This document provides information about antibodies (immunoglobulins). It discusses the structure of antibodies, including the variable and constant regions of the heavy and light chains. The five classes of antibodies (IgG, IgA, IgM, IgD, IgE) are described along with their functions. IgG is the most abundant antibody and can activate complement and mediate phagocytosis. IgA is found in secretions and provides mucosal immunity. IgM is the first antibody produced during infection and is a potent complement activator. Abnormal immunoglobulins produced in diseases are also mentioned.
1. Dr. Meenakshi Sharma
Assistant Professor
Department of Microbiology
Mayo Institute Of Medical Sciences
Antibodies - Immunoglobulins
2. Index
Introduction
Structure of Antibody
Functions of Immunoglobulins
Immunoglobulin classes
Antigenic determinants of
Immunoglobulins
Abnormal Immunoglobulins
3. Introduction
Antibodies are
Glycoprotein molecules that recognise a particular
epitope on an antigen
Bind specifically to it and facilitates its clearance.
Present on B cell membrane and secreted by
plasma cells.
Secreted antibodies circulate in blood, where they
eliminate/neutralise the antigen
Sera having high antibody levels following
infection or immunisation –IMMUNE SERA
4. Fractionation of immune sera separates the
serum proteins by half saturation with ammonium
sulphate separates the serum proteins into:-
Soluble albumin
Insoluble globulins
Globulins can be separated into water soluble
Pseudoglobulins and water insoluble Euglobulins.
Most antibodies have been found to be
euglobulins
6. Most antibodies were found in gamma globulin
frations, hence named immunoglobulins (Ig).
7. In 1964, WHO endorsed the term
‘immunoglobulin’ which was internationally
accepted .
The definition includes besides antibody
globulins, the abnormal protiens found in
myeloma, macroglobulinemia, cryoglobulinemia
and the naturally occuring subunits of
immunoglobulins.
All antibodies are immunoglobulins but all
immunoglobulins may not be antibodies.
8. Immunoglobulin (Ig) constitute 20-25% of total
serum proteins.
Based on physiochemical and antigenic
differences 5 classes: IgG, IgA, IgM, IgD and IgE
9. Structure of Antibody
Y shaped heterodimer,
composed of 4
polypeptide chains
2 identical light (L)chains, of
molecular weight 25000 Da each
and
2 identical heavy (H) chains
each having molecular weight
50000 Da or more
10. All 4 H and L chains are bound to each other by
disulfide bonds and noncovalent interactions such
as salt linkages, hydrogen bonds and
hydrophobic bonds All chains have 2 ends
an amino terminal end (NH3) and
a carboxy terminal end (COOH)
There are 5 classes of H chains
and 2 classes of L chains
11. H chains
5 classes
Structurally and antigenically distinct
Each designated by Greek letter corresponding
to immunoglobulin class
5 classes of Ig (IgG, IgA, IgM, IgD and IgE)
classified based on AA sequence of heavy chains
12. L chains
2 types
Kappa (κ) and lambda (λ) named after Korngold
and Lapari
In humans, L chains, 60% kappa and 40%
lambda
Both light chains of Ab molecule should be same
type, either κ or λ, never both
13. Type of heavy chain in each Ig class
Immunoglobulin class Heavy chain type
IgG γ (gamma)
IgA α (alpha)
IgM μ (mu)
IgD δ (delta)
IgE ε (epsilon)
14. Variable and constant regions
Each H and L chain – comprises of 2 regions
Variable region
Constant region
Depending upon whether AA sequences of the
regions show variable or uniform pattern among
different antibodies
15. Variable region
The 1st 110 AA residues near amino terminal end
(NH3) end of both L and H chains constitute
variable region – V L and VH respectively.
Hypervariable region: maximum sequence
variation is concentrated in a few discrete regions
called hypervariable regions
Less variable stretches are termed as framework
regions
16. HV regions form the antigen binding site which
are complementary to the structure of the epitope
and are called complementarity determinig
regions (CDR’s)
Each Fab fragment has six CDR’s (three in H and
3 in L)
The framework region acts as a scaffold support
to six CDR loops.
Paratope: site on hypervariable regions that make
actual contact with epitope
17. Constant region
The sequence of aminoacids beyond the variable
region is realtively constant throughout the
antibody molecule and is caleed the C constant
region .
Length = 104 AA for L chain, 330 AA for γ, α, and
δ heavy chains and 440 AA for μ and ε heavy
chains.
Carbohydrate moieties are linked to constant
region of H chains
Carboxy terminal constant region of heavy chains
mediates the effector functions.
The C region of light chains does not attach to the
cell membrane and does not participate in its
effector functions.
18. Immunoglobulin domains
H and L chain are further
folded into
domains
Within a domain, a loop like
structure of 60 AA is present
which is formed due to an
intrachain disulfide bond
Light chain – contains one
variable (VL) and one constant
domain (CL)
Heavy chain – one variable
(VH ) and 3-4 constant domains
(CH)
19. Hinge region
In heavy chain (γ,α,δ), the
junction formed between CH1
and CH2 domain = Hinge region
Rich in proline and cysteine
Very flexible, allows Ig
molecule to assume different
positions, helps Ab reach
towards Ag
Sensitive to various enzymatic
digestions
21. In the presence of cysteine, Cleave Ig above
disulfide bridge of hinge region into two fractions :
Insoluble fraction which crystallises in cold called Fc for
crystallisable
Soluble fraction which while unable to precipitae can still
bind the antigen called Fab (antigen binding)
Results in 3 fragments each
Two Fab fragments
One Fc fragment
Papain digestion
22. Pepsin digestion
Cleaves Ig molecule at point below
disulfide bridge of hinge region
One F(ab’)2 fragment; 2 Fab subunits
bound together
Many smaller fragments
23. Functions of Immunoglobulins
Antigen binding (by Fab region)
Primary function of an antibody; protects host
Fab fragment – bears variable region; involved in interaction
with Ag
Valency of Ab = No. of Fab region it possesses.
Effector functions (by Fc region)
Variety of secondary “effector functions” are produced
like
Fixation of complement
Binding to various cell types
25. IgG
70-80% of total Ig in body
Maximum daily production, longest half life of 23
days
Highest serum concentration
Has 4 subclasses – IgG1, IgG2, IgG3 and IgG4
Subclasses vary in biological function, length of
hinge region and No. of disulfide bridges
26. Functions
Can cross placenta: provide immunity to fetus and
newborn
Complement fixing: Fc region can bind to
complement factors, activate classical pathway of
complement system
Phagocytosis: IgG1 and IgG3 bind to Fc receptors
on phagocytes with high affinity and enhances the
phagocytosis of antigen bound to it
Mediate precipitation and neutralisation reaction
Plays major role in neutralisation of toxins as it can
easily diffuse into extravascular space
Is raised after long time following infection and
represents chronic or past infection
27. IgA
Second most abundant
Constitutes 10-15% of total serum Ig
Half life of 6-8 days
2 subclasses: IgA1 and IgA2
Occurs in two forms :Serum IgA and Secretory
IgA
Serum IgA : monomeric 7 S molecule, Interacts
with Fc receptors expressed on immune effector
cells, to initiate various functions like ADCC,
degranulation of immune cells, etc
28. Secretory IgA
Dimeric in nature, larger molecule than serum IgA
2 IgA monomeric units joined by J chain
Also secretory component present,belived to
protect IgA from denaturation by bacterial
proteases
Major Ig in colostrum, saliva & tears
29. Functions
Secretory IgA provides an important defence
mechanism against organisms like salmonella,
vibrio cholera and viruses like polio, influenza etc.
Breast milk rich in IgA – protects the newborn
against infection during first month of life.
IgA does not fix complement but can activate the
alternative complement pathway.
Promotes phagocytosis and intracellular killing of
microorganisms.
31. IgM
Highest MW (Millionaire molecule)
Present only in intravascular compartment
Not in body fluids or secretions
Exists in monomeric and pentameric forms (10
Fab regions and 10 valencies)
32. Functions
Acute infection: 1stAb to be produced following
infection.
Represents acute or recent infection. Also called
primary immune response Ab
Complement fixing: most potent activator of
classical complement pathway. Multiple
complement binding sites (5 Fc regions)
Present on B cell surface. Serves as B cell
receptor for Ag binding
Acts as Opsonin: binds as antigen which is then
easily recognised and removed
33. Functions
Protection against intravascular organisms
Mediate agglutination: 20 times more effective in
bacterial agglutination than IgG
Fetal immunity: 1stAb to be synthesised in fetal
life. Indicator of IU infection
34. IgE
Lowest serum concentration, shortest half life and
minimum daily production
Affinity for surface of tissue cells (mast cells)
Functions
Potent mediator of Type 1 hypersensitivity
Elevated in helminthic infection
35. IgD
Found as membrane Ig on surface of B cells
Acts as B cell receptor along with IgM
36.
37. Antigenic determinants of
Immunoglobulins
Since Ab are glycoproteins, they can themselves
function as potent immunogens
Can induce Ab response in hosts other than
parent host
Not entire Ig molecule is immunogenic, contains
antigenic determinants at specific sites
Based on location of Ag determinants: Ig
molecules divided into isotype, idiotype and
allotype
38.
39.
40. Abnormal Immunoglobulins
Bence Jones proteins
Produced in neoplastic condition of plasma cells
called Multiple Myeloma
Also called light chain disease
Cancerous plasma cells produce excess light chain
(BJP) accumulated in pt serum and urine
Waldenstrom’s Macroglobulinemia :B cell
lymphoma, produce excess IgM
Heavy chain disease
Cryoglobulinemia