Antigens & Antibody

Structure, types and functions

Antigens (Ag)
• Antigens are any substance that binds specifically to an
antibody or a T cell receptor

• When an antigen is introduced into the host’s body, it activates

either T cells or B cells

• If T cell is activated, it undergoes division to form effector cells

and react with antigen in a process called cell mediated
immunity

• If B cells are activated, they secrete antibodies, and respond

by humoral mode

• Exogenous antigen, Endogenous antigens, Autoantigens

Characteristics of Antigen
Foreignness
• Higher the degree of foreignness, higher the antigenicity

Physico-chemical characteristics

• Molecular size
• Higher the MW, higher the antigenicity
• Less than 5 kDa poor Ag

• Chemical complexity
• Higher the chemical complexity, higher the antigenicity
• Proteins are better Ag than carbohydrates, lipids & NA

• Molecular shapes
• Compounds that lack a stable configuration are poorly recognized by the immune system and hence less
antigenic
• E.g. Gelatin structural instability non Ag

• Degradability
• Compounds that degrade rapidly are quickly eliminated from the body
• Hence, sufficient quantities not available poor Ag
Antigen vs Immunogen
• An immunogen refers to a molecule that is capable of eliciting an immune response by an
organism’s immune system

• Antigen refers to a molecule that is capable of binding to the product of that immune response

• So, an immunogen is necessarily an antigen, but an antigen may not necessarily be an

immunogen
Haptens
• A.k.a partial Ag

• Small molecules incapable of inducing humoral / cellular immunity by themselves

• But these when complexed with a protein / carrier molecule (e.g. serum protein), stimulate
immune response

• E.g. drugs, chemicals

• Penicillin, when bound to a body protein induces drug allergy

Epitopes
• The part of the antigen to which an antibody binds is called the epitope.
• The epitope is a short amino acid sequence that the antibody is able to
recognize.
• Epitope is the antigenic determinant region of an Ag that is recognized by
the B cell receptor or T cell receptor

• Each Ag contain different epitopes possessing different configuration each can elicit different
immune response (humoral cellular)
• One virus or microbe may have several antigenic
determinant sites, to which different antibodies
may bind.
Cross Reactions
• Sometimes different Ag possess identical epitopes.

• So, Ab generated against one Ag, may react with another Ag

• This is known as cross reaction

• This can be used to study relationships between molecules or organisms

Antibody
• An antibody is defined as “an immunoglobulin capable of specific combination with the antigen
that caused its production in a susceptible animal.”

• Antibodies are produced by plasma cells which are differentiated by B cells, in response to the
foreign proteins, called antigens.

• All antibodies are proteins known as immunoglobulins.

• Antibodies contain four polypeptide chains and are arranged in Y-shape.

• Each antibody has at least two identical sites that bind antigen: Antigen binding sites.

• Valence of an antibody: Number of antigen binding sites.

• Most are bivalent.

Basic Structure
• Antibody molecule contain 4 polypeptide chains, among them two are identical heavy chains and two
identical light chains.
• These chains are held together by disulphide bonds, some are inter-chain and some are intra-chain.

• Monomer: A flexible Y-shaped molecule with four protein chains:

• 2 identical light chains
• 2 identical heavy chains

• Variable Regions: Two sections at the end of Y’s arms.

Contain the antigen binding sites (Fab). Identical on the same antibody, but vary from one antibody
to another.

• Constant Regions: Stem of monomer and lower parts of Y arms.

• Fc region: Fragment that crystallizes

• The Fc fragment is where complement binds and also the anti-antibodies (anti-IgG) will bind. The
amino acid sequences present here are constant.
Antibody Classes
• There are five different isotypes of antibody depending on their difference in heavy chain.

• These includes:

• IgG, IgM, IgA, IgD and IgE.

• Heavy chains of IgG, IgM, IgA, IgD, and IgE, are known as gamma, mu, alpha, delta, and
epsilon, respectively.

Variation in:

• Amino acid sequences

• Location and no. of di-sulphide bonds

• MW

• Hinge region
IgG
• This is the principle antibody found in blood and body fluids.
• Nearly 75% of the antibody circulating in the blood is IgG.
• IgG is a monomeric immunoglobulin
• This is the only isotype that can pass through the placenta, thereby
providing protection to the fetus in its first weeks of life before its own
immune system has developed.

IgD
• It is monomeric in nature.

IgA
• IgA is involved in mucosal immunity and prevent the colonization of
bacteria in the digestive and respiratory tracts. It does not activate
complement.
IgM
• The IgM isotype is expressed on the surface of B cells and it is also
secreted by plasma cells.
• IgM is a pentamer
• Each monomer has two antigen binding sites, so an pentameric IgM has
10 antigen binding sites,
• It is also a "natural antibody" where it is found in the serum without any
prior contact with antigen.

IgE
• IgE is a monomeric immunoglobulin which is heat labile and plays an
important role in defending against parasitic worms.
• IgE is mainly responsible for allergies and this is through their ability to
trigger the release of chemicals from the granulocytes when the antibody
reacts with specific antigen.
Fish Antibodies
• Fishes are the lowest group of vertebrates that can synthesize Abs.

• Major group of antibodies found in fishes are IgM

• It is present in Tetrameric form, but also occur in monomeric or dimeric form.

• Functions also similar

• 2 heavy chains 70 kDa

• 2 light chains 25 kDa

Some rare types of Abs also present:

• IgR, IgN

• IgW in Sharks and skates

Antibody functions
Antigen-Antibody Complex: Formed when an antibody binds to an antigen it recognizes.

Affinity: A measure of binding strength.

1) Agglutination: Antibodies cause antigens (microbes) to clump together.

2) Opsonization: Antigen (microbe) is covered with antibodies that enhances its ingestion and
lysis by phagocytic cells.

3) Neutralization: inactivates viruses by binding to their surface and neutralize toxins by blocking
their active sites.

4) Antibody-dependent cell-mediated cytotoxicity: Used to destroy large organisms (e.g.:

worms). Target organism is coated with antibodies and bombarded with chemicals from
nonspecific immune cells.

5) Complement Activation: Both IgG and IgM trigger the complement system which results in
cell lysis and inflammation.
How Do B Cells Produce Antibodies?
• B cells develop from stem cells in the bone marrow of adults (liver of fetuses).

• After maturation B cells migrate to lymphoid organs (lymph node or spleen).

• Clonal Selection: When a B cell encounters an antigen it recognizes, it is stimulated and

divides into many clones called plasma cells, which actively secrete antibodies.

• Each B cell produces antibodies that will recognize only one antigenic determinant.
Clonal Selection of B Cells is Caused by Antigenic
Stimulation
Monoclonal antibody
• Each B lymphocyte in an organism synthesizes only one kind of antibody and a huge population of
different types of B cells can be seen in an organism capable of producing specific antibodies to the
various antigens that the organism had been exposed to.

• Monoclonal antibodies are single B lymphocyte generating antibodies to one specific epitope.

• In 1975 Kohler and Milstein developed a technology to fuse immortal Myleoma cells with B cells,
using poly ethyl glycol (PEG).

• The resulting cell type is called a hybridoma.

• This hybridoma takes on the characteristics of both the B cell (that produce antibody) and Myeloma
cell (which is immortal), creating an immortal cell with the ability to produce antibody.

• This hybridoma cells are selectively grown using HAT media.

• As the hybridoma cells grow they produce a specific antibody which is called a Monoclonal antibody.
Monoclonal antibody
Advantages

• Once hybridomas are made it is a constant and renewable source and all batches will be
identical

• They are less likely to cross-react with other proteins

Disadvantages

• High technology required.

• Training is required for the technology used.

• Time scale is long for hybridomas.

Polyclonal antibody
• Bacterial cell possess different epitopes, upon injection each epitope will stimulate the
proliferation and differentiation of B-cell to produce antibody, each particular antibody derived
from a B cell that recognizes a particular epitope.

• Hence large numbers of antibodies are produced with different specificities and epitope
affinities, The resulting antibodies in the serum are heterogeneous in nature, each specific for
one epitope.

• Thus polyclonal antibodies are antibodies that are obtained from different B cell resources and
are combination of immunoglobulin molecules secreted against a specific antigen, each
identifying a different epitope.

• So these antibodies are purified from the serum of immunised animals were the antigen of
interest stimulates the B-lymphocytes to produce a diverse range of immunoglobulin's specific
to that antigen.
Polyclonal antibody
Advantages

• Inexpensive to produce

• Technology and skills required for production low

• Production time scale is short

Disadvantages:

• Prone to batch to batch variability.

• Multiple epitopes may cross-reactivity and give false result.

Pattern of Antibody Levels During Infection
Primary Response:

• After initial exposure to antigen, no antibodies are found in serum for several days.

• A gradual increase in titer, first of IgM and then of IgG is observed.

• Most B cells become plasma cells, but some B cells become long living memory cells.

• Gradual decline of antibodies follows.

Secondary Response:

• Subsequent exposure to the same antigen displays a faster and more intense antibody response.

• Increased antibody response is due to the existence of memory cells, which rapidly produce
plasma cells upon antigen stimulation.