1. Submitted to :
Dr. Gautam Kumar
Assistant Professor
Centre of Biological Science
Central University Bihar
Presented by :
P.K. Aditya
M.Sc.Life science
Roll no. - 09

2. Some terms
Antibodies :-It is also called immunoglobulins, large Y-
shaped proteins, a specialized immune protein, produced
because of the introduction of an antigen into the body, and
which possesses the remarkable ability to combine with the
very antigen that triggered its production
Antigens :- An antigen is any substance that causes your
immune system to produce antibodies against it. An
antigen may be a foreign substance from the environment,
such as chemicals, bacteria, viruses, or pollen. An antigen
may also be formed inside the body, as with bacterial
toxins or tissue cells.
Plasma cell :- is the pale yellow liquid component
of blood that normally holds the blood cells in whole
blood in suspension, this makes plasma the extracellular
matrix of blood cells. It makes up about 55% of the body's

3. Immunoglobulin structure

4. Monoclonal antibodies (mAb) are antibodies
that are identical because they were
produced by one type of immune cell, all
clones of a single parent cell.
Polyclonal antibodies are antibodies that
are derived from different cell lines. They
differ in amino acid sequence.

5. History of mAb
development
• 1964 Littlefield developed a way to isolate
hybrid cells from 2 parent cell lines using
the hypoxanthine-aminopterin-thymidine
(HAT) selection media.
• 1975 Kohler and Milstein provided the
most outstanding proof of the clonal
selection theory by fusion of normal and
malignant cells
• 1990 Milstein produced the first
monoclonal antibodies

6. Discovery of Monoclonal
Antibodies
• Monoclonal
antibodies were
produced in mice
using a technique
described by Köhler
and Milstein et al..
They were awarded
the Nobel Prize in
1984 (along with
Jerne) fortheir
work.

7. Nobel prize in Medicine and Physiology was
awarded to Köhler, Milstein and Jerne in
1984

8. Characters of Monoclonal
Antibodies
• Monoclonal antibodies (mAb) are a single
type of antibody that are identical and are
directed against a specific epitope
(antigen, antigenic determinant) and are
produced by B-cell clones of a single
parent or a single hybridoma cell line. A
hybridoma cell line is formed by the fusion
of a one B-cell lymphocyte with a
myeloma cell. Some myeloma cells
synthesize single mAb antibodies naturally

9. Principles in Hybridoma
technology
• Inject the protein into a
mouse.
• - remove the spleen.
• - identify which spleen
cells are producing
antibodies.
• - separate these cells and
grow in tissue culture
tubes.
• - screen each Ab forcross
reactivity.
• - select the Ab which
doesn't cross react with
any otherprotein.

10. Strategy to generate
monoclonal antibodies

11. HAT selection is used to select for growth of hybrids and against the growth of
the parental myeloma.

12. • HAT medium (Hypoxanthine Aminopetrin
Thymidine) is used for preparation of
monoclonal antibodies,because it contains
hypoxanthine, aminopterin, and thymidine. This
medium is selective for fused (hybridoma) cells.
Unfused myeloma cells and unfused spleen
cells cannot grow because they lack HGPRT
( hypoxanthine-guanine-phosphoribosyl
transferase), and thus cannot replicate their
DNA and because of their limited life span
respectively.

13. Types of Monoclonal Antibodies

14. Murine antibody
• Whole of the antibody is of
murine origin
• Major problems associated
with murine antibodies include
 reduced stimulation of
cytotoxicity
 Formation of complexes after
repeated administration
 allergic reactions
 anaphylactic shock

15. Chimeric antibodies
 Chimeric antibodies are
composed of murine
variable regions fused onto
human constant regions.
 Antibodies are
approximately 65% human.
 This reduces
immunogenicity and thus
increases serum half-life.

16. Humanised Mab
• Humanised antibodies
are produced by grafting
murine hypervariable
amino acid domains into
human antibodies.
• This results in a molecule
of approximately 95%
human origin

17. Applications of Monoclonal Antibodies
 Diagnostic Applications
- Detects protein of interest either
by blotting or
immunofluorescence
 Therapeutic Applications
Transplant rejection
Cancer
Autoimmune disorders
Inflammatory disease

18. • A monoclonal
antibody can be used
to detect pregnancy
only 14 days after
conception. Other
monoclonal antibodies
allow rapid diagnosis
of hepatitis, influenza,
herpes, streptococcal,
and Chlamydia
infections.
• Monoclonal antibodies
can also be used to
purify a substance with
techniques called
immunoprecipitation
and affinity
chromatography.

19. Advantages :
• 1) Homogeneity:
• Monoclonal antibody represents a single antibody
molecule that binds to antigens with the same
affinity and promote the same effectors functions.
• 2) Specificity:
• The product of a single hybridoma reacts with the
same epitope on antigens.
• 3) Selection:
• It is possible to select for specific epitope
specificities and generate antibodies against a
wider range of antigenic determinants.
• 4) Antibody Production:
• Unlimited quantities of a single well-defined

20. Disadvantages:
• 1) Affinity:
• Average affinity of monoclonal antibodies are generally
lower than polyclonal antibodies.
• 2) EffectorFunctions:
• Because antibody is monoclonal, it may not produce the
desired biologic response.
• 3) Specificity:
• Monoclonals against conformational epitopes on native
proteins may lose reactivity with antigens.
• 4) Cross reactions:
• Antibodies sometimes display unexpected
crossreactions with unrelated antigens.
• 5) Time and effort commitment:

24. Monoclonal antibodies for cancer treatment
There are three mechanisms that could be responsible for the
cancer treatment:
1. mAbs act directely when binding to a cancer specific
antigens and induce immunological response to cancer cells.
Such as inducing cancer cell apoptosis, inhibiting growth, or
interfering with a key function.
2.mAbs may be modified for delivery of a toxin, radioisotope
[RADIOIMMUNOTHERAPY], cytokine or other active
conjugates.
3.it is also possible to design bispecific antibodies that can
bind with their Fab regions both to target antigen and to a
conjugate or effector cell.

26. When a monoclonal antibody attaches to a cancer
cell, it can:
• Make the cancer cell more
visible to the immune
system.
ex. Rituximab
• Block growth signals ex.
Cetuximab used for colon
cancer and block epidermal
growth factors.
• Stop new blood vessels formation
(angiogenesis) Ex. Devacizumab (Avastin)
acts on vascular endothelial growth factor
(VEGF).
• Deliver radiation to cancer cells by binding to
radioactive element ex. Zevalin used for
non-hodgkin lymphoma.