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Blood Film Technique
Making Blood film
Practical Parasitology
2nd StageLab 12: Blood Film
University of Sulaimani
School of ScienceDepartment of
Biology
University of Sulaimani
School of ScienceDepartment of
Biology
OBJECTIVES
• Students should be able to:– Make a good blood film– Stain blood film – Identify the diagnostic stages of a
blood-dwelling parasites
Blood Film• The most commonly used technique for blood
examination is blood film.• A very thin layer of blood spread over a
microscope slide• Allows the various types of blood cells to be
seen and identified. • Blood smear plays an important role in
diagnosing a wide range of illnesses.• Includes detection of blood-borne parasites, like
malaria.
Collection of Blood Smears
5.Touch the drop of blood to the slide from below.
4.Slide must always be grasped by its edges.
2.Puncture at the side of the ball of the finger.3.Gently squeeze toward the puncture site.
1.The second or third finger is usually selected and cleaned.
Preparing thick and thin films1. Touch one drop of blood to a clean slide.2. Spread the
first drop to make a 1 cm circle.3. Touch a fresh drop of blood to the edge of another slide.
6. Wait for both to dry before fixing and staining.
5.Pull the drop of blood across the first slide in one motion.
4.Touch the drop of blood by spreader slide at 45degree angle.
Blood Film Preparation
Thick Blood Film preparation
Thin Blood film preparation
Thick and Thin Films
• THICK FILM– lysed RBCs– larger volume– 0.25 μl
blood/100 fields– more difficult to
diagnose species– parasite density
• THIN FILM– Intact RBCs– smaller volume– 0.005 μl
blood/100 fields– good species
differentiation– low density
infections can be missed
Staining
• Staining is a biochemical technique of adding a specific dye to a substrate (DNA, proteins, lipids, carbohydrates) to qualify or quantify the presence of a specific compound.
• Stains and dyes are frequently used in biology and medicine to highlight structures in biological tissues for viewing, often with the aid of different microscopes.
Staining• Romanowsky stains are based on a
combination of eosine and methylene blue
• Wright's stain, Leishman stain and Giemsa stain.
• All are used to examine blood or bone marrow samples.
• Nuclei stained dark blue/violet, erythrocytes pale pink & cytoplasm pale blue
• All are also suited to examination of blood to detect blood-borne parasites like malaria.
Staining Procedure • Thin smear are air dried.• Overflow the smear with Leishman stain
(1 ml) for 1-5 min. • Add a double amount of distilled water
and mix the stain by blowing the fluid.• Leave the mixture on the slide for 10-15
min. • Wash off by slow-running water to
remove the extra liquid stain.• Stand slide on end, and let dry in air.• Examination
Methylene blue• Basic dye• Blue-purple colour• Stains nuclei,
ribosomes & rough ER (DNA & RNA - acidic)
• Structures that stain with methylene blue are termed basophilic
Eosin• Acid dye• Pink-red colour• Stains most
cytoplasm proteins which are mostly basic
• Structures that stain with eosin are termed eosinophilic
Leishman stain
Examination
Microfilaria Microfilaria
References
• http://www.dpd.cdc.gov/dpdx/ • http://www.stanford.edu/group/parasites/Parasit
es2006/• http://www.bsieducation.org/Education/14-19/to
pic-areas/applied-science/standard-procedure/sp-0002-1.shtml
• http://www.ruf.rice.edu/~bioslabs/studies/studies.htm
• Clodfelter, R.L. (1986). The peripheral smear. Emerg. Med. Clin. North. Am. 4(1):59-74.
Next LabCiliates:
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