Download Monoclonal Antibodies

Monoclonal Antibodies
• Antibodies have important uses beyond
fighting infections in the body.
• Production of long-lasting monoclonal
antibodies is a recent invention and it is used
in both medicine and research.
• Monoclonal Antibody: a stable antibody
which can be used over a period of time
Producing Monoclonal Antibodies
1. Inject a mouse with a specific antigen to
stimulate its immune system to produce
necessary antibodies.
2. Extract mouse spleen cells (containing Blymphocytes) and culture them in the lab.
3. Extract mouse tumour cells, which grow
continuously, and culture them in the lab.
4. Mix spleen cells and tumour cells on the
same plate and culture.
Producing Monoclonal Antibodies
5. Add polyethylene glycol – this causes some Blymphocytes to fuse with tumour cells to
produce a hybrid cell called a hybridoma.
6. Grow the cells under conditions that allow only
hybridoma cells to survive.
7. Extract the cells, culture them separately and
test the medium around each cell for the
specific antibody of interest.
8. Culture the cells making the desired antibody
and use as needed.
http://youtu.be/7ymKofaHCoY
Pregnancy Tests
• A pregnant woman has the hormone human
chorionic gonadotrophin (HCG) in her urine.
• Monoclonal antibodies to HCG have been
produced. These have been attached to
enzymes which can later interact with a dye
molecule and produce a colour change.
Pregnancy Tests
• Pregnancy test contains three regions – reaction
region, test region and control region
– Reaction region: contains monoclonal anti-HCG
antibodies linked to enzyme
– Test region: contains polyclonal anti-HCG antibodies,
which will bind to HCG molecules bound to
monoclonal anti-HCG antibodies. Also contains dye
molecules which will be activated if monoclonal
antibodies bind to polyclonal anti-HCG antibodies
– Control region: contains anti-mouse antibodies and
dye molecules which will be activated if monoclonal
antibodies bind here
Pregnancy Tests
• When pregnant woman’s urine travels up the
pregnancy test, HCG will bind to monoclonal
antibodies in reaction region
• Movement of the urine will move the monoclonal
antibodies up to the test region
• Monoclonal antibodies with bound HCG will bind
to antibodies in test region and activate dye
molecules, producing a colour change
• Any unbound monoclonal antibodies will
continue to travel to control region and will bind
to anti-mouse antibodies, activating dye
molecules and producing a colour change
Pregnancy Tests
Diagnosis of HIV Infection
• The test of HIV
infection is based on
detecting the
presence of HIV
antibody in the
patient’s blood
serum.
a) HIV antigen is attached to the plate.
b) Patients serum passed over the plate. Any HIV
antibody in the patients serum will attached to
the antigen already on the plate.
c) A second antibody which is specific to the HIV
antibody is passed over the plate. This
antibody will attach to the concentrated HIV
antibody on the plate. This second antibody
has an enzyme attached to its structure.
d) Chromagen dye is passed over the complex of
concentrated HIV antibody/conjugated
antibody.
e) The enzyme will turn the chromagen to a more
intense colour. The more intense the colour,
the greater the HIV antibody level. This would
be the a positive result for a HIV test.
Treatment of Cancer
• Cancer cells carry specific tumour-associated
antigens (TAA) on their plasma membrane.
• Monoclonal anti-TAA antibodies have been
produced.
• Drugs which kill tumour cells or inhibit key
proteins in tumour cells are attached to
monoclonal anti-TAA antibodies.
• Cancer cells are specifically targeted, avoiding
damage to healthy host cells.
Problems…
• Many patients develop immune response to
monoclonal antibodies produced in mice, as
these are foreign proteins.
• Genetically engineered antibodies are being
perfected to avoid triggering immune
response.