HEMATOLOGICAL

STAINING
TECHNIQUES


TAREMWA Ivan M
HTM, 2014.
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Clinical
Assessment
Investigations
Definitive
diagnosis
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Blood Composition
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Diagnostic trends
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Any challenges with these slides?
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Forensic Characterization of Blood
O Hemoglobin possesses peroxidase like activity which
when mixed with phenolphthalein and hydrogen
peroxide it will cause the formation of a deep pink
color

O Once blood is found it must be determined to be
human
O Precipitin test is the standard test
O Reagents are available to type if blood is dog,cat or
others
O A positive test is a cloudy ring or band at the point
where the two liquids meet
O Only a small amount of sample is required
O Precipitin test is very sensitive and can test positive on
a sample as old as 10 years


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O Blood is examined
in a smear

O Smears are
stained
using
Romanowsky
stains;
Giemsa, Jenner,
Wright, Field and
Leishman.
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Reticulocyte stains and Count
O Better and more reliable results are obtained with
New
methylene blue, (? different from methylene blue)
Brilliant cresyl blue and Azure B. -
Reticulofilamentous material.
Method
O Deliver 2 or 3 drops of the dye solution in a tube-
Supravital staining.
O Add 24 volumes of the patients EDTA-
anticoagulated blood to the dye solution and mix.
O Keep the mixture at 37
0
C for 1520 min.
Resuspend the red cells by gentle mixing and make
films on glass slides the usual way. When dry,
examine the films without fixing or counterstaining.
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O Place a small drop of blood from
the frosted end of a slide.
The end of a second slide is
used to spread the blood drop
out on the first slide.

Pull the end of the slide
through the blood drop toward
you and then push in the
opposite direction forcing the
blood in your direction.

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Staining the blood smear:
The smear should be stained as soon as the slide dries.

If the slide cannot be stained immediately, it can be immersed in
methanol (fixed) and stained later.




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Staining blood and BM films
O Romanowsky stains are used universally for
routine staining of blood films; with satisfactory
results.

O These offers subtle distinctions in shades of
staining between nucleus & cytoplasmic contents.


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Color patterns of blood cells

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Cell Morphology
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Lymphocyte
Segmented Neutrophil
Platelet
rbc

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neutrophil eosinophil
basophil
small lymphocyte
monocyte

The Peripheral Smear
Schistocytes Hemolytic Anemia, Burns
Spherocytes AIHA, Hereditary Spherocytosis
Target cells Alcoholics, Hbpathies
Sideroblasts Myelodysplasia, Alcoholics
Teardrop cells Myelofibrosis/Myeloid metaplasia,
Thalassemia
Burr cells Uremia
Howell Jolly
bodies
Splenectomy, Functional asplenia
Hypersegmented
PMN
Megaloblastic Anemia
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What are these?



Blood smear in a 43 year old man with history of a motor vehicle
accident 12 years ago.

What is this?
What may have
happened?

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Additional tests
Questions?
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