[Histonet] Cell block processing

[Tony Henwood (SCHN)]

Hi Charles,

I have had excellent success with lysing the red blood cells (using  Isotonic Ammonium Chloride) prior to cell block preparation with thromboplastin-plasma. 
The lysing solution contains EDTA so you will need to add a few drops of 1% calcium chloride. Method as follows:

Lysis solution
Ammonium Chloride        4.5g
Potassium carbonate      0.5g
EDTA                     0.0186g
Distilled water          500mls

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.     Rinse in Hanks or RPMI, centrifuge.    
6.	Mix pellet in a few drops of plasma.
7.     Add thromboplastin and a few drops  of 1% Calcium Chloride, mix gently and allow clot to form.
8.     Add 10% buffered formalin and fix and process as usual.

Reference:  
Kuenen-Boumiester etal (1996) Acta Cytolog 40:475-479

The lysis solution can also be purchased commercially from several companies (eg Biolegend). It is commonly used for sample preparation for flow cytometry. Check the SDS to make sure it does not contain formaldehyde.


Regards
Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC), FFSc(RCPA)
Principal Scientist, the Children’s Hospital at Westmead
Adjunct Fellow, School of Medicine, University of Western Sydney
Tel: 612 9845 3306
Fax: 612 9845 3318
Pathology Department
the children's hospital at westmead
Cnr Hawkesbury Road and Hainsworth Street, Westmead
Locked Bag 4001, Westmead NSW 2145, AUSTRALIA

________________________________________
From: Charles Riley via Histonet <histonet at lists.utsouthwestern.edu>
Sent: Friday, 25 October 2019 23:12
To: Histo List
Subject: [Histonet] Cell block processing

Does anyone have any tips or suggestions on how to better process extremely
bloody FNA  specimens?    Is there anyway to clear out some or all of the
blood without destroying the other tissues?

--

Charles Riley BS  HT, HTL(ASCP)CM

Histopathology Coordinator/ Mohs
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