[Histonet] Formalin fixation of cells for paraffin embedding?

[Henry, Charlene]

We process a lot of cell lines for research and have found that if you
intend to perform IHC or ISH on the FFPE cells, the following protocol
works great. The PBS washes the cells of excess medium used to grow the
cells and yields clean IHC. The Hist Gel yields a cell pellet with an
even distribution of cells through-out without the cells being clumped
together. 
1. Suspend cells in 10% NFB to fix cells.
2. Spend down suspended cells and draw off 10% NBF.
3. Wash cells in PBS by adding 2-3 ml of PBS and re-suspend cells with
vortex.
4. Spend down cells and draw off PBS.
5. Wash cells in PBS one more time as in step 3 & 4.
6. Add volume of Histo Gel equal to the amount of cells and re-suspend
cells with vortex.
7. Allow gel time to become firm (usually 5-10 minutes.
8. Remove gel pellet carefully, cut in half, place in lens paper, and
process. 

Hope this helps because we tried multiple protocols before this one.

Thanks,

Charlene Henry, HT QIHC
Histology/IHC Section Head
Department of Pathology
St. Jude Children's Research Hospital
332 North Lauderdale
Memphis, TN  38105
Phone: 901-495-3349
FAX: 901-495-3100


-----Original Message-----
From: Bill Blank [mailto:bill501 <@t> mindspring.com] 
Sent: Friday, May 21, 2004 9:55 AM
To: histonet <@t> lists.utsouthwestern.edu
Cc: histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] Formalin fixation of cells for paraffin
embedding?

At 8:52 AM -0500 5/21/04, Marsha Linville wrote:
>Wrap the pellet in lens paper,and put it in a cassette to fix and
process
>with your tissues.

We use those papers the hairdressers use when curling hair. Much 
cheaper than lens paper.

Bill
-- 
______________
Bill Blank, MD
Heartland Lab, Inc

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