[ad_1]
Immunology 382
Lab #1
Blood Constituents
Leukocytes’ morphology, function & count
Introduction
• Definitions:
– Immunology:is the study of the immune system and is
a very important branch of the medical and biological
sciences.
– Antigens: are substances, which can trigger the
immune system to fight infection or diseases by the
producing antibodies.
– Antibody: also called immunoglobulin, a protective
protein produced by the immune system in response
to the presence of a foreign substance (antigen).
Antibodies recognize and attach onto antigens in
order to remove them from the body.
Types of
Immunity
Innate (natural)
immunity
Protection always
present and ready to
fight microbes
Adaptive
(acquired)
immunity
Humoral immunity
(Protection against
extracellular microbes)
Antibodies produced
by plasma cells.
Cell-mediated immunity
(Protection against microbes
within phagocytic or nonphagocytic cells)
T-lymphocytes
http://missinglink.ucsf.edu/lm/immunology_module/prologuefireworksimages/obj2slice_r1_c2.gif
https://wickedbiology.files.wordpress.com/2016/01/blood-components.png
http://antranik.org/wp-content/uploads/2011/12/Picture1.jpg
White Blood Cells (WBCs)
• White blood cells (WBCs) are also
known as leukocytes.
• They can be divided into:
– Granulocytes: the cytoplasm of
the cells contains organelles that
appear as colored granules
through light microscopy.
• They consist of neutrophils,
eosinophils and basophils.
– Agranulocytes: the cytoplasm of
cells does not contain granules.
• They consist of lymphocytes and
monocytes.
Granulocytes
http://www2.highlands.edu/academics/divisions/scipe/biology/faculty/harnden/2121/images/blood.jpg
Agranulocytes
Granulocytes
• Neutrophils or poly-morphonuclear
leukocytes:
– These cells constitute 70% of
leukocytes
– Usually have 2 to 5 nuclear lobes
connected by fine filaments of
chromatin.
– The most numerous of all leukocytes.
– The cytoplasm is pink to grey because
of the neutral staining of specific
granules (i.e. they don’t stain).
– They function as scavengers within
extravascular tissue, destroying
bacteria or other infectious organisms
that invade the body.
http://1.bp.blogspot.com/-H2sT1-e63U0/UxcjmenLZXI/AAAAAAAAAK0/4s5uOtedGWU/s1600/Norm+Neutrophils.jpg
• Eosinophils:
– Represent about 2-4 % of
leukocytes.
– They contain segmented bi-lobed
nuclei and specific granules.
– They function specifically as
phagocytes to destroy larvae of
parasites that have invaded tissues,
and appear to play a role in allergic
responses.
– Other functions of eosinophils
include phagocytosis of antigen
antibody complexes.
– Their number increases during
parasitic and allergic conditions.
• Basophils:
– Represent about 1 % of leukocytes.
– They contain specific granules in cytoplasm
and contain lots of deep blue staining granules
(basic) and a bi-lobed nucleus.
– They secrete histamine which increases tissue
blood flow via dilating the blood vessels, and
also secrete heparin which is an anticoagulant
that promotes mobility of other WBCs by
preventing clotting.
• Mast cells:
– Similar to basophils, only they associate with
tissues instead of circulating.
– Their granules with histamines.
– Non-lobed nucleus .
– Released as undifferentiated cells, maturing in
their tissues.
Agranulocytes
• Lymphocytes:
– Represent 25-30 % of leukocytes in adult
human.
– They are distinguished into
• B-lymphocytes (Differentiation in bone marrow) and
• T-lymphocytes (differentiation in thymus) and
• NK-cells (natural killer cells).
– They destroy cancer cells, cells infected by
viruses, and foreign invading cells.
– Present antigens to activate other cells of
the immune system. T
– Coordinate the actions of other immune
cells, secrete antibodies and serve in
immune memory.
A lymphocyte in a blood smear. Most of the lymphocytes are small;
a bit bigger than red blood cells, at about 6-9µm in diameter. The
rest (around 10%) are larger, about 10-14µm in diameter. These
larger cells have more cytoplasm, more free ribosomes and
mitochondria. Lymphocytes can look like monocytes, except that
lymphocytes do not have a kidney-bean shaped nucleus, and
lymphocytes are usually smaller. Larger lymphocytes are commonly
activated lymphocytes.
Lymphocytes
• Monocytes:
– They are the largest of the
formed elements.
– Their cytoplasm tends to be
abundant and relatively clear.
– They enlarge into macrophages
(which are large phagocytic
cells ) and migrate into tissues
by amoeboid motion.
• They digest pathogens, dead
neutrophils, and the debris of
dead cells.
– Like lymphocytes, they also
present antigens to activate
other immune cells.
Monocytes
A monocyte in a blood smear. These are the largest type of
white blood cells, and can be up to 20µm in diameter. They
have a large kidney bean shaped nucleus.
They have abundant cytoplasm, and some fine pink/purple
granules in cytoplasm.
https://d2gne97vdumgn3.cloudfront.net/api/file/7ceaitOXTvWcYlwbTL9T
Lab Activity 1
Differential leukocyte count
• The biological functions of leukocytes include:
– Defense and protection against microbial infections or any
foreign substances, and
– The production of immunoglobulins.
• Differential Count:
– In disease, either the normal percentage of WBCs is
disturbed or/and abnormal or immature WBCs are seen in
peripheral blood.
– The purpose of the WBC differential count is to determine
the percentage of each type of WBC from a stained blood
film.
Lab Activity 1
Differential leukocyte count
• Preparation of Blood Smear
1. Clean slide with ETOH.
2. Place drop of blood or cells onto
slide on one end (the cells should
be somewhat (concentrated).
3. Place 2nd slide onto edge of drop
(start from center of slide and back
slide into drop) and once fluid is
pulled across 2nd slide push the 2nd
slide across the 1st slide.
4. Let cells dry, and warm fixing next
the flame.
http://mt-lectures.blogspot.com/2017/08/lecture-13-
morphological-examination-of.html
Figure 1–2 Well-made
peripheral blood smear.
Figure 1–3 Unacceptable peripheral blood films. Slide appearances associated with
the most common errors are shown, but note that a combination of causes may be
responsible for unacceptable films. A, Chipped or rough edge on spreader
slide. B, Hesitation in forward motion of spreader slide. C, Spreader slide pushed too
quickly. D, Drop of blood too small. E, Drop of blood not allowed to spread across the
width of the slide. F, Dirt or grease on the slide; may also be caused by elevated
lipids in the blood specimen. G, Uneven pressure on the spreader slide. H, Time
delay; drop of blood began to dry.
(From Rodak BF, Fritsma GA, Keohane EM: Hematology: clinical principles and applications, ed 4, St. Louis, 2012, Saunders.)
Giemsa Stain Procedure
1. Flood smears with Giemsa stain for 5 minutes
(do not let slides dry out).
2. After 5 minutes, add an equal amount of
distilled water and let stand 3 minutes.
3. After 3 minutes, gently rinse slide by washing
with water from one end until water runs
clear.
4. Blot dry carefully using filter paper
5. Examine under high (oil) power.
Giemsa stain
• Principle And Interpretation
– The polychromatic staining solutions such as Giemsa
stain contain basic dyes (methylene blue, azure) and
acidic dye (eosin).
– These basic and acidic dyes induce multiple colors
when applied to cells.
• Methanol acts as fixative and also as solvent. The fixative
does not allow any further change in the cells and makes
them adhere to the glass slide.
• Basic Part: azure blue or methylene blue which bind to the
acidic part of the cell (nucleus) and stain it blue.
• Acidic Part: eosin Y which binds to the basic parts of the cell
(proteins, cytoplasm) giving them a red color.
• The neutral parts of the cell are stained by both the dyes
Differential leukocyte count
• Identify and count 25 consecutive
leukocytes and record each cell
type separately.
– Begin at the thin end of the smear
and count the white cells observed
as the slide is moved in a vertical
direction.
– When near the edges of the smear,
move the slide horizontally for a
distance of about two fields, then
proceed vertically back across the
smear.
– Continue this “snake-like”
movement until 25 leukocytes have
been counted and classified.
Results after staining
• Neutrophils : Dark purple nucleus, reddish lilac granules, pale pink cytoplasm
• Eosinophils : Blue nuclei, red to orange red granules, blue cytoplasm
• Basophils : Purple to dark blue nucleus, dark purple granules
• Lymphocytes & Monocytes: Dark purple nuclei, sky blue cytoplasm
• Platelets : Violet to purple granules
https://www.researchgate.net/publication/346962804/figure/fig1/AS:9726
[email protected]/Microscopic-images-of-WBC.png
Results:
calculate the number of cells of
each type/give percentage of cells (this should be
done by counting at least 25 cells from smears)
Cell type Neutrophils Lymphocytes Monocytes Eosinophils Basophils
Drawing
Total cells
Percent
Discussion:
1. Compare your count with standard count.
2. Explain the variations in the WBC counts.
3. Reasons for false counts
Reference Range
Differential blood count gives relative percentage of each type of white blood cell
and also helps reveal abnormal white blood cell populations (e.g., blasts,
immature granulocytes, or circulating lymphoma cells in the peripheral blood).
Reference ranges for differential white blood cell count in normal adults is as
follows:
Neutrophils: 40% to 60%
Lymphocytes: 20% to 40%
Monocytes: 2% to 8%
Eosinophils: 1% to 4%
Basophils: 0.5% to 1%
The reference ranges may vary depending on population studies, the individual
laboratory, instruments, and methods.
https://medlineplus.gov/ency/article/003657.htm
[ad_2]
Testimonials
