1.

Bleeding Time (BT)

Definition:

●​ Bleeding time measures the duration required for bleeding to stop after a small incision
is made in the skin. It evaluates the primary phase of hemostasis, particularly platelet
function and capillary integrity.

Purpose:

●​ To assess platelet function.

●​ To evaluate the vascular and platelet response to injury.
●​ Used as a diagnostic tool for bleeding disorders.

Normal Range:

●​ 2 to 7 minutes (depending on the method used).

Test Method:

Several methods can be used to measure bleeding time:

1.​ Duke's Method:

o​ A small puncture is made in the earlobe or fingertip.
o​ The blood is blotted every 30 seconds with filter paper until bleeding stops.
o​ The time is recorded from the puncture until the bleeding ceases.
2.​ Ivy’s Method (more standardized):
o​ A blood pressure cuff is inflated on the upper arm (typically at 40 mmHg) to
standardize the pressure.
o​ Small standardized incisions (2 to 3 mm deep) are made on the forearm.
o​ The blood is blotted every 30 seconds until bleeding stops, and the time is
recorded.
3.​ Modified Ivy’s Method:
o​ Same procedure as Ivy’s method, but a lancet is used to make multiple small cuts
instead of a blade incision.

Clinical Relevance:

●​ Prolonged Bleeding Time can indicate:

o​ Thrombocytopenia (low platelet count).
o​ Platelet dysfunction due to inherited disorders (e.g., Bernard-Soulier
syndrome, Glanzmann’s thrombasthenia) or acquired causes (e.g., aspirin and
NSAIDs affecting platelet function).
o​ von Willebrand disease, a disorder where platelets cannot adhere properly due to
a deficiency in von Willebrand factor.
o​ Scurvy (vitamin C deficiency), which weakens blood vessels.
 o​ DIC (Disseminated Intravascular Coagulation) or other coagulopathies.
●​ Shortened Bleeding Time is rarely seen but may indicate conditions such as
polycythemia or hypercoagulable states.

Limitations:

●​ Bleeding time is a non-specific test.

●​ It may vary based on operator technique, location of incision, and patient factors such as
skin thickness and vascular conditions.
●​ It does not reflect abnormalities in coagulation factors, making it less reliable in
diagnosing coagulation disorders.

2.Clotting Time (CT)

Definition:

●​ Clotting time measures the time required for blood to clot once it has been withdrawn
from the body. It evaluates the secondary phase of hemostasis, particularly the efficiency
of the coagulation cascade and the involvement of clotting factors.

Purpose:

●​ To assess the functionality of the intrinsic and common pathways of the coagulation
cascade (involving factors I, II, V, VIII, IX, X, XI, and XII).
●​ Helps identify disorders related to coagulation factor deficiencies (e.g., hemophilia).

Normal Range:

●​ 8 to 15 minutes (varies depending on the method and temperature).

Test Method:

There are different methods for measuring clotting time:

1.​ Capillary Tube Method (Wright's Method):

o​ A drop of blood is collected in a capillary tube.
o​ The tube is gently broken at intervals to check for clot formation.
o​ The time from the start of blood collection to the formation of a clot is recorded.
2.​ Lee and White Method (Whole Blood Clotting Time):
o​ Blood is collected in a glass tube and maintained at 37°C.
o​ The time required for the blood to clot is measured manually by tilting the tube
until a solid clot forms.
o​ This method measures clotting time as a reflection of intrinsic pathway activation.
3.​ Siliconized Tube Method:
 o​ Blood is collected in silicon-coated tubes to prevent early clotting due to surface
contact.
o​ The clotting time is measured in a similar manner to other methods.

Clinical Relevance:

●​ Prolonged Clotting Time can indicate:

o​ Hemophilia (deficiency in clotting factors VIII, IX, or XI).
o​ von Willebrand disease.
o​ Liver disease (since clotting factors are produced in the liver).
o​ Vitamin K deficiency, which affects the synthesis of clotting factors II, VII, IX,
and X.
o​ Heparin therapy or other anticoagulant therapies.
o​ Disseminated Intravascular Coagulation (DIC), which leads to widespread
clotting factor consumption.
●​ Shortened Clotting Time:
o​ Rarely seen, but may occur in conditions of hypercoagulability, where blood clots
excessively, increasing the risk of thrombosis.

Limitations:

●​ Clotting time is influenced by temperature and surface contact (glass vs. siliconized
tubes).
●​ It is less precise compared to more advanced coagulation tests like Prothrombin Time
(PT) and Activated Partial Thromboplastin Time (aPTT), which provide more
detailed information on specific coagulation pathways.

Comparison of Bleeding Time and Clotting Time:

Feature Bleeding Time Clotting Time

Coagulation factor activity (intrinsic
Assesses Platelet function, capillary integrity
pathway)
Normal Range 2-7 minutes 8-15 minutes
Common Methods Duke’s, Ivy’s, Modified Ivy’s Capillary tube, Lee-White method
Prolonged Time Platelet dysfunction, vascular defects, Hemophilia, liver disease, DIC,
Indicates von Willebrand disease anticoagulation therapy
Detects platelet disorders, vascular
Clinical Use Detects clotting factor deficiencies
issues
3.Determination of Hemoglobin (Hb)

Definition:

●​ Hemoglobin (Hb) is a protein in red blood cells responsible for carrying oxygen from the
lungs to the tissues and transporting carbon dioxide from the tissues back to the lungs.
The measurement of hemoglobin levels helps assess the oxygen-carrying capacity of the
blood.

Purpose:

●​ To diagnose and monitor conditions such as anemia, polycythemia, and blood loss.
●​ To assess the oxygen-carrying capacity of the blood.

Normal Range:

●​ Males: 13.8 – 17.2 g/dL

●​ Females: 12.1 – 15.1 g/dL
●​ Newborns: 14 to 24 g/dL

Test Methods:

1.​ Sahli's Method (Acid Hematin Method):

o​ Principle: Hemoglobin is converted to acid hematin by adding dilute HCl, and
the color intensity is compared to a reference standard.
o​ Procedure:
▪​ Add 0.1N HCl to the graduated Sahli tube up to the mark.
▪​ Add a few drops of blood (using a pipette) to the acid solution.
▪​ The hemoglobin is converted to brown-colored acid hematin.
▪​ The color is matched against a reference standard using a
hemoglobinometer.
o​ Advantages: Simple and economical.
o​ Disadvantages: Subjective, as it depends on color matching and is less accurate
compared to modern methods.
2.​ Cyanmethemoglobin Method (most commonly used in laboratories):
o​ Principle: Hemoglobin is converted into cyanmethemoglobin by the addition of
potassium cyanide and potassium ferricyanide. The concentration is measured
spectrophotometrically.
o​ Procedure:
▪​ Blood is mixed with Drabkin’s reagent (potassium cyanide and
ferricyanide).
▪​ Hemoglobin is converted to cyanmethemoglobin.
▪​ The absorbance of the solution is measured at a wavelength of 540 nm
using a spectrophotometer.
o​ Advantages: Accurate, reliable, and standardized.
 o​ Disadvantages: Requires specialized equipment (spectrophotometer) and
handling of hazardous chemicals (cyanide).
3.​ Automated Hemoglobinometry:
o​ Automated analyzers can measure hemoglobin levels rapidly and precisely, using
light absorption and other principles like hemiglobincyanide formation.

Clinical Relevance:

●​ Low Hemoglobin (Anemia):

o​ Causes include nutritional deficiencies (iron, vitamin B12, folate), chronic
diseases, bone marrow disorders, and acute or chronic blood loss.
o​ Symptoms include fatigue, weakness, pale skin, shortness of breath, and
dizziness.
●​ High Hemoglobin (Polycythemia):
o​ Causes include dehydration, chronic lung disease, smoking, living at high
altitudes, and polycythemia vera.
o​ Symptoms include headaches, dizziness, and a ruddy complexion.

4.Erythrocyte Sedimentation Rate (ESR)

Definition:

●​ Erythrocyte Sedimentation Rate (ESR) measures the rate at which red blood cells
(erythrocytes) settle in a vertical tube over a specified period (usually one hour). It is a
non-specific marker of inflammation and is often used to detect inflammatory processes,
infections, and autoimmune diseases.

Purpose:

●​ To detect and monitor inflammatory conditions, infections, autoimmune diseases, and

certain cancers.
●​ To assess disease activity and monitor the response to treatment.

Normal Range:

●​ Men: 0-15 mm/hour

●​ Women: 0-20 mm/hour
●​ Children: 0-10 mm/hour
●​ Newborns: 0-2 mm/hour
●​ Elderly: Values tend to increase with age, often considered normal up to 30 mm/hour.

Test Methods:

1.​ Westergren Method (most widely used):

 o​ Procedure:
▪​ Blood is drawn and mixed with an anticoagulant (usually sodium citrate).
▪​ The blood is placed in a vertical, graduated Westergren tube.
▪​ The distance the RBCs settle in the tube (in millimeters) is measured after
one hour.
o​ Advantages: Simple, sensitive, and the international standard method for ESR
determination.
o​ Disadvantages: Non-specific and may be influenced by various factors such as
anemia, pregnancy, or other physiological changes.
2.​ Wintrobe Method:
o​ Similar to the Westergren method, but uses a shorter tube (100 mm).
o​ The blood sample is directly drawn into the tube without any dilution with an
anticoagulant.
o​ Less sensitive compared to the Westergren method, and thus not used as
commonly in clinical practice.

Factors Affecting ESR:

●​ Increased ESR:
o​ Inflammatory diseases (e.g., rheumatoid arthritis, systemic lupus
erythematosus).
o​ Infections (e.g., bacterial endocarditis, tuberculosis).
o​ Autoimmune diseases.
o​ Cancers (e.g., lymphoma, multiple myeloma).
o​ Pregnancy (ESR is physiologically elevated due to increased fibrinogen).
o​ Anemia (low RBC count leads to less aggregation and faster settling).
●​ Decreased ESR:
o​ Polycythemia (increased RBC count).
o​ Sickle cell anemia (abnormal RBC shape).
o​ Congestive heart failure.

Clinical Relevance:

●​ Increased ESR can be indicative of a wide range of conditions such as infections,

inflammatory diseases (e.g., lupus, rheumatoid arthritis), and some cancers.
o​ It is a sensitive but non-specific test, meaning it can detect inflammation but
cannot pinpoint the exact cause.
●​ Normal ESR does not rule out disease, as some conditions like angina, viral hepatitis,
and thyroid disease may show normal ESR despite active disease.
●​ ESR vs CRP (C-reactive protein): While both tests measure inflammation, CRP is more
specific for acute inflammation and infection, whereas ESR reflects more chronic
inflammation.

Comparison Between Hemoglobin Determination and ESR:

 ESR (Erythrocyte Sedimentation
Feature Hemoglobin Determination
Rate)
Measures the oxygen-carrying Measures the rate of RBC sedimentation,
Purpose
capacity of blood indicates inflammation
Males: 13.8–17.2 g/dL; Females:
Normal Range Males: 0-15 mm/h; Females: 0-20 mm/h
12.1–15.1 g/dL
Cyanmethemoglobin, Sahli's,
Methods Westergren Method, Wintrobe Method
Automated Methods
Conditions Inflammatory diseases, infections,
Anemia, polycythemia, blood loss
Indicated autoimmune disorders
Oxygen-carrying capacity
Common Use Inflammation detection and monitoring
assessment

4.Packed Cell Volume (PCV) (Hematocrit)

Detailed Procedure:

1.​ Sample Collection: Blood is drawn from a patient and mixed with an anticoagulant
(usually EDTA) to prevent clotting.
2.​ Centrifugation:
o​ The blood is placed in a capillary tube or a specialized test tube.
o​ The tube is spun in a centrifuge at high speed (usually 10,000-12,000 revolutions
per minute) for 5-10 minutes.
o​ This separates the blood into three distinct layers:
▪​ Bottom layer: Packed RBCs.
▪​ Middle layer (Buffy coat): A thin layer of white blood cells and platelets.
▪​ Top layer: Plasma.
3.​ Measurement:
o​ After centrifugation, the percentage of the total blood volume that consists of
packed red cells is measured using a special hematocrit reader or by comparing
the height of the packed RBC column with the total column height.

Clinical Significance of PCV:

●​ Low PCV (below normal):

o​ Anemia: Decreased RBC production (e.g., iron deficiency, B12 or folate
deficiency).
o​ Blood loss: Acute hemorrhage.
o​ Chronic disease: Chronic kidney disease, where erythropoietin (EPO) levels are
reduced.
o​ Hemolysis: Premature destruction of RBCs (e.g., in hemolytic anemia).
●​ High PCV (above normal):
o​ Polycythemia vera: A bone marrow disorder causing an overproduction of
RBCs.
 o​ Dehydration: Reduced plasma volume makes PCV appear falsely high.
o​ Chronic hypoxia: Conditions like chronic obstructive pulmonary disease
(COPD) or living at high altitudes can stimulate the body to produce more RBCs.
●​ Interferences:
o​ Pregnancy: PCV tends to decrease due to hemodilution.
o​ Acute stress or exercise: Can cause temporary increases in PCV.

5.Total RBC Count

RBC Count Detailed Procedure:

1.​ Sample Collection: A venous blood sample is collected in an anticoagulant tube.

2.​ Automated Counting:
o​ In modern laboratories, RBC count is part of a complete blood count (CBC),
performed by automated analyzers.
o​ These analyzers use optical light scatter or electrical impedance methods to
count RBCs as they flow through a small aperture.
3.​ Manual Counting:
o​ If an automated analyzer is not available, a manual count can be done using a
hemocytometer.
o​ Blood is diluted and placed in a counting chamber under a microscope, and the
number of RBCs is counted in a specific grid area, followed by calculations to
estimate the total count per microliter.

Clinical Implications:

●​ Low RBC Count:

o​ Iron-deficiency anemia: The most common form of anemia worldwide.
o​ Aplastic anemia: Bone marrow failure leads to decreased RBC production.
o​ Hemorrhagic anemia: Due to acute or chronic blood loss (e.g., trauma,
gastrointestinal bleeding).
●​ High RBC Count:
o​ Primary polycythemia (Polycythemia vera): A myeloproliferative disorder
leading to excessive RBC production.
o​ Secondary polycythemia: Due to chronic hypoxia (e.g., COPD, sleep apnea),
high altitudes, or EPO-producing tumors.

6.Total WBC Count

Detailed Procedure:

1.​ Sample Collection: Blood is collected in an EDTA tube to prevent clotting.

 2.​ Automated Counting:
o​ Modern hematology analyzers perform a complete blood count (CBC), which
includes the total WBC count.
o​ These machines use either flow cytometry, optical light scatter, or electrical
impedance to identify and count WBCs in a specific volume of blood.
3.​ Manual Counting (rarely used now):
o​ Using a hemocytometer, blood is diluted with a special diluent that lyses the red
cells but preserves the white cells for counting under a microscope.

Clinical Implications:

●​ Leukocytosis (High WBC count):

o​ Infections: Most common cause of elevated WBC count, especially in bacterial
infections.
o​ Leukemia: A significantly high WBC count may indicate blood cancer, such as
acute leukemia.
o​ Inflammation: Conditions like rheumatoid arthritis or inflammatory bowel
disease (IBD) may cause increased WBC counts.
o​ Stress or Trauma: Physical stress, injury, or burns can lead to elevated WBCs.
●​ Leukopenia (Low WBC count):
o​ Viral infections: Commonly seen in viral infections like influenza, HIV, or
hepatitis.
o​ Bone marrow suppression: Chemotherapy, radiation, or certain medications can
decrease WBC production.
o​ Autoimmune diseases: Conditions like lupus or rheumatoid arthritis can lead to
low WBC counts.

7.Differential Count of WBCs

Detailed Procedure:

1.​ Manual Differential:

o​ A blood smear is prepared by placing a drop of blood on a glass slide and
spreading it thinly.
o​ The slide is stained with Wright-Giemsa stain, which helps differentiate various
WBCs based on their size, nucleus shape, and cytoplasmic granules.
o​ Under a microscope, a technician manually counts 100 WBCs and records the
percentage of each type.
2.​ Automated Differential:
o​ Modern analyzers use techniques like flow cytometry to differentiate between
different types of WBCs based on their size, granularity, and other physical
characteristics.

Clinical Implications of WBC Subtypes:

 1.​ Neutrophils (40-70%):
o​ Neutrophilia (High neutrophils): Commonly seen in bacterial infections,
trauma, and stress.
o​ Neutropenia (Low neutrophils): Seen in viral infections, bone marrow
suppression, and certain medications (e.g., chemotherapy).
2.​ Lymphocytes (20-40%):
o​ Lymphocytosis (High lymphocytes): Common in viral infections (e.g.,
mononucleosis, hepatitis) and in chronic lymphocytic leukemia (CLL).
o​ Lymphopenia (Low lymphocytes): Can occur in HIV/AIDS, steroid use, or
severe systemic infections.
3.​ Monocytes (2-8%):
o​ Monocytosis (High monocytes): Seen in chronic infections (e.g., tuberculosis,
syphilis), recovery from acute infections, and certain cancers like Hodgkin’s
lymphoma.
4.​ Eosinophils (1-4%):
o​ Eosinophilia (High eosinophils): Associated with allergic conditions (e.g.,
asthma, eczema), parasitic infections, and some autoimmune diseases.
5.​ Basophils (0.1-1%):
o​ Basophilia (High basophils): Seen in allergic reactions, chronic myeloid
leukemia (CML), and some inflammatory conditions (e.g., ulcerative colitis).

Additional Special Considerations

●​ Factors Influencing Total WBC and Differential Count:

o​ Medications: Some drugs (e.g., steroids, chemotherapy) can affect WBC counts
and their differentials.
o​ Physiological Stress: Factors like pregnancy, surgery, or trauma can temporarily
alter WBC counts.
o​ Smoking: Chronic smokers often have elevated WBC counts due to ongoing
low-grade inflammation.
8.Hemostasis

Definition: Hemostasis is the process that prevents and stops bleeding, or hemorrhage. It
involves a complex interaction between blood vessels, platelets, and plasma proteins to form a
stable blood clot at the site of vascular injury.

Stages of Hemostasis:

1.​ Vascular Spasm (Vasoconstriction):

o​ Immediate response to blood vessel injury.
o​ Blood vessels constrict to reduce blood flow to the affected area.
o​ Mediated by the release of endothelin from endothelial cells and other factors
such as serotonin and thromboxane A2.
o​ This temporary reduction in blood flow minimizes blood loss.
2.​ Platelet Plug Formation:
o​ Adhesion: Platelets adhere to exposed collagen fibers in the damaged vessel wall
through the interaction of von Willebrand factor (vWF), which binds platelets
and collagen.
o​ Activation: Adhered platelets become activated and release granules containing
ADP, thromboxane A2, and other mediators that recruit more platelets to the
site.
o​ Aggregation: The activated platelets express surface receptors (e.g., GPIIb/IIIa)
that bind fibrinogen, leading to platelet aggregation and the formation of a
temporary platelet plug.
3.​ Coagulation Cascade:
o​ This stage involves a series of enzymatic reactions leading to the conversion of
fibrinogen (a soluble plasma protein) into fibrin (an insoluble protein).
 o​ Intrinsic Pathway: Triggered by damage to the blood vessel, involving factors
XII, XI, IX, and VIII.
o​ Extrinsic Pathway: Triggered by tissue factor (TF) released from damaged
tissues, involving factor VII.
o​ Both pathways converge on the common pathway, leading to the activation of
thrombin (factor IIa), which converts fibrinogen to fibrin.
o​ Fibrin strands form a mesh that stabilizes the platelet plug.
4.​ Clot Retraction and Repair:
o​ The clot contracts through the action of platelets, which pull on the fibrin threads,
resulting in clot retraction.
o​ This process helps to reduce the size of the damaged area and aids in tissue repair.
o​ Tissue growth factors released by platelets and endothelial cells promote healing
of the blood vessel wall.
5.​ Fibrinolysis:
o​ Once the vessel is repaired, the clot is removed by a process called fibrinolysis.
o​ Plasminogen is activated to plasmin, which digests fibrin and dissolves the clot.
o​ This ensures that normal blood flow is restored.

Clinical Relevance:

●​ Proper hemostasis is crucial to prevent excessive bleeding (hemorrhage) or inappropriate

clotting (thrombosis).

9.Bleeding Disorders in Humans

Bleeding disorders can be classified into two main categories: inherited and acquired.

Inherited Bleeding Disorders:

1.​ Hemophilia:
o​ A genetic disorder where blood doesn't clot properly due to a deficiency of
clotting factors.
o​ Hemophilia A: Caused by a deficiency of factor VIII.
o​ Hemophilia B: Caused by a deficiency of factor IX.
o​ Symptoms: Excessive bleeding, easy bruising, joint pain, and swelling.
2.​ Von Willebrand Disease:
o​ The most common inherited bleeding disorder caused by a deficiency or
dysfunction of von Willebrand factor (vWF).
o​ Symptoms: Mucosal bleeding (nosebleeds, gum bleeding), easy bruising, heavy
menstrual periods.
3.​ Factor V Leiden:
o​ A genetic mutation that increases the risk of thrombosis but can also lead to
bleeding disorders when combined with other factors.
4.​ Other Rare Disorders:
 o​ Conditions like Bernard-Soulier syndrome and Glanzmann's thrombasthenia
affect platelet function.

Acquired Bleeding Disorders:

1.​ Vitamin K Deficiency:

o​ Vitamin K is crucial for the synthesis of several clotting factors (II, VII, IX, and
X).
o​ Can result from malabsorption disorders, liver disease, or certain medications
(e.g., anticoagulants).
2.​ Liver Disease:
o​ The liver produces most clotting factors, so liver dysfunction can lead to
coagulopathy.
3.​ Anticoagulant Medications:
o​ Medications like warfarin and heparin are used to prevent clotting but can also
lead to excessive bleeding if not monitored properly.
4.​ Disseminated Intravascular Coagulation (DIC):
o​ A serious condition characterized by widespread activation of the coagulation
cascade, leading to both bleeding and thrombosis.
o​ Can be triggered by infections, trauma, or pregnancy complications.

Symptoms of Bleeding Disorders:

●​ Easy bruising.
●​ Prolonged bleeding from cuts.
●​ Nosebleeds (epistaxis).
●​ Heavy menstrual periods (menorrhagia).
●​ Joint pain and swelling in hemophilia.

10.Hemolytic Disease of the Newborn (HDN)

Definition: Hemolytic Disease of the Newborn (HDN) is a condition in which a mother’s

antibodies attack her baby’s red blood cells, leading to hemolysis (destruction of red blood cells)
and resulting anemia.

Causes:

1.​ Rh Incompatibility:
o​ Occurs when an Rh-negative mother carries an Rh-positive fetus.
o​ During delivery or if there is any bleeding during pregnancy, the mother may be
sensitized to the Rh antigen, producing antibodies against it.
o​ In subsequent pregnancies, these antibodies can cross the placenta and attack the
fetal red blood cells.
2.​ ABO Incompatibility:
 o​ Occurs when a mother with type O blood has a fetus with type A, B, or AB blood.
o​ Maternal antibodies against A or B antigens can cross the placenta and cause
hemolysis.

Clinical Features:

●​ Anemia: Due to the destruction of red blood cells.

●​ Hyperbilirubinemia: Increased levels of bilirubin in the blood, leading to jaundice
(yellowing of the skin and eyes).
●​ Hydrops Fetalis: Severe cases can result in fluid accumulation in the fetus (edema).
●​ Kernicterus: A serious complication where excess bilirubin deposits in the brain,
causing neurological damage.

Diagnosis:

●​ Maternal Blood Tests: Check for Rh antibodies.

●​ Fetal Blood Tests: Blood type and direct Coombs test to check for hemolysis.
●​ Ultrasound: To assess for signs of hydrops or other complications.

Management:

1.​ Prevention:
o​ Rh immunoglobulin (RhoGAM) is given to Rh-negative mothers during
pregnancy and after delivery to prevent sensitization.
2.​ Intrauterine Blood Transfusion:
o​ For severely affected fetuses, an intrauterine transfusion may be performed to
replace the destroyed red blood cells.
3.​ Postnatal Treatment:
o​ Phototherapy: To reduce bilirubin levels.
o​ Exchange transfusion: In severe cases, to remove bilirubin and antibodies from
the newborn’s blood.

Prognosis:

●​ With proper management, most babies recover well. However, severe cases of HDN can
lead to significant morbidity and mortality.

Platelet Count

Definition: The platelet count measures the number of platelets in a volume of blood, typically
expressed in thousands per microliter (x10³/µL). Platelets (thrombocytes) are small cell
fragments critical for blood clotting and hemostasis.

Normal Range:

●​ 150,000 to 450,000 platelets per microliter of blood.

Procedure:

1.​ Sample Collection:

o​ A blood sample is collected in a tube containing an anticoagulant (e.g., EDTA).
2.​ Automated Counting:
o​ Most platelet counts are performed using automated hematology analyzers that
utilize methods like optical light scatter or electrical impedance to count and
size platelets.
3.​ Manual Counting (if necessary):
o​ A blood smear is prepared, stained (e.g., with Wright’s stain), and examined under
a microscope. The number of platelets in several fields is counted and averaged to
estimate the count.

Clinical Significance:

●​ Low Platelet Count (Thrombocytopenia):

o​ Causes: Bone marrow disorders (e.g., aplastic anemia), increased destruction
(e.g., immune thrombocytopenic purpura (ITP), drug-induced), or sequestration in
the spleen (e.g., hypersplenism).
o​ Symptoms: Increased bleeding tendencies, easy bruising, petechiae (small red or
purple spots), and prolonged bleeding from cuts.
●​ High Platelet Count (Thrombocytosis):
o​ Causes: Reactive thrombocytosis (due to infection, inflammation, iron deficiency,
or after surgery) or primary thrombocytosis (e.g., essential thrombocythemia).
o​ Symptoms: Increased risk of thrombosis (blood clots), which can lead to stroke
or heart attack.

11.Reticulocyte Count

Definition: The reticulocyte count measures the number of reticulocytes (immature red blood
cells) in the blood, indicating bone marrow activity and the body’s response to anemia.

Normal Range:

●​ Typically 0.5% to 2.5% of total RBCs in adults. This can be higher in children and
newborns.

Procedure:

1.​ Sample Collection:

o​ A blood sample is drawn into an anticoagulant tube (EDTA).
2.​ Staining:
o​ The sample is prepared as a smear and stained using a special stain (e.g., New
Methylene Blue or Supravital stain) that highlights reticulocytes.
 3.​ Counting:
o​ The stained slide is examined under a microscope, and the reticulocytes are
counted and reported as a percentage of the total RBC count.
o​ Automated analyzers can also measure reticulocyte counts.

Clinical Significance:

●​ Low Reticulocyte Count:

o​ Suggests insufficient bone marrow response, which may be seen in aplastic
anemia or chronic disease.
●​ High Reticulocyte Count:
o​ Indicates increased production of RBCs, often due to:
▪​ Hemolytic Anemia: The body compensates for the loss of red blood cells.
▪​ Acute Blood Loss: Following trauma or surgery.
▪​ Recovery from Iron Deficiency Anemia: After treatment.

12.Absolute Eosinophil Count

Definition: The absolute eosinophil count measures the number of eosinophils (a type of white
blood cell involved in allergic responses and parasitic infections) in a microliter of blood.

Normal Range:

●​ Typically 0 to 500 eosinophils per microliter of blood.

Procedure:

1.​ Sample Collection:

o​ A blood sample is collected in an anticoagulant tube (EDTA).
2.​ Automated Counting:
o​ Most counts are part of a complete blood count (CBC) performed by automated
analyzers that measure eosinophils directly.
3.​ Manual Counting (if needed):
o​ A blood smear is prepared, stained, and examined under a microscope.
Eosinophils are identified by their distinct bilobed nucleus and prominent
granules.

Clinical Significance:

●​ High Absolute Eosinophil Count (Eosinophilia):

o​ Causes:
▪​ Allergic Reactions: Conditions like asthma, eczema, or allergic rhinitis.
▪​ Parasitic Infections: Especially helminthic infections.
 ▪​ Autoimmune Diseases: Conditions such as eosinophilic granulomatosis
with polyangiitis (Churg-Strauss syndrome).
▪​ Certain Cancers: Some leukemias and lymphomas.
●​ Low Absolute Eosinophil Count:
o​ Usually not of clinical significance and may occur in stress or acute infections.

Summary Chart

Parameter Normal Range Clinical Implications

150,000 – Thrombocytopenia (bleeding risks), Thrombocytosis
Platelet Count
450,000/µL (thrombosis risks)
0.5% – 2.5% of Low: Insufficient marrow response; High: Increased
Reticulocyte Count
total RBCs RBC production
Absolute Eosinophil Eosinophilia (allergic reactions, parasitic infections),
0 – 500/µL
Count Low: Typically not significant

13.Blood Banking Technology

Definition: Blood banking technology encompasses the processes involved in the collection,
testing, processing, storage, and distribution of blood and blood products for transfusion.

Key Components of Blood Banking:

1.​ Collection:
o​ Donor Recruitment:
▪​ Potential donors are screened for eligibility based on health history,
lifestyle factors, and risk of infectious diseases.
▪​ Education is provided about the donation process and its benefits.
o​ Types of Blood Donations:
▪​ Whole Blood Donation: Donors give a full unit of blood, which can be
separated into components (RBCs, plasma, platelets).
▪​ Apheresis: A process where specific blood components (e.g., platelets,
plasma) are collected, and the remaining blood is returned to the donor.
This method allows for targeted collection of the needed components.
▪​ Autologous Donation: Patients donate their own blood before a scheduled
surgery, minimizing the risk of transfusion reactions.
o​ Collection Procedure:
▪​ A sterile needle is inserted into a vein (usually in the arm) to draw blood.
▪​ Blood is collected into sterile bags containing anticoagulants to prevent
clotting.
▪​ Donation typically takes about 8-10 minutes, with a total volume of
approximately 450-500 mL for whole blood.
 o​ Donor Monitoring:
▪​ Vital signs (blood pressure, pulse, temperature) are monitored before and
after donation to ensure donor safety.
▪​ Donors are given post-donation care, including snacks and fluids to
replenish lost volume.
▪​
2.​ Testing:
o​ Screening: All donated blood is screened for infectious diseases (HIV, hepatitis B
and C, syphilis, West Nile virus, etc.) and blood type.
o​ Blood Typing: ABO and Rh typing are performed to identify the blood group and
ensure compatibility for transfusions.
o​ Crossmatching: A test is performed between donor blood and recipient serum to
ensure compatibility before transfusion.
3.​ Processing:
o​ After collection, whole blood is processed to separate it into its components: red
blood cells (RBCs), plasma, platelets, and cryoprecipitate.
o​ This is typically done using a centrifuge, which spins the blood to separate
components based on density.

Storage:

1.​ Storage Conditions:

o​ Red Blood Cells (RBCs):
▪​ Stored at 2-6°C (refrigerated).
▪​ Shelf life is typically 42 days with proper preservation techniques.
o​ Platelets:
▪​ Stored at 20-24°C with constant agitation.
▪​ Shelf life is typically 5-7 days.
o​ Plasma:
▪​ Frozen within hours of collection at -18°C or colder.
▪​ Can be stored for up to 1 year.
o​ Cryoprecipitate:
▪​ Derived from plasma and stored at -18°C or colder.
▪​ Shelf life is also 1 year.
2.​ Storage Systems:
o​ Blood banks use specialized refrigerators, freezers, and platelet incubators that
maintain optimal temperatures and conditions for blood components.
o​ Monitoring Systems: Temperature monitoring devices ensure that storage
conditions remain stable. Alarms are set for deviations to prevent spoilage.
3.​ Inventory Management:
o​ Blood banks employ inventory management systems to track blood products,
ensuring that the supply meets the needs of patients and minimizing wastage.
o​ Expiration dates are closely monitored, and older blood products are often
prioritized for use.

Clinical Application:
●​ Blood transfusions are critical for various medical conditions, including:
o​ Trauma and surgical patients.
o​ Patients with anemia, bleeding disorders, or chronic diseases.
o​ During pregnancy complications (e.g., severe hemorrhage).