COMPLETE BLOOD COUNT (CBC) COMPOSITION OF BLOOD

• Plasma:
BLOOD – connective tissue with fluid matrix
- 55% of total blood
(plasma = 91% water and 9% formed elements)
- pale, yellow liquid that surrounds
**out of the 9%, 7% are proteins and the
cells
remaining 2% are other solutes (organic and
- 91% water, 7% proteins, and 2%
inorganic substances)
other
SERUM – fluid matrix from clotted blood • Formed Elements:
(absence of fibrinogen) - 45% of total blood
- cells and cell fragments
✓ What is CBC for? Gives idea of possible - erythrocytes, leukocytes,
underlying conditions like anemia and thrombocytes
infection
✓ What are measured? PLASMA PROTEINS
1. Hematocrit • Albumin:
2. Hemoglobin - 58% of plasma proteins
3. RBC count - helps maintain water balance
4. WBC count • Globulins:
5. WBC Differential count - 38% of plasma proteins
- helps immune system
FUNCTIONS OF BLOOD • Fibrinogen:
• Transport of gases, nutrients and waste - 4% of plasma proteins
products - aids in clot formation
• Transport of processed molecules
• Transport of regulatory molecules ERYTHROCYTES
• Regulation of pH and osmosis
• Maintenance of body temperature • Red blood cells (RBC)
• Protection against foreign substances • Disk-shaped with thick edges
• Clot formation • Nucleus is lost during development
o Mature RBCs don’t have
CHARACTERISTICS OF BLOOD nucleus
• Type of connective tissue o Presence of nucleus on RBC
• Sticky circulation may indicate a
• Heavier than water disease (or is only minimal)
• O2 content determines color ▪ Severe blood loss
• Temp. slightly higher than rest of body ▪ Immature RBCs are
• Males (5-6 L), females (4-5 L) released from the
bone
▪ Anemia
• Live for 120 days
• Function: transport O2 to tissues
• RBC count: 4.6-6.2 x 106/uL

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 B. AGRANULOCYTES
• No granules on cytoplasm

1. Monocytes:
• Largest
• Produce
macrophages

LEUKOCYTES 2. Lymphocytes
• White blood cells (WBC) • Immune
• Lack hemoglobin response
• Larger than erythrocytes • Several
• Contain a nucleus different types
• Functions: o T cells:
✓ fight infections from
✓ remove dead cells and debris Thymus
by phagocytosis o B cells: from bursa
• Normal Value: 5000 -10000/cumm or 5- • Production of antibodies
10 x 103/uL
PLATELETS
TYPES OF LEUKOCYTES • What are they?
A. GRANULOCYTES - blood clotting cells
• Contain granules on cytoplasm - produced in red bone marrow
• Platelet count: not a part of CBC
1. Neutrophils (estimated value lang ng platelets nasa
• Most common CBC: increased/decreased); separate
• Remain in blood for test
10-12 hours then
move to tissues
• Phagocytes
• Appear segmented
•
2. Eusinophils
• Reduce
inflammation
o Increase in
number in
cases of
allergies or
parasitism
• Reddish-orange granules
3. Basophils
• Least common
• Release histamine
(during allergies)
and heparin
(natural anti-
coagulant)
• Blue-black granules

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HEMATOCRIT • Transports O2
• Aka packed cell volume (PCV) • Consists of globin chain and heme
• Volume of packed red cells after pigment
centrifugation • Each globin protein is attached to
o Proportion of RBC to plasma a heme molecule
• Normal Values: • Each heme contains one iron
✓ Male: 47  7 % (40-54) atom
✓ Female: 42  5 % (37-47) • O2 binds to iron
✓ Newborn: about 56% • Oxyhemoglobin:
✓ Increased in: Polycythemia, hemoglobin with an O2
Shock, and severe dehydration attached
(high RBC concentration) • To be delivered to
✓ Decreased in: Leukemia, body tissues, then
Anemia, Hyperthyrodism, picks up CO2 in
Cirrhosis (low RBC count) blood, making them
*Capillary tubes: into:
• Red: heparinized carboxyhemoglobin
o After Centrifugation: RBC + • Normal Values:
Plasma ✓ Male: 14 – 16.5 g/dL
▪ Presence of fibrinogen ✓ Female: 12-15 g/dL
(protein responsible for ✓ Increased: high altitudes,
blood clotting) obstructive pulmonary disease,
• Blue: no anti-coagulant present congestive heart failure,
o After centrifugation: RBC + Serum polycythemia
o Serum: liquid portion of clotted ✓ Decreased: Severe hemorrhage,
blood without fibrinogen anemia, hyperthyroidism, liver
▪ Fibrinogen is consumed cirrhosis
during blood clotting.
▪ Contains only Albumin & ACID-HEMATIN METHOD
Globulin • Comparator block/ Sahli's
Hemoglobinometer
ADAM’S MICROHEMATOCRIT METHOD o determine the quantity of
PROCEDURE: hemoglobin in the blood
✓ Anticoagulated purple blood tube – o Sahli’s pipette
use blue ringed (non-additive) ▪ marked at 20 microliters
✓ Skin prick – use red ringed 0.02 ml
(heparinized) ▪ like a graduated cylinder
✓ Venipuncture – anti-coagulated
syringe
Process:
• Fill capillary tube, atleast ¾ full → Seal with
clay (3mm) + paraffin wax → centrifuge
blood at 10,000 rpm for 4-5 minutes using
microhematocrit centrifuge (sealed portion
outer portion of centrifuge) → use
microhematocrit reader

HEMOGLOBIN
• Pigment in red blood cells that has
affinity to oxygen
• Responsible in carrying blood’s
oxygen
• Main component of erythrocytes

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WBC and RBC COUNT • Platforms contain ruled area
• number of cells in 1 cubic millimeter of o 9 large square = 1 mm x 1 mm =
blood total of 9 square mm
• WBC pipette: has white bead inside ▪ Primary Squares: 9 squares
• RBC pipette: has red bead inside & ▪ Secondary squares: for
bigger mouth WBC (contain 16 squares,
• Normal Value: reading is done in 4 corner
✓ WBC Count: 5-9 x 103/uL squares)
o Lymphocytes: 20% - 25% (↑ in anti- ▪ Tertiary Squares: 25 squares
body rxn) • RBC is counted in
o Monocytes: 3% - 8% ( ↑ in chronic R1, R2, R3, R4
infection) o Volume on one platform = 0.9uL
o Neutrophils:60%-70% (↑ in acute
infections)
o Eosinophils: 2% - 4% (↑ in allergic
rnx)
o Basophils: 0.5% - 1% ( ↑ in chronic
infection)
✓ RBC Count: 4.6-6.2 x 106/uL

WBC Diluting Fluid:

1. Hypotonic Solution
2. Easily Prepared
3. Cheap
4. Readily Available
5. Good Preservative

RBC Diluting Fluid:

1. Isotonic Solution Procedure:
2. With high specific gravity 1. Aspirate blood up to .5 mark
3. Easy to prepare 2. Wipe off excess blood at the tip of pipet with
4. Cheap tissue paper
5. Good preservation 3. Aspirate diluting fluid up to “11 mark” for wbc
6. Has a buffer action and “101 mark” for rbc
7. Does not initiate growth of molds 4.Mix the tube using figure of 8 dilution factor
✓ WBC: 20
WBC COUNT ✓ RBC: 200
• Number of WBCs in 1 liter (L) or 1 5.Compute for dilution factor
microliter (mL) of blood
Sample used: Discharging..
• Ethylenediaminetetraacetic acid (EDTA) 1. Place cover slip on top
on whole blood 2. Discard 2-3 drops
• Blood from a skin puncture 3. Place a drop to fill the chamber
• Fluid: Diluted with 1% buffered
ammonium oxalate or a weak acid
solution (3% acetic acid or 1%
hydrochloric acid).
o To hemolyze RBC, in order to not
see RBC under microscope

Materials Used:
Counting chamber/hemocytometer:
• Has 2 identical ruled platforms

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4. Focus under LPO.. For WBC count in 4 W
squares.. For RBC count in 5 R squares with HPO

Procedure:
1. Make a blood smear by placing one drop of
blood on a clean glass slide
2. Using another slide at a 50 degrees angle,
ADAM’S MICROHEMATOCRIT METHOD push the drop of blood towards the end
1. Fill heparinized tube ¾ full 3. Allow blood smear to dry
2. Cover with sealing clay 4. Stain the blood smear w/ giemsa stain
3. Centrifuge at 10k RPM for 5 mins. (sealed end 5. Count WBC on the blood smear’s feathery
should be outside) edge
4. Read in a reader - count 100 WBC, assuming that there are
120 of it
FORMULA:
Manual Differential Counter
WBC/cumm =
𝒄𝒆𝒍𝒍𝒔 𝒄𝒐𝒖𝒏𝒕𝒆𝒅 𝒙 𝒅𝒊𝒍𝒖𝒕𝒊𝒐𝒏 𝒇𝒂𝒄𝒕𝒐𝒓 - Rings when it reaches 100
𝒂𝒓𝒆𝒂 𝒄𝒐𝒖𝒏𝒕𝒆𝒅 (𝒎𝒎𝟐) 𝒙 𝒅𝒆𝒑𝒕𝒉

DIFFERENTIAL COUNT
- provides relative proportion of leukocyte
types
- Normal Values:
✓ Neutrophils: 60-70%
DIAGNOSTIC BLOOD TESTS
✓ Lymphocytes: 20-25%
• Complete blood count:
✓ Monocytes: 3-8%
o provides information such as RBC
✓ Eosinophils: 2-4%
count, hemoglobin, hematocrit,
✓ Basophils: 0.5-1%
and WBC count
- done in order for the physician to know
• Hematocrit:
what type of infection the patient has
o % of total blood volume
o Bacterial infection: ↑ Neutrophils
composed of RBC
value (up to 90%) = low
• Hemoglobin:
lymphocyte value
o determines amount of
o Viral infection: ↑ Lymphocyte
hemoglobin
value
- Granulocytes:
1. Neutrophils
2. Eosinophils
3. Basophils
- Agranulocytes:
1. Lymphocytes
2. Monocytes

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 - indicate anemia ABO blood group consist of:
• Two antigens (A & B) on the surface of the
RBCs
• Type A: A antigens
• Type B: B antigens
• Type AB: A&B antigens
• Type O: NO antigens
• Two antibodies in the plasma (anti-A & anti-
B)
• Type A: anti-B antibodies
• Type B: anti-A antibodies
• Type AB: NO antibodies
• Type O: anti-A&B antibodies

• Type O are universal donors because they

• Prothrombin time: have no antigens
- time it takes for blood to begin clotting • Type A can receive A and O blood
(9-12 sec.) • Type B can receive B and O blood
• White blood cell count: • Type AB can receive A, B, AB blood
- total number of WBC • Type O can only receive O blood
• White blood cell differential count:
- Determines the % of each 5 kinds of
leukocytes
- neutrophils: 60-70%
- lymphocytes: 20-25%
- monocytes: 3-8%
- eosinophils: 2-4%
- basophils: 0.5-1%

White Blood Cell Disorders

• Leukopenia:
- low wbc count
- caused by radiation, chemotherapy
drugs, tumors, viral infections
• Leukocytosis: ANTIGENS AND ANTIBODIES
- high wbc count • In the case of the ABO blood groups, the
- caused by infections and leukemia antigens are present on the surface of the
red blood cell, while the antibodies are in
the serum.

ABO ANTIGENS & CORRESPONDING ANTIBODIES

ABO BLOOD GROUPS

• Most well-known & clinically important blood
group system.
• Discovered by Karl Landsteiner in 1900
• He was awarded the Nobel Prize in
Physiology or Medicine in 1930

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Classification of ABO blood group is done with
the use of:
✓ Anti-A (yellow) and Anti-B (blue) serum
o (Type A/B/AB/O)
✓ Anti-D serum
o for Rh blood group
▪ Positive
▪ Negative

ABO Blood Group and Rh Blood Group

Systems

ABO ANTI-A ANTI-B Serum

GROUP Serum

A Agglutination No
Agglutination

B No Agglutination
Agglutination
AB Agglutination Agglutination
O No No
Agglutination Agglutination
Rh Blood ANTI-D
Group
Rh Agglutination
Positive
Rh No Agglutination
Negative

Rh BLOOD GROUP
• Rh positive means you have Rh antigens
• 85-95% of the population is Rh+
• Antibodies only develop if an Rh- person
is exposed to Rh+ blood by transfusion or
from mother to fetus

THE RHESUS (Rh) Blood Group Systems

- Rh Genetics: The genes that control the
system are autosomal codominant
located on the short arm of chromosome
- The presence or absence of D Ag
determines if the person is Rh+ or Rh-
o D Antigen: 85%  Rh Positive
o d antigen: 15%  Rh Negative
o C antigen: 70%
o c antigen: 80%
o E antigen: 30%
o e antigen: 98%

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An introduction of Rh-positive blood(antigen) to HEMOSTASIS OR HAEMOSTASIS
Rh-negative individual, it will initiate production - Is a complex process which causes the
of antibody. bleeding process to stop. It refers to the
- No adverse reaction process of keeping blood within a
damaged blood vessel.
HEMOLYSIS - Following an injury to blood vessels
• If an individual is transfused with an several actions may help prevent blood
incompatible blood group, destruction of loss, including:
the red blood cells will occur. a) Vascular spasm
• This may result in the death of the recipient b) Platelet Plug formation
c) Blood clotting
HEMOLYTIC DISEASE OF THE NEWBORN (HDN)
➢ How it Occurs
A. Child is Rh positive, Mother is Rh
negative
B. During pregnancy fetal Rh positive
rbc’s escape into maternal
circulation
C. Mother produces antibodies to Rh (D)
pos child, Ab (Anti-D from the mother)
results in destruction of fetal D positive
RBCs Hemostasis is maintained in the body via three
➢ What is it? mechanisms:
o occurs when mother produces
anti-Rh antibodies that cross • Vascular Spasm – Damaged blood vessels
placenta and agglutination and constrict.
hemolysis of fetal erythrocytes • Platelet Plug Formation – Platelets adhere to
occurs damaged endothelium to form platelet plug
o can be fatal to fetus (primary hemostasis) and then degranulate.
o prevented if mother is treated with • Blood Coagulation – Clots form upon the
RhoGAM which contains conversion of fibrinogen to fibrin, and its
antibodies against Rh antigens addition to the platelet plug (secondary
➢ Right after birth of first baby, mother is hemostasis).
given a treatment
o Rh-immune Globulin (RhoGAM) BLEEDING TIME
▪ Coat D antigen, to not • Measures the ability of small blood
initiate any anti-D vessels to control free flow of blood after
production on mother’s injury
blood circulation, so that • Measure of platelet function and
on the next pregnancy, the vascular integrity
baby will not be hemolytic. • In vivo measurement of platelet
participation in small vessel hemostasis
✓ Screening test for detecting
disorders of platelet function and
von Willebrand Disease
✓ Directly affected by platelet
count and the ability of the
platelets to form a plug
• PURPOSE: provide a measure of the
platelet function in small vessel
hemostasis
• A prolonged bleeding time is not in itself
diagnostic of underlying platelet

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 disorders. It indicates the need for more DROP OR SLIDE METHOD
quantifiable test. Procedure:
1. Perform a skin puncture. Discard the first drop
DUKE’S METHOD of blood
- Not really reliable because it has many 2. Place 2 drops of blood on a clean glass slide.
Start timing
factors affecting prolonged bleeding
3. Draw the tip of the lancet across the drop of
time blood at 30-second interval until fibrin
o Thick/Vascularity of skin = prolonged threads cling to the tip
bleeding time 4. Stop timing and record the clotting time
▪ Prolonged bleeding time can
also mean low platelet count
or may indicate a disease
WHAT CAUSES THE ABSENCE OF INTRAVASCULAR
o Less thick/vascularity of skin = CLOTTING?
increase in bleeding time • In the absence of any vascular injury (intact
o Quality of blood vessels blood vessels), the intravascular clotting is
o Depth of puncture prevented through the presence of
endogenous anticoagulants (heparin) that
circulates in the body.
Procedure:
1. Cleanse the 3rd or 4th finger with 70% isopropyl
alcohol and allow it to dry WHAT IS THE IMPORTANCE OF DETERMINING THE
2. Make a relatively deep puncture with a CLOTTING TIME?
sterile blood lancet and start timing (3mm) • Screening test for all stages of the intrinsic
3. Blot the blood using filter paper every 30 coagulation pathway
seconds. • Monitor heparin therapy
NOTE: do not allow the filter paper to touch the
wound as this will hasten the bleeding time. HEMOSTASIS
(START TIMING IMMEDIATELY AFTER THE SKIN • Process in which several factors work
PUNCTURE) together or in sequence to stop the flow
4. Stop timing as bleeding ceases and record of blood from an injured blood vessel.
the bleeding time • Combination of clotting and lysing
Normal Value: 2-4 minutes
mechanisms that maintain the integrity of
the vascular system.
CLOTTING TIME
• Measure of all intrinsic clotting factors
3 ASPECTS OF HEMOSTASIS
in the absence of tissue factors.
1. Extravascular:
• Least effective test in the diagnosis of • Physical constriction of injured skin and
actual hemostasis failure tissues resulting in the release of tissue
• It has been replaced by PTT. (Partial juice which contains tissue
Thromboplastin Time) thromboplastin
• Used as a screening test to measure 2. Vascular:
all stages in the intrinsic coagulation • Constriction of injured blood vessel
system and to monitor heparin brought about by reflex constriction and
therapy. by the serotonin released by
o Heparin therapy is done if disintegrated platelets
patient have experienced 3. Intravascular:
heart attack/vascular system • Physical and biochemical changes
undergone by platelets and the
failure.
interaction of the different coagulation
• Takes longer time than bleeding time: factors
4-10mins or 3-6 mins ALL THESE ASPECTS REACT SIMULTANEOUSLY TO STOP
BLEEDING.

3 STEPS OF NORMAL HEMOSTASIS

• Vasoconstriction
• Platelet aggregation
• Fibrin formation

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