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Blood Smear

1. Haematology is the study of blood, which is useful for human health and done routinely in hospitals. Blood consists of erythrocytes, leucocytes, thrombocytes, and plasma. 2. Blood smears allow examination of blood cells under a microscope. They are made by placing a blood drop on a slide and using another slide to spread the blood. The slide is then stained using Leishman's stain. 3. Leishman's stain uses acidic and basic dyes that induce different colors in cells. Neutrophils, eosinophils, basophils, lymphocytes, and monocytes can be identified by their nuclear shape and cytoplasmic staining.

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Hussein Ala
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376 views

Blood Smear

1. Haematology is the study of blood, which is useful for human health and done routinely in hospitals. Blood consists of erythrocytes, leucocytes, thrombocytes, and plasma. 2. Blood smears allow examination of blood cells under a microscope. They are made by placing a blood drop on a slide and using another slide to spread the blood. The slide is then stained using Leishman's stain. 3. Leishman's stain uses acidic and basic dyes that induce different colors in cells. Neutrophils, eosinophils, basophils, lymphocytes, and monocytes can be identified by their nuclear shape and cytoplasmic staining.

Uploaded by

Hussein Ala
Copyright
© © All Rights Reserved
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Download as PDF, TXT or read online on Scribd
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Haematology:- is the study of blood ,this usefull for health of human and a routine hospital laboratory .

Blood is composed of 4portions:- 1-Erythrocytes(red blood cell) 2- Leucocytes (white blood cells)

3-thermobocytes(platelets) 4- plasma (fluid medium)

A-(capillary blood ) : there are 3 sites of puncture to collect blood by lancet:

1-the lope of the ear 2- the ball of the finger(in adult) 3- the base of the heel or great toe (in babies)

B- Venous blood :puncturing a vein with a needle attached to a syringe is called venipuncture .this use
when the large volumes of blood are required.

Blood smear

Fig. 1 blood film

A blood film or peripheral blood smear is a microscope slide made from a drop of blood, that allows the cells to be
examined microscopically. Blood films are usually done to investigate the normal blood cells ( R.B.C, W.B.C, platelets)

Materials
1- Lancet , ethanol 70% , cotton
2- Slides , spreader slide
3- Drop of blood
4- Methanol
5- Leishman’s stain

1
6- Preparation

Blood films are made by placing a drop of blood on one end of a slide, and using a spreader slide to disperse
the blood over the slide's length. The aim is to get a region where the cells are spaced far enough apart to
be counted and differentiated.
The slide is left to air dry, after which the blood is fixed to the slide by immersing it briefly in methanol. The
fixative is essential for good staining and presentation of cellular detail. After fixation, the slide is stained to
distinguish the cells from Leishman stain .

Characteristic red blood cell abnormalities are anemia, sickle cell anemia and spherocytosis.
White blood cells are classified according to their propensity to stain with particular substances, the shape
of the nuclei and the granular inclusions.

Types of white blood cells :-


A-GRANULOCYTES:- as they contain small granules in their cytoplasm,there are 3 types:-

1-Neutrophil granulocytes usually make up close to 80% of the white count. They have
multilobate( 2-5 lobes)nuclei and lightly staining granules. They assist in destruction
of foreign particles by the immune system by phagocytosis and intracellular killing.

2
2-Eosinophil granulocytes have(2 lobes) granules that stain with bright red or orange and play a role in
allergy and parasitic disease.itis 1-4% of total W.B.Cs

3-Basophil granulocytes areirregular nucleusdivided 2 lobes (s-shape) and their granules stain dark with
alkaline stains,it less than 1 of total W.B.Cs.

B- AN GRANULOCYTES
1-Lymphocytes have very little cytoplasm and a large nucleus (high NC ratio) and are responsible for
antigen-specific immune functions, either by antibodies (B cell) or by direct cytotoxicity (T cell). have
one small nucleus and 20-45%of total W.B.Cs
2- Monocytes: it is the largest white cells with nucleus of irregular shape large and centrally placed
with in the cell (kidney shape),the cytoplasm stain blue colour (2-10)%.

Leishmans stain:

Principle
The polychromic staining solutions (Wright, Leishman Giemsa) contain methylene blue and eosin. These
basic and acidic dyes induce multiple colors when applied to cells. Methanol acts as fixative and also as a
solvent. The fixative does not allow any further change in the cells and makes them adhere to the glass
slide. The basic component of white cells (i.e. cytoplasm) is stained by acidic dye and they are described as
eosinophilic or acidophilic. The acidic components (e.g. nucleus with nucleic acid) take blue to purple shades
by the basic dye and they are called basophilic.The neutral components of the cell are stained by both the
dyes.

Procedure
A thin smear is prepared by spreading a small drop of blood evenly on a slide.

Making the film


1. Take a clean dry (grease free) slide
2. Transfer a small drop of blood near the edge of the slide.
3. Place the spreader slide at an angle of 30°. Pull back the spreader until it touches the drop of blood. Let
the blood run along the edge of the spreader

3
4. Push the spreader forward to the end of the slide with a smooth movement.
5. Dry the blood smear at room temperature. Adequate drying is essential to pre serve the quality of the
film.

Precautions
1. Dirty slides do not give an even smear
2. Use an appropriate size of blood drop
3. After putting the drop on the slide, make the smear immediately for even distribution of white blood cells
on the slide.
4. The thickness of the smear depends on the angle of the spreader. If the angle is less than 300 a thinner
smear is obtained and if the angle is more than 300 a thicker smear is obtained.
5. The film must be smooth at the end. There should be no lines extending across or down through the film
and it should not contain holes.

Identification marking :-By using a lead pencil or a ball pen, write the identification number on the slide.

Fixing the Smear :


The slide should be stained after making the smear. Methanol present in the stain fixes the smear. If the
staining is to be done later, the blood smear must be fixed with methanol for 2 to 3 minutes to prevent
distortion of cells.

Staining the Film


1. Cover the smear with the staining solution by adding 10-15 drops on the smear. Wait exactly for one
minute.
2. Add equal number of the drops of buffer solution. Mix the reaction mixture adequately by blowing on it
through a pipette. Wait for 10 minutes.
3. Wash the smear by using tap water (tap water is preferred because sometimes if distilled water is not
stored properly it becomes slightly acidic and gives unsatisfactory results).
4. Stand the slide in a draining rack or on the laboratory counter to dry.

W.B,C and R.B.C Cells in microscope after staining

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