Re: lysing buffer

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From:"Tony Henwood" <henwood@mail.one.net.au>
To:"'histonet'" <histonet@Pathology.swmed.edu>, "Berger, Jennifer" <jberger@lrri.org>
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Date:Fri, 9 Jul 1999 10:50:46 +0000
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Dear Jennifer,

> Does anyone have a recipe for or know where I can order Lysing Buffer?

Try the following technique:
 
Haemolysing haemorrhagic specimens causes fewer problems in 
identifying individual cells (1). The removal of red blood 
cells can be achieved using Ficoll-Hypaque gradient separation 
or using a lysis solution such as isotonic ammonium chloride 
(1):

SOLUTIONS:

1.  Lysis solution
         Ammonium Chloride        4.5g
         Potassium carbonate      0.5g
         EDTA                     0.0186g
         Distilled water          500mls
2.  Hanks medium

METHOD:

1.  Centrifuge bloody fluid.
2.  Remove supernatant and add equal volume of lysis solution.
3.  Resuspend and incubate for 5 minutes at 4oC.
4.  Centrifuge, if blood still remains, then repeat from step
         2.
5.  Resuspend pellet in Hanks.

(1)  Kuenen-Boumiester etal (1996) Acta Cytolog 40:475-479.
Tony Henwood
Senior Scientist
Anatomical Pathology
Royal Prince Alfred Hospital
Sydney, AUSTRALIA

http://www2.one.net.au/~henwood
http://www.pathsearch.com/homepages/TonyHenwood/default.html



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