[Nhcoll-l] Preserving frozen fish

[John E Simmons]

I suggest you separate this into two issues, (1) preserving tissues for DNA
and (2) preserving the whole fish as a fluid preparation.

For DNA preservation, I think that the best procedure will be to remove a
tissue sample from the frozen specimen and store the sample at -80C (for
example, in liquid nitrogen). Ultracold temperature is a much better
long-term preservative than 100% ETOH.

The problems with preserving directly in 100% alcohol are that:
(1) 100% ETOH is chemically dehydrated and may have traces of the dehydrant
chemicals in it that may interfere with future molecular work (not to
mention that 100% ETOH is more expensive than 95.6% ETOH).
(2) As the alcohol penetrates the specimen it dehydrates the tissues, which
impedes the penetration of alcohol to deeper tissues. Using 70% alcohol
means less dehydration of the tissues (allowing deeper penetration) while
still providing a good biocide to protect the specimen during the
preservation process.
(3) Once the specimen is preserved you will need to change the alcohol
anyway because it will have become diluted with water extracted from the
specimen. The chance of diluting the 100% ETOH to 70% by extraction of
water from the specimen is slim, so there is no real gain in starting with
100%.

Preservation in 70% ETOH without using a formaldehyde fixative works fine
as long as the alcohol can get deep enough into the specimen. This is why I
recommend thawing the specimen in 70% ETOH and injecting the alcohol or
cutting into the specimen as it thaws to get the alcohol deeper into the
specimen, and after 24 hr to change the specimen to fresh alcohol. Using
100% or even 95.6% alcohol will produce a more dehydrated specimen, not a
better preserved one, and is not likely to produce very good DNA.

If you thaw the specimen in a 1:9 buffered formaldehyde and water solution,
the recommendation to inject the fixative into (or cut into) the specimen
as it thaws still holds, but you can still get good results even if the
formaldehyde mixture becomes diluted (formaldehyde penetrates better than
alcohol).

Formaldehyde has only been available since the mid 1890s, so specimens
preserved prior to that were preserved directly in alcohol. It is
instructive that almost all of the pre-formaldehyde instructions for
preservation call for the use of alcohol at a concentration below full
strength and to change to fresh alcohol preservative after 24 hr.

I hope this information is helpful.

--John

John E. Simmons
Writer and Museum Consultant
Museologica
*and*
Investigador Asociado, Departamento de Ornitologia
Museo de Historia Natural, Universidad Nacional Mayor de San Marcos, Lima


On Tue, Aug 19, 2025 at 2:13 PM Rebecca Abrams <ridabrams at berkeley.edu>
wrote:

> Hello all,
> I have an ever increasing number of frozen fishes coming to me from a
> local aquarium that I will be preserving in either ethanol or formalin then
> ethanol. Our usual protocol is to use 100% ethanol for reasons of DNA
> preservation (I know this is an unusual protocol) if the specimen will fit
> into one of our jars, otherwise it goes into formalin then into 70% ethanol.
> Both ASIH's online resources and John Simmons's *Fluid Preservation* say
> to avoid preserving frozen specimens and if one has to then to thaw them in
> the preservative and inject with preservative as they thaw.
>
> If anyone has tips and tricks for this in particular for fish that would
> be very helpful. I have some rather large friends waiting for me to prep
> that I would like to not ruin by turning them to mush first. I know that I
> can skeletonize them, but I would very much like to have at least one in
> fluid.
>
> Thank you,
> Rebecca Abrams
> MA, M.Ed
> Collections Manager- Fishes
> Museum of Vertebrate Zoology
> 3101 Valley Life Science Building
> University of California, Berkeley, CA, 94720
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