department of immunology room 225, building of basic medicine
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IntroductionTRANSCRIPT
Department of Immunology Room 225, Building of Basic
Medicine
Chapter 4 Immunoglobulin (Antibody) Xing-cheng WEI () Tel (office)
Department of Immunology Room 225, Building of Basic Medicine
Introduction von Behring (German) Found Ab [1890] Passive
immunization
Nobel prize [1901] 1890Koch,
BehringKitasato,,antibody.1891BehringBehring1901 von Behring
(German) Found Ab [1890] Passive immunization
-Throughout history, diphtheria was a leading cause of death among
children, and it was once referred to as the strangling angel of
children. 173517401080% -If diphtheria has become a rare disease
today, we owe it to the efforts of Emil von Behring, who discovered
the antitoxin to cure diphtheria. Found Ab [1890] Passive
immunization Nobel prize [1901] 1890Koch,
BehringKitasato,,antibody.1891BehringBehring1901 Bacillus
diphtheria and the disease
1890Koch, BehringKitasato,,antibody.1891BehringBehring1901
Difficulty of breathing with diphtheria In 1940, USA, elementary
school children Against diphtheria (A) goat anti-rabbit as the
secondary antibody (green),
*cryptocidin (epithelia ofintestineantimicrobial peptides. To
sterilize lumen (the killing mechanisms not known).
Immunocytochemistry analysis of HeLa cells detected withSTAT6
Recombinant Rabbit MAb and using (A) goat anti-rabbit as the
secondary antibody (green), (B) the nuclear stain (blue) (C) stain
actin (red). (D) A composite image of cells shows cytoplasmic
localization of STAT6. Antibody (Ab) A group of structurally
related glycoproteins produced by B cells, whichspecificallybindto
their correspondingAgsandmediate specific humoral immune
responsesto eliminate the bound Ags. *cryptocidin (epithelia
ofintestineantimicrobial peptides. To sterilize lumen (the killing
mechanisms not known). Immunoglobulin (Ig) *Ig:
The molecules that perform Ab activity or that no Ab function but
possess the similar structure with an Ab. *Ig: -Ab (functional, in
blood & other fluids) -BCR (B Cell Receptor), mIg (Membrane
Immunoglobulin) -sIgA (Secretory Immunoglobulin) *cryptocidin
(epithelia ofintestineantimicrobial peptides. To sterilize lumen
(the killing mechanisms not known). Immunoglobulins Plasma cells
Ab-Ag BCR/mIg-Ag BCR/mIg Memory B Sec.1 Molecular Structure of Ig
I. Basic Structure of Ig 1. Heavy Chain And Light Chain (1) H
Chain: -450-550 amino acid residues.
(2) L Chain : -214 amino acid residues. H-L linked by disulfide
bond. N Linked by S-S Linked by S-S *cryptocidin (epithelia
ofintestineantimicrobial peptides. To sterilize lumen (the killing
mechanisms not known). C (3) Domain of Ig Structure of IgG revealed
by X-ray crystallography
[Ig folding] -A globular motif. -2 Anti-parallel b sheet. -110AA.
-IgSF: any molecule that contains the Ig folding. Domain=functional
area
[L Chain] VLCL [H Chain] (IgGIgAIgD) VHCH1CH2 CH3 (IgMIgE) CH3CH4
CH4 (4) Class IgA IgD IgE IgG IgM a a d d g g e e m m
-Class is distinguished according to the structure of H chain CH of
Ig molecules. -There are 5 different H chain (g, a, m, d, e) in a
body, witch make up 5 classes of Ig molecule (IgA, IgD, IgE, IgG,
IgM). 5 Classes of Ig Processes of B Cell Mediated Immune
Responses
Activation Proliferation Differentiation (5) Subclass -IgG1 -IgG2
-IgG3 -IgG4 IgG -IgA1 IgA -IgA2 Ig
-IgG and IgA are divided into subclass. -According to the
difference of H chain constant region (CH). -IgG1 -IgG2 -IgG3 -IgG4
IgG -IgA1 -IgA2 IgA Ig (6) Type k k l l Type of the Ig is k Type of
the Ig is l
-According to the difference of CL, L chains are divided into 2
kinds (k or l). -The L chain of a Ig decides its type. k k l l Type
of the Ig is k Type of the Ig is l Class Subclass CH CL IgG IgG1-4
k/l IgA IgA1-2 IgM - IgD IgE
Type CH CL IgG IgG1-4 k/l IgA IgA1-2 IgM - IgD IgE 2. V and C
Region -Ag-binding specificity of a Ab is determined
1 2 3 4 -Ag-binding specificity of a Ab is determined by Ag-binding
sites of the Ab molecule, witch complementary bind to Epitopes.
-Basic structures of Abs are similar, but the amino acid sequences
that compose Ag-binding sites are different. -Thus in a individual,
Abs are mixture with different structures. (1) V regions: The
regions of variability in
amino acidsequence. (2) C regions: The regions of constancy in
amino acid sequence. VH: H chain 1/4 from N end VL: L chain 1/2
from N end CL: L chain 1/2 from C end CH: H chain 3/4 or 4/5 from C
end *cryptocidin (epithelia ofintestineantimicrobial peptides. To
sterilize lumen (the killing mechanisms not known). VAAIg Ig The
investigators who make clear the basic structure of the Ig
Rodney Porter Nobel Prize in 1972 Gerald Edelman Nobel Prize in
1972 (3) HVR / CDR: -The AA residues binding directly to an
epitope.
-The most variable portion in a V region. -An epitope-binding HVR
consists of 6 short AA sequences. *cryptocidin (epithelia
ofintestineantimicrobial peptides. To sterilize lumen (the killing
mechanisms not known). 3. Hinge Region Hinge Region -between CH1
and CH2, -a proline rich region.
-permitting flexibility to fit the structure of Ag. N *cryptocidin
(epithelia ofintestineantimicrobial peptides. To sterilize lumen
(the killing mechanisms not known). C II. Other Components of Ig 1.
J chain (Joining chain)
-A polypeptide chain that links 5 IgM monomers to form a pentameric
IgM. -A polypeptide chain that link 2 IgA monomers to form a
dimeric IgA. Structure of IgM and IgA 2. SP (Secretory piece) -A
polypeptide chain that bound to the CHs of a dimeric IgA to protect
against proteolytic cleavage. -To form a SIgA (secretary IgA) that
enters mucus layers to protects the mucosa. Structure of IgM and
IgA Digesting Fragment of Ig
III. Digesting Fragment of Ig Rodney Porter Nobel Prize in 1972
1.Papain Digest Products
-2 Fab(fragment of antigen-binding) -1 Fc (fragment crystalizable)
2.Pepsin Digest Products -1F(ab)2 -n pFc 3.significance -Clarify
the functions of Ig. -Prepare functional fragments for clinical or
research usage. Sec.2 Functions of Ab Major Functions of Ab
1Neutralization 2Opsonization
3Complement activation 4Mediation of ADCC Activate Complement Bind
to FcR
AgBinding Activate Complement Bind to FcR FcR Immune Cell
Mechanisms of Neutralization CDR/HVR of a Ab binds to:
-Microbial surface molecules to inhibit the microbe to invade into
host cell. -Microbial toxins to inhibit the toxins to interact with
host cell. Mechanisms of Ab-Mediated
Opsonization -CDR/HVR of Ab specifically bind to the Ag of microbe.
-Fc of Ab(IgG) bind to FcR on Phagocytes (Mf/Neutrophil). -The Abs
enhance phagocytosis efficiently. -The Abs mediate a innate immune
cell to kill the microbe specifically. Ab Microbe Phagocyte
Mechanisms for Abs to Activate Complement
After an Ab binding to an Ag, the CH changes
theconformationthatcanbeboundby C1,so as
toinitiatecomplementactivation.
Complementactivationgeneratesimportant effectors of both innate and
adaptive immune responses. Mechanisms of ADCC (Ab-Dependent
Cell-Mediated Cytotoxicity)
-CDR/HVR of Ab specifically bind to Ag of target cell (e.g. virus
infected cell). -Fc of the Ab binds to FcR on NK cell. -The Abs
enhance NK to kill the target cell efficiently and specifically. NK
Cell FcR Mf or NK Catch Target Cell
by FcR-Ab-Ag Binding Phagocyte or NK cell Sec.3 Features &
Functions of Each Ig Class Major Characteristics
Class Serum conc [mg/ml] Half life [day] Major Characteristics IgA
3.5 6 SIgA is the only Ab that is secreted into mucus layer to
protect mucosa (mediate mucosal immune) -External secretion
alimentary tract respiratory tract lacteal gland salivary gland
tear gland -Colostrum (neonate) Generation & Secretion
of SIgA Secreted IgA is transported through
epithelial cells by the poly-Ig receptor SIgA is transported into
the intestinal lumen by an IgA specific Fc receptor Characteristics
Class Serum conc [mg/ml] Half life [day] IgD trace 3
mIg (BCR) Marker of mature B cell -IgM+IgD+ B cell. IgE 0.05 2
Mediate type I Hypersensitivity BCRs on mature B Cell Consist of
both IgM and IgD Characteristics Class Serum conc [mg/ml] Half life
[day] IgG 13.5 23
Most rich in blood. Longest half life. Major Ab in 2nd immune
response. Highest Affinity. Pass placenta. Primary and Secondary
Humoral Responses ()
Log 100000 10000 1000 100 10 1 IgG IgM Placental barrier Placental
barrier FcRn transmit IgG through placental or intestinal
cells
*FcRn: FcgR, intracellular trafficking receptor n: neonate Pentamer
is strong in: The marker of immature B cell
IgM 1.7 10 Produced earliest in immune responses -1st line defense
against infection. -Early diagnosis of infection. Produced earliest
in ontogenesis -diagnosis of intra-uterine infection (Cord Blood)
Pentamer is strong in: -Ag binding ability. -Complement activation.
The marker of immature B cell -IgD-IgM+ B is a immature cell that
resides in bone marrow. Primary and Secondary Humoral
Responses
IgM Log - - 100000 10000 1000 100 10 1 IgG IgM Only 1 IgM can
initiate complement Activating reaction
10 Ag-binding sites of 1 IgM Only 1 IgM can initiate complement
Activating reaction Pentamer IgM is strong in Ag binding and
complement activation The Cell that Only Expresses IgM on surface
is a Immature B Cell
IgD has not yet been expressed on the cell surface Sec.4
Preparation of Ab -Polyclonal Ab [pAb] -Monoclonal Ab [mAb]
-Genetic engineering Ab mAb [Hybridomas] -Proliferation in vitro or
in vivo
By cell fusion between a normal B cell and a myeloma cell to get
the hybridomas that secret specific Abs to one kind of epitopes.
[Hybridomas] -Proliferation in vitro or in vivo -Secret Ab in vitro
or in vivo [mAb] Single epitope specific Uniform High purity Low
cost of preparation Genetic Engineering Antibody
The Abs expressed by recombined Ig gene that was deleted, spliced
or revised before transferred into cells. -human-mouse chimeric
antibody. - -reshaped humanized antibody. -small molecular antibody
Fab fragment Fcfragment Fvfragment -bispecific antibody -Ig-fusion
protein Ig- -antibody-directed enzyme [The main points] 1.Concept:
(1) Monoclonal Ab;
(2) Genetic engineering Ab; 2.Functions of Ab (Ig).
3.Characteristics of: (1) IgA (2) IgD (3) IgE (4) IgG (5) IgM