[Histonet] Cytology CSF Cell Pellets made from Histogel

Kim Merriam kmerriam2003 <@t> yahoo.com
Wed Feb 22 07:23:15 CST 2012

I forgot to mention, we prepare these specifically for IHC; so yes, you can do IHC on them.

Kim Merriam, MA, HT(ASCP)QIHC
Cambridge, MA

From: Kim Merriam <kmerriam2003 <@t> yahoo.com>
To: "Turner, Leandra" <LTurner1 <@t> seton.org>; "histonet <@t> lists.utsouthwestern.edu" <histonet <@t> lists.utsouthwestern.edu> 
Sent: Wednesday, February 22, 2012 8:21 AM
Subject: Re: [Histonet] Cytology CSF Cell Pellets made from Histogel

I work in a research lab, but I use histogel all the time to make FFPE blocks of cell culture material.  Here is my procedure:

1.     you need about 5X10^ cells per pellet
2.     spin cells @ 2000 rpm for 5 minutes in a 50 ml conical tube
3.     aspirate and resuspend in 15ml NBF and fix overnight @ RT
4.     spin @ 2000 rpm for 5 minutes and aspirate the NBF
5.     microwave the histogel (thermo #HG-4000-012) for 15-20 seconds (loosen cap before microwaving)
6.     histogel should now be at liquid for and about 60C
7.     resuspend cell pellet in about 350ul of histogel
8.     place cell pellet in freezer or on ice for about 20 minutes to solidify
9.     using spatula, remove histogel pellet and place into cassette
10.                  post-fix in NBF for 2 hours (this step is very important); histogel will dissolve in processor if not fixed
11.                  process as usual

Good luck!

Kim Merriam, MA, HT(ASCP)QIHC
Cambridge, MA

From: "Turner, Leandra" <LTurner1 <@t> seton.org>
To: histonet <@t> lists.utsouthwestern.edu 
Sent: Tuesday, February 21, 2012 12:26 PM
Subject: [Histonet] Cytology CSF Cell Pellets made from Histogel

Hello Everyone,

    I am trying to find out a few of things about making cell pellets on
cerebrospinal fluids.  I would like to know: 

    1. If anyone has ever made cell pellets from CSF's using Histogel
and has any tips or procedures they could share?

    2. How to process the CSF pellets made with Histogel, do we need a
routine or stat process?  (we use a Sakura Tissue Tek)

    3. Can you do IHC staining on the pellets?

Thank you for any and all help in advance.

Leandra Turner, HT (ASPC)CM

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