CHAPTER 5: INTRODUCTION TO HEMATOLOGY & HEMOSTASIS

● Blood - nutritive fluid circulating the body that carries nutrients, oxygen and waste materials to

and from tissues.

● Hematology - the study of the formed elements of blood and blood-forming tissues; “haima”

(blood) + “logos” (study)

● Hemostasis - complex interaction between blood vessels, platelets, and biochemical factors

in plasma in a way that they create and dissolve clots.

Functions of Blood

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● Respiration. Oxygen is carried throughout the body by the red blood cells (RBCs).

● Nutrition. Nutrients are carried throughout the body.

● Excretion. Blood has an excretory function in the kidneys; excreted as urine.

● Buffer. Sudden change in pH levels is prevented.

● Transportation. Hormones and other endocrine substances are transported throughout the

body.

● Maintenance. Constant body temperature and slight alkalinity of tissues is maintained.

● Protection. White blood cells protect the body from infections.

● Hemostasis. Platelets and coagulation factors help stop bleeding.

Characteristics of Blood

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● Composition. Blood is 6-8% of total body weight.

● Amount. 6 liters in normal adults

● Acidity. 7.35-7.45 pH; if blood has a pH <7.35, it is considered acidosis. If pH is >7.45, it is

alkalosis.

● Consistency. Thick and viscous due to protein and cells.

● Fluid. Blood is fluid due to heparin, a natural anticoagulant.

● Color. Red due to hemoglobin.

● Smell. Fishy odor due to iron found in red blood cells.

● Taste. Sweet and salty due to sodium chloride and glucose.

● Volume. 50-60% of blood volume is plasma, which contains water, proteins, carbohydrates,

vitamins, hormones, enzymes, lipids, salt, minerals, electrolytes, and non-protein hydrogen.

Blood = Formed elements (45%) + Liquid elements (55%)

Ratio: 1 (WBCs): 30 (Platelets): 500 (RBCs)

Origin of Blood Cells

All blood cells are derived from stem cells that replicate and differentiate. Hematopoiesis is the

production of blood.

Cellular Component/Formed Elements of Blood

Red Blood Cells White Blood Cells Platelets

(Erythrocytes) (Leukocytes) (Thrombocytes)

Functions Respiration Protection Hemostasis

● Biconcave ● Grouped ● Small, blue or

● Central pallor is according to colorless bodies
⅓ cell diameter, nuclear with red or
which is 7-9 um structure, purple
in diameter. granulocyte granules.
Characteristics
content/ ● 1-2 um
● function diameter,
varied in
shape; round
to
cigar-shaped

Anemia - decreased Leukocytosis - increased Thrombocytosis -

production, abnormal production increased production
Associated maturation, increased Leukopenia - decreased Thrombocytopenia -
disorders
destruction, blood loss production decreased production
Erythrocytosis - Leukemia - abnormality/ Thrombasthenia -
increased production malignancy abnormal function

Neutrophils,
Subgroups None lymphocytes, None
monocytes, eosinophils,
basophils

White Blood Cells

Neutrophils Lymphocytes Monocytes Eosinophils Basophils

Present in Bacterial Viral infections Bacterial Allergic/parasi Allergic

infections infections t-ic infections reactions

Releases None None None Antihistamine Histamine

Polymorphonu Mononuclear, Mononuclear, Polymorphonu Polymorphonu

- clear with no visible amorphous -clear with clear with
Characteristics lavender granules, cytoplasm uniform-sized large,
granules scanty, with ground orange-red irregular-sized,
light-to-dark glass granules in blue-black
blue appearance cytoplasm granules
cytoplasm
Hemostasis

3 Components of Hemostasis

● Extravascular - tissues; release proteins to initiate coagulation

● Vascular - blood vessels

● Intravascular - platelets/plasma proteins

Coagulation - plasma proteins, tissue factors, and calcium interact on the surface of platelets to form

a fibrin clot.

Fibrinolysis - fibrin clot is dissolved

4 Phases of Hemostasis

1. Vasoconstriction - regulates blood flow to wound.

2. Platelet plug formation

3. Coagulation - stabilizes platelet plug and forms fibrin clot.

4. Fibrinolysis - fibrin clot is dissolved

The Hematology Laboratory

Capillary or venous blood are the possible specimens used. Meanwhile, ethylenediaminetetraacetic

acid (EDTA) and 3.2% buffered sodium citrate are the most common anticoagulants.

Complete Blood Count (CBC) - most frequently-ordered test. An improved neubauer is used to count

the cells, while a thoma pipette is used to dilute blood.

● Red blood cell count

● Hemoglobin - hemoglobin determination; absorbent and hemoglobin determination are

directly proportional.

● Hematocrit - collected using a capillary pipette and processed in microhematocrit centrifuge.

● White Blood Cell count

● WBC Differential

● RBC Morphology

○ Mean Corpuscular Volume (MCV) - measures sizes of RBCs

○ Mean Corpuscular Hemoglobin (MCH) - measures color of RBCs

○ Mean Cell Hemoglobin Content (MCHC) - measures concentration of whole cell

● Estimated platelet count

Other tests:

● Platelet count - normal values are 140-440 x 10^9/L

● Erythrocyte sedimentation rate - test for inflammation; non-specific test

● Coagulation tests - prothrombin time, activated partial thromboplastin time, clotting/bleeding

time
CHAPTER 6: INTRODUCTION TO IMMUNOLOGY-SEROLOGY

● Immunology - study of the immune system and its response to any foreign substance

introduced into the body.

○ Roles of the immune system - recognize and protect body from pathogens, remove

abnormal/damaged host cells, repair damaged tissues

● Serology - the study of antigen-antibody reactions.

● Antigen/Immunogen - a substance that triggers a disease.

● Antibody/Immunoglobulin - neutralizes antigens and destroys infected cells.

○ IgG - most abundant in blood; present in chronic infections

○ IgM - present in acute infections

○ IgA - present in secretions

○ IgE - found in mast cells (basophils)

○ IgD - found on the surface of B-cells

Immune System

● Primary immune organs - bone marrow and thymus (responsible for T-cell production)

● Secondary immune organs - lymph nodes, spleen, Mucosa-associated lymphoid tissue

(MALT), tonsils, Cutaneous-associated lymphoid tissue (CALT)

2 Branches of Immunity

Cellular Humoral

White blood cells Acute phase proteins,

lysozymes, complement
Natural/innate/non-specific
(present at birth) Processes:
● Inflammation
● Phagocytosis
● Complement activation

Lymphocytes - T and B Antibody

Acquired/adaptive/specific lymphocytes
(not present at birth)
Processes:
● T-cell response
● B-cell response

Body’s Defense Mechanism

First Line of Defense Second Line of Defense Third Line of Defense

● Physical barriers - skin ● Phagocytosis ● T-cell activation

and hair ● Inflammation ● Antibody production
● Complement by B cells
activation ● Antibody neutralization
● Natural killer cell
activation
● Interferons

Host factors affecting immune system: nutritional status, hormones, genetics, age, race, exercise level,

disease/injury
Immunologic Diseases

● Immunodeficiency - loss/impaired function of one or more major components of the immune

system; increased risk of unusual or recurrent infections.

● Hypersensitivity - exaggerated response to an innocuous antigen resulting in gross tissue

changes dangerous to the host.

● Autoimmune diseases - immune responses are directed against antigens found in the host's

body.

Serologic Test Principles

● Agglutination - forms a visible reaction caused by large particles.

● Precipitation - caused by soluble antigens.

● Labeled immunoassay

● Molecular biology techniques - detect specific bacteria or viruses.

Examples of Tests Performed in Laboratory

● HBsAg Test - test for Hepatitis B

● Rapid Plasma Reagin - Syphilis

● Antistreptolysin O test - test for antibodies against Streptolysin O

● Tests for HIV - ELISA (screening), Western Blott (confirmatory)

CHAPTER 7: INTRODUCTION TO BLOOD BANK & TRANSFUSION MEDICINE

● Blood banking - collecting, storing, and processing blood and distribution of RBCs and blood

components.

● Immunohematology - the study of the immunologic responses to blood components.

● Transfusion medicine - practice and techniques involving the replacement of RBCs and blood

components.

Blood Group Systems - a group of antigens produced at a single gene locus or closely linked loci.

Each antigen is given a six-digit numerical terminology by the International Society of Blood

Transfusion (ISBT).

● Major Blood Groups

a.) ABO

b.) Rh

● Minor Blood Groups

Blood Transfusion

1. Donor Screening. 2-3 months interval.

a.) Medical history questionnaire

b.) Physical - general appearance, weight, temperature, pulse, blood pressure, hemoglobin,

skin lesions

c.) Serological tests - ABO/Rh, antibody screen, HBsAg, Anti-HCV, Anti-HIV ½, Anti-HTLV I/II

(human T-lymphocytropic virus), serologic test for syphilis, malarial smear

 2. Blood Collection. Collection of 450mL of blood anticoagulated with CPDA-1

(Citrate-phosphate-dextrose-adenine) which allows up to 35 days of storage after collection.

Special Blood Donation

1. Autologous - donor and recipient are the same person.

2. Apheresis - whole blood is withdrawn, desired component is separated and retained,

and remainder of blood is returned to the donor.

3. Therapeutic - removal of a blood component to alleviate symptoms caused by an

underlying disease.

3. Component Preparation. Component therapy is the transfusion of the specific blood

component needed by the patient.

● This is done primarily to treat anemia or blood loss and insufficient coagulation

proteins or platelets to provide adequate hemostasis.

● Components: Whole blood (21-35 days, 6-8oC in ref), packed red blood cells (21-35

days, 6-8oC in ref), platelets (5 days, 21-25oC), Fresh Frozen Plasma (FFP),

cryoprecipitate, factor IX concentrate, albumin, immune globulin

4. Compatibility Testing

● ABO and Rh typing of donor and recipient

○ Agglutination indicates presence of antigen

○ Anti-B (yellow), Anti-A (blue), Anti-D (colorless)

● Unexpected antibody screening

● Cross-matching - testing of patient’s serum with donor’s red blood cells

5. Blood Transfusion. Goal is for 75% of transfused RBCs to remain viable for 24 hours.

6. Investigation of Blood Transfusion Reactions

CHAPTER 8: INTRODUCTION TO BACTERIOLOGY

● Bacteriology - identification and the study of bacteria cultivation.

General Characteristics of Bacteria

● Unicellular

● Prokaryotic, meaning that they have no nucleus or cell-bound organelles.

● Smallest free-living organisms

● Spore-forming; considered the hardiest microorganisms

Structural Components

● Cell wall - rigid structure that maintains the shape of the cell.

● Cell membrane - a semi-permeable layer that controls what goes inside and outside of the

cell.

● Nucleoid - bacterial chromosome; single circular piece of DNA

● Plasmid - extrachromosomal DNA

● Capsule - protective layer superficial to the cell wall

● Flagella - aids in the movement of the cell

○ Lophotrichous - multiple flagella

○ Polar - one flagella

○ Peritrichous - all over the flagella

 ● Pili - used for attachment and genetic exchange.

● Fimbriae - used for attachment.

Classification According to General Morphology

● Cocci - round-shaped bacteria

○ Streptococci - round-shaped bacteria in chains

○ Staphylococci - round-shaped bacteria in clusters

● Bacilli - rod-shaped bacteria

● Spirochete - spiral-shaped bacteria

● Spirilla - spiral-shaped bacteria; shorter version of spirochete

Classification According to Cell-Wall Structure

● Gram-positive - has a thick peptidoglycan layer

● Gram-negative - has a thin peptidoglycan layer

● Acid-Fast - mycolic acid is present

● Cell Wall deficient - has no cell wall; irregularly-shaped

Important Factors That Aid in Virulence

● Virulence - cause of damage bacteria can do to a host; cause of disease

○ Endotoxin - cell wall component that acts as a toxin; exclusive for gram-negative

bacteria.

○ Exotoxins - toxic substances secreted by bacterial cells.

○ Invasiveness - ability of bacteria to enter deep into tissues by disrupting cell

membranes and tissue matrix.

○ Genetic recombination - bacteria are able to accomplish horizontal exchange of

genes.

■ Transformation - uptake of naked DNA

■ Transduction - transfer of gene material via bacteriophage

■ Conjugation - transfer of genes via pili

○ Antibiotic resistance - result of drug exposure and results from the overuse of

antibiotics.

Common Procedures in Bacteriology

● Gram-staining - a microscopic exam that divides bacteria into four major groups: gram

positive/negative cocci, gram positive/negative staphylococci

○ 4 coagulants: crystal violet (primary stain), gram’s iodine (mordant), acetone alcohol

(decolorizer), safranin (counter stain)

● Acid-fast staining - detects acid-fast bacteria such as Mycobacterium tuberculosis

○ 3 reagents: carbolfuchsin (primary stain), acid alcohol (decolorizer), methylene blue

(counterstain); heat is used as mordant.

● Culture - specimens are inoculated onto artificial agar and incubated to allow the growth of

bacteria; bacteria are then identified through biochemical tests.

● Susceptibility tests - bacteria are tested against antibiotics; the higher the zone of inhibition,

the more effective the antibiotic.

CHAPTER 9: INTRODUCTION TO MYCOLOGY & VIROLOGY

● Mycology - study of fungi. Mycosis is the study of fungi that can cause disease.

● Virology - study of viruses.

General Characteristics of Fungi

● Unicellular or multicellular. They need oxygen to survive. They are gram positive when stained.

● Eukaryotic. Fungi have membrane-bound organelles and a nucleus.

● Achlorophyllous. Fungi have no chlorophyll and are usually filamentous.

Reproduction - accomplished through spores

● Sexual spores - requires two organisms for reproduction.

● Asexual spores - requires only one organism for reproduction.

Morphologic Types

● Yeast - unicellular; reproduce through budding; no mycelium

● Mold - multicellular; consists of thread-like lyphae; contains mycelium

● Dimorphic fungi - may exist in yeast/mold form

Pathogenicity of Fungi

● Diseases are primarily a result of the host’s immune reaction.

● A few species are able to produce toxins (mycotoxins).

Classification According to Infection

● Cutaneous mycoses - fungal infection on hair and skin; most common type of fungal

infection.

○ Superficial mycoses - not recognized by the immune system; minor tissue damage.

○ Dermatophytosis (ringworms) - recognized by the immune system.

● Subcutaneous mycoses - fungal infection that reaches the muscles, connective tissues and

bones.

● Deep-seated mycoses - most serious type of fungal infection affecting tissues and organs.

● Opportunistic mycoses - fungal infection that can cause disease among people with weak

immune systems.

Laboratory Diagnosis of Fungi

● KOH mount - performed by adding 10% KOH; most common method of direct microscopic

examination; used to confirm a diagnosis of cutaneous mycoses.

● Culture and susceptibility - specimens are inoculated into artificial media and allowed to

grow; helps physicians prescribe correct type/dosage of medicine.

General Characteristics of Viruses

● Smallest infectious agents

● Obligate intracellular parasites - requires a host for cell growth.

General Structural Components

● Genome - contains either DNA or RNA, but never both.

● Capsid - protein shell that protects the genome.

● Envelope - derived from a host cell membrane.

● Peplomers/Spikes - used for attaching to the host cell prior to penetration.

Viral Infectious Cycle (Replication)

1. Attachment - peplomers attach to the envelope of the cell.

2. Penetration - attachment to a specific receptor; induces conformational changes in viral

capsid proteins.

3. Uncoating - capsid is removed/degraded by viral enzymes.

4. Eclipse

5. Synthesis

6. Release - virus is released outside the host cell; will cause lysis of cell, while some remain in the

cell.

Pathogenicity of Viruses

● Damage or death of host cells

● Changes due to the incorporation of viral genes

Laboratory Diagnosis of Virus

● Electron microscopy - obtains high-resolution of bio or non-biological specimen.

● Antibody detection - i.e. rapid antigen testing.

● Viral culture and isolation - i.e. RT-PCR

Prions

● Proteinaceous material devoid of nucleic acids; misfolded proteins; always fatal.

CHAPTER 10: INTRODUCTION TO PARASITOLOGY

Groups of Parasites

Helminths Protozoans Arthropods

Examples Worms Single-celled Insects like mosquitoes

eukaryotes (vectors of diseases)

Cell Types Multicellular, Unicellular, eukaryotic none

eukaryotic

Nematodes - Amoeba - i.e. e.coli

roundworms Ciliates - i.e. b. Coli,
Cestodes - tapeworms paramecium
Major Groups Trematodes - flukes Flagellates - have none
flagella
Sporozoa - reproduce
via sporozoic
formations

Pathogenicity

● Mechanical destruction of tissue

● Competition for essential metabolites

● Hypersensitivity reaction of the host

● Toxic reaction to the parasite’s metabolic products

Laboratory Diagnosis

● Microscopy is the most common method of identifying parasites.

● Common procedures: routine stool microscopy/malarial smear

CHAPTER 11: INTRODUCTION TO CLINICAL CHEMISTRY

● Clinical chemistry - analysis of body fluids or tissue specimens to provide information for the

diagnosis or treatment of disease.

○ Analyte - substance for which analyses are performed.

○ Normal value - range of values for a given constituent in a healthy individual.

○ Clinical significance - consideration when interpreting the result of an analysis.

Clinical Chemistry 1

A.) Laboratory safety, Reagent Preparation and Laboratory Mathematics

B.) Quality Control - analytical sensitivity is the ability of an analytical method to measure the

smallest concentration of the analyte of interest. Meanwhile, analytical specificity measures

only the analyte of interest.

C.) Analytical Methods, Instrumentation and Automation - involves energy, wavelength, and

frequency. Wavelength is inversely related to frequency and energy.

D.) Patient Separation, Specimen collection and Handling - prior to blood collection, patients

must be properly briefed on how to prepare for each laboratory test. Patient identification,

the first step in sample collection, is the prime factor to obtain accurate results in a clinical

laboratory.

E.) Carbohydrates - hydrates of aldehyde or ketone derivatives based on the location of the

carbonyl functional group.

F.) Lipids - primary sources of fuel; provide stability to cell membrane and allow for

transmembrane transport.

G.) Proteins - amphoteric, meaning that they can bear positive and negative charges; can

also be both a weak base or weak acid, making it an effective blood buffer.

H.) Kidney Function Test/Non-Protein Nitrogen - urea is the most sensitive test for kidney

function, which is the end product of amino acid. Creatinine is the waste product formed in

the muscle tissue after energy production, which is the most specific test for kidney function.

Clinical Chemistry 2

I.) Liver Function Tests - Bilirubin is the breakdown product of hemoglobin.

J.) Enzymology - study of proteins produced by living cells that hasten chemical reactions in

organic matter; measure

K.) Electrolytes - ions capable of carrying an electric charge

L.) Blood Gases and pH - CO2, HCO3-, H+

Clinical Chemistry 3

M.) Endocrinology - the endocrine system is a network of ductless glands of internal

secretions; considered to be the regulatory system of the body.

N.) Therapeutic Drug Monitoring - analysis, assessment and evaluation of circulating

concentration of drugs in serum, plasma, or whole blood.

O.) Toxicology