IMMUNOLOGY AND SEROLOGY EXTERNAL DEFENSE SYSTEM

 designed to keep microorganisms from entering the

IMMUNOLOGY body.
 host’s reac�on to foreign substances when introduce to  Structural barriers
the body.  Prevent most infec�ous agents.
 An�gen – a foreign substance that induces an immune  Unbroken skin/ intact skin – major structural barrier
response.  Lac�c acid – sweat
 Faty acids – sebaceous glands, maintain skin pH of
IMMUNITY approx. 5.6 – acid pH keeps microorganisms from
 condi�on of being resistant to infec�on. growing.
 Respiratory tract
CROSS IMMUNITY  Mucous secre�ons and mo�on of cilia lining the
 condi�on at which exposure to one agent produces nasopharyngeal passages clears the away almost
protec�on against another agent. 90% of deposited material.
 Flushing of urine – acidity, helps remove many poten�al
Ancient Chinese Variola�on pathogens in genitourinary tract.
Edward Jenner Immunity to smallpox  Produc�on of Lac�c acid in female genitalia keeps
Vaccina�on vagina at pH of about 5.
Louis Pasteur 1st atenuated vaccine  Acidity of stomach
Ellie Metchnikoff Phagocytosis  Produc�on of hydrochloric acid – pH as low as 1 and
Almoth Wright Linked two theories serves to halt microbial growth.
(cellular & humoral)  Lysozyme – found in many secre�ons such as tears and
saliva. Atacks cell wall of microorganism, especially
OPSONINS gram-posi�ve.
 acted to coat on bacteria so they can become more
suscep�ble to phagocytosis. INTERNAL DEFENSE SYSTEM
 can be categorized into cellular and humoral factors.
ANTIBODIES • process of inflamma�on brings cells and humoral
 specific proteins, produced by plasma cells in response to factors to the area in need of healing.
an An�gen.  recognize molecules that are unique to infec�ous
organisms.
ANTIGEN  Phagocytosis – engulfment of cells or par�culate mater
 s�mulate An�body produc�on, it is a foreign substance, by leukocytes, macrophages,
usually a protein or polysaccharides. and other cells. Most important func�on of the internal
defense system. Enhanced by soluble factors called acute
ACUTE PHASE REACTANTS phase reactants.
 nonspecific factors that increase nonspecifically in any
infec�on. ACUTE PHASE REACTANTS
 normal serum content that increases rapidly by at least
NATURAL/ INNATE IMMUNITY ACQUIRED IMMUNITY 25% due to infec�on, trauma, or injury to the �ssues.
Normally present An�body Specific for each individual  Produced primarily by hepatocytes (liver parenchymal
pathogen.
cells) within 12-24 hours in response to an increase in
Nonadap�ve or Nonspecific
certain intercellular signaling polypep�des known as
Ability to remember a prior
No prior exposure is required. exposure. CYTOKINES - cell messenger, mainly produced by
monocytes & macrophages.
Response does not change with Increase response upon C-Reac�ve Protein Opsoniza�on, complement
subsequent exposures repeated exposure. ac�va�on
Serum amyloid A Removal of cholesterol
NATURAL DEFENSE SYSTEM Alpha, an�trypsin Protease inhibitor
 EXTERNAL DEFENSE SYSTEM Fibrinogen Clot forma�on
 INTERNAL DEFENSE SYSTEM Haptoglobin Binds hemoglobin
• Both systems work together to promote Ceruloplasmin Binds copper and oxidizes
iron
phagocytosis, which results in the destruc�on of
Complement C3 Opsoniza�on, lysis
foreign cells and organisms.
Mannose-binding protein Complement ac�va�on
CELLULAR DEFENSE MECHANISM
NEUTRPHIL 50-70%, 10-15 um, 2-5 lobes
First cell to response.
Phagocytosis
Half of the total popula�on is found in
margina�ng pool on blood vessel walls,
the rest freely circulate for approx. 6-10
hours.
Diapedesis – allow neutrophils to move
from the circula�ng blood to �ssues.
Selec�ns – receptor that help make
neutrophils s�cky and enhance
adherence to endothelial cells.
Chemotaxins – chemical messenger that
cause cells to migrate in par�cular
direc�on.
EOSINOPHIL 1-3%, 12-15 um, bilobed, reddish-orange
granules
Parasi�c infec�ons and allergic reac�on
Most important role is to neutralize
basophil and mast cell products and
killing certain parasites.
BASOPHIL <1%, coarse densely staining deep bluish-
purple granules that obscure their
nucleus.
Allergic reac�ons
Granules:
Histamine – vasoac�ve amine, contract
smooth muscle
Heparin – an�coagulant
• S�mulate B cells to produce IgE
MAST CELLS Connec�ve �ssue cells of mesenchymal
origin
Lifespan: 9-18 months
Plays a key role in hypersensi�vity
reac�ons by binding IgE.
Works as an an�gen presen�ng cells.
MONOCYTE 12-22 um irregularly folded/ horseshoe-
shaped nucleus, grayish blue, ground
glass appearance
Stay in PB for up to 70 hours then
migrate to �ssue to became
macrophages.
MACROPHAGE No peroxidase
Not as efficient as Neutrophils for
phagocytosis
Plays an important role in ini�a�ng &
regula�ng immune response.
Microbial killing, tumoricidal ac�vity,
intracellular parasite eradica�on,
secre�on of cell mediators and an�gen
presenta�on.
Ac�vated by contact with
microorganisms or cytokines which
released by T lymphocytes during
immune response.
DENDRITIC Covered with long membranous
CELLS extensions that make them resemble
nerve cell dendrites.
Phagocytose an�gen and present it to
helper T lymphocytes.
Most potent phagocy�c cell in the
�ssue.
TOLL-LIKE Each receptor recognizes different
RECEPTORS microbial products.
Once receptor binds to par�cular
substance or ligand, phagocytosis may be
s�mulated or cell produces cytokines
that enhance inflamma�on and eventual
destruc�on of microorganisms.
TLR 1, 2, 4, 5.6 – cell surface
TLR 3, 7, 8, 9 – endosomal
TLR 5 – flagellin
TLR 1 – lipoprotein
TLR 2 – pep�doglycan (gram +)
TLR 3 – endosomal dsRNA
TLR 4 – lipopolysaccharide (gram -)
TLR 6 – mycoplasma
TLR 7, 8 – viral ssRNA
TLR 9 – bacterial & viral DNA
NATURAL 1st line of defense against tumor cells
KILLER CELL and cells infected by viruses.
5-10% of the circula�ng lymphoid pool,
mainly found in spleen & PB.
PHAGCYTOSIS (ICED – Ini�a�on, Chemotaxis, Engulfment,
Diges�on)
1. Physical contact between WBCs and the foreign par�cle
2. Forma�on of phagosome
3. Fusion with cytoplasmic granules to form
phagolysosome.
4. Diges�on and release of debris to the outside -
Exocytosis
PHAGOSOME – vacuole formed within a phagocy�c cell.
Pseudopodia fuse to completely enclose the pathogen
forming this structure.
PHAGOLYSOSOME – fusion of lysosomal granules and
phagosomes.
INTERFERONS – group of molecules that limit the spread of
viral infec�ons by blocking transla�on of viral proteins.
- Interferes with replica�on of virus.
 Alpha Interferon
 Beta Interferon
COMPLEMENT – principal soluble mediator of inflammatory
response.
INFLAMMATION
 Vascular response – injured mast cells → release of
Histamine → dilate blood vessel = ↑blood flow =
Redness (Rubor) & Heat (Calor) → contrac�on of
endothelial cells in blood vessel = ↑permeability =
leaking of plasma in �ssues = Swealing (Tumor) & Pain
(Dolor)
 Cellular response – Neutrophil is mobilized (lasts 24-28 Can be recognized by the presence of
hrs.), Macrophage & Monocyte (a�er 16-48 hrs.) membrane-bound Ab; IgM & IgD
 Cellular prolifera�on & Repair – Fibrinogen forms clot to Other surface proteins: CD19, CD21,
close the site of injury. MHC Class II
T CELLS 61-80% (majority of circula�ng
ADAPTIVE IMMUNITY lymphocyte)
 Specificity for each individual pathogen or microbial Express unique surface markers that
agent allow them to recognize foreign Ag
 ability to remember a prior exposure. bound to cell membrane proteins called
 An increased response to that pathogen upon repeated MHC molecules.
exposure Role: produce cytokines that s�mulate
 more tailored response. It takes a longer �me to be B cells to produce Ab, assis�ng in
ac�vated, but it is more specific and longer las�ng. killing tumor cells or infected target
cells, and helps to regulate both innate
and adap�ve immune response – Cell
CELLULAR IMMUNITY HUMORAL IMMUNITY
mediated immunity.
Cell mediated Ab mediated
Helper T cells – CD4
Acts against intracellular Extracellular pathogens
Cytotoxic T cells – CD8+
pathogens
Regulatory T cells – CD4
Virus, fungi, parasites Bacteria
CD4:CD8+ = 2:1
mycobacteria, tumor cells
NK CELLS 10-15%
T lymphocyte B lymphocyte
Can kill the target cells without prior
Produce CYTOKINES Produce ANTIBODY exposure to them.
Elimina�on of tumor cell, An�body- dependent cell- CD16 & CD56
gra� rejec�on, mediated cytolysis (ADCC) Plays an important role as a
hypersensi�vity reac�on transi�onal cell bridging the innate &
adap�ve immune response against
ACTIVE PASSIVE pathogens.
NATURAL ARTIFICIAL NATURAL ARTIFICIAL IMMUNOGEN – any agent capable of INDUCING an immune
Infec�on Vaccina�on Transfer in Infusion of response.
vivo or serum or ANTIGEN – any agent capable of BINDING to components of
colostrum plasma immune response such as lymphocyte & Ab.
injec�on
• IMMUNOGEN SUBSTANCE ARE ALWAYS ANTIGENIC
Produced by YES NO NO
host • ANTIGENS ARE NOT NECESSARILY IMMUNOGENIC
NOT an NO YES YES
immediate EPITOPES/ ANTIGENIC DETERMINANTS
response  Por�on that binds specifically with the binding site of an
Dura�on: Long Short Short Ab or to lymphocyte.
Long  4-6 amino acids or 5-7 monosaccharides
• Paratope – por�on of Ab that binds with the binding site
PRIMARY RESPONSE of an Ag.
 latent period: approx. 5-15 days IgM – 1st Ig to appear. • LINEAR EPITOPES – amino acids following one another
SECONDARY RESPONSE on a single chain.
 Rapid response IgG – most predominant Ig. • CONFORMATIONAL EPITOPE – results from folding of
one chain or mul�ple chains, bringing certain amino
CELLS OF THE ADAPTIVE IMMUNITY acids from different segments of linear sequence or
LYMPHOCYTES 20-40% sequences into close proximity with each other so they
Arise from an HSC and then further can be recognized together.
differen�ated in the primary lymphoid
organs such as the BM & Thymus. CLASSES OF IMMUNOGEN
Can be divided into 3: 1. ACCORDING TO DEPENDENCY TO T-CELLS
T cells, B cells, NK cells
THYMIC DEPENDENT
B CELLS 10-20%
 Requires the help of T-Cells for forma�on of
Go through a developmental process
An�body.
that prepares them for their role in Ab
produc�on and restricts the type of Ag  T-helper Cell: Helps to s�mulate the immune
to which one cell can respond. response.
 IgG class
THYMIC INDEPENDENT
 An�body produc�on without interac�ng T Cells.
 IgM class
2. ACCORDING TO THE EPITOPES
 Unideterminant, Univalent An�gen
• 1 Kind and 1 Number of Epitope
 Unideterminant, Mul�valent An�gen
• 1 Kind of Epitope; High number of Epitope
 Mul�determinant, Univalent An�gen
• Many kinds of Epitope; for every kind, only 1
 Mul�determinant, Mul�valent
• many kind, many epitope
HAPTEN – incomplete/ par�al Ag, gives Ag its Ab provoking
abili�es.
CARRIER – proteins which give the hapten the required size.
CROSS REACTIVITY – situa�on in which two or more
compounds that may have various degree of dissimilarity,
share an�genic determinants.
- condi�on at which exposure to one agent produces
protec�on against another agent.
TOXOID – form of toxin may have one or more an�genic
determinants in common with na�ve toxin.
ADJUVANTS – substance when mixed with an immunogen
enhances the immune response against the immunogen.
- Helps to make immuniza�on more effec�ve.
REQUIREMENTS OF IMMUNOGENECITY
A. FOREIGNNESS
1. AUTOLOGOUS Ag: same individual; Autogra�
2. SYNGENEIC Ag: gene�cally iden�cal individuals: Isogra�
3. ALLOGENEIC/ HOMOLOGOUS Ag: different individual
same species; Homogra�/ Allogra�
4. XENOGENEIC/ HETEROLOGOUS: different individual,
different species, Xenogra�
5. HETEROGENETIC/ HETEROPHILIC Ag: unrelated animal
and plant species.
B. CHEMICAL COMPLEXITY
1. PROTEIN – majority of immunogens, strongest
immunogens
- high no. of epitopes = high no. of Ab
- amino acids, varia�on of subunit = highly immunogenic
• Synthe�c Polymers: Nylon & Teflon – have few simple
repea�ng units = NONIMMUNOGENIC
– use for ar�ficial heart valves, elbow replacement.
2. CARBOHYDRATES – less immunogenic because of smaller
& limited no. of sugar.
GLYCOPROTEINS – Rh and Lewis BGS
GLYCOLIPIDS – ABO BGS
3. POLYSACCHARIDE – most are HAPTENS
- do not possess sufficient chemical diversity
Pure Polysaccharide & Lipopolysaccharides –
Immunogenic
4. NUCLEIC ACIDS
Nucleic acid + Protein = Nucleoprotein = Immunogenic
5. LIPIDS
Lipids + Protein/ Polysaccharide = Immunogenic
C. MOLECULAR SIZE
<5,000 Daltons = NON-IMMUNOGENIC
>10,000 Daltons = Reasonable Immunogenic
>1,000,000 Daltons = Excellent Immunogenic
D. GENETIC COMPOSITION
- Major Histocompa�bility Complex (MHC) exerts gene�c
influence of an individual’s ability to respond to an
immunogen.
LYMPHOID SYSTEM!
ORGANS OF THE IMMUNE SYSTEM
PRIMARY LYMPHOID ORGANS – where matura�on of B and T
lymphocytes takes place.
- Came from pluripoten�al hematopoie�c stem cells.
 BONE MARROW
 Main source of hematopoie�c stem cells
 Center for Ag-independent lymphopoiesis
 Lymphocytes are released from the marrow and
travel to other primary lymphoid organs where
further matura�on takes place.
 THYMUS
 Where T cells develop
 Matura�on of T cells takes place over a 3-week
period as cells filter through the thymic cortex to the
medulla (this is where the mature T cells are
released).
SECONDARY LYMPHOID ORGANS – it provides loca�on where
contact with foreign Ag can occur.
 Where main contact with foreign an�gens takes place.
 Spleen – removes old, damaged cells, and foreign Ag
from the blood.
 Red pulp – destroys RBC’s.
 White pulp – produces WBC’s.
 Lymph nodes – serve as the collec�ng points for lymph
fluid from adjacent �ssues.
 It allows contact between lymphocytes and foreign
an�gens from the �ssues to take place.
 Appendix (high conc. of Lymphocytes), Tonsils
 MALT (mucosal associated �ssue) – these mucosal
surfaces represent some of the main ports of entry of
foreign Ag, and so numerous macrophages &
lymphocytes are localized here.
 PRIMARY BONE MARROW Produces - Clonal dele�on – process of elimina�on of clones of
hematopoie�c stem T cells that would be capable of autoimmune
cells, matura�on of response.
B and NK cells.
Matura�on of T MATURE T CELLS
THYMUS cells. CD4+ T CELLS
SECONDARY Spleen Filters blood  Class II MHC protein
Lymph nodes Places where
 T helper (Th) cells
MALT contact between T
 Approx. 2/3
CALT cells, Ag & B cells
occur.  Th 1 cells – IFN y, IL-2, TNF-B
– protect cells against intracellular pathogens by
SURFACE MARKERS ON LYMPHOCYTES ac�va�ng cytotoxic lymphocytes and macrophage.
 Proteins that appear on the cell surface can be used as  Th 2 cells – IL-2,-5,-6,-9,-10,-13
markers to differen�ate T cells and B cells. – help B cells to produce Ab and regulate B-cell ac�vity.

LYMPHOCYTE – mononuclear, phagocy�c WBCs found in the CD8+ T CELLS

blood, lymph, and lymphoid �ssues.  Class I MHC protein
 Cytotoxic T cells
SUBPOPULATIONS OF LYMPHOCYTES  1/3 express CD8
Thymus cortex: Early precursor of T cell
Medulla: Mature T cells (takes over a 3-week period) T REGULATORY (Treg) CELLS
• Early surface markers on thymocytes are commited to  Possess CD4 as well as CD25 Ag.
being T cells include CD44 & CD25.  Comprise approx. 5% of all CD4+ T cells.
 Suppress the immune response to self-an�gens.

STAGES IN T CELL DIFFERENTIATION

DOUBLE-NEGATIVE STAGE STAGES IN B-CELL PROLIFERTAION

 Early thymocytes lack CD4 & CD8 markers, which are
important to their later func�on.
 Ac�vely proliferates in the outer cortex under the
influence of IL-7.
 Rearrangement of the genes that code for the an�gen
receptor known as the T-cell receptor begins at this
stage CD3.
 Express gamma (y) and delta (d) chains.
DOUBLE-POSITIVE STAGE
 Thymocytes express both CD4 and CD8 an�gens.
 Begins to rearrange the genes that code for alpha chain.
 CD3-aB receptor complex (TCR) is complete and
expressed on the cell surface.
• POSITIVE SELECTION
- allows double posi�ve cells with func�onal TCR
receptors to survive.
• NEGATIVE SELECTION
- Takes place among surviving double posi�ve T cells.
- Second selec�on process takes place in the
cor�comedullary region and medulla.
- Strong reac�ons to self-pep�des other than MHC
an�gens trigger apoptosis.
PRO-B CELLS
 Rearrangement of genes that code for the heavy and
light chains of an Ab.
 Rearrangement of genes on chromosome 14, which code
for the heavy-chain part of the an�body molecule, takes
place first in a random fashion.
 C-Kit, a receptor on the pro-B cell, interacts with a cell
surface molecule called stem cell factor found on stromal
cells. This interac�on triggers the ac�va�on process.
 Differen�a�on of pro-B cells into pre-B cells occurs upon
successful rearrangement of heavy-chain genes on one
of the number 14 chromosomes.
 Only pro-B cells that successfully rearrange one set of
heavy-chain genes go on to become pre-B cells.
PRE-B CELLS
 Once heavy-chain genes are rearranged, then these
genes are transcribed to make the protein that will be
part of an an�body molecule.
 When synthesis of the heavy-chain part of the an�body
molecule occurs, the pre-B stage begins.
 first heavy chains synthesized are the μ chains, which
belong to the class of immunoglobulins called
immunoglobulin M (IgM).
 The combina�on of the two heavy chains along with Ig-α,
Ig-β and the surrogate light chains form the pre-B cell
receptor (pre-BCR)
 Only pre-B cells expressing the μ heavy chains in
associa�on with surrogate light chains (an unusual light
chain) survive and proceed to further differen�a�on.
IMMATURE B CELLS
 Dis�nguished by appearance of complete IgM Ab
molecules on the cell surface.
 Varia�on of light chains and heavy chains, determine the
specificity for an�gen.
 Preexis�ng diversity of receptors for an�gen is a hallmark
of the adap�ve immune system.
 Surface proteins that appear on the Immature B cell; CD
21, CD 40, Class I MHC
 Evidence that self-an�gens give a nega�ve signal to
immature B cells. Immature B cells that �ghtly bind self-
an�gens through cross-linking of surface IgM molecules
receive a signal to halt development, resul�ng in
arrested matura�on and cell death.
 Central tolerance - elimina�on of B cells that bear self-
reac�ve receptors.
 Immature B cells that survive this selec�on process leave
the bone marrow and proceed to the spleen, where they
become mature B cells.
MATURE B CELLS
 Immature B cells develop in the Spleen known as either
Marginal zone B cells or Follicular B cells.
 Marginal B cells – remain on the spleen to respond
quickly to any blood-borne pathogens.
 Follicular B cells – migrate to lymph nodes and other
secondary organs.
 In addi�on to Ig M, mature B cells also exhibit Ig D
(prolong lifespan of mature b cells)
 When B cell is s�mulated by an�gen, it undergoes
transforma�on to a blast stage that eventually forms
memory cells and an�body-secre�ng plasma cells this
process is known as the an�gen-dependent phase.
 Produc�on of an�bodies by plasma cells is called
HUMORAL IMMUNITY.
PLASMA CELLS
 Represent the most fully differen�ated lymphocyte and
their main func�on is An�body produc�on.
 In the bone marrow, plasma cells can survive in the
niches around by stromal cells.
 Stromal cells provide chemical s�mula�on by cytokines,
which allow plasma cells to be long-lived and con�nually
produce an�bodies.
NATURAL KILLER CELLS
 Small percentage of lymphocytes that do not express
markers of either T or B cells.
 Arise form common lymphocyte precursor (CLP) and
differen�ate into T or NK cells.
 CD16 – receptor for the fragment crystallizable por�on,
or nonspecific end, of the Ig molecule IgG.
 Surface markers: CD16, CD56
 Development: Il-15
 S�mulated by Il-12, INF y & B
 Half-life: 7-10 days
ADCC (An�body- dependent cell-mediated cytolysis)
 process of destroying An�body-coated target cells by NK
cells, monocyte, macrophage, neutrophils, all of which
have specific receptors for an�body.
LABORATORY IDENTIFICATION
 Cell Flow Cytometry – scatering of light as the cells in
the string fluid flow – labelled monoclonal An�body.
 Fluorescence Microscopy
 Other Methods: Rosete Test
CYTOKINES
 Chemical messengers that regulate the immune system,
orchestra�ng both innate and adap�ve response.
 Small proteins produce by several different types of cells
that influence the hematopoie�c and immune systems
through ac�va�on of cell-bound receptors.
TYPES OF ACTION:
AUTOCRINE – same cell that secreted it
PARACRINE – affec�ng target cell in close proximity.
ENDOCRINE – systemic
CHARACTERISTICS:
PLEIOTROPY – single cytokine can have many different
ac�ons.
REDUNDANCY – different cytokines ac�vate some of the
pathways and genes. Many cytokines share receptor subunits
SYNERGISTIC – cytokines o�en act in networks if the effects
complement and enhance each other.
ANTAGONISM – one cytokine may counteract the ac�on of
another cytokine.
CASCADE INDUCTION – cytokine secreted by a specific type
of cell can ac�vate target cells to produce addi�onal cytokine.
Ex. T helper (Th) cells secrete IFN-gamma which ac�vates
macrophages to secrete IL-12 which then ac�vates Th cells to
produce other cytokines.
CYTOKINES IN INNATE IMMUNE RESPONSE
INTERLEUKIN-1 (IL-1)
 induced by the presence of microbial pathogens,
bacterial lipopolysaccharides, or other cytokines.
 IL-1 induces the produc�on of vascular cell-adhesion
molecules as well as chemokines and IL-6. These
chemokines and cell-adhesion molecules atract and
assist leukocytes to enter the inflamed area.
IL-1α IL-1β
Proinflammatory cytokines Proinflammatory cytokines
- remains within the cells - responsible for most of
that produce it and is rarely the systemic ac�vity
found outside these cells. atributed to IL-1, including
- can be released a�er cell fever, ac�va�on of
death and can help atract phagocytes, and produc�on
inflammatory cells to areas of acute-phase proteins
 where cells and �ssues are - acts as an endogenous
being killed or damaged. pyrogen (induces fever) in
the acute-phase response
through its ac�ons on the
hypothalamus
IL-1RA (RECEPTOR ANTAGONIST)
 produced by monocytes and macrophages, is the best
characterized cytokine inhibitor.
 acts as an antagonist to IL-1 by blocking the IL-1
receptor, which helps to regulate the physiological
response to IL-1 and turn off the response when no
longer needed.
TUMOR NECROSIS FACTOR
 first isolated from lymphocytes and macrophages
 named because they induced lysis in tumor cells.
 Both endocrine and paracrine
 Vasodila�on and increase vasopermeability
 Direct cytotoxic effect, modula�on of cell growth and
differen�a�on, has a role in chronic inflammatory
condi�on and infec�on.
 Increase phagocytosis, degranula�on, and respiratory
burst ac�vity through s�mula�on of neutrophils.
 • RANTES – ligand for CCR5
3. Conforma�onal changes in GP130-exposes tyrosine
residues in the intracellular por�on of the molecule
4. Acute phase reactants and Interferon regulatory factor
1 (IRF-1), B and T genes are ac�vated.
CHEMOKINES
 enhance mo�lity and promote migra�on of many types
of WBCs toward the chemokine source via a process
known as chemotaxis.
 regulate response to infec�ous diseases, autoimmune
inflamma�on, cancer, and the homing of lymphocytes to
all the lymphoid �ssues.
• alpha, or CXC, chemokines—contains a single amino
acid between the first and second cysteines.
• beta, or CC, chemokines—has adjacent cysteine
residues.
• C chemokines—lacks one of the cysteines.
• CX3C - the last major group, has three amino acids
between the cysteines.
• HIV uses the chemokine receptors CXCR4 and CCR5
as coreceptors for infec�on of CD4+ T lymphocytes
and macrophages.
• SDF1 – ligand for CXCR4

TNF-α TRANSFORMING GROWTH FACTOR-BETA (TGF-B)

 exists in both membrane-bound and soluble forms and
causes vasodila�on and increased vasopermeability.
 the main trigger for TNF-α produc�on is the presence of
lipopolysaccharide, which is found in gram-nega�ve
bacteria.
 par�ally regulated by soluble forms of both TNF
receptors.
 present in rheumatoid synovial fluids and synovial
membranes of pa�ents with rheumatoid arthri�s.
 factor that induces an�prolifera�ve ac�vity in a wide
variety of cell types.
 primarily a regulator of cell growth, differen�a�on,
apoptosis, migra�on, and the inflammatory response.
 acts as a control to help downregulate the inflammatory
response when no longer needed.
 regulates the expression of CD8 in CD4–CD8–
thymocytes and acts as an autocrine inhibitory factor for
immature thymocytes.

TNFR-1 (TUMOR NECROSIS TNFR-2 (TUMOR NECROSIS INTERFERONS

FACTOR RECEPTOR 1)
- cons�tu�vely expressed
on most �ssues and binds
soluble TNF-a
- primary mediator of TNF-α
signal transduc�on in most
cell types of soluble TNF-α.
FACTOR RECEPTOR 2)
- usually expressed in
epithelial cells and cells of
the immune system and is
ac�vated by the
membrane-bound form of
TNF-a.
 interferes with the viral replica�on.
 Limit infec�on to small area of the body.
Type I IFN
 ac�vates natural killer (NK) cells and enhances the
expression of class I MHC proteins, thus increasing the
recogni�on and killing of virus-infected cells.

INTERLEUKIN-6 CYTOKINES IN ADAPTIVE IMMUNE RESPONSE

 single protein produced by both lymphoid and
nonlymphoid cell types.
 part of the cytokine cascade released in response to
lipopolysaccharide and plays an important role in acute-
phase reac�ons.
 this pleiotropic cytokine affects inflamma�on, acute-
phase reac�ons, immunoglobulin synthesis, and the
ac�va�on states of B cells and T cells.
 Only one IL-6 receptor has been iden�fied that consists
of IL-6Rα (the IL-6-specific receptor) and gp130 (the
common signal-transducing receptor subunit used by
several cytokines).
1. Binding of IL6 to IL-6Rα
2. Dimeriza�on of gp130 and α-subunits
 cytokines involved in the adap�ve immune response are
mainly secreted by T cells, especially Th cells, and affect
T- and B-cell func�on more directly.
 Th1 lineage is driven by the expression of IL-12 by
dendri�c cells and is primarily responsible for cell-
mediated immunity.
 Th2 cells drive an�body-mediated immunity and are
developmentally regulated by IL-4.
 Treg cells are derived from naïve T cells in response to
TGF-β. Tregs help to regulate the ac�vi�es of Th1 and
Th2 cells.

Th1 CYTOKINES
IL-12
- binds to its receptor on naïve T cells and causes the
expression of a new set of genes, including those that
determine matura�on into the Th1 lineage.
- increases the cytoly�c ability of NK cells; it serves as an
important link between the innate and adap�ve immune
responses by enhancing defenses against intracellular
pathogens.
INTERFERON-γ (IFN-γ)
 principal molecule produced by Th1 cells, affects the
RNA expression levels of more than 200 genes.
 Major func�on of IFN-γ is s�mula�on of an�gen
presenta�on by class I MHC and class II MHC
molecules.
 ↑expression = ↑Ag capture =↑lymphocytes
 most potent ac�vator of macrophages and boosts
their tumoricidal ac�vity.
INTERLEUKIN-2 (IL-2)
 also known as the T-cell growth factor, drives the
growth and differen�a�on of both T and B cells and
induces ly�c ac�vity in NK cells.
 can ac�vate prolifera�on of Th2 cells and helps to
generate IgG1- and IgE-producing cells.
Th2 CYTOKINES
- primarily responsible for an�body-mediated immunity.
INTERLEUKIN-4 (IL-4)
 key cytokines regula�ng Th2 immune ac�vi�es and
helps drive an�body responses in a variety of diseases.
 IL-4 ac�vity on naïve T cells turns on the genes that
generate Th2 cells and turns off the genes that promote
Th1 cells.
 induces produc�on of MHC-I, IL-4, IL-5, IL-13, and the
cos�mulatory molecules CD80 and CD86.
 also s�mulates the produc�on of IgG2a and IgE and,
along with IL-5, drives the differen�a�on and ac�va�on
of eosinophils in both allergic immune responses and
response to parasi�c infec�ons.
 IL-13 is a cytokine with many of the same proper�es as
IL-4; both cytokines induce worm expulsion and favor
IgE-class switching.
INTERLEUKIN-10 (IL-10)
 Its major effects is the inhibi�on of IFN-gamma
produc�on via the suppression of IL-12 synthesis by
accessory cells and the promo�on of a Th2 cytokine
patern.
 IL-10 serves as an antagonist to IFN-gamma—it is a
down regulator of the immune response.
 Has primarily inhibitory effects on the immune system.
 Produced by monocytes, macrophages, CD8+ T cells, and
Th2 CD4+ T cells and has an�-inflammatory and
suppressive effects on Th1 cells.
 also inhibits an�gen presenta�on by macrophages and
dendri�c cells.
CYTOKINES ASSOCIATED WITH T REGULATORY CELLS
 third major subclass of CD4+ T cells are the T regulatory
(Treg) cells.
 Tregs are CD4+ CD25+ T cells
 Key role in establishing peripheral tolerance to a wide
variety of self-an�gens, allergens, tumor an�gens,
transplant an�gens, and infec�ous agents.
 CD4+ CD25+ Tregs affect T-cell ac�vity primarily through
the ac�ons of TGF-β.
TGF-β – induces expression of Foxp3, a transcrip�on factor
that causes Treg cells to suppress the ac�vity of other T cells.
HEMATOPOIETIC GROWTH FACTORS
COLONY STIMULATING FACTORS (CSFs)
- primary mediators of hematopoiesis, they s�mulate the
forma�on of colonies of cells in the bone marrow.
- Includes IL-3, EPO, G-CSF, M-CSF, and GM-CSF
IL-3
– is a mul�lineage CSF that induces bone marrow stem cells
to form T and B cells.
– in conjunc�on with IL-3, CSFs direct immature bone
marrow stem cells to develop into RBCs, platelets, and the
various types of WBCs.
GM-CSF
– acts to drive differen�a�on toward other WBC types.
M-CSF
– if ac�vated, the cells become macrophages, it also increases
phagocytosis, chemotaxis, and addi�onal cytokine produc�on
in monocytes and macrophages
G-CSF
– if ac�vated, the cells become neutrophils, it also enhances
the func�on of mature neutrophils and affects the survival,
prolifera�on, and differen�a�on of all cell types in the
neutrophil lineage.
EPO
– regulates RBC produc�on in the bone marrow but is
primarily produced in the kidneys.
ANTI-CYTOKINES
 Disrupts interac�on between cytokines and their
receptors.
Types: Murine monoclonal
Recombinant DNA techniques
CLINICAL ASSAYS FOR CYTOKINES
 The most basic test is the ELISpot assay, which employs
the enzyme-linked immunosorbent assay (ELISA Assay)
standard.
MAJOR HISTOCOMPATIBILITY COMPLEX
 originally referred to as Human Leukocyte An�gen (HLA)
– Dausset (gave this name, first defined by discovering
an Ab response to circula�ng WBCs)
 now known as MHC molecules because they determine
whether transplanted �ssue is histocompa�ble and thus
accepted or recognized as foreign and rejected.
 MHC molecules can func�on as an�gens when
transplanted from one individual to another.
 their main func�on is to bring an�gen in the body to
the surface of cells for recogni�on by T cells.
 T-cell ac�va�on will occur only when an�gen is combined
with MHC molecules on the surface of other cells. The
genes that encode these cell-surface molecules are the
system of genes known as the Major Histocompa�bility
Complex.
GENES CODING FOR MHC MOLECULES (HLA ANTIGENS)
 Genes coding for the MHC molecules in humans are
found on the short arm of chromosome 6 and are
divided into three categories or classes.
 CLASS I GENES are found at three different loca�ons or
loci, termed A, B, and C
 CLASS II GENES are situated in the D region, and there
are several different loci, known as DR, DQ, and DP
- Have one gene that codes for the code for the α
chain and one or more genes that β chain.
• Class I and Class II genes involved in an�gen
recogni�on; in this role, they influence the
repertoire of an�gens to which T cells can respond.
 CLASS III GENES lies between the class I and class II
regions on chromosome 6. Genes code for the C4A,
C4B, C2, and B complement proteins, as well as
cytokines such as tumor necrosis factor (TNF).
• Class III secreted proteins that have an immune
func�on, but they are not expressed on cell
surfaces, and have a completely different structure.
ALLELES – alternate forms of a gene that code for slightly
different varie�es of the same product.
CODOMINANT – same alleles at a given loca�on, all alleles
that an individual inherits code for products that are
expressed on cells.
HAPLOTYPE – total set of MHC alleles inherited from one
parent.
GENOTYPE – sum of haplotype.
HISTOCOMPATIBILITY GENES – genes that determines the
outcome of gra�.
HISTOCOMPATIBILITY ANTIGENS – protein encoded by those
genes.
CLASS I MHC/ CLASS I ANTIGENS
 highest on lymphocytes and myeloid cells and low or
undetected on liver hepatocytes, neural cells, muscle
cells, and sperm.
 POLYMORPHIC HEAVY CHAIN - Alpha Chain
- Encoded within the MHC locus on Chromosome 6
- a chain – folded into 3 domains; a1, a2, a3, and is
inserted into the cell membrane via transmembrane
segment that is hydrophobic.
 NON-POLYMORPHIC LIGHT CHAIN – B2-microglobulin
- Encoded within the MHC locus on Chromosome 15
- B2-microglobulin – does not penetrate cell membrane
but is essen�al for proper folding of a chain.
CLASS II MHC/ CLASS II ANTIGENS
 found on APCs that include B lymphocytes, monocytes,
macrophages, dendri�c cells (have highest levels of class
II molecules on their surface), and thymic epithelium.
 DP, DQ, and DR—consist of two noncovalently bound
polypep�de chains that are encoded by a and b. They
are normally expressed on an�gen presen�ng cells.
 DR is expressed at the highest level because it accounts
for about one-half of all the class II molecules.
IMPORTANT ASPECTS OF MHC
 The maximum number of Class I MHC gene products
expressed in an individual is six. That for Class II MHC
product can exceed six but is also limited.
 Mature T cells respond to foreign an�gens, but not self-
protein. The repertoire of an�gen recogni�on is based
on selec�on processes involving MHC molecules that
occur mainly in the thymus. This is another level of
control of immune responses.
 Only a single binding site exists on a class I or class II
MHC molecules; all pep�des that are capable of binding
must bind to the same site.
ROLE OF CLASS I AND CLASS II MOLECULES IN THE IMMUNE
RESPONSE
 Main role: ANTIGEN PRESENTATION
 T cells can only “see” and respond to an�gens when they
are combined with MHC molecules.
 Class I molecules present an�gen to CD8+ T cells,
triggering a cytotoxic reac�on. Watchdogs of viral,
tumor, and certain parasi�c an�gens that are
synthesized within the cell.
 Class II molecules present an�gen to CD4+ T cells, which
are helper cells involved in an�body produc�on. Help to
mount an immune response to bacterial infec�ons or
other pathogens found outside cells.
  Heterophile An�bodies
ACCORDING TO REACTION WITH ITS ANTIGEN
AGGLUTININS Ab involved in agglu�na�on
PRECIPITINS Ab involved in precipita�on
AGGLUTINOIDS Agglu�nins that are modified by heat
HEMAGGLUTININS Ab involved in hemagglu�na�on
LYSINS Ab capable of lysis
ALLERGIC ANTIBODY Ab that reacts with allergens
HLA Typing Technique
1. As a rule, HLA typing is performed using the ACCORDING TO IN VITRO BEHAVIOUR
lymphocytotoxicity technique. COMPLETE INCOMPLETE
2. HLA-A, HLA-B, HLA-C, (Class I) series an�gen, a ANTIBODY
lymphocyte suspension consis�ng of T + B lymphocytes. Synonyms Bivalent, saline Univalent, blocking,
3. HLA-DR, HLA-DQ an enriched B Cells. This is because class ac�ng conglu�na�ng
II an�gens are not found in res�ng T-Cells. Response to Thermolabile Thermostable
temp.
Importance of HLA typing Ability to cross Cannot cross Can cross
Tissue/Organ transplant placenta placenta
Paternity studies Occurrence Early in late
Studies of racial ancestry and migra�on immuniza�on
For diagnos�c and gene�c counselling reac�on Saline ac�ng Albumin ac�ng

ANTIBODIES TETRAPEPTIDE STRUCTURE OF IMMUNOGLOBULINS

 serum factors in the blood formed in response to foreign  All immunoglobulin molecules are made up of a basic
substance exposure. four chain tetrapep�de unit that consists of two large
 end product when B lymphocytes are s�mulated by chains heavy or H chains and two smaller chains light or
an�gen and undergo differen�a�on. L chains.
 also known as IMMUNOGLOBULINS  Fc fragment (fragment crystallizable) – had no an�gen-
 immunoglobulins are glycoproteins found in the serum binding ability and is known to represent the carboxy-
por�on of the blood, in response to immuogen terminal halves of two H chains that are held together by
 considered to be the main humoral element of the S–S bonding. Important in effector func�ons of
adap�ve immune response. immunoglobulin molecules.
 play an essen�al role in an�gen recogni�on and in  Fab fragment (fragment an�gen binding) – remaining
biological ac�vi�es related to the immune response such two iden�cal fragments, each fragment represented one
as opsoniza�on and complement ac�va�on. an�gen-binding site. Each thus consists of one L chain
 Structurally all an�bodies are immunoglobulins but and one-half of an H chain held together by disulfide
func�onally, not all immunoglobulins are an�bodies. bonding.
• The four-chain unit of the immunoglobulin molecule
CLASSIFICATION OF ANTIBODIES indicated that each L chain was bonded to an H chain by
ACCORDING TO SEDIMENTATION CONSTANT means of an S–S bond and the H chains were joined to
Immunoglobulin Sedimenta�on Molecular weight each other by one or more S–S bonds.
constant (Daltons) • An�body structure was described by Gerald Edelman
IgG 7s 150,000 & Rodney Porter.
Serum IgA 7s 160,000 • Monomer – basic structural unit of an an�body.
Secretory IgA 9s 170,000 • Ig’s monomer “Y” shape molecule consists of 4
IgM 19s 900,000 polypep�de chain – 2 iden�cal heavy chain & 2
IgD 7s 180,000 iden�cal light chain.
IgA 8s 180,000
ACCORDING TO TEMPERATURE LIGHT CHAINS
Cold An�bodies IgM  composed of 23,000 Dalton & more than 200 amino acid
Warm An�bodies IgG  2 main types:
ACCORDING TO OCCURRENCE • kappa (k) chain – 214 amino acids
Natural An�bodies IgM Chromosome 2
Immune An�bodies IgG • lambda (λ) chains – 213-216 amino acids
Chromosome 22
Region: Amino Terminal – variable region
ACCORDING TO SPECIE WHICH PRODUCE THEM
- responsible for An�gen binding
 Autoan�bodies
Carboxy Terminal – constant region
 Isoan�bodies/ Alloan�bodies
- determine serological & physical chain of an Ab

HEAVY CHAINS
 composed of 50,000 Dalton & 110 amino acids at the
terminal end.
 Determine the serological & physical characteris�cs of
the an�body.
• Gene: Chromosome 14
• Types: IgG (γ), IgM (μ), IgA (α), IgD (δ), IgE (ε)
• Region: Amino terminal, Carboxy terminal
DISULFIDE BONDS
 Bonds that hold the 4 polypep�de chains in normal
immunoglobulin molecules.
 Types:
• INTERCHAIN BONDS – occurs between Heavy chain
(H-H) – Hinge region
Heavy chain & Light chains (H-L)
Light chains (L)
• ITRACHAIN BONDS – forms in the basis of division of
each immunoglobulin into domains.
REGIONS
 Each heavy and light chains consist of 2 segments:
• Constant (C) – amino acid sequence is fixed and
unchanging.
- carboxyl terminal (COOH)
- concerned with binding to host �ssue
• Variable region (V) – shows wide variety of amino
acid sequence in the amino terminal por�on of the
molecule.
- amino acid sequence subject to change
- amino terminals (NH2)
- concerned with binding to an�gen
High variability: Variable region of Heavy chain (VH)
Variable region of Light chain (LH)
DOMAINS
 Globular regions on polypep�de chain stabilized by
intrachain disulfide bonds.
• Domain on heavy chain: VH, CH1, CH2, CH3, CH4
• Domain on light chain: VL, CL
HINGE REGION
 Interchain disulphide bonds form between the arms of
Fab fragments.
 Por�on of heavy chain between CH1 and CH2 domain.
 High flexible & allows movement of fab arms in rela�on
to other.
 ability to bend lets the two an�gen binding sites operate
independently and engage in an angular mo�on rela�ve
to each other and to the FC stem.
IMMUNOGLOBULIN VARIABILITY
ISOTYPIC VARIATION
 Type of heavy chain that is unique to each Ig class.
 IgG : have gamma chain
 IgA : have alpha chain
 IgM: have mu heavy chain
 IgE: have epsilon heavy chain
 IgD: have delta chain
ALLOTYPIC VARIATION
 Gene�c varia�on in the constant region of Ig molecule
 Minor varia�ons of sequences that are present in some
individuals but not others.
IDIOPATHIC VARIATION
 Diversity at the binding site and relates to the hyper
variable segments of the an�body combining site
(Paratope).
FRAGENTATION OF ANTIBODIES
 Degrade immunoglobulin molecules into definable
fragments to facilitate study of their structure.
CLEAVES WITH PAPAIN
2 Fab Can bind two an�gens.
Recognize specific foreign an�bodies.
CH1- involve with binding for c4b fragments.
1 Fc Composed of two heavy chains.
Mediates different physiological effects including
opsoniza�on, cell lysis, and degranula�on of
mast cells, basophils & eosinophil.
PEPSIN DIGESTION
F(ab)’2 Both can ini�ate agglu�na�on and precipita�on
2 Fc Can integrate into smaller places.
TYPES OF IMMUNOGLOBULINS
IgG
 predominant immunoglobulin in humans, comprising
approximately 70% to 75%
 has the longest half-life of any immunoglobulin class,
approximately 23 days.
 four major subclasses with the following distribu�on:
 IgG1, 66%; IgG2, 23%; IgG3, 7%; and IgG4, 4%
 IgG3 has the largest hinge region and the largest number
of interchain disulfide bonds; therefore, it is the most
efficient at binding complement, followed by IgG1. IgG2
and IgG4 – poor mediators
 has a high diffusion coefficient that allows it to enter
extravascular spaces more readily than other Ig.
 plays a major role in neutralizing toxins and viruses.
 response appears later than IgM in primary response,
but they form the major an�body of the secondary
immune response.
Major func�ons of IgG include the following:
• Providing immunity for the newborn because IgG is the
only an�body that can cross the placenta.
• Fixing complement
• Coa�ng an�gen for enhanced phagocytosis (opsoniza�on)
• Neutralizing toxins and viruses
• Par�cipa�ng in agglu�na�on and precipita�on reac�ons
IgM
 PRIMITIVE ANTIBODY MACROGLOBULIN
 largest of the immunoglobulin molecule, accoun�ng for
5-10% of the total immunoglobulin pool.
 Found mainly in the intravascular pool and not in other
body fluids or �ssues.
 Earliest Ab to appear in primary response but doesn’t
persist for long.
 most efficient of all immunoglobulins at triggering the
classical complement pathway.
 serves as a surface receptor for an�gen.
Func�ons of IgM include:
• Complement fixa�on
• Agglu�na�on
• Opsoniza�on
• Toxin neutraliza�on.
IgA
 10% to 15% of all circula�ng Immunoglobulin
 IgA2 is the predominant form in secre�ons at mucosal
surfaces, sero-mucous secre�ons of respiratory tract,
genito-urinary tract, and GI tract, whereas IgA1 is mainly
found in serum.
 Major role: an�-inflammatory agent
 Serum IgA – appears to downregulate IgG-mediated
phagocytosis, chemotaxis, bactericidal ac�vity, and
cytokine release. They cannot ac�vate complement
system.
 Secretory IgA – known as the “an�sep�c paint” of
mucous membranes. It can ac�vate the bacterioly�c
ac�vity through the alternate pathway of complement
system and only the presence of lysozyme.
IgD
 extremely scarce in the serum, represen�ng less than
0.001% of total immunoglobulins.
 Most of the IgD is found on the surface of
immunocompetent but uns�mulated B lymphocytes.
 ANTI-IDIOTYPIC ANTIBODY
 second type of immunoglobulin to appear (IgM being
the first) and it may play a role in B-cell ac�va�on,
although its func�on is not completely understood.
 those with both IgM and IgD receptors can respond to T-
cell help and switching to synthesis of IgG, IgA, or IgE.
 may play a role in regula�ng B-cell matura�on and
differen�a�on.
THEORIES OF ANTIBODY
ERLICH’S SIDE CHAIN THEORY
 certain cells have had specific surface receptors for Ag
that were present before contact with Ag occurred.
 Once Ag was introduced, it would select proper
receptors. Combina�on would take place and receptors
will break off and enter circula�on as Ab molecules.
 New receptors will be formed in place of those broken
off and this process could be repeated.
TEMPLATE THEORY
 Ab-producing cells are capable of synthesizing
generalized type of Ab, and when in contact with an Ag
occurs. Ag serves as a template and alters protein
synthesis so that Ab with specific fit is made.
 The molded Ab the enters circula�on, while Ag remains
behind to direct further synthesis.
SELECTIVE THEORY
 Ab are synthesized in a manner similar to other proteins
 Instruc�on for their synthesis are provided by gene�c
elements in the nucleus of cell rather from the Ag.
CLONAL SLECTION THEORY
 Individual lymphocytes are gene�cally pre-programmed
to produced on type of Immunoglobulin. That specific
Ag finds or select cells capable of responding to it,
causing prolifera�on.
 Repeated contact with Ag would con�nually increase
lymphocyte pool.

IgE
 best known for its very low concentra�on in serum
 REAGENIC ANTIBODY/ NUISANCE ANTIBODY
 has the ability to ac�vate mast cells and basophils.
 Least abundant immunoglobulin in the serum
 most heat-labile
 does not par�cipate in typical immunoglobulin reac�ons
such as complement fixa�on, agglu�na�on, or
opsoniza�on.
 shortly a�er synthesis it ataches to basophils,
Langerhans cells, eosinophils, and �ssue mast cells by
means of specific surface proteins, termed high-affinity
FC ε RI receptors, which are found exclusively on these
cells.
 Associated with immediate hypersensi�vity reac�ons &
with immunity to certain helminthic parasites.