3. Immunochemistry
•Immunochemistry is the study of the chemistry of
the immune system.
•The immune system is a network of biological
processes that protect an organism from diseases.
•This involves the study of the properties, functions,
interactions and production of the chemical
components (antibodies/immunoglobulins, toxin,
Antigen etc.)
APPLIED BIOCHEMISTRY
INTRODUCTION
5. Introduction
• Antigen and antibodies are important components of the
immune system when a foreign antigen enters the body.
• Immunity is a complex biological system, it refers to the
body's ability to prevent the invasion of pathogens. Pathogens
are foreign disease-causing substances, such as bacteria and
viruses, and people are exposed to them every day.
APPLIED BIOCHEMISTRY
INTRODUCTION
6. TYPES OF IMMUNITY
•Immunity is broadly divided into two types:
1.Innate or natural or (nonspecific) immunity
2. Adaptive or acquired or (specific) immunity
APPLIED BIOCHEMISTRY
INTRODUCTION
8. • Innate or natural immunity is the inborn immunity that is
already present at the time of birth. It is genetic in origin and
also depends upon the constitutional makeup of any
individual.
• It is independent of any previous exposure to foreign
substances.
• It includes barriers like skin, mucous membrane, earwax etc.
APPLIED BIOCHEMISTRY
INTRODUCTION
Innate immunity- Types
9. Innate immunity- Types
• Species Immunity - Some diseases occurs only in certain specific
species such as pathogen of plants can nit cause diseases in humans.
• Racial Immunity - Different races within a particular species may
show certain differences in the immunity. For example, negroes are
more resistant to malaria caused by plasmodium falciparum due to in
structural difference of their RBCs.
• Individual Immunity - Different individuals in a species or a race
shows resistance to diseases. Factors affecting individuals immunity
are: age, diet, hygiene, individuals sex etc.
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INTRODUCTION
10. • Innate immunity is nonspecific and represents the inherent capability to fight against
diseases. This defense system is of two types:
APPLIED BIOCHEMISTRY
INTRODUCTION
First line of defence
• Skin is the largest organ and provides mechanical barren to prevent the entrance of
microorganisms and viruses.
The sweat contains an enzyme lysozyme that destroys bacterial all well.
Second line of defence
• Despite, skin is the first line of barrier, still microorganism enter our body and the body
protect itself by the nonspecific mechanisms such as sneezing, secreation of mucus.
11. • Immunity that is not present by birth in an individual but its developed with
age after exposure to an pathogen.
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INTRODUCTION
Acquired immunity
12. IMMUNOGLOBULINS
APPLIED BIOCHEMISTRY
INTRODUCTION
• Immunoglobulins are also called
antibodies.
• It is a large, Y-shaped protein used by the
immune system to identify and neutralize
foreign objects such as pathogenic
bacteria and viruses.
• Antibodies are proteins that your immune
system makes to fight germs, such as
viruses and bacteria. When you're
exposed to germs, your body makes
unique antibodies that are specifically
designed to destroy only those germs
13. STRUCTURE OF IMMUNOGLOBULINS
APPLIED BIOCHEMISTRY
INTRODUCTION
Immunoglobulins have four polypeptide
chains
(2 heavy chains and 2 light chains) linked
with each other by disulfide bonds.
Heavy chains are designated as gamma
alpha(α), mu (µ), delta (γ), and epsilon
(ε).
Light chains are designated as Kappa
(K) and Lambda (λ)
15. STRUCTURE OF IMMUNOGLOBULINS
APPLIED BIOCHEMISTRY
INTRODUCTION
Variable region:
It is present in both the
heavy and the light
chains.
Constant region:
For a single type of heavy
chain or light chain, the constant
region has the same amino acid
sequences.
Hinge region:
It provides flexibility and
facilitates binding of
antibody with antigens.
FC region:
It comprises only of heavy chains.
It interacts with cell surface
receptors called FC receptors.
Fab region: It contains antigen binding site
and is composed jf the variable region of
both heavy and light chains.
18. TYPES OF IMMUNOGLOBULINS- IgG
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INTRODUCTION
• 75% of total
• Found in blood, lymph, and intestines
• Active against bacteria, its toxins and viruses
• enhances phagocytosis, crosses placenta
• Defends the body fluids
• Major antibodies found in blood
• Function: Protection of newborn as means of naturally immunity
• Protection against many bacteria and virus
19. TYPES OF IMMUNOGLOBULINS- IgA
APPLIED BIOCHEMISTRY
INTRODUCTION
• 10-15% of total
• Found in Saliva, tears, bronchial, Gl and vaginal secretions
• Provides local protection on exposed mucous membrane surfaces and
potent antiviral activity
• Prevents absorption of antigens from food, and protects against
respiratory, Gl,
• Function: neutralizes allergens
• Protection against virus
20. TYPES OF IMMUNOGLOBULINS- IgM
APPLIED BIOCHEMISTRY
INTRODUCTION
• 5-10% of total
• Levels decrease during stress
• Found in blood and lymph First antibody produced with primary
immune response
• High concentrations early in infection, decrease within about week
• Function: defends the blood stream
• Serum helps in diagnosing infection
21. TYPES OF IMMUNOGLOBULINS- IgD
APPLIED BIOCHEMISTRY
INTRODUCTION
• Less then 1% of total
• Unknown function, found in blood and lymph
• Function- activates the beta cells during fetal growth and normal
adult.
22. TYPES OF IMMUNOGLOBULINS- IgE
APPLIED BIOCHEMISTRY
INTRODUCTION
• Less then 0.1% of total
• Found on mastcells and basophils
• Involved in immediate hypersensitivity response
• Provide immunity against helminthic infection
• Cannot cross the placenta
24. ANTIGEN-ANTIBODY REACTION
• The interaction between antigen and antibodies is reversible
• The pressure of specific antibody can be detected using many
different assays.
• Some assays measure direct binding of the antibody to its antigen and
are termed primary interactions.
• For example, radioimmunoassay (RIA) and enzyme linked
immunosorbent (ELISA).
• Secondary interaction includes the determination of amount of
antibody present by the changes which induces in the physical state
of the antigen, in precipitation or the clumping of antigen.
APPLIED BIOCHEMISTRY
INTRODUCTION
25. Enzyme linked immunosorbent Assay(ELISA)
• The ELISA is an antigen-antibody reaction
• That is done on a specific 96 wells plate (of polystyrene) for analyzing
the presence of antigens, antibodies or hormones in a patient’s
serum.
• This analysis is done by making use of the adhesive nature of the
surface of the ELISA plate wells that binds and fixes certain molecules
and the unbound are washed away in the process.
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INTRODUCTION
26. Enzyme linked immunosorbent Assay(ELISA)
• ELISA enzymes and the substrates are described in Table
APPLIED BIOCHEMISTRY
INTRODUCTION
27. Process of ELISA
• Coating: The target antigen/antibody containing sample is coated on
to the wells of the ELISA plate that leads to the attachment of the
Ag/Abs on the adhesive wells, if present.
• 2.Blocking: The unattached sites are blocked by binding with an inert
blocking agent.
• 3.Washings: The blocking is followed by a series of washing steps so
that any unbound material is removed out of the wells.
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INTRODUCTION
28. Process of ELISA
• .Addition of the enzyme-conjugated molecules: The next step is the
addition of an enzyme-conjugated molecule (Ab/Ag) that has the
capacity to attach to the bound Ag/Ab molecules respectively, on the
wells surfaces of the ELISA plate.
• 5.Incubation: The whole material is incubated to ensure the
combination of the bound molecules with their enzyme-conjugated
counter parts.
• 6.Washings: A series of washing is done toagain ensure the removal of
any unbound molecules from the inside of the wells.
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INTRODUCTION
29. Process of ELISA
• 7.Addition of the substrate: In this step, the enzyme-specific substrate
molecules are added into the wells that are acted upon bythe bound
enzymes.
• 8.Observation
• 9.Interpretation
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INTRODUCTION
32. METHODS
APPLIED BIOCHEMISTRY
INTRODUCTION
Also known as inhibition ELISA or competitive immunoassay, competitive ELISA assays measure the concentration
of an antigen by detection of signal interference. Each of the previous formats can be adapted to the competitive
format.
33. METHODS
APPLIED BIOCHEMISTRY
INTRODUCTION
It requires two antibodies specific for different epitopes of the antigen. These two antibodies are
normally referred to as matched antibody pairs. One of the antibodies is coated on the surface of
the multi-well plate and used as a capture antibody to facilitate the immobilization of the
antigen. The other antibody is conjugated and facilitates the detection of the antigen.
