RE: [Histonet] Lycing of RBC on Bone Marrow Aspirate Slides

From:"Tony Henwood"



If you want an easy way to lyse rbc's from a slide the following method
works quite well for cytologists who wish to have a 95% ethanol
fixed-like appearance:

Ng et al Re-hydration Method for Red Blood Cell Lysis


Principle:

Rehydration of air dried smears with normal saline for 30 seconds before
fixation in alcohol has been found to produce slides of a quality
superior or at least equivalent to that of immediate fixed smears. It
allowed the cells to adhere better to the slide and lysis of red blood
cells provided a clean background for better assessment.

Solutions:

1.  Normal Saline
              Sodium Chloride     9g
              Distilled water 1000ml

2.  95% Ethanol

Procedure:

1.  After smearing material, air dry slides using a hair dryer.

2.  Place slides in Normal Saline for 30sec.

3.  Immediately place slides in 95% ethanol and fix for 10 minutes.

4.  Stain slides via PAP.


Results:

RBC were effectively lysed but epithelial and mesothelial cells were
retained. In general, smears showed a decrease in chromaticity of both
nuclear and cytoplasmic staining. Cytoplasmic vacuoles were less
distinct and epithelial fragments, such as acini, appeared flattened and
could be more easily focussed on the same plane. Nuclear and cellular
moulding was exaggerated and more appreciable when present. 

There was a slight enlargement of cells. Degenerated specimens, even in
the absence of polymorphs, often resulted in disappointing re-hydrated
smears. Cell clusters were easier to see probably because of the higher
transparency of the cytoplasm.


Reference:  

Acta Cytolog (1994) 38(1):56-64.

Regards

Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: 612 9845 3306
Fax: 612 9845 3318




-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of Matthew
Lunetta
Sent: Thursday, 17 April 2008 7:05 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Lycing of RBC on Bone Marrow Aspirate Slides


Hello Histonetters,

I am looking at doing some research into the lycing of Red Blood Cells
after the Bone Marrow aspirate is already collected onto the slide. I
was wondering if there has already been some work in this area. If
anyone has any info please let me know.

thanks
Matt Lunetta
Longmont United Hospital
Colorado

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