Posted January 1, 2005
The Laboratory Report series is written for those who collect
patient specimens. By providing this educational background on
medical tests, it is intended that fewer errors in specimen collection
will be made.
Examination of the blood smear is an important part of the hematologic
evaluation. The reliability of the information obtained depends on
well-made and well-stained films that are systematically examined.
Peripheral blood or potassium EDTA anticoagulated blood (1-2 mg EDTA/1
mL blood) may be used. Smears of peripheral blood must be made immediately.
Smears made from EDTA-anticoagulated blood should be made within
2 hours of collection. There are 2 main manual methods described;
the 2-slide or wedge method and the coverglass method.
The wedge method: First, place a 2-3 mm drop of blood about 1 cm
from the frosted end of a clean slide that is on a flat surface.
Second, with the thumb and forefinger of the right hand, hold the
end of a second slide or coverglass (“spreader slide”)
against the surface of the first slide at an angle of 30-45 degrees.
Third, draw it back to contact the drop of blood. Allow the blood
to spread and fill the angle between the 2 slides. Finally, push
the “spreader slide” at a moderate speed forward until
all of the blood has been spread into a moderately thin film.
The coverglass method: First, place a 2-3 mm drop of blood on 1
coverglass and then place another coverglass crosswise over the drop
so that the corners appear as an 8-pointed star. Just as it stops
spreading, pull the coverglasses quickly but firmly apart on a plane
parallel to their surfaces. The coverglasses should then be dried
film side up.
Using either method, the films should be fixed in absolute methanol
for 1 to 2 minutes. The slide is next exposed to undiluted Wright
stain solution for 2 minutes. Then, without removing the stain from
the horizontal slide, an equal amount of buffer is added and mixed
by blowing gently. The stain is flushed from the slide using water
for about 30 seconds. The slide is allowed to air dry in a tilted
position. Slides can be cover-slipped or coverglasses can be mounted
(2 on each slide) with standard mounting media.
The film should not cover the entire surface of the slide. In a
good film, there is a thick portion and a thin portion and a gradual
transition from one to the other. The film should have a smooth,
even appearance and be free from ridges, waves or holes. The end
of the smear (the “feathered edge”) should be smooth
and even. A well-stained smear will have pink-colored red cells and
the nuclei of the leukocytes will be purple, with well-differentiated
chromatin and parachromatin.
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NewsPath® Editor: Megan
J. DiFurio, MD, FCAP
This newsletter is produced in cooperation with the College of American
Pathologists Public Affairs Committee and may be reproduced in whole or
in part as a service to the medical community. Copyright © 2006 by
the College of American Pathologists.
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