It was just pointed out to me in a private message that there are many
different kinds of EDTA. Looking in the Sigma catalogue, I count 16
different species (?) of it.  I use the di sodium salt.  This person who
contacted me also recommended using ACETIC ACID to adjust pH with rather
than HCl.  I use HCl because that's the way I was trained to do it. 
Considering that EDTA is a tetraacetic acid, maybe I should rethink my
practices... acetic acid seems to be more logical... and yet I've had
wonderful success with the formula that I recommended (included below). 
I'll have to try acetic acid and see how this works.

Connie McManus 



Connie McManus wrote:
> 
> I just completed a whopping big project of adult trout heads (looking
> for whirling disease) which EDTA decal was necessary.  Below is my
> method for preparing 1 liter of EDTA decal solution.
> 
> I.  Prepare a PBS solution.  I used Na phosphates.  I cannot vouch for
> how K phosphates will perform in EDTA.
>         dibasic.... 6g/liter DI water
>         monobasic.... 4.5g/liter DI water
>         NaCl ........ 8.1g/liter DI water
> 
> In approx. 700 mL of PBS, add 100 g EDTA and begin stirring. Bring pH up
> to 7.2-7.4 by adding pellets of NaOH (this may take aprox. 5 g or more
> to do).  Use either a pH meter or litmus paper to determine the pH.  I
> used litmus paper.  After the EDTA is COMPLETELY dissolved, add DI water
> qs to 1 liter.  If you overshoot the pH (more alkaline that 7.40),
> adjust with HCl. If you use conc HCL, you can do it with just about 1 or
> 2 drops (the voice of experience).  Try to NOT overshoot the pH, but if
> you do, minimize the amount of HCl you need to use. If you overshoot big
> time, just add more EDTA/PBS to the mix and go from there. I made my
> solution up in huge quatities (2 x 5 gallon containers full), so I just
> dumped the NaOH in by the spoonfulls (each spoonfull weighs 11.5g), but
> if you're just making up 1 or 2 liters, be carefull with the NaOH.
> 
> This whirling disease project was the first time I had ever used EDTA as
> a decal solution and I found that i really like it.  The tissues take a
> bit longer to decal, but the soft tissues are preserved much nicer than
> my regular decal solution and they also cut softer.  I also found that
> for those tissues that have really hard crunchies in them (like a kidney
> with kidney stones), soaking in EDTA decal solution for about 1-2 hours
> really softens them up and they cut like butter. I love this stuff.
> 
> Hope this helps.
> 
> Connie McManus
> 
> jane weeks wrote:
> >
> > Histonetter's -
> >         Does anyone have a protocol for making EDTA - based decalcifying solution?
> >
> > Thanks!
> > Jane Weeks
> > McArdle Lab for Cancer Research - UW
> > Madison, WI
> 
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