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Immunology and Serology Part 1
Immunology and Serology Part 1
Part 1
MTAP 2
IMMUNOLOGY
□ Study of our immune system
□ Study of a host’s reactions when a foreign
substances are introduced into the body
□ Study of all aspects of body defenses,
such as antigens, antibodies,
hypersensitivity, graft rejection and
autoimmune diseases.
H ISTORY
YEAR SCIENTIST
Edward Jenner, an English countryside
physician, demonstrated that protection from
1798 cowpox could be generated by the transfer of
postural material from a cowpox lesion instead of
a more hazardous smallpox lesion
Elie Metchnikoff demonstrated that certain blood
1888 cells ingest foreign material, a concept now
known as phagocytosis
1894 Jules Bordet discover complement
□ Liver
□ CNS
□ Bone
□ Lung
□ Placenta
□ Spleen
□ Kidney
□ Skin
□ Tissue
□ Synovium
C ELLS
□ Cells involved are from the myeloid series
⚫ Neutrophils
⚫ Basophils
⚫ Eosinophils
o PRIMARY FXN:
⚫ Monocyte
⚫ Dendritic cells
□ BEST ANTIGEN PRESENTOR
□ Covered with long membranous extensions
cells
B. P HAGOCYTOSIS
□ Concept was credited to Elie Metchnikoff
□ Process of engulfing and killing extracellular organisms and
foreign particles
□ STEPS: (I.C.E.D)
1. INITIATION
□ Results from tissue damage or multiplication of
microorganisms
□ Increased surface receptors of phagocytes for
subsequent adherence
1. CHEMOTAXIS
□ Migration of phagocytes in a certain direction under the
stimulation of chemotaxins*
□ Positive chemotaxis
□ Negative chemotaxis
□ Random movement: happens in the
absence of chemotactic substances
● JOB’S SYNDROME
□ Random Movement: Normal
□ Directional/Chemotactic Movement:
Abnormal
● LAZY LEUKOCYTE SYNDROME
□ Random Movement: Abnormal
□ Directional/Chemotactic Movement:
Abnormal
□ BOYDEN-CHAMBER ASSAY
● test for chemotaxis
B. P HAGOCYTOSIS
3. ENGULFMENT
□ Process of enclosing the pathogen into a
phagocytic vacuole (phagosome or
phagolysosome)
□ Facilitated by amoeboid movement of
phagocytes
□ Opsonins: includes antibodies and complement
proteins which interact with bacterial surfaces and
makes them more susceptible to engulfment and
phagocytosis
□ Opsonization is downgraded by virulence factors
such as bacterial capsules.
B. P HAGOCYTOSIS
4. DIGESTION
□ Facilitated by digestive enzymes enclosed in cell
particles
□ As foreign materials or pathogens are digested, the cells
degranulate
□ CGD (Chronic Granulomatous Disease)
□ Affects neutrophil microbicidal function leading to
inability of the cell to kill ingested organisms
□ Characterized by impaired NADPH production
□ Normal: Blue formazan formation
□ CGD (+): colorless
□ G6PD deficiency, MPO deficiency and glutathione
synthetase/reductase deficiency may also show
positive result
B. PHAGOCYTOSIS
4. DIGESTION
□ Negative APRs:
D. I NTERFERONS
□ Glycoprotein family produced by cells which exert a virus-
nonspecific but host-specific antiviral response
1. TYPE 1
● Non-immune; produced in the initial innate response to
viral infection
a) INTERFERON-ALPHA
□ also known as leukocyte interferon
□ produced by virus-induced leukocyte culture
□ Major producers: NK cells
b) INTERFERON-BETA
□ a.k.a. fibro/epithelial/ fibroepithelial interferon
□ produced by double-stranded fibroblasts
□ Major producer: epithelial cells and fibroblasts
D. I NTERFERONS
2. TYPE 2
● Immune; produced as a component of the specific
immune response to pathogens and viruses
● INTERFERON-GAMMA
□ Also known as immune interferon
□ Produced by immunologically-stimulated
lymphocytes
□ MAJOR PRODUCER: T-CELLS and NK cells
□ activates macrophage
E. T UMOR N ECROSIS F ACTOR
□ Exhibit cytotoxic effects against tumor cells and virally infected
cells
□ PRINCIPAL MEDIATOR of acute inflamm. response to
gram(-) bacteria. NEGATIVE SIDE EFFECT: septic shock
1. TNF-ALPHA
● Cachectin
● Produced by macrophages
2. TNF-BETA
● Lymphotoxin
● Produced by CD4+ and CD8+ cells
F. P ROPERDIN
•Exerts bactericidal effect in the presence of C’3
and magnesium
G. C OMPLEMENT P ROTEINS
□ Discovery was credited to Jules Bordet
□ Functions:
● Immune system activation
● Mediates inflammation(Anaphylatoxin)
● Opsonization
● Cellular lysis
H. B ETALYSINS
□ Released by platelets during coagulation
□ Heat-stable cationic substances with bactericidal activity
M UST KNOW !
Cytokines
● General term for the soluble mediators secreted by
cells to regulate the immune system
□ Interleukins
● Cytokines secreted by macrophages and other
leukocytes which has a wide range of functions
depending on the secreting and target cell.
□ Defensins
● Cysteine-richcationic proteins which acts against
pathogen’s cell wall lysis
ACQUIRED
IMMUNITY
ACQUIRED/ ADAPTIVE/
NONSPECIFIC
IMMUNITY
THIRD LINE OF DEFENSE
Specialized Lymphocytes
-T lymhocytes
-B lymphocytes and plasma cells
Humoral
-Antibodies
TYPES OF ACQUIRED IMMUNITY
ACTIVE 1. NATURAL, ACTIVE
-acquired Ag-Ab -INFECTION with pathogens and
production is DONE subsequent production of
by the body antibodies specific to the antigen
2. ARTIFICIAL, ACTIVE
Advantage: -VACCINATION
LONG-TERM a. Live organisms:smallpox
immunity b. Attenuated or weakened
organisms: Bacillus of Calmette
Disadvantage: and Guerin for MTB =
SLOW immune attenuated M. bovis
response c. Dead organisms: cholera,
typhoid
d. Toxins: tetanus
e. Modified virus: poliovirus
M UST KNOW !
Inactivated vaccines: composed of micro-organisms that
have been killed with chemicals and/or heat and are
no longer infectious.
o Whole cell viral: polio, rabies, and hepatitis
o Fractional: made from pieces of bacteria or viruses
o Toxoids: modified toxins (diphtheria and tetanus)
o Subunit: hepatitis B, human papillomavirus,
inactivated influenza, acellular pertussis, anthrax, and
Lyme disease
o Polysaccharide
o Conjugate: Haemophilus influenzae type b,
meningococcal, and pneumococcal
M UST KNOW !
Live, attenuated/weakened antigen: composed of micro-
organisms that have been cultivated under conditions
which disable their ability to induce disease. These
responses are more durable and do not generally require
booster shots.
● Viral: measles, mumps, rubella, yellow fever,
vaccinia (smallpox), rotavirus, live attenuated
influenza vaccine (LAIV), varicella, and zoster
(shingles)
● Bacterial: oral typhoid and Bacillus Calmette-Guérin
(BCG)
TYPES OF ACQUIRED IMMUNITY
PASSIVE 3. NATURAL, PASSIVE
-acquired Ag-Ab -transfer of antibodies in vivo
production is NOT -transfer of antibodies through
done by the body ingestion of c olostrum*
4. ARTIFICIAL, PASSIVE
Advantage: -IMMUNIZATION or administration
IMMEDIATE immune of immune serum IgG antibodies
response via injection
Disadvantage:
SHORT-TERM
immunity
L YMPHOID O RGANS
1. PRIMARY/CENTRAL LYMPHOID ORGANS
● Maturation site of T and B cells
● Site of ANTIGEN-INDEPENDENT
LYMPHOPOIESIS
a) Bone Marrow
□ Bursa of Fabricius
□ Maturation site for B cells (If BM is not in choices,
Select fetal liver)
b) Thymus
□ Maturation site for T cells
□ Become hypoplastic as age increases
A. L YMPHOID O RGANS
2. SECONDARY LYMPHOID ORGANS
● Site of proliferation and differentiation of T and B
cells
● Functions:
□ Trapping site of pathogens
□ Stand by areas of T cells, B cells and phagocytes
□ Site of cell-pathogen encounter and phagocytosis
□ Site of antibody and lymphokine production
□ Site of ANTIGEN-DEPENDENT LYMPHOPOIESIS
A. L YMPHOID O RGANS
SECONDARY LYMPHOID ORGANS
a) Spleen
□ Filters antigen found in the blood
□ LARGEST secondary lymphoid organ
b) Lymph nodes
□ Filters antigen in tissue fluids
c) Tonsils and adenoids
d) MALT, BALT, SALT, GALT
e) Peyer’s patches
f) Appendix
A. C ELLULAR C OMPONENT
□ LYMPHOCYTES are the cells involved in
acquired immunity
□ Marker: Terminal deoxynucleotidyl
transferase
● For DNA polymerase immunoperoxidase
● (+) color reaction: NONE
● (+) ALL (lymphoid cells)
● (-) AML (myeloid cells
□ 20-40% of circulating WBCs
T CELLS B CELLS
•Cellular-mediated immunity •Humoral-
•60-80% mediated immunity
•Long-lived (4-10 yrs) •20-35%
•Develop in the thymus •Short-lived (3-4days)
• Produce •Develop in the BM
lymphokines •Produce antibodies
(cytokines) •Markers: CD19, CD20,
•Markers: CD2, CD3, CD4, CD21, CD40, MHC Class II
CD8 •Identified by surface Igs
•Identified by Erythrocyte
Rosette Assay
•Locations: •Locations:
-medullary, -follicular and medullary
perifollicular and or germinal centers of
paracortical region of lymph nodes
lymph nodes -primary follicles and
-periarteriolar regions red pulp of spleen
of spleen -follicular region of GALT
-thoracic duct of the
circulatory system
T LYMPHOCYTES
□ Involved in CELLULAR-MEDIATED IMMUNITY
□ Most circulating T lymphocytes express 3 of the following
CD markers:
● CD2 – sheep R B C receptor, CLASSICAL T cell RECEPTOR
● CD3 – part of T cell antigen-receptor complex
● CD4 – receptor for MHC class II molecule
● CD8 – receptor for MHC class I molecule
□ Helper-inducer T cells: CD4+
□ Suppressor-cytotoxic T cells: CD8+
□ CD4+: CD8+ normal ratio = 2:1
□ In HIV, ratio is : 0.5-1:2
□ In AIDS, CD4+ cell count drops to <200/uL (N.V. 500-
1,300/uL)
T L YMPHOCYTE D EVELOPMENT
1. DOUBLE-NEGATIVE THYMOCYTES
● CD2, CD5, CD7 (+)
Unsensitized T
● CD4 (-) AND CD8 (-)
cell
2. DOUBLE-POSITIVE THYMOCYTE S -Memory T cell
● CD4 (+) AND CD8 (+)
3. SINGLE-POSITIVE THYMOCYTE (MATURE T CELL)
● CD4 (+) OR CD8 (+)
4. ACTIVATED T LYMPHOCYTE
● Found in the 2° lymphoid organs
● CD25(+) : receptor for IL-2 which causes lymphocyte
proliferation
5. SENSITIZED/SECRETORY T LYMPHOCYTE
● Exposed to antigens
● Secretes lymphokines
TC R
S UBSETS OF T L YMPHOCYTES
1. T-helper/T-inducercells (Th)
● 60%
● help B cells to produce antibodies
2. T-suppressor cells (Ts)
● 30%
● suppress B cells NOT to produce antibodies.
3. T-cytotoxic cells (Tc)
● lyse and destroy foreign cells (cancer cells) and
virus-infected cells reject grafts & transplants
4. T-delayed type hypersensitivity (Tdth)
● involved in some delayed allergic reactions
SUBSETS OF T-HELPER CELLS
Th1:
◆ enhance inflammation(pro-inflammatory)
● IL-2:
● IFN-gamma:
○ stimulates B cell production of
○macrophage
● IL-12:
Th2:
◆ suppress inflammation(anti-
inflammatory)
● IL-4:
● IL-5:
Source:
Turgeon
B L YMPHOCYTES AND ITS
D EVELOPMENT
□ Involved in HUMORAL-MEDIATED IMMUNITY
□ Bone-marrow derived and precursor cells for antibody
production
1. PRO-B CELL (PROGENITOR B CELL)
● No antibodies yet
● There is rearrangement of genes coding for the formation of
heavy chains (Ch.14)
2. PRE-B CELL (PRECURSOR B CELL)
● No surface antibodies yet Light Chains:
● Mu chains are present in the cytoplasm Lambda: Ch. 22
● Heavy chains are present on the surface Kappa: Ch. 2
● There is rearrangement of genes coding for light chains
B L YMPHOCYTES AND ITS
D EVELOPMENT
3. IMMATURE B CELL
● Has complete antibodies on the surface
● First antibody on the surface of B cell: monomeric
IgM
● CD21 and CD35 markers are expressed
4. MATURE B CELL
● Increased IgM density
● IgD is expressed
● Cells are releases from the BM and seed in
peripheral lymphoid organs
B L YMPHOCYTES AND ITS
D EVELOPMENT
5. ACTIVATED B CELL
● CD25 triggered by IL-2 attachment enhanced
lymphocyte proliferation
□ a.k.a lymphocyte capping
6. PLASMA CELL
● Activated B cell
● Surface markers on B cell surface disappear
● WITHOUT surface immunoglobulins but WITH
cytoplasmic immunoglobulins which are later
released as antibodies
B CELL MATURATION
Plasma
Cell
Memory B
cell
CD M ARKERS ON T AND B C E LLS
ANTIGEN FUNCTION
CD2, CD3,
pan T-cell markers
CD5, CD7
CD19,
B-cell markers
CD20, CD21
CD21 receptor for Epstein-Barr Virus
CD2 sheep R BC receptor
marker for T-helper cells, receptor for MHC
CD4
class II molecule, receptor for HIV
marker for T-cytotoxic cells, receptor for
CD8
MHC class I molecule
CD10 CALLA marker
CD28 Binds B7 (CD80) of B cells
CD33 Myeloid receptor
CD152 a.k.a. CTLA-4; Binds B7 and B7.2 of B cells
CD154 CD40 ligand; Binds CD40 of B cells
ANTIGEN FUNCTION
CD16, CD56 NK cell markers
CD16 receptor for Fc portion of IgG
Regulator of IgE synthesis;
CD23
triggers release of GM-CSF from monocytes
CD25 receptor for interleukin 2
CD41, CD61 platelet and megakaryocyte markers
Adhesion molecule in most lymphocytes which mediate
CD44
homing to peripheral lymphoid organs
Essential in T and B cell antigen-stimulated proliferation
CD45
Formerly leukocyte common antigen
Decay accelerating factor (DAF) (GPI-linked), decays C3
CD55
and C5 convertases
CD56 Found in NK cells with no known function
Membrane of inhibitory reactive lysis (MIRL), inhibits MAC
CD59
formation
CD71 Glycophorin A, transferrin and erythroid receptor
CD94 Involved in inhibition of Nk cell cytotoxicity
REMEMBER!
□ CD4+ T cells recognize antigens ONLY in
the context of MHC class II antigens
□ MITOGENS
substances that initiate DNA synthesis and
mitosis or production of antibodies
□ T-cell blastogenesis is induced by:
● Pokeweed mitogens*
● Phytohemagglutinin
● Concanavalin A
2. DISEASES ASSOCIATION
● HLA-B27: (90% chance) ankylosing spondylitis
● HLA-DR3: SLE
● HLA-DR4: Rheumatoid arthritis
● HLA-B8:
□ Grave’s disease
□ Myasthenia gravis
□ Addison’s disease
I MPORTANCE OF HLA T YPING
3. PATERNITY TESTING
● An allele present in the child but not in the mother is
referred to as the obligatory paternal gene (OG)
● Direct exclusion
□ If the tested man does not have the possibility of passing
the obligatory paternal gene and the marker for the gene is
absent in the presumed mother BUT the child has it
● Indirect exclusion
□ absence of an expected genetic marker in the child when
the parent in question appears to be homozygous (one
allele present) for the gene
□ sometimes also known as reverse homozygosity
M ETHODS OF HLA T YPING
I. LYMPHOTOXICITY ASSAY/
MICROCYTOTOXICITY ASSAY)
□ (-) Unstained
M ETHODS OF HLA T YPING
II. MIXED LYMPHOCYTE REACTION (MLR)/
MIXED LEUKOCYTE REACTION
● Cellular assay
● Can be used to quantify the degree of class II
MHC compatibility between potential donors and
a recipient.
● Advantage over microcytotoxicity typing:
it gives better indication of the degree of TH cells
activation generated in response to the Class II
MHC antigens of the potential graft
P R O CEDURE :
1. Lymphocytes from potential donor that have been x-
irradiated or treated with mitomycin C serve as the
stimulator cells, and lymphocytes from the recipient
serve as responder cells.
2. Proliferation of the recipient T cells, which indicates T
cell activation, is measured by the uptake of
thymidine into cell DNA.
□ ANTIGEN
● Substance that reacts with a specific antibody but may not
be able to evoke an immune response in the first place
● Capable of combining with an antibody
● MAIN PARTS (of a complete antigen)
□ Carrier
immunogen)
□ CHEMICAL COMPOSITION & COMPLEXITY
● Protein> Polysaccharide>Lipids and Nucleic Acids
F ACTORS A FFECTING IMMUNOGENICITY
□ ROUTE, DOSAGE AND TIMING
● ↑ Dose = ↑ immune response
● IV and IP> ID> IM and SQ
□ GENETIC COMPOSITION
□ DEGRADABILITY
● ↑ degradability = ↓ immune response
□ STRUCTURAL STABILITY
● ↑ stability = ↑ immune response
□ ADJUVANTS
● Substances added to vaccines with less
immunogenic substances
● Added to enhance immune response of the body
A DJUVANTS
1. CFA (Complete Freund’s adjuvant)
water-in-oil emulsion of M. butyricum
●
Stimulates T cell activity = enhaced CMI
●
2. LPS (Lipopolysaccharides)
● Stimulates B cell activity = enhanced HI
3. Synthetic Muramil Dipeptide
● Stimulates T cell activity = enhanced CMI
4. Alum Adjuvant
MOST COMMONLY USED adjuvant in vaccination
●
enhances phagocytosis
●
5. Squaline (MF59)
● derived from shark oil, for HIV vaccine
T YPES OF A NTIGEN
□ Autoantigen
● derived from the same individual
□ Alloantigen/ Homologous antigen
● Derived from different individual of the same
species
□ Heteroantigen/ Xenogeneic antigen
● Derived from different species
● Triggers the greatest immune reponse
□ Heterophile Antigen
● Found in unrelated plants or animals which cross-
reacts with other antibodies
T YPES OF G RAFTS
□ Autograft
● Derived from the same individual
□ Isograft/Syngraft
● Derived from different class but identical
individual or twins
□ Allograft
● Found on different individual of the same
species
□ Heterograft/ Xenograft
● Derived from totally different species
I MMUNOGENECITY OF D IFFERENT
T RANSPLANT T ISSUES
□ Bone Marrow: MOST IMMUNOGENIC
□ Skin
□ Islets of Langerhans
□ Heart
□ Kidney
□ Liver
□ Bone
□ Xenogeneic valve replacements
□ Cornea: LEAST IMMUNOGENIC
● Privileged site not reached by the immune system
● Avascular
C ATEGORIES OF G RAFT R EJECTION
TYPE TISSUE MECHANISM CAUSE
DAMAGE
Hyperacute Within Humoral Preformed cytotoxic
minutes antibodies to donor
antigens
Accelerated 2-5 days Cellular- Previous
mediated sensitization to
donor antigens
Acute 7-21 days Cellular- Dev’t of allogeneic
mediated reaction to donor
antigens
Chronic >3 mos Cellular- Disturbance of
mediated host/graft tolerance
ANTIBODY
□ a.ka.
**subjected to electrophoresis, pH
□ 65%
● Lambda
□ Encoded in Chromosome 22
□ 35%
□ Isotype
● Heavy chain that determines immunoglobulin class
□ Allotype
● Variation
in constant region of both the heavy chain
heavy and light chains
□ Idiotype
● Variation in the variable region of both heavy and light
chain
M UST KNOW !
□ Monomer
● Basic immunoglobulin structure
● Composed of 2HC+ 2LC
□ IgG, IgD, IgE, Serum IgA and B cell surface IgM
□ Dimer
● Consists of 2 monomers
□ Secretory IgA : monomers are connected by a
□ Secretory component
● Prevents enzyme degradation of IgA
M UST KNOW !
✔ MOST PRIMITIVE
✔ First to appear in phylogeny and the last to leave in
senescence
✔ First antibody to appear on B-cells
✔ Major Ig class produce in primary immune response
✔ Does NOT have a hinge region
✔ Best agglutinating and complement-fixing antibody due to its
multiple binding sites
IgM
• Its heavy chain has an additional domain (CH4)
which serves as the receptor for complement
• Due to its large size, it is confined within
intravascular space and not found in C S F
• Free state = “STAR-LIKE”
• Attached to an antigen = “CRAB-LIKE”
• First effective defense against bacteremia
• Antibody most often formed in response to
stimulus by Gram (-) bacteria
• Consists of two subclasses:
• IgM1
• IgM2
F UNCTIONS :
1. Complement fixation
● BEST COMPLEMENT FIXER
2. Agglutination
● BEST AGGLUTINATING ANTIBODY
3. Opsonization
4. Neutralization of toxins
Examples of IgM:
● Wasserman antibodies,
● Heterophil antibodies
● Rheumatoid factor
● Cold agglutinins and allohemaglutinins
IgD
• Second to appear on B-cell
• Primarily a cell membrane immunoglobulin
found in the surface of MATURE B lymphocytes
in association with IgM
✔Postulated to be anti-idiotypic antibody
✔MOST SENSITIVE TO ENZYME
DEGRADATION
• Function: IMMUNOREGULATION
IgE
✔Least abundant immunoglobulin in serum
✔REAGINIC ANTIBODY
• Heat-labile antibody, originally called reagin
• Binds strongly to a receptor on mast cells and basophils
(via Fcεr) and together with antigen, mediates the
release of histamines and heparin from these cells
• Mediates some type of hypersensitivity reactions
(Type 1 hypersensitivity), allergies and anaphylaxis and
is generally responsible for an individuals immunity to
invading parasites
Serum: mono
Structure Mono Penta Mono Mono
Sec: dimer
Heavy
γ α μ δ ε
Chain
Serum
23 days 5-6 days 5 days 1-3 days 2-3 days
half-life
150,000 160,000 900,000 180,000 190,000
MW
Daltons Daltons Daltons Daltons Daltons
Sedimentation
coefficient 7S 7S 19S 7S 8S
A NTIBODY /POLYMER
R EDUCTION
□ Conversion of polymeric antibody into a
monomeric antibody
1. 2-mercaptoethanol (2-ME)
2. Dithiothreitol (DTT)