[Nhcoll-l] fluid specimens and iodine staining

Sat Jul 1 04:44:52 EDT 2017

Hi Christopher,

this is a rather theoretical reply after looking into available 
literature which you surely also consulted (e.g. Butters et al 2014 & 
Balint et al 2016 on Plos One).

Both papers basically say that the staining results are hard to predict 
and largely dependent on perfusion rates and tissue thickness (which of 
course is no surprise but pure chemistry and physics). Obvious issues 
effecting the scan results are over-staining or under-staining, and the 
optimal incubation time is hard to predict and depends on specimens 
size, tissue composition and - surely too - deposits in and on the 
specimens (the cholesterol you already noticed surely is one of the most 
obvious ones).

Other influencing factor are is the x-ray source and data recovery.

Personally (gut feeling rather then proofed experience), exposure to 
water based solutions for weeks or even months surely damages the 
specimens and weakens the tissues, and my next question would be what 
happens will all those (dissolved) fats in the specimens if they are 
immersed in water for such a long time. Also, Balint et al (2016) report 
that over-staining can lead to deformation and tissue shrinkage .

The other method you mention, exposure to nearly 100% EtOH has just the 
effects that you describe, and the question for me is what would happen 
especially to those tissues near the body surface (e.g. in fish 
increasing the EtOH strength may cause cell rupture in the scale pockets 
and consequently may loosen the fish scales).

So it might be worth to explore two alternatives:

1. It might be worth to compare the imaging capacities of different 
ct-scanners; colleagues at the MNHN Paris for example used the 
synchroton in Geneva for imaging of or Lungfish larvae with amazing results.

2. You mentioned that targeted specimens in your collection were 
preserved +100 yrs, some probably used pre-formalin fixatives. If this 
is the case, it might be worth to test if any mercury salts have been 
added. Balint et al (2016) mention, that "Mercury (II) chloride provide 
the best contrast between different soft tissue types".

Around 1900, Mercury salts were not rarely added to enhance the "fixing" 
properties of the ethanol. Perhaps some of your specimens have already 
been treated with a contrast enhancing agent?

Hope this helps, even though it is not a direct answer to your question.
All the best
Dirk

Am 30.06.2017 um 19:59 schrieb Milensky, Christopher:
>
> Hello,
>
> I am hoping that some folks in the NHCOLL community have experience 
> with iodine staining and would be willing to share their knowledge.  
> We are receiving more and more requests to use our fluid specimens 
> (birds) for CT scanning projects, some of which require staining.  In 
> the recent past, we have been asked to approve two different staining 
> techniques.  One is a ‘water’ based staining methodand the other an 
> ‘ethanol’ based method.  We would like to hear opinions about which 
> method is best for the short and long term care of the specimens.  As 
> with any old collection, the exact methods of fixation used over the 
> last 100+ years have been variable, but typically our birds were fixed 
> in formaldehyde and then moved to storage in 70% ETOH.  I’m sure some 
> of our oldest specimens were never properly ‘fixed’ since formaldehyde 
> did not come into use until the early 1900’s.
>
> The two methods:
>
> I2KI (potassium iodine and iodide) is dissolved in water and makes a 
> naturally antimicrobial solution in which the specimen is removed from 
> ETOH storage and soaked for weeks to months, depending on the size of 
> the animal, in this water solution.  Pickled tissues (fixed in 
> formalin and preserved in 70% ethanol) should remain pickled even 
> after beingremoved from preservative due to the chemical changes that 
> occur during fixation, and iodine prevents bacterial growth, but it 
> still involves putting a pickled specimen in water for an extended 
> period.
>
> I2E is elemental iodide dissolved in 100% ethanol (200 proof), which 
> raises another set of issues for specimens stored in lower 
> concentrations of ethanol. Increasing the concentration of ethanol 
> further dehydrates and shrinks the specimen, but it has been found to 
> be the most effective concentration of ethanol for staining soft tissues.
>
> So, the question is:  Would you rather have a specimen removed from 
> 70% and stored in a water/iodine solution for weeks/months or stored 
> in 100% ETOH for a similar period or neither?  What are the pro/cons 
> of each?  Would you allow a specimen collected pre-1910, and 
> presumably not fixed, go into water/iodine for a month or more?  Are 
> you aware of other methods?
>
> I look forward to hearing your thoughts!
>
> Chris
>
> *Christopher Milensky*
>
> Collections Manager
>
> Division of Birds 
> <https://urldefense.proofpoint.com/v2/url?u=http-3A__vertebrates.si.edu_birds_&d=DwMFAg&c=cjytLXgP8ixuoHflwc-poQ&r=LpYc_Z_iN1KRw0hheb3x6-8MJUMu482qfHowpGYJqwc&m=_ng57dMSlpRtwx104LoLynttAJTY-hHusbDzZvPHtcM&s=GUIwF2jcK-a5sAF-jpiFUFCe2_UmI54Q-akmUeSa6xE&e=>
>
> *w*202.633.0794 milenskyc at si.edu <mailto:milenskyc at si.edu>
>
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