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Antigen is a substance usually protein in nature and sometimes polysaccharide, that generates a
specific immune response and induces the formation of a specific antibody or specially
sensitized T cells or both.
Although all antigens are recognized by specific lymphocytes or by antibodies, only some
antigens are capable of activating lymphocytes. Molecules that stimulate immune responses are
called Immunogens.
Epitope is immunologically active regions of an immunogen (or antigen) that binds to antigen-
specific membrane receptors on lymphocytes or to secreted antibodies. It is also called antigenic
determinants.
Autoantigens, for example, are a person’s own self antigens. Examples: Thyroglobulin, DNA,
Corneal tissue, etc.
Alloantigens are antigens found in different members of the same species (the red blood cell
antigens A and B are examples).
Heterophile antigens are identical antigens found in the cells of different species. Examples:
Forrssman antigen, Cross-reacting microbial antigens, etc.
Adjuvants are substances that are non-immunogenic alone but enhance the immunogenicity of
any added immunogen.
Chemical Nature of Antigens (Immunogens)
A. Proteins
The vast majority of immunogens are proteins. These may be pure proteins or they may be
glycoproteins or lipoproteins. In general, proteins are usually very good immunogens.
B. Polysaccharides
Pure polysaccharides and lipopolysaccharides are good immunogens.
C. Nucleic Acids
Nucleic acids are usually poorly immunogenic. However, they may become immunogenic when
single stranded or when complexed with proteins.
D. Lipids
In general lipids are non-immunogenic, although they may be haptens.
These antigens enters the body or system and start circulating in the body fluids and
trapped by the APCs (Antigen processing cells such as macrophages, dendritic cells, etc.)
The uptakes of these exogenous antigens by APCs are mainly mediated by the
phagocytosis
Examples: bacteria, viruses, fungi etc
Some antigens start out as exogenontigens, and later become endogenous (for example,
intracellular viruses)
These are body’s own cells or sub fragments or compounds or the antigenic products that
are produced.
The endogenous antigens are processed by the macrophages which are later accepted by
the cytotoxic T – cells.
Endogenous antigens include xenogenic (heterologous), autologous and idiotypic or
allogenic (homologous) antigens.
Examples: Blood group antigens, HLA (Histocompatibility Leukocyte antigens), etc.
3. Autoantigens
Possess antigenic properties denovo, i.e. ther are able to generate an immune response by
themselves.
High molecular weight (more than 10,000)
May be proteins or polysaccharides
2
Determinants of Antigenicity
The whole antigen does not evoke immune response and only a small part of it induces B and T
cell response.
The small area of chemical grouping on the antigen molecule that determines specific immune
response and reacts specifically with antibody is called an antigenic determinant.
Property of antigens/ Factors Influencing Immunogenicity
1. Foreignness
2. Molecular Size
The most active immunogens tend to have a molecular mass of 14,000 to 6,00,000 Da.
Examples: tetanus toxoid, egg albumin, thyroglobulin are highly antigenic.
Insulin (5700 ) are either non-antigenic or weakly antigenic.
In general, the more complex the substance is chemically the more immunogenic it will
be.
Antigens are mainly proteins and some are polysaccharides.
It is presumed that presence of an aromatic radical is essential for rigidity and
antigenicity of a substance.
4. Physical Form
5. Antigen Specificity
Antigen Specificity depends on the specific actives sites on the antigenic molecules
(Antigenic determinants).
Antigenic determinants or epitopes are the regions of antigen which specifically binds
with the antibody molecule.
6. Species Specificity
7. Organ Specificity
The autologous or self antigens are ordinarily not immunogenic, but under certain
circumstances lens protein, thyroglobulin and others may act as autoantigens.
Some substances are immunogenic in one species but not in another .Similarly, some
substances are immunogenic in one individual but not in others (i.e. responders and non-
responders).
The species or individuals may lack or have altered genes that code for the receptors for
antigen on B cells and T cells.
They may not have the appropriate genes needed for the APC to present antigen to the
helper T cells.
10. Age
11. Degradability
Generally the subcutaneous route is better than the intravenous or intragastric routes.
The route of antigen administration can also alter the nature of the response.
Antigen administered intravenously is carried first to the spleen, whereas antigen
administered subcutaneously moves first to local lymph nodes.
14. Adjuvants
Substances that can enhance the immune response to an immunogen are called adjuvants.
The use of adjuvants, however, is often hampered by undesirable side effects such as
fever and inflammation.
Example: aluminum hydroxide.
Superantigens
When the immune system encounters a conventional T-dependent antigen, only a small
fraction (1 in 104 -105) of the T cell population is able to recognize the antigen and
become activated (monoclonal/oligoclonal response).
However, there are some antigens which polyclonally activate a large fraction of the T
cells (up to 25%). These antigens are called superantigens.
Examples of superantigens include: Staphylococcal enterotoxins (food poisoning),
Staphylococcal toxic shock toxin (toxic shock syndrome), Staphylococcal exfoliating
toxins (scalded skin syndrome) and Streptococcal pyrogenic exotoxins (shock).
Although the bacterial superantigens are the best studied there are superantigens
associated with viruses and other microorganisms as well.
The diseases associated with exposure to superantigens are, in part, due to hyper
activation of the immune system and subsequent release of biologically active cytokines
by activated T cells.
Structure
Antibodies are heavy (~150 kDa) globular plasma proteins. The size of an antibody molecule is
about 10 nm. They have sugar chains (glycans) added to conserved amino acid residues. In other
words, antibodies are glycoproteins. The attached glycans are critically important to the structure
and function of the antibody. Among other things the expressed glycans can modulate an
antibody's affinity for its corresponding Fc receptors.
Anatomy of light (L) and heavy (H) chain:
L- chain:
L- chain of antibody is composed of about 220 aminoacids.
Around 100-110 aminoacids are located at N-terminal (amino-terminal) and the
aminoacids sequences varies among antibodies. This region of L-chain is known as
variable (V) region.
Remaining 110 aminoacids located at C-terminal (carboxyl-terminal) of L-chain are
almost constant among antibodies. This region of L-chain is known as constant (C) region.
Two types of constant region sequences are found ie. Lambda (λ) and Kappa (κ). In a
particular antibody either2lambda or 2 kappa chains are present but not 1 lambda and
kappa.
In human 60% light chain are kappa and 40% are lambda whereas in mice 95% of light
chain are kappa and 5% are lambda.
H-chain:
In H-chain about 110 aminoacids are located at N-terminal which shows great variation
among antibody. This region is known as Variable (V) region.
Remaining aminoacid sequences of H-chain is somewhat constant but reveals five
different types of constant (C) heavy chain region ie. µ, α, δ, ε and γ.
The length of constant region of H-chain is 330 aminoacids for α, γ and δ and 440
aminoacids for µ and ε.
Antibodies molecules are classified into five class on the basis of constant region of H-chain.
The γ, δ and α heavy chain contains an extended peptide sequence between CH1 and
CH2 domain that has no homology with other domain, this region is known as hinge
region.
Hinge region is rich in proline residue and is flexible. Therefore IgG, IgD and IgA are
flexible.
The flexibility given by hinge region enable Fab region to assume various angle to bind
antigen.
Characteristics of antibodies;
Classes/Types of Antibody
Serum containing antigen-specific antibodies is called antiserum. The 5 types – IgG, IgM, IgA,
IgD, IgE – (isotypes) are classified according to the type of heavy chain constant region, and are
distributed and function differently in the body.
1. IgG:
Molecular weight: 150,000 Da
H-chain type: gamma (53,000 Da)
IgG is the most abundant class of Immunoglobulin in serum and constitute of about 80%
of total serum immunoglobulin.
IgG molecule consists of two gamma (γ) heavy chains and two kappa (k) or two lambda
(λ) light chains.
There are four sub class of IgG ( IgG1, IgG2, IgG3 and IgG4) on the basis of decreasing
serum concentration.
It has longest half-life among other antibodies. Half-life is about 23 days.
IgG is the only antibody that can cross placenta. It cross placenta and provide immunity
to fetus upto 6 month of age. The immunity is known as natural passive immunity.
It can also activate complement.
Biological functions:
IgG is the major antibody produced in secondary immune response.
Ig, IgG3 and IgG4 readily cross the placenta and play important role in protecting the
fetus.
IgG3 is the most effective complement activator followed by IgG1 and IgG2. IgG4 is not
able to activate complement at all.
IgG1 and IgG3 binds with high affinity to Fc receptor on phagocytic cell and thus
mediate opsonization.
IgG helps in bacterial immobilization.
IgG neutralize toxin and viruses.
2. IgM:
Molecular weight: 900,000 Da
H-chain type: mu (65,000 Da)
IgM accounts for 5-10% of total serum Immunoglobulin with an average serum
concentration of 1.5mg/dl.
IgM is secreted by plasma cell and it exists in pentameric form in which five IgM
mononers are linked together by disulphide bond (J-chain).
Due to large size, IgM is also known as millionare molecule.
There are 10 antigen binding site (Fab) in pentameric IgM molecule but it cannot bind to
10 complete antigen due to steric hindrance.
It is the major antibody produced during primary immune response.
Monomeric form IgM (180000 Da) is also expressed as membrane bound receptor on B-
cell.
Biological functions:
IgM is the first antibody produced in primary immune response and it is also the first Ig
produced by neonate.
IgM has higher valency (antigen binding ste) due to its pentameric form.
Due to pentameric form, IgG is very effective in agglutination reaction.
IgM is more efficient than IgG in complement activation.
IgM plays important accessory role as secretory immunoglobulin due to J-chain.
3. IgA:
Molecular weight: 320,000 Da
H- chain type: Alpha (55000 Da)
IgA constitute 10-15% of total serum immunoglobulin.
It is the predominant Immunoglobulin n external secretions such as breast milk, saliva,
tears and mucus of bronchial, genitourinary and digestive tracts.
IgA primarily exists as monomeric form but dimeric, trimeric and some tetrameric form
are also present.
IgA in blood occurs in monomeric form whereas those in body secretion occurs in
dimeric or multimeric forms.
Dimeric form of IgA contains J-chain and secretory chain. Secretory chains helps in
transcytosis.
IgA can cross epithelial layer and enter into body secretion. The process of crossing
epithelial layer by IgA is known as transcytosis.
There are two sub-class of IgA ie. IgA1 and IgA2.
Biological functions;
IgA can cross the epithelial layer and enter into body secretion and provides local
immunity in GI tracts, respiratory tract, genital tract etc
In body secretion IgA neutralize viruses and prevent attachment on host surface.
4. IgD:
Molecular weight: 180,000 Da
H-chain type: Delta (70000 Da)
IgD is present in extremely low concentration and it constitute 0.2% of total serum
immunoglobulin.
IgD together with IgM is the major membrane bound immunoglobulin expressed on
mature B-cell.
There are two sub-classes of IgD (IgD1 and IgD2)
IgD plays important role in maturation and proliferation of B-cell.
5. IgE:
Molecular weight: 200,000 Da
H-chain type: epsilon (73,000Da)
IgE accounts for 0.3% of total serum Immunoglobulin.
IgE is also known as reagenic antibody due to its involvement in allergic reaction. IgE
mediate immediate hypersensitivity reaction and responsible for symptoms like hey fever,
asthma, anaphylactic shocks, etc.
Fc region of IgE binds on blood basophils and tissue mast cells. The cross reaction with
antigen to Fc region bound IgE causes degranulation of mast cell and basophils releasing
histamine. Histamine is responsible for symptoms of allergy.
Biological functions;
IgE provides immunity against parasite by Antibody dependent cell mediated cytotoxicity
(ADCC).
Level of IgE antibody in blood of normal individual is very low and its level increases
during parasitic infection and in allergic reactions.
For example, in humans there are 5 different types of antibodies; IgM, IgA, IgG, IgE and IgD
based on the class of heavy chain they have (µ, α, γ, ε and δ respectively). That means we have
specific set of genes which codes for these heavy chains. So there are specific sequences on
constant regions of heavy and light chain (κ and λ) which decides what class or subclass of
antibody it is.
It is called isotype (iso = same) because all members of a given species expresses all the isotypes
in the serum.
For example, all immunologically normal individuals will have all five types of antibodies
present in their serum i.e. IgM, IgA, IgG, IgE and IgD.
Therefore, different species inherits different constant regions genes and thus expresses different
isotypes.
So if take antibodies from one species and inject it in another speciesit would result in anti-
isotypic antibody production.
Location: Constant region of heavy chain and light chain.
Observed: In all the individuals of a species (immunologically normal individuals).
Importance: To measure Ig levels,
To check for immunodeficiency,
In detection of B cell tumors.
2. Allotype: Allo = Different
Definition: Allotype antigenic determinants are specified by the allelic forms of the Ig genes.
Although all the members of a species inherit the same set of Ig genes, there would be multiple
alleles present for these genes which code for different amino acids. That means the amino acid
sequence for the same antibody heavy chain would be slightly different in you than in me.
So if we take antibodies from one member of the species and inject it into another member
of the same species it will result in production of antibodies against allotypic determinants.
Location: Constant region of heavy chain and light chain
Observed: During blood transfusion
During pregnancy
Importance: Monitoring bone marrow grafts
Paternity testing
Forensic
3. Idiotype: Idio = Own or Peculiar
Definition: Idiotypes are the unique antigenic determinants present on variable heavy chain and
variable light region of individual antibody molecules.
When we encounter any Ag, the variable region of our Ab recognizes the Ag and our B cells
produce specific antibodies (during affinity maturation by somatic hypermutaion) against that
particular Ag. So when we encounter an Ag, our B cell makes very specific or peculiar Abs
against it.
Let’s say for example, I encounter two different antigens i.e. Ag a and Ag b and produce IgG1
against both. In this case the idiotypic determinants for the IgG1 against Ag a will be different
than IgG1 against Ag b.
Location: Variable region of heavy chain and light chain.
Observed: When we inject antibodies from a donor who is genetically identical to a
recipient, it induces production of anti-idiotypic antibodies. In genetically identical twins the
isotype and allotype will be the same but the idiotype will be different.
Importance: Treatment of B cell tumors
Vaccines
MHC class-I:
MHC class 1 refers to one class of major histocompatibility complex molecules
found on the surface of all nucleated cells in mammals. MHC class 1 molecule is
composed of three alpha domains (alpha 1, alpha 2, and alpha 3) and a single beta
domain. Alpha domains are encoded by the chromosome 6 while the beta domain
is encoded by chromosome 11. The alpha 3 domain serves as the membrane-
spanning domain. The alpha 1 and alpha 2 domains consist of most variable amino
acid sequences and antigens are bound to these two domains.
α-chain of MHC-I:
The α-chain is a transmembrane glycoprotein encoded by polymorphic gene within A, B and
C region of Human HLA complex
The α-chain is anchored in the plasma membrane by its hydrophobic trans-membrane
segment and hydrophilic cytoplasmic tail.
α-chain is made up of 3 domains (α1,α2 and α3). Each domain containing approximately 90
aminoacids, a transmsmbrane domain of about 25 hydrophobic aminoacids followed by short
stretch of charged (hydrophilic) aminoacids of cytoplasmic tails of 3o aminoacids.
α1 and α2 domains interacts to form a deep groove on the top which is a peptide binding
clift. It can binds antigen of 8-10 animoacids long.
α3 and β2 are organized into β-pleated sheets, each formed by antiparallel β-strand of
aminoacids, this structure is known as immunoglobulin fold. Because of this structure α-chain
and β2 microglobulin are classified as member of immunoglobulin super-family receptor.
β2 microglobulin of MHC-I:
β2 microglobulin is a protein encoded by a highly conserved gene located on different
chromosome
β2 microglobulin is similar in size and organization to α3 domain.
Β2 microglobulin does not contain transmembrane region and is non-covalently linked with
α-chain.
MHC class 1 molecules are expressed on almost every nucleated cell in the body. Hence,
they present endogenous antigens that originate from the cytoplasm. However, these
endogenous antigens can be either self-proteins or foreign protein such as viral proteins
produced within the cell. Generally, viral proteins are produced inside animal cells with the
help of cellular machinery of the host. Upon presentation on the cell membrane, antigens are
recognized by cytotoxic T cells. MHC class 1 molecules are involved in the presentation of
antigens that belong to every type of protein produced inside the cell. These antigens are
monitored by killer T cells. This identification serves as a part of the surveillance system that
destroys over-abundant or unfamiliar antigen presenting cells. Thus, malignant cells, as well
as virus-harboring cells, can be destroyed.
Functions of MHC class I:
Major function of MHC-I is to bind peptide antigens and present to CD8+ T cells (T helper
cells)
CD8 T cells are specific for MHC-I antigen
MHC-I binds endogenous antigen and present to T helper cells.
MHC-I molecules are found on surface of all nucleated cells.
MHC class-II:
MHC class 2 refers to a class of major histocompatibility complex molecules mainly found on antigen
presenting cells such as macrophages, dendritic cells, and B cells. MHC class 2 molecule is composed of
two alpha (alpha 1 and alpha 2) and two beta (beta 1 and beta 2) domains. Both alpha and beta domains
of the MHC class 1 molecules are encoded by chromosome 6. The alpha 2 and beta 2 domains serve as
the membrane-spanning domains while alpha 1 and beta 1 domains serve as antigen-presenting
domains.
α-chain and β-chain of MHC-II:
α-chain and β-chain of MHC-II is a membrane bound glycoprotein that contains external
domains, atransmembrane segment and acytoplasmic tail.
α-chain and β-chain are made up of two domains (α1 and α2) and (β1 and β2) respectively.
The peptide biding cleft is a open ended groove formed between α-chain and β-chain at
proximal end. The cleft can bind antigenic peptide of 13-18 aminoacids long.
MHC class 2 molecules are expressed on specialized antigen-presenting immune cells, including
macrophages, dendritic cells, and B cells. Macrophages are the most professional phagocytes that
engulf bacteria and virus-like foreign particles. Dendritic cells are also a type of phagocytes that
present antigens to T cells. B cells produce antibodies during humoral immunity. MHC class 2
molecules present exogenous antigens. The exogenous antigens are originated extracellularly from
bacteria-like foreign particles. The phagocyted pathogens are degraded inside the antigen
presenting cells and peptide fragments are presented on the cell membrane with the help of MHC
class 2 molecules. These antigens are recognized by helper T cells, activating them. The activated
helper T cells release lymphokines, attracting other cells that destroy the antigenic material.
Functions of MHC class II:
Major function of MHC-II is to bind peptide antigen and present to CD4 T cells.
MHC-II are found on surface of Antigen presenting cells (APCs).
CD4+T-cells are specific for MHC-II
Activates B cells for antibody production
MHC-II plays a significant role in graft versus host response and in mixed lymphocyte
reaction (MLR) because the immune response gene is identical to MHC-II in human.
MHC class-III:
MHc-III are diverse group of molecules that serves a wide variety of functions in immune
system.
MHC-III are not a marker on cell surface.
Functions of MHC class-III:
Involved in complement activation
Involved in inflammation caused by cytokines, tumor necrosis factors etc
References:
Kuby immunology Owen, Judith A; Punt, Jenni; Stranford, Sharon A; Jones, Patricia P;
Kuby, Janis.7th ed. New York : W.H. Freeman, c2013. NLM ID: 101607504 [Book]
https://www.onlinebiologynotes.com/antibody-structure-classes-functions/
https://microbiologyinfo.com/antibody-structure-classes-and-functions/