Lesson 48 - Monoclonal antibodies and ELISA tests

Question 1

What are 3 industrial/medical uses of monoclonal antibodies?

Answer 1

Targeting medication to specific cell types
Medical testing
Medical diagnosis using ELISA

Question 2

Describe the use of monoclonal antibodies in targeting medication to specific cell types by attaching a therapeutic drug to an antibody?

Answer 2

Direct monoclonal antibody therapy:
- Monoclonal antibodies are produced specific to the antigens on the target cells (e.g. cancerous cells)
- The antibodies are given to a patient
- The antibodies attach to the antigens on the surface of the target cells & prevent their uncontrolled growth (by blocking signalling pathways in them)

Indirect monoclonal antibody therapy:
- A cytotoxic drug (drug that kills cells) is attached to the monoclonal antibodies
- The antibodies are given to the patient
- When the antibodies attach to the antigens on the target cells it kills the cells

Question 3

How are monoclonal antibodies used in medical tests, e.g. pregnancy test

Answer 3

• Pregnant women contain the hormone hCG in their urine
• The application area contains antibodies that are complementary to the hCG protein, bound to a coloured bead (blue)
• when urine is applied to the application area, any hCG will bind to the antibody on the beads, forming an antigen-antibody complex.
• The urine moves up the stick to the test strip, carrying any beads with it.
• The test strip contains (immobilised) antibodies to hCG
• If there’s hCG present, the test strip turns blue because the immobilised antibody binds to any hCG - concentrating the hCG-antibody complex with the blue beads attached.
• If no hCG is present, the beads will pass through the test area without binding to anything, so it won’t go blue

Question 4

How are monoclonal antibodies produced?

Answer 4

1. A mouse is injected with an antigen.
2. This triggers an immune response in the mouse and its plasma cells will begin producing antibodies specific to the injected antigen.
3. These cells are collected from the mouse spleen.
4. These plasma cells don’t divide easily so are fused with tumour cells that divide rapidly. These new cells are called hybridomas.
5. Hybridomas divide rapidly and create lots of cells that all create the same antibody. These antibodies can be collected, purified and used.

Question 5

How are monoclonal antibodies unethical?

Answer 5

• Mice used to produce the monoclonal antibodies & the tumour cells - deliberately inducing cancer in mice
• Testing the safety of new drugs in trials can be dangerous - in 2006 6 healthy volunteers took part in a trial for a new monoclonal antibody - within minutes they had multiple organ failures. All survived but raises concerns.
• Although monoclonal antibodies have successfully treated a number of diseases there have also been some deaths to do with their use (e.g. in multiple sclerosis - affects the brain)
• It’s important people are clear on the risks as well as the benefits before agreeing

Question 6

What are ELISA tests used for?

Answer 6

They allow you to see if a patient has any antibodies to a certain antigen, or any antigens to a certain antibody. E.g. for pathogenic infections or allergies.

Question 7

How do direct ELISA tests work?

Answer 7

• Uses a single antibody that’s complementary to the antigen you’re testing for
• Antigens (from a patient sample) are bound to the inside of a well in a well plate.
• A detection antibody (with an attached enzyme), that’s complementary to the antigen of interest, is added.
• If the antigen of interest is present in the patient sample, it will be immobilised on the inside surface of the well and the detection antibody will bind to it
• Well is washed out to remove any unbound antibody and a substrate solution is added.
• If the detection antibody is present, the enzyme reacts with the substrate to give a colour change. Positive result for the presence of the antigen.

Question 8

How do indirect ELISA tests work?

Answer 8

• It uses 2 different antibodies, and can be used to see if a patient possesses antibodies to HIV.
• HIV antigen is bound to the bottom of a well in a well plate
• A patient’s blood plasma is added to the well. Any HIV-specific antibodies in the plasma will bind to the antigen stuck to the well. Well is then washed out to remove any unbound antibodies.
• A 2nd antibody, with a specific enzyme attached, is added to the well. This secondary antibody can bind to the HIV specific antibody (primary antibody). Well is washed out again. If there’s no primary antibody in the well, all the secondary antibody will be washed away, due to there being nothing for it to bind to.
• A solution is added to the well, which contains a substrate which is able to react with the enzymes attached to the secondary antibody and produces a coloured product.
• If the solution changes colour, the patient is infected with HIV.