Transcript Lect 04 Mab1x

Dr. sarah I bukhari
Assistant Professor of pharmaceutical
microbiology Microbiology
www.themegallery.com
Microbiology section
The Latin term “IMMUNIS” means EXEMPT, referring to
protection against foreign agents.
DEFINITION: - The integrated body system of organs,
tissues, cells & cell products that differentiates self from
non – self & neutralizes potentially pathogenic
organisms.
(The American Heritage Stedman's Medical Dictionary)
The Immune System consists of
1. Innate Immunity
2. Acquired Immunity
Response
Primary Response
Secondary
FUNCTIONING OF THE IMMUNE SYSTEM
HUMORAL (ANTIBODY MEDIATED)
IMMUNE RESPONSE
CELL MEDIATED IMMUNE RESPONSE
ANTIGEN (1ST EXPOSURE)
ENGULFED BY
MACROPHAGE
FREE
ANTIGENS
DIRECTLY
ACTIVATE
ANTIGENS
DISPLAYED
BY
INFECTED
CELLS
ACTIVATE
BECOMES
APC
STIMULATES
B CELLS
STIMULATES
HELPER
T CELLS
STIMULATES
MEMORY
HELPER T
CELLS
GIVES RISE TO
STIMULATES
STIMULATES
CYTOTOXIC
T CELL
GIVES RISE TO
STIMULATES
ANTIGEN (2nd EXPOSURE)
PLASMA
CELLS
MEMORY
B CELLS
SECRETE ANTIBODIES
STIMULATES
MEMORY
T CELLS
ACTIVE
CYTOTOXIC T
CELL
IMMUNOTHERAPY
Treatment of the disease by Inducing, Enhancing or
Suppressing the Immune System.
Enhancing the
immune system
by
Active Immunotherapy: -
Passive Immunotherapy: -
It stimulates the body’s own
immune system to fight the
disease.
It does not rely on the body to
attack the disease, instead they
use the immune system
components ( such as
antibodies) created outside the
body.
•1895 Jules Bordet, Complement and antibody
activity in bacteriolysis
•1926 Lloyd Felton & GH Bailey, Isolation of pure antibody
preparation
•1958-62 Jean Dausset et al., Human leukocyte antigens
•1934-8 John Marrack, Antigen-antibody binding hypothesis
•1959-62 Rodney Porter et al., Discovery of antibody
structure
•1900 Paul Erlich, Antibody formation theory
•1936 Peter Gorer, Identification of the H-2 antigen in mice
•1959 James Gowans, Lymphocyte circulation
•1901 Karl Landsteiner, A, B and O blood groupings
•1940 Karl Lansteiner & Alexander Weiner, Identification of
the Rh antigens
•1961-62 Jaques Miller et al., Discovery of thymus
involvement in cellular immunity
•1941 Albert Coons, Immunofluorescence technique
•1961-62 Noel Warner et al., Distinction of cellular and
humoral immune responses
•1901-8 Carl Jensen & Leo Loeb, Transplantable
tumors
•1902 Paul Portier & Charles Richet, Anaphylaxis
•1903 Almroth Wright & Stewart Douglas,
Opsonization reactions
•1906 Clemens von Pirquet, coined the word allergy
•1862 Ernst Haeckel, Recognition of phagocytosis
•1877 Paul Erlich, recognition of mast cells
•1879 Louis Pasteur, Attenuated chicken cholera vaccine
development
•1883 Elie Metchnikoff Cellular theory of vaccination
•1798
Edward
Jenner,
•1942
Jules Freund
& Katherine McDermott,
Adjuvants
•1942
Karl
Landsteiner
&
Merill
Chase,
Cellular
transfer of
Smallpox vaccination
sensitivity in guinea pigs (anaphylaxis)
•1944 Peter Medwar, Immunological hypothesis of allograft
rejection
•1948 Astrid Fagraeus, Demonstration of antibody
production in plasma B cells
•1948 George Snell, Congenic mouse lines
•1949 Macfarlane Burnet & Frank Fenner,
Immunological tolerance hypothesis
•1963 Jaques Oudin et al., antibody idiotypes
•1964-8 Anthony Davis et al., T and B cell cooperation in
immune response
•1965 Thomas Tomasi et al., Secretory immunoglobulin
antibodies
•1967 Kimishige Ishizaka et al., Identification of IgE as
the reaginic antibody
•1971 Donald Bailey, Recombinent inbred mouse strains
•1974 Rolf Zinkernagel & Peter Doherty, MHC restriction
•1975 Kohler and Milstein, Monoclonal
•1950 Richard Gershon and K Kondo, Discovery of
•1975
and Milstein, antibodies used in genetic analysis
suppressorKohler
T cells
•1888 Pierre Roux & Alexandre Yersin, Bacterial toxins
Robert Good, Failed treatment of severe combined
•1952 Ogden and Bruton, discovery of
antibodies used in genetic •1984
analysis
•1888 George Nuttall, Bactericidal actionMonoclonal
of blood
immunodeficiency
(SCID, David the bubble boy) by bone
agammagobulinemia (antibody immunodeficiency)
•1885 Louis Pasteur, Rabies vaccination development
•1891 Robert Koch, Delayed type hypersensitivity
marrow grafting. 1985 Tonegawa, Hood et al.,
Identification of immunoglobulin genes
•1894 Richard Pfeiffer, Bacteriolysis
•1953 Morton Simonsen and WJ Dempster, Graftversus-host reaction
•1907 Svante Arrhenius, coined the term immunochemistry
•1953 James Riley & Geoffrey West, Discovery of
histamine in mast cells
•1910 Emil von Dungern, & Ludwik Hirszfeld, Inheritance of
ABO blood groups
•1953 Rupert Billingham, Leslie Brent, Peter Medwar, &
Milan Hasek, Immunological tolerance hypothesis
•1990 Yamamoto et al., Molecular differences between
the genes for blood groups O and A and between those
for A and B
•1910 Peyton Rous, Viral immunology theory
•1955-1959 Niels Jerne, David Talmage, Macfarlane
Burnet, Clonal selection theory
•1990 NIH team, Gene therapy for SCID using cultured T
cells.
•1957 Ernest Witebsky et al., Induction of autoimmunity
in animals
•1993 NIH team, Treatment of SCID using genetically
altered umbilical cord cells.
•1957 Alick Isaacs & Jean Lindemann, Discovery of
interferon (cytokine)
•1985-onwards Rapid identification of genes for immune
cells, antibodies, cytokines and other immunological
structures.
•1914 Clarence Little, Genetics theory of tumor
transplantation
•1915-20 Leonell Strong & Clarence Little, Inbred mouse
strains
•1917 Karl Landsteiner, Haptens
•1921 Carl Prausnitz & Heinz Kustner, Cutaneous reactions
•1924 L Aschoff, Reticuloendothelial system
•1985-7 Leroy Hood et al., Identification of genes for the T
cell receptor
Immunoglobulins






Glycoproteins
Bind specifically to antigens that induced
their formation
Present in gamma globulin fraction of
plasma proteins
Present in extravascular compartment
Five classes: IgG IgA IgM IgE IgD
Subclasses also present: IgG1, IgG2,
IgG3, IgG4
Antibody Structure
 Two
identical
light chains
 Two
identical
heavy chains
 Linked
bonds
by S-S
Light Chains





200 amino acids
each
MW: 25kDa
May be kappa (κ)
or lambda (λ)
Both types found
among all isotypes
One type only per
antibody
Heavy chains




400 amino acids
MW: 50 – 75 kDa
Determine isotype of
antibody:
◦ Gamma (γ) → IgG
◦ Alpha (α) → IgA
◦ Mu (μ) → IgM
◦ Delta (δ) → IgD
◦ Epsilon (ε) → IgE
2 heavy chains
joined by
S-S bonds in hinge
region
Domains

Variable domains:
widely variable
amino acid sequence

Constant domains:
uniform amino acid
sequence
Light chain:
one VL + one CL
Heavy chain:
one VH + 3-4 CH


The structure of antibodies

http://www.path.cam.ac.uk/~mrc7/igs/mikeimages.html
Hypervariable Regions
 Variability
of
amino acid
sequence in
VH and VL
domains is
restricted to
short
segments

Hypervariable
regions of H and
L chains are
folded together
single →
hypervariable
surface:
PARATOPE

Paratope
= antigenbinding site

Paratope
complementary
to epitope
Proteolytic cleavage of antibody gives:
I.
Two identical
Fab
fragments:
(fragment
antigen
binding)
II.
One Fc
fragment:
(fragment
crystallizable)
Biological activities:
• Complement fixation
• Attachment to various
cells
• Placental transfer
NOTE:
Different isotypes
different Fc fragments
different functions
Structure of Antibody
POLYCLONAL vs
MONOCLONAL ANTIBODIES
EPITOPE = ANTIGENBINDING
SITE
• Protein Antigens have many epitopes
• B cells make Ab to a single epitope
• Different clones of B cells make Ab
to different epitopes
Polyclonal and Monoclonal
Antibodies
Adapted from Milstein (1980) Scientific American, Oct. p.58
31 2 4
Antigen
1
2
BA
LB /
c
Immunization
3
m
4
m
B cell
1
2
3
31 2 4
m
2 3
4
1
2
Spleen cells
+
Myeloma
1
4
x
m
3
Cell fusion
4
Monoclonal
antibodies
Antiseum
A mixture of all Ab
Pure single Ab
If a single
Each
B cellB produces
cell was picked
only one
up kind
and of
cultured,
antibody,
then
which
it willbinds
produce
to itsonly
specific
one kind
antigen.
of antibody.
After
Myeloma
Each
immunization,
hybridoma
cell
canline
bethe
can
cultured
mouse
produce
inspleen
the
pure
test
contains
single
tube,
antibody,
but
B cells
canbenot
producing
called
produce
monoclonal
specific
useful
antibody.
antibodies.
antibody.
If
B
cell
is
fused
with
myeloma,
the
fused
cell
might
cultured
and
produce
antibody.
Conventional
But
B cell canantiserum
not surviveiswell
the mixture
in
culture.
of all antibodies produced by B cells from spleen.
Derived from different B
Lymphocytes cell lines
Derived from a single B
cell clone
Batch to Batch variation
affecting Ab reactivity &
titre
mAb offer Reproducible,
Predictable & Potentially
inexhaustible supply of Ab
with exquisite specificity
NOT Powerful tools for
clinical diagnostic tests
Enable the development of
secure immunoassay
systems.
Functions of antibodies
1. Antibodies neutralize bacterial
toxins and complete virus
particles and bacterial cells.
2. Antibodies coating an antigen
render it recognizable as foreign
by phagocytes (macrophages
and neutrophils), which then
ingest and destroy it; this is
called opsonization.
3. Antibodies activate of the
complement system by coating
a bacterial cell.
4. Antibody-dependent cellular
cytotoxicity (ADCC)
Fig. 1.24 Antibodies can participate
in host defense in three main
ways.
1- Neutralization
 Prevents
attachment to
cell receptors
prevents
infectivity or
toxicity
2- Opsonisation
 Binding
of antibodycoated pathogen to
Fc receptor on
phagocyte
→ better
phagocytosis and
killing
3- Complement Activation
This may lead to:
 Opsonization
 Lysis
of bacterial cell
Complement activation followed
by opsonization
Complement activation leading
to lysis
4- Antibody-dependent cellular
cytotoxicity (ADCC)
Thank you