[Histonet] DNA extraction from Tissue Blocks

[Monson, Frederick]

Afternoon Phyllis,
	Your problem may be that almost all methods are hydrolytic and
the 'force' needed for a specimen will vary with the degree of
crosslinking of the DNP (deoxyribonucleoprotein).  In methods for in
situ hybridization, the DNA must be exposed in order to be 'melted'.
This often requires, for HCHO-fixed material, that the DNP protein be at
least disrupted.  Pronase is most often suggested.  As it is a very
active peptidase, each batch/lot will behave differently AS will each
specimen.  Thus, it is almost always necessary to perform an empiric
test that establishes the concentration of pronase and time of action.
i.e., I have discovered no easier way, though I have not done ISH for 4
years.  The problem is to remove the RNP with pronase and then the DNA
with dnase without removing most of the histology in the process.

If I were looking for more selective methods to expose DNA, I would look
to enzymes that modify histones.  Look here:

http://web.wi.mit.edu/young/pub/histone_modifiers.html

Hope this helps,

Fred

 Frederick C. Monson, PhD
Center for Advanced Scientific Imaging
Mail to Geology
West Chester University of Pennsylvania
Schmucker II Science Center, Room SS024
South Church Street and Rosedale Avenue
West Chester, PA, 19383
Phone/FAX:  610-738-0437
eMail:  fmonson <@t> wcupa.edu
CASI Page and Scheduling
	http://darwin.wcupa.edu/CASI/


-----Original Message-----
From: phyllis [mailto:pxs76 <@t> po.cwru.edu] 
Sent: Wednesday, September 24, 2003 9:28 AM
To: Histonet <@t> pathology.swmed.edu
Subject: [Histonet] DNA extraction from Tissue Blocks




I would like to try using a DNA extraction method on formalin fixed
tissue 
blocks. Can anyone recommend a method or kit? we  have had difficulty
with 
consistent results.


Phyllis Scalzo, HT(ASCP)
Case Western Reserve University
2085 Adelbert Rd.
Cleveland, Ohio 44106
Ph: 216-368-0822
Fax: 216-368-2546


_______________________________________________
Histonet mailing list
Histonet <@t> lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet