An alternative to Paraffin Wax for sectioning specimens

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Dave S
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An alternative to Paraffin Wax for sectioning specimens

#1 Post by Dave S » Wed Aug 21, 2019 4:05 pm

Although Paraffin wax has been a tried and trusted method of wax embedding for many years, it is a byproduct of oil refining in the production of Petroleum.

As such it does contain harmful chemicals, that are released when burned (as in candles). It releases Toluene, and Benzene, both known Carcinogens.

Of course for wax embedding, we would only melt it, not burn it. However, it seems that there may be a totally safe alternative, in the form of Beeswax.

The question is, has anybody here actually tried it, and if so, was it successful?

It is a cheap, and readily available commodity.
Suffolk, UK

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MicroBob
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Re: An alternative to Paraffin Wax for sectioning specimens

#2 Post by MicroBob » Wed Aug 21, 2019 4:44 pm

Hi Dave,
an other alternative is glycerin soap, at least for simpler tasks. This has been in use in the beginning of microtomy, mid 1800s, but has been forgotten since. The big alternative is that you don't need Xylene to remove the embedding material, but just water. In my view the solvents are much more problematic than the traces of dangerous stuff in paraffine.

Bob

Dave S
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Re: An alternative to Paraffin Wax for sectioning specimens

#3 Post by Dave S » Wed Aug 21, 2019 6:32 pm

Not heard of Glycerin Soap Bob.

We now live in a world of well publicised health and safety issues, and reagents such as Xylene, Toluene, Benzene, and Formaldehyde etc, are now proven, and classified hazards.

That said, used in the quantities that an experienced amateur microscopist would use, probably not a significant issue. A different situation where children are concerned, who would be unlikely to be aware of the hazards. Hence governments placing restrictions on the availability of such substances.

I can remember as a schoolboy, being told about the wonder material that is Asbestos, even my Mum's ironing board had a square of it, to stand the iron on. Now we know just how deadly it can be, and too late for many. :o

Apparently, its not the dangerous stuff contained within Paraffin Wax, but when it is released during combustion. As I said, I we are only going to be melting it, so it shouldn't be a problem.

That said, Beeswax does apparently offer a good alternative.
Suffolk, UK

Brunel SP100 (with 4x, 10x, 40x,60x, and 100x (oil) plan objectives), and Canon EOS 4000d Camera (microscopy use only)

Hobbyst46
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Re: An alternative to Paraffin Wax for sectioning specimens

#4 Post by Hobbyst46 » Wed Aug 21, 2019 7:52 pm

DaveS wrote:...
The first comparative study of possible replacement of paraffin with beeswax for histology - NOT botany - is recent - believe it or not:

International Journal of Orofacial Biology
ORIGINAL ARTICLE
Year : 2017 | Volume : 1 | Issue : 1 | Page : 13-15

Impregnation and embedding using bees wax and paraffin wax in oral tissue samples: A comparative study

J Vini Mary Antony, Pratibha Ramani, N Anuja, Herald J Sherlin, S Gheena, R Abilasha, Gifrina Jeyaraj, KR Don, S Archana
Department of Oral and Maxillofacial Pathology, Saveetha Dental College, Saveetha University, Chennai, Tamil Nadu, India

Reading fast through it, I saw that the authors claim that the two substances are fairly similar to each other, in several respects.
However, I think that they performed the same step of dehydration before embedding. That could mean (though not expicitly stated) that they used xylene in both cases.

I do not know if xylene is absolutely necessary, but if it is, then in agreement with MicroBob's comment above, I would be concerned with it more than by any potential toxicity of vapors released by the waxes. Alternatively, I would check out if anyone has yet demonstrated that vapors from molten beeswax (at a temperature of 62-67C or so) are more hazardous than vapors from molten paraffin (at 45C, say)or vice versa. In my humble opinion, the fact that a substance is a natural product rather than synthetic does not guarantee that it is always safe for a specific use.
These comments are not meant to un-motivate anyone to try embedding in beeswax.
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Roldorf
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Re: An alternative to Paraffin Wax for sectioning specimens

#5 Post by Roldorf » Wed Aug 21, 2019 8:17 pm

Yes I agree, natural products are not necessarily safe. Asbestos is a naturally occurring element. As long as we leave it in the ground we shouldn't worry too much about it.
Location: Northern Germany

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MicroBob
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Re: An alternative to Paraffin Wax for sectioning specimens

#6 Post by MicroBob » Wed Aug 21, 2019 8:57 pm

I think it is always good to look for safer and environment friendly methods, but in this case I don't really get the point. We would try to avoid to melt paraffin occasionally while we atually burn it in higher amounts in form of candles, most from dubious sources. On my personal priorty list of avoidable dangers this would rank last place. For children this is not relevant anyway as they wont like to spend the time and effort needed for slide from a paraffin embedded object.
If somebody is interested in paraffin embedding, this is a proven method that avoids dangerous chemicals: http://www.klaus-henkel.de/isomethode.pdf

Beeswax: Alternative materials would have to prove themselves as dependable to used as paraffin. Here a natural material has more variation. I have never done any paraffin embedding so far, but my guess is that beeswax would give trouble. From the feel of it I would say that it is a mixture of different substances, some more oily. It might be perfect if one wishes to drive an experienced microscopists crazy! :lol:

Bob

MichaelG.
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Re: An alternative to Paraffin Wax for sectioning specimens

#7 Post by MichaelG. » Fri Aug 23, 2019 11:35 am

Given that Paraffin is a major constituent of Epidermal ointments / emollients, I think the toxicity concerns are probably being over-thought [in the context of embedding].

MichaelG.
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Lab-Rat
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Re: An alternative to Paraffin Wax for sectioning specimens

#8 Post by Lab-Rat » Thu Aug 27, 2020 9:06 pm

Hi. I just discovered your forum and this thread and I felt I had to comment.
I'm the manager of a large diagnostic histology lab and I've been involved with histology and microscopy for 30 years. I've seen a few changes over the years and I can tell you a little about beeswax.
Beeswax as an embedding medium was broadly considered to be too soft to give adequate support for animal (and human) specimens. Paraffin wax, in its most common form, was a rather harder material but was consequently harder to cut and many hours were lost on sharpening microtome knives before the advent of disposable blades. In Western Europe and in North America, synthetic additives were mixed into the paraffin to soften it a little for histology, a situation that remains till today. In Eastern European countries, where access to reagents was limited for a long period, labs sometimes mixed their paraffin with beeswax for the same effect. This was still being done quite often until about 20 years ago. Since that time standardized protocols for immunohistochemistry and molecular biology have meant that beeswax had to go as it contained various proteins as well as RNA and DNA that can really mess up your results.... Pity really, it smelled good.
As for toxicity, histology grade paraffin is barely toxic at all. The lethal dose is very high and you'd need to eat well over a kilogram of it for a worse problem than having to spend a few days near the toilet.
Thanks again for a fascinating topic.... I may just stick around.
Matt

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