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 B-lymphocytes) 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 B-lymphocytes 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.