Phenol Protocol

David L. Haviland, Ph.D. dhavilan at IMM2.IMM.UTH.TMC.EDU
Mon Feb 3 11:48:55 EST 1997


At 17:54 1/31/97 GMT, Paul N Hengen wrote:
>I burned my hands once with phenol, and therefore will never again distill
>or extract large volumes of it. Be very careful if you do this and make
>sure you are using the correct tubes, especially if you decide to centrifuge
>to separate phases. Cleaning up a phenol spill in a centrifuge rotor is no
>fun, and if you leave any phenol in there without knowing it, you risk
>burning other people when they use it. Now I just buy phenol from Life
>Technologies or Boehringer and use it directly from the bottle. There
>was an article somewhere (BRL Focus?) that compared unextracted phenol
>with that extracted with Tris buffer, water, etc. and the conclusion was
>that the water worked just fine for DNA. I think this is the reference,
>but I'm not sure...

I "second" Paul's comments.  I,too, had a mis-hap with phenol, a somewhat
bigger one I might add.  Despite having gloves, etc.,  I was removing a
freshly prepared 500g bottle from a shoulder-height shelf in the 4'C room.
Even though I had a hold on the bottle, the condensation on the outside
provided enough lubrication that it just slipped right through my hands.
The safety floor unfortunately worked to my disadvantage - instead of
having the bottle shatter as I would have prefered, the energy of the fall
rebounded from the rubber mat back to the phenol and not the bottle.  The
phenol blew the lid off (in 4 pieces I might add...)  and I received a
shower of phenol.  The bottle was still intact.  I saw what was about to
happen so I was at least able to turn my head away.  Fortunately in long
sleeve shirt and pants, they took the brunt of it but I none the less was
in the ER getting my face, arm, one eye, and chest washed extensively with
what was the coldest water and saline... (it was in the mid west in
January...)  I couldn't shave for a week and had both arms wrapped in gauze
for 3-4 days....  then about a 5-7 days later, it peeled like the worst
sunburn you can think of...  

I now keep phenol on the lowest shelf possible and deal with as small a
quantity as possible.   When I read of folk redistilling phenol, that alone
gives me cause for gastric distress.  I have also made it a point that no
matter how hot it is, shorts and open toed shoes are never worn in the lab
by any personel.
And I do still use gloves... ::grin::

Although I do still equilibrate my phenol, I would be interested in
pre-equilibrated material.

I too have used water saturated phenol for DNA and have observed very
little if any loss of material. Yet the methods books claim that phenol at
pH 8.0 is absolutely necessary.  However, I have tried using Tris buffered
phenol in the acid-phenol protocol for RNA isolation and it simply doesn't
work...

Hope this is of help,
David


=============================
 David L. Haviland, Ph.D.
 Asst. Prof. Immunology 
 University of Texas - Houston, H.S.C.
 Institute of Molecular Medicine  
 2121 W. Holcombe Blvd.  
 Houston, TX  77030 
 Internet:"dhavilan at imm2.imm.uth.tmc.edu" 
 Voice: 713.500.2413  FAX: 713.500.2424
 ------------------------------------------------------  
" Sometimes you're the windsheild, sometimes you're the bug."
=============================




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