[Histonet] Soaking artifact

Mon Jan 6 07:18:43 CST 2014

With respect, this may work in a non-industrial setting, but in Pharma, with high through-put, it is generally not possible to have individualized processing programs for a lot of different tissues.  You have to find one or two that work for the majority and compensate with some soaking.  Experience will help you know which tissues can't get left on too long.  Again, you can usually get past areas that look over-soaked just by cutting through them as you also mentioned. Just be careful, everyone, with small lesions so you don't ace through them.

Phil.
Philip Manfre, B.A., HT (ASCP)
Associate Principal Scientist
Merck Research Laboratories
WP45-251
PO Box 4
West Point, PA 19486

215-652-9750
215-993-0383 (fax)
philip_manfre <@t> merck.com

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Lee & Peggy Wenk
Sent: Monday, January 06, 2014 8:07 AM
To: Deanna Leslie; histonet <@t> lists.utsouthwestern.edu
Subject: Re: [Histonet] Soaking artifact

The only "soaking" artifacts that I can think of would be caused by:
- soaking too long in water (minutes instead of a few seconds)
- soaking under-processed tissue in water

In both cases, the tissue is supposed to be "protected" by the wax, and if 
it is not (under-processed), or if the faced block is in water too long, the 
tissue can start re-absorbing water. The tissue then turns white and swells 
out of the block. So all that "swelled out" tissue is cut away and lost when 
the tissue is put back on the microtomy for sectioning the ribbon.

If you soak for just a few seconds, such as a gauze with water being held 
against the block on the microtome, after it has been faced, then you will 
get a little bit of water absorbed into just a few layers of cells. Just 
enough to cut 2-4 sections. And you won't see that swelling artifact.

For those of you saying - "but I have to face all the blocks, put them back 
on ice and/or water while I cut a bunch more blocks, and then go back and 
cut each block" - that is an artifact also. You have over-dehydrated your 
tissue during processing, and you are putting back the water that you should 
not have taken out. Processing is supposed to remove the unbound water (not 
attached to proteins), and some of the bound water (attached to proteins), 
and leave some of the bound water (attached to proteins) in the tissue. If 
you HAVE to soak EVERY block for more than a couple of seconds, then you are 
wasting time rehydrating and wasting time while microtoming. Cut down the 
time in the alcohols on the tissue processor, to leave a little bound water 
in the tissues. And you can NOT processing little biopsies on the same long 
processing cycle as the larger pieces of tissue (uterus, breast, etc.). 
Those little biopsies will be over-dehydrated. They HAVE to be run on a 
separate cycle of much shorter time intervals (10-20 minutes in each 
solution (once fixed), instead of 45-60 minutes in each solution).

You should be able (on nearly every tissue block) to rough trim the tissue, 
and immediately start cutting ribbons. Possibly, you will need to put an ice 
cube on some of the harder tissue (cervix, uterus, bone, lens, etc.) just to 
get the paraffin hardness to match the hardness of the tissue. That being 
said, some tissues are naturally brittle or crumbly, and always need some 
water put back in the tissue, such as spleen or bloody tissue, but again, 
some wet gauze on the faced block for a few seconds should be enough time to 
get 2-4 sections. And that's all the tissue we usually need from those 
blocks. If you need more for IHC, put the wet gauze back on the faced block, 
and cut a few more sections.

Peggy A. Wenk, HTL(ASCP)SLS

-----Original Message----- 
From: Deanna Leslie
Sent: Sunday, January 05, 2014 4:42 PM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Soaking artifact

Has anybody in histoland ever heard of this?  I have been cutting tissue
for 25 yrs and until recently I had never heard of this!
I am under contract to a facility and the supervisor there does not want
anybody to soak their tissue or use ice!  Your are supposed to use the cold
plate, because as I have stated soaking them causing an artifact. I have
not disputed this because it is not my place or in my job discription as a
traveler.  I am not even sure what it is supposed to look like or what type
of problems it causes.

Thanks for listening!
Deanna Leslie HT ASCP
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