[Histonet] Isopropyl Alcohol in the histology lab

[Rene J Buesa]

Teri:
It is a wise decision to substitute EthOL with Isopropyl alcohol (IPA) and to your questions:
1- although it is not advisable to keep tissues for long periods of time in EthOL sometimes this is done. EthOL will sometimes make the tissue brittle mostly because its chemical properties but IPA dehydrates more gently and is a larger molecule that EthOL so, if you use to keep tissues in 70EthOL you will be able to leave them in 70IPA more safely.
2- with a different dehydrating quality you will probably need to adjust your times in the auto-stainer or in your hand staining sequence. Your best option though is to totally eliminate xylene and the alcohols in your staining sequence. How? Dewax sections with 2% aq. dishwasher soap at 90ºC → distilled water → staining protocol → dry your stained sections in an oven for 5 minutes at 60ºC → coverslip as usual, but I am sure this may seem "too drastic" for you but I can assure you it works. Ask me if you want to.
3- Immnuno reactivity depends on how the tissue is FIXED and not in how it is processed. Had processing have any impact there would be scores of different protocols for all those who dehydrate or clear in different reagents. The BondMax IHC stained was developed in conjunction with the Peloris instruments and this later instrument was able to use EthOL, later IPA and even it has a "dry-out" step before infiltration (which I am not very fond of).
4- IPA is a safe substitute to EthOL but you have to consider other aspects such as costs and availability. Some labs lave "alcohol licenses" allowing them to use "200 grade EthOL", other can use only denatured EthOL but IPA will be always "pure" (about 98-99%).
Recycling is also an issue: I never recycled EthOL because it takes much more time and energy than recycling xylene, which I used to recycle. You will have to determine if recycling IPA is worth the cost and time.
René J.


________________________________
From: Teri Johnson <TJohnson <@t> gnf.org>
To: "histonet <@t> lists.utsouthwestern.edu" <histonet <@t> lists.utsouthwestern.edu> 
Sent: Monday, August 12, 2013 5:18 PM
Subject: [Histonet] Isopropyl Alcohol in the histology lab


Dear colleagues,

I am entertaining the notion of switching over to isopropyl alcohol (IPA) for our tissue processing and have a few questions. I know the xylene-free processing and microwave processing protocols use this universally, but at this time I am only interested in getting rid of the ethanol/methanol/isopropyl alcohol blend.

1 - Are there any known issues with holding tissues in 70% IPA rather than 70% ethanol/reagent alcohol?
2 - Is there any impact on the stain line if you use it for H&Es and hydration/dehydration of histochemically stained slides?
3 - Have any of you who use IPA found any impact on immunoreactivity when compared to using other dehydrants?
4 - Is there anything else I have overlooked?

We have pure ethanol available to us if needed for particular stain applications.

Best wishes, and thanks in advance.

Teri Johnson
Manager, Histology
GNF - San Diego, CA
858-332-4752

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