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• Functions:
Respiratory
Excretory
Body Temperature
Transport of Hormones
Defense
BASIC TERMINOLOGIES IN BLEEDING
DISORDERS
• Petechiae
• Purpura
• Ecchymosis
• Hemarthosis
• Hematemesis
• Hematoma
• Hematuria
• Hemoglobinuria
CHARACTERISTICS OF BLOOD
Two types:
Skin puncture
Venipuncture
SKIN PUNCTURE
• Used only when small quantities of blood are required
• Avoid pressure/squeezing
• Recommended depth of skin puncture: _________
• Why do we need to discard 1st drop of blood?
• ____________________________________
• Remove dead epidermal cells
• Remove excess tissue fluid
Red cell count, platelets haematocrit and haemoglobin LOWER in capillary blood
HIGHER WBC count (15-20%) as compared to venous blood
SKIN PUNCTURE
• Puncture sites:
Infants - _______________
For older children and adults - ________________________
VENIPUNCTURE
• Major veins
Cephalic vein
Basilic vein
Median Cubital vein
5 times If plastic
• >Globin chains
• Produced on the specific ribosomes in the cytoplasm of RBCs. Globin in each hgb
molecule contains _______________ which determine the hemoglobin formed.
GLOBIN CHAINS IN HEMOGLOBIN
GREEK NAME NO OF AMINO COMMENTS
ACIDS
Alpha 141
Beta 146
Delta 146 Differs from beta
chain by 10 amino
acids
Gamma 146 Differs from beta
chain by 39 amino
acids
Epsilon 146 Embryonic only
Zeta 146 Embryonic only
OXYHEMOGLOBIN DISSOCIATION CURVE
• Chromosome 16
• Chromosome 11
NORMAL HUMAN HEMOGLOBINS
• Carboxyhemoglobin
• Methemoglobin
• Sulfhemoglobin
CARBOXYHEMOGLOBIN
• Carbon monoxide will bind with hemoglobin even if its concentration in the air is
extremely low
• Cannot bind and carry oxygen
• Causes anoxia
METHEMOGLOBIN/HEMIGLOBIN (HI)
• At birth: 15-20g/dL
• Women: 12-16 g/dL
• Men: 13-18 g/dL
•
• Higher in the morning and lower in the evening
• Increased in strenuous muscular activity
HEMOGLOBIN DETERMINATION
Reagent: ______________
• Color intensity is measured at _____
• All forms of hgb are measured except ________
• ___________ does not affect result
• ___________ will cause falsely elevated result
• High WBC count _____________
• Lipemic blood ____________
RULE OF THREE
> 3 X RBC = Hb
> 3 X Hb = Hct +/- 3%
HEMATOCRIT
• At birth = 45-60%
• Female = 36 -48%
• Male = 40-55%
• Decreased in anemia
• Increased in polycythemia
MACROMETHODS
• Adams
ERYTHROCYTE SEDIMENTATION RATE
Elevated ESR: acute and chronic infections, pregnancy (after 3rd month),
rheumatic fever, rheumatic arthritis, menstruation
• MCV
• MCH
• MCHC
MCV (MEAN CORPUSCULAR VOLUME)
NV: 80-100 fl
Macrocytic: megaloblastic anemia, nonmegaloblastic anemia (liver disease,
hypothyroidism
Microcytic: IDA, defective iron utilization (chronic disease), thalassemia
MCV = Hct %
------------- x 10
RBC
MCH (MEAN CORPUSCULAR HEMOGLOBIN)
• NV : 27-31 pg
Increased: found in macrocytic anemias because RBCs are larger and carry more Hb
Decreased: found in hypochromic anemias and microcytic anemias unless RBCs are
also spherocytic
• Cross sectional/Crenellation
• Longitudinal
• Battlement
BLOOD SMEAR
PREPARATION
Wedge Method
BLOOD SMEAR PREPARATION
Specimen : EDTA blood within 2 to 3 hours &
collected to the mark on tube.
Note : May change RBCs morphology such as
Spiculated (crenated) cells if :
1. Excessive amount of anticoagulant to specimen
2. Old blood - long standing.
3. Warm environment (room temperature) may
hasten changes.
BLOOD SMEAR PREPARATION
Specimen : AC-free blood
Advantages:
Made at the patient’s side
Some artifacts may be prevented
Disadvantages:
Platelet clumping
Few films can be made
STEPS FOR BLOOD FILM
CHARACTERISTICS OF A
GOOD SMEAR
1. Thick at one end, thinning out to a smooth rounded
feather edge.
3. Failure to keep the entire edge of the spreader slide against the
slide while making the smear.
4. Failure to keep the spreader slide at a 30° angle with the slide.
7. Holes in film
AUTOMATED METHODS
Miniprep
Centrifugal (Spinner Type)
SLIDE FIXATION AND
STAINING
II- FIXING THE FILMS
To preserve the morphology of the cells, films
must be fixed as soon as possible after they
have dried.
Methyl alcohol (methanol) is the choice,
although ethyl alcohol ("absolute alcohol") can
be used.
To fix the films, place them in a covered
staining jar or tray containing the alcohol for 2-
3 minutes.
NOTES
• It is important to prevent contact with
water before fixation is complete.
PRINCIPLE
The main components of a Romanowsky stain
are:
A cationic or basic dye
Platelet ( Thrompocytes)
• Nucleus: No nucleus.
• Cytoplasm: small amount bluish cytoplasm &
contains reddish – purple granules
WHITE BLOOD CELLS
Granulocytes
Neutrophils, eosinophils, basophils
Agranulocytes
Lymphocytes and monocytes
Leukocytosis
Leukocytosis, a WBC above 10,000 is usually
due to an increase in one of the five types of
white blood cells and is given the name of the
cell that shows the primary increase.
Diameter:12-16
Cytoplasm : pink
Granules: primary,
secondary
Nucleus: dark purple
blue
dense chromatin
Segmented Neutrophil
Diameter: 12-16
Cytoplasm : pink
Granules: primary,
secondary
Nucleus: dark purple blue,
dense chromatin, 2-5
lobes.
Increased neutrophils count (neutrophilia)
1. Acute bacterial infection.
2. Granulocytic leukemia.
Diameter: 14-16
Cytoplasm : full of granules
Granules: large refractile
orange-red.
Nucleus: blue, dense
chromatin, 2 lobes like a
pair of glass.
The most common reasons for an
increase in the eosinophil count are :
1. Allergic reactions such as hay fever, asthma,
or drug hypersensitivity.
2. Parasitic infection
3. Eosinophilic leukemia
3. Basophil
Diameter: 14-16
Cytoplasm : pink
Granules: dark blue –black
obscure nucleus
Nucleus: blue
4. Lymphocyte
• RSVP
MARROW CELLULARITY
• No hemoglobin
• Nucleus 12 um
• Contain nucleoli
BASOPHIL ERYTHROBLAST
• Early normoblast
• Nucleoli disappear
• Show mitosis
• Hemoglobin starts
appearing – Little Hb
POLYCHROMATOPHIL ERYTHROBLAST
• Late normoblast
• Nucleus smaller
• Coarse Chromatin
• Hemoglobin increase
ORTHOCHROMATIC ERYTHROBLAST
• Normoblast
• Nucleus smaller
• Pyknosis
• Nuclear extrusion
RETICULOCYTE
• Reticulum
• Remnant of ER & GA
• Synthesize Hb
• Few Mitochondria
• Young RBCs
• 1 % of Red Cells
ERYTHROCYTES
1-Microcytosis:
Morphology:
- Decrease in the red cell size.
- Found in:
- Iron deficiency anemia.
- Thalassemia.
- Sideroblastic anemia.
- Lead poisoning.
- Anemia of chronic disease.
1-VARIATION IN ERYTHROCYTE SIZE
(ANISOCYTOSIS)
1-VARIATION IN ERYTHROCYTE
SIZE (ANISOCYTOSIS)
2-Macrocytosis:
Morphology:
Found in:
Any situation with reticulocytosis -
for example bleeding, hemolysis or
response to haemostatic factor
replacement.
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
RBCs may have different shapes:
1- Spherocytosis:
Morphology:
2-Target Cells:
Morphology:
Red cells have an area of increased
staining which appears in the area of
central pallor.
Found in:
-Obstructive liver disease
- Severe iron deficiency
- Thalassaemia
- Haemoglobinopathies (S and C)
- Post splenectomy
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
3- Ovalocytes:
Morphology:
oval shape red blood cell
Found in:
- Thalassaemia major.
- Hereditary ovalocytosis.
- Sickle cell anemia
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
4- Elliptocytosis:
Morphology:
- Hereditary elliptocytosis
- Megaloblastic anemia
- Iron deficiency
- Thalassaemia
- Myelofibrosis
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
6- Blister cell:
Morphology:
Have accentric hallow area.
Found in:
Microangiopathic hemolytic
anemia
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
7- Schistocytosis:
Morphology:
8- Stomatocytosis:
Morphology:
Red cells with a central linear slit
or stoma. Seen as mouth-shaped
form in peripheral smear.
Found in:
- Alcohol excess
- Alcoholic liver disease
- Hereditary stomatocytosis
- Hereditary spherocytosis
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
11- Acanthocytosis:
Morphology:
are red blood cells with irregularly
spaced projections, these
projections very in width but
usually contain a rounded end
Found in:
- Liver disease
- Post splenectomy
- Anorexia nervosa and starvation
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
- Hyperfibrinogenaemia
- Hyperglobulinaemia
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
- Warm autoimmune
hemolysis
III- VARIATION OF RED CELLS
SHAPE (POIKILOCYTOSIS)
15- Nucleated red blood cells.
These red blood cells are
released from the bone
marrow early into the blood
stream, due to the need for
oxygen. Normal red blood
cells do not contain a
nucleus on a peripheral
smear.
IV -ERYTHROCYTE INCLUSION
BODIES
1- Howell-Jolly Bodies:
Morphology:
Small round cytoplasmic
red cell inclusion with
same staining
characteristics as nuclei
Found in:
- Post
splenectomy
- Megaloblastic anemia
IV -ERYTHROCYTE INCLUSION
BODIES
2- Siderotic Granules
(Pappenheimer Bodies)
RBCs which contain no
hemoglobin iron granules.
They appear as dense blue,
irregular granules which are
unevenly distributed in
Wright stained RBCs.
Pappenheimer bodies can be
increased in hemolytic
anemia, infections and post-
splenectomy.
IV -ERYTHROCYTE INCLUSION
BODIES
3- Basophilic stippling:
Morphology:
Considerable numbers of
small basophilic inclusions in
red cells.
Found in:
- Thalassaemia
- Megaloblastic anemia
- Hemolytic anemia
- Liver disease
- Heavy metal poisoning.
IV -ERYTHROCYTE INCLUSION
BODIES
4- Heinz Bodies:
Represent denatured hemoglobin
(methemoglobin - Fe+++) within
a cell. With a supravital stain
like crystal violet, Heinz bodies
appear as round blue
precipitates. Presence of Heinz
bodies indicates red cell injury
and is usually associated with
G6PD-deficiency.
IV -ERYTHROCYTE INCLUSION
BODIES
5- Cabot Rings:
Reddish-blue threadlike rings
in RBCs of severe anemia's.
These are remnants of the
nuclear membrane and
appear as a ring or figure 8
pattern. Very rare finding in
patients with Megaloblastic
anemia, severe anemia's,
lead poisoning, and
dyserythropoiesis.
IV -ERYTHROCYTE INCLUSION
BODIES
.
WHITE BLOOD CELLS
Nucleated cells that function in body’s defense.
Reference range: 4.5 -11.5 x 10^9/L (SI) or 4,500- 11,500/mm^3
(conventional)
WBC Classifications:
• Granulocyte /Agranulocyte
• Polymorphonuclear/Mononuclear
• Phagocyte/Immunocyte
WBC ANOMALIES
SMUDGE CELLS
• HYPOSegmented neutrophils,
inherited condition characterized by
granulocytes with dumbbell-shaped
(Pince-nez/spectacle) appearance of
nucleus
LE CELL
• Seen in:
Hunter and Hurler Syndrome
Mucopolysaccharidoses
San Filipo Syndrome
CHEDIAK HIGASHI SYNDROME
• Enzyme deficiency:________________
• Also called Foam Cells/Pick cells
• Appears to be swollen
HAIRY CELLS
Seen in ________________
• Has cerebriform nucleus “brain-like”,
lymphocytic in origin.
LAZY LEUKOCYTE SYNDROME
• Neutropenia
• Neutrophils that respond poorly to
chemotactic agents
• abnormal inflammatory response
JOBS SYNDROME
• Abnormal neutrophil
• Poor directional motility
CHRONIC GRANULOMATOUS DISEASE
• Characterized by ganulocyte/monocyte
having FAT containing granulations
• Seen in ichthyosis, muscle dystrophy
DOHLE BODIES
• Malignant neoplasm of the blood forming tissues of the bone marros, spleen and
lymphatic system
• General rule: More blasts Shorter, more fatal disease
• FAB Classification:
Based on morphology of cells in Romanowsky stained smear
Based on cytologic and histochemical characteristics of cells involved
LYMPHOMA
• Hodgkins Lymphoma
NLPHL
Popcorn cells
• Classical Lymphoma
• Characterized by ______________ (owl’s eye appearance)
• Non-Hodgkin’s Lymphoma
1. Burkitt’s Lymphoma
2. Cutaneous T Cell Lymphoma – most common subtype
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