one before counts too. It is possible to make a not too bad permanent slide in just a few minutes if perfection is not the goal.

Bob

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Re: Slide mounting media for a beginner - observations

#12 Post by Hobbyst46 » Sat Jun 08, 2019 9:41 pm

Sauerkraut wrote:... Not all of them are presentable but a lot of them are decent, and to think they can mass produce those slides with acceptable results is impressive. It's not easy to slice samples thin enough and to stain them well enough to get decent slides from my failed attempts.
AFAIK, mass produced slides are made by robots (automatic machines, not human robots... :) ).
Last edited by Hobbyst46 on Sat Jun 08, 2019 9:55 pm, edited 1 time in total.
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Re: Slide mounting media for a beginner - observations

#13 Post by mrsonchus » Sat Jun 08, 2019 9:55 pm

MicroBob wrote:For me it is important to learn to prepare slides, be it temporary or permanent. When microscopy consists of observing a set of ready made slides interest will faint soon. (Unless you have a really big slide collection of cause). It is like 3D-printing without the ability to design ones own 3D-models. I was told that the availability of prepared slides of good quality has dwindeled in the past decades in general. ........

Bob
Must agree wholeheartedly there Bob - the ability to make one's own slides gives the freedom to choose exactly what you would like to add to your selection, be it plant, animal or mineral etc.. It gives me enormous pleasure, engagement and satisfaction to now, after 1-2 years of endeavour, be able go to my garden, a hedgerow or a nice plant-anatomy book and choose a plant to make into slides. It's not easy , but as has been said, a start at a basic level is really within the reach of most everyone I think.
John B

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Re: Slide mounting media for a beginner - observations

#14 Post by MicroBob » Sun Jun 09, 2019 6:05 am

Hi John,
your slide making is on an extraordinary quality level and I always enjoy your posts where you openly explain your methods!

For the beginner it is good to understand that scientific value and personal enjoyment can already be gained from slides that are very crude compared to what the best slide preparators acheive. No mountant available - make a temporary slide with the specimen in water. No microtome - cut wedges by hand.... The lack of equipment, capability and free time shouldn't lead the beginning amateur to not start at all.

Bob

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Re: Slide mounting media for a beginner - observations

#15 Post by coltrane » Sun Dec 01, 2019 10:41 pm

Hi,
Interesting thread.
For a school project with sugar and steromicroscope, I need a mounting medium with the following characteristics:

- Permanent
- Not toxic
- Colorless: very transparent.
- Inert: which does not destroy or alter the composition or color of sugar.
- Slow or semi-slow drying: to allow the manipulation of sugar crystals and create simple patterns / drawings with a fine needle, to finally place the coverslip on top. You could also create the pattern / drawing on the slide. We are not sure whether to create the drawing first and then pour the mounting medium or create the drawing with the sugar and the mounting medium already mixed.

It is important that it be a permanent assembly, since the slides are kept in a school closet and it would be nice to see them within a decade. We have tested the dry assembly, but the crystals move and lose their position.
Maybe I ask too much ... excuse me.

Thanks

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Re: Slide mounting media for a beginner - observations

#16 Post by MicroBob » Mon Dec 02, 2019 12:44 pm

Hi,

in dry mounts you might fix the crystals to the slide with shellac. A very thinly mixed shellac in water free ethanol leaves a thin clear shellac layer on the cover slip. By heating the mount to 60°C you can soften the shellac so the crystals would stay in their pattern.
A different approach would be a natural resinous mountant like Euparal (solvent Isopropylalcohol) Another Idea would be UHU Hart glue which would solidify quickly.
The crystals will become invisible it there is not at least a difference in refractive index between crystal and mountant of 0,1.

One general problem with thick mounts like this is that when the solvent of the mountant evaporates air bubbles move between cover slip and slide from the edges.

Bob

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Re: Slide mounting media for a beginner - observations

#17 Post by coltrane » Mon Dec 02, 2019 9:41 pm

MicroBob wrote:
Mon Dec 02, 2019 12:44 pm
Hi,

in dry mounts you might fix the crystals to the slide with shellac. A very thinly mixed shellac in water free ethanol leaves a thin clear shellac layer on the cover slip. By heating the mount to 60°C you can soften the shellac so the crystals would stay in their pattern.
A different approach would be a natural resinous mountant like Euparal (solvent Isopropylalcohol) Another Idea would be UHU Hart glue which would solidify quickly.
The crystals will become invisible it there is not at least a difference in refractive index between crystal and mountant of 0,1.

One general problem with thick mounts like this is that when the solvent of the mountant evaporates air bubbles move between cover slip and slide from the edges.

Bob
Thanks for your reply Bob.
Undiluted sugar has a refractive index of 1.56.
Today we have tried different transparent non-toxic and solvent-free glues, but the sugar has diluted after a few minutes.
On the other hand, with superglue type acrylics sugar does not resist either.
¿Euparal damage sugar?
The option of mounting sugar crystals in non-dry permanent mode seems complicated, but we will continue testing.
However, dry assembly with Shellac (or nail polishes) seems the most reasonable but does not meet the characteristic of non-toxic and I am not sure that sugar resists.
We must experiment with nonpolar compounds.
Difficult matter!

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Re: Slide mounting media for a beginner - observations

#18 Post by Hobbyst46 » Tue Dec 03, 2019 11:58 am

coltrane wrote:
Mon Dec 02, 2019 9:41 pm
Undiluted sugar has a refractive index of 1.56.
Today we have tried different transparent non-toxic and solvent-free glues, but the sugar has diluted after a few minutes.
On the other hand, with superglue type acrylics sugar does not resist either.
¿Euparal damage sugar?
The option of mounting sugar crystals in non-dry permanent mode seems complicated, but we will continue testing.
However, dry assembly with Shellac (or nail polishes) seems the most reasonable but does not meet the characteristic of non-toxic and I am not sure that sugar resists.
We must experiment with nonpolar compounds.
Difficult matter!
Indeed, sugars dissolve to some extent in alcohols (and other polar solvents), so mounting sugar crystal within a thick layer of alcohol-based mountant seems unlikely to succeed.
Perhaps if the shellac layer is so thin that only a small surface area of the crystal touches it, mounting will work, but with the slight attachment, for how long will they stay bonded to the slide ?
BTW, shellac is not toxic and its solvent (ethanol) is also not toxic. They are quite harmless, unless used in high quantities in a confined, non-aerated space. Shellac has been used in the furniture industry for ages...shellac is a natural product, produced by insects.

That said, as you say, the best direction would be a non-polar stuff. But here is the catch: most non-polar compounds are more hazardous than ethanol or isopropanol.
Beeswax and paraffin wax could be used, if the slide can be kept on a hotplate and the crystals are arranged while the wax is hot. After cooling the wax solidifies. The problem is that solid wax is not very transparent.

Here is another, maybe crazy, idea: Try to mount the sugar crystals in a saturated solution of sugar in water (a syrup). When the solution is saturated the crystals will not dissolve. Although the mountant does not become solid, it is so viscous that objects will not detach or move. I would prepare a layer of the syrup on the slide, arrange the crystals with a needle then cover with a slip. Sealing with nail polish will prevent evaporation.
The potential problem with this idea is that the RI's of the crystals and mountant will be close to each other. So, as a preparatory step, must check if the crystals are at all visible...
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Re: Slide mounting media for a beginner - observations

#19 Post by MicroBob » Tue Dec 03, 2019 12:15 pm

The shellac is applied to the cover slip and fully dried. This allows to move the objects on the shellac layer. The bonding takes place by softening the dried shellac layer with heat. So the sugar crystals only have contact to shellac, not to ethanol. In diatom arranging a thin layer of skin fat is applied on top of the dried shellac to allow to slightly fix the frustules in place until the shellac is melted to let the bonding take place.

Bob

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Re: Slide mounting media for a beginner - observations

#20 Post by MicroBob » Tue Dec 03, 2019 12:39 pm

One more idea:
A strip of scotch tape could keep the arrangement together, act as a cover slip and is safe to use.

Bob

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Re: Slide mounting media for a beginner - observations

#21 Post by coltrane » Tue Dec 03, 2019 12:42 pm

Hobbyst46 wrote:
Tue Dec 03, 2019 11:58 am
Indeed, sugars dissolve to some extent in alcohols (and other polar solvents), so mounting sugar crystal within a thick layer of alcohol-based mountant seems unlikely to succeed.
Perhaps if the shellac layer is so thin that only a small surface area of the crystal touches it, mounting will work, but with the slight attachment, for how long will they stay bonded to the slide ?
BTW, shellac is not toxic and its solvent (ethanol) is also not toxic. They are quite harmless, unless used in high quantities in a confined, non-aerated space. Shellac has been used in the furniture industry for ages...shellac is a natural product, produced by insects.

That said, as you say, the best direction would be a non-polar stuff. But here is the catch: most non-polar compounds are more hazardous than ethanol or isopropanol.
Beeswax and paraffin wax could be used, if the slide can be kept on a hotplate and the crystals are arranged while the wax is hot. After cooling the wax solidifies. The problem is that solid wax is not very transparent.

Here is another, maybe crazy, idea: Try to mount the sugar crystals in a saturated solution of sugar in water (a syrup). When the solution is saturated the crystals will not dissolve. Although the mountant does not become solid, it is so viscous that objects will not detach or move. I would prepare a layer of the syrup on the slide, arrange the crystals with a needle then cover with a slip. Sealing with nail polish will prevent evaporation.
The potential problem with this idea is that the RI's of the crystals and mountant will be close to each other. So, as a preparatory step, must check if the crystals are at all visible...
Thank you very much for your comments I am a newbie in the permanent slide montage.
Excuse me, I confused the Shellac with nail polish. I will investigate about this gum.
We agree with the crazy idea: I had planned to mount the sugar in high-grade fructose syrup. The problem is that it is no longer sold, the current Karo is very low in fructose. I have only found it in China and in the Philippines but it must be bought in tons!
The other option is to buy fructose in the supermarket and prepare the mounting medium as indicated by Walter Dioni. But ... beginner's doubt ... the little water in the saturated solution: will it end up destroying sugar?
We will do the experiment. It is an economical method and we will know if their respective RI's allow to differentiate sugar from fructose and if sugar remains intact.
Other candidates are: beeswax and Damar gum.
Last edited by coltrane on Tue Dec 03, 2019 1:21 pm, edited 1 time in total.

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Re: Slide mounting media for a beginner - observations

#22 Post by coltrane » Tue Dec 03, 2019 1:09 pm

MicroBob wrote:
Tue Dec 03, 2019 12:15 pm
The shellac is applied to the cover slip and fully dried. This allows to move the objects on the shellac layer. The bonding takes place by softening the dried shellac layer with heat. So the sugar crystals only have contact to shellac, not to ethanol. In diatom arranging a thin layer of skin fat is applied on top of the dried shellac to allow to slightly fix the frustules in place until the shellac is melted to let the bonding take place.
Understood thank you!

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Re: Slide mounting media for a beginner - observations

#23 Post by Hobbyst46 » Tue Dec 03, 2019 1:27 pm

coltrane wrote:
Tue Dec 03, 2019 12:42 pm
Thank you very much for your comments I am a newbie in the permanent slide montage.
Excuse me, I confused the Shellac with nail polish. I will investigate about this gum.
We agree with the crazy idea: I had planned to mount the sugar in high-grade fructose syrup. The problem is that it is no longer sold, the current Karo is very low in fructose. I have only found it in China and in the Philippines but it must be bought in tons!
The other option is to buy fructose in the supermarket and prepare the mounting medium as indicated by Walter Dioni. But ... beginner's doubt ... the little water in the saturated solution: will it end up destroying sugar?
We will do the experiment. It is an economical method and we will know if their respective RI's allow to differentiate sugar from fructose and if sugar remains intact.
Other candidates are: beeswax and Damar gum. Oh, and work with Numount directly?
You are most welcome.
AFAIK the solvents for Numount are toluene or xylene - both much more hazardous than alcohols. In the end, it all depends on the quantities in use and aeration of the workspace. If you have access to a fume hood, then no problem with any mountant.
About Karo etc: which sugar crystals are to be mounted ? if sucrose (ordinary table sugar) I would prepare a solution of sucrose. If fructose - a solution of fructose. But very likely a saturated solution of fructose will not dissolve sucrose. Worth a trial IMHO.
I prepared a saturated solution of fructose myself, without Karo. Simply weigh the ingredients (water and sugar) into a small jar that can be stoppered tightly. Allow for air headspace. Place the jar in a hot water bath. Mix from time to time. Continuous magnetic stirring is best if available. With patience, the syrup is obtained. Cool, weight and check for undissolved crystals. Anyway, the liquid above such crystals is saturated so ready for use. Fructose is very cheap.
These syrups are transparent, only slightly yellowish, and are resistant against mold and bacteria.

But ... beginner's doubt ... the little water in the saturated solution: will it end up destroying sugar?Honestly, I do not know the answer.
Another concern is long term storage: if the slides are stored at a temperature that is significantly different from that of the preparation temperature, crystals might dissolve - or new unwanted crystals might precipitate.

Please let us know which mounting method worked for you !
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Re: Slide mounting media for a beginner - observations

#24 Post by Hobbyst46 » Tue Dec 03, 2019 1:31 pm

P.S. since Numount is a replacement for Canada Balsam, I would check about its RI.
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Re: Slide mounting media for a beginner - observations

#25 Post by coltrane » Tue Dec 03, 2019 1:52 pm

Hobbyst46 wrote:
Tue Dec 03, 2019 1:27 pm
AFAIK the solvents for Numount are toluene or xylene - both much more hazardous than alcohols. In the end, it all depends on the quantities in use and aeration of the workspace. If you have access to a fume hood, then no problem with any mountant.
About Karo etc: which sugar crystals are to be mounted ? if sucrose (ordinary table sugar) I would prepare a solution of sucrose. If fructose - a solution of fructose. But very likely a saturated solution of fructose will not dissolve sucrose. Worth a trial IMHO.
I prepared a saturated solution of fructose myself, without Karo. Simply weigh the ingredients (water and sugar) into a small jar that can be stoppered tightly. Allow for air headspace. Place the jar in a hot water bath. Mix from time to time. Continuous magnetic stirring is best if available. With patience, the syrup is obtained. Cool, weight and check for undissolved crystals. Anyway, the liquid above such crystals is saturated so ready for use. Fructose is very cheap.
These syrups are transparent, only slightly yellowish, and are resistant against mold and bacteria.
Please let us know which mounting method worked for you !
It is ordinary table sugar (sucrose). Ok, we will do the fructose test.
The Numount is out, not suitable for school work.
Thanks again!

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