[Histonet] VIP issue

[Rene J Buesa]

Gudrun:
I have used/recommended for 10 years now graded isopropyl (2-propanol) alcohol to dehydrate and before paraffin ONLY and when I went to Australia in 2011 found out that they had been using the same protocol for years.
You just grade 2-propanol in the same way you grade ethanol (60%→80%→90%→100%) and continue DIRECTLY to paraffin.
Usually I dedicate one "hot" (paraffin) VIP station with a 1:1 of 2-propanol + paraffin mixture to facilitate the infiltration or (better yet) one "hot" station with a 1:1 mixture of 2-propanol + pure mineral oil (= paraffin of low molecular weight) and the infiltration is really excellent.
In this way you will be able to eliminate BOTH xylene and any other "friendly" clearing agent (ALL are toxic lat different evels). 2-propanol is NOT toxic AT ALL!
I have ALSO stopped using xylene to de-wax sections before staining and substituted it with a 2% aq. sol. of dishwashing soap and go to water → staining after (I can send you the detailed protocol / publications).
AFTER staining I just rinse the stained slides in 2-propalo → shake it well → to dry oven at 60ºC for 10 minutes → coverslip.
My lab was TOTALLY xylene free, and you can do the same thing. If you are hesitant just TRY these methods with only few blocks / sections / stained sections and convince yourself.
Best regards
René 

    On Friday, October 12, 2018 1:13 PM, Gudrun Lang via Histonet <histonet at lists.utsouthwestern.edu> wrote:
 

 Thank you for all the good hints and advices.

It has to be a reagens-issue, because the second instrument was also
concerned. 
But we had no similar problems for decades, although we use 96% as
cleaning-ethanol after ShellSol. 
We change the reagenses after 5 runs, once a week. 

We soon are forced to end using ShellSol, because the company stops the
production. We don't like to switch to xylene. 
Which xylen-substitutes are recommended for the VIP besides of Tissue-Clear
(Sakura)?

Best wishes
Gudrun 


-----Ursprüngliche Nachricht-----
Von: Gudrun Lang via Histonet [mailto:histonet at lists.utsouthwestern.edu] 
Gesendet: Donnerstag, 11. Oktober 2018 18:58
An: histonet at lists.utsouthwestern.edu
Betreff: [Histonet] VIP issue

Dear all!

I have a question for those, who are familiar with the VIPs from Sakura. 

Last time we changed the reagenses the cleaning-ethanol (96%) was very milky
and even was full of many small particles. It was a paraffin-soup.

What is the cause for such a case?  We have been using the organic solvent
(ShellSol) for decades as xylensubstitute. 

Maybe the quality has suffered and the ability of solving paraffin has
decreased. But are there other explanations? Maybe a malfunction of the
instrument?

 

Thanks in advance

Gudrun Lang

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