Cells: Cell Recognition & the Immune System - ELISA Tests

Question 1

What is an ELISA test?

Answer 1

• The Enzyme-Linked Immunosorbet Assay (ELISA) allows you to see if a patient has any antibodies to a certain antigen and vice versa

Question 2

What can an ELISA test be used for?

Answer 2

• It can be used in medical diagnosis to test for pathogenic infections, allergies etc. (anything you can make an antibody for)

Question 3

How do ELISA tests work?

Answer 3

• In an ELISA test, an antibody is used which has an enzyme attached to it
• This enzyme can react with a substrate to produce a coloured product
• This causes the solution in the reaction vessel to change colour
• If there’s a colour change, it shows the antibody/antigen of interest is present
• In some types of ELISA, the quantity of this antigen/antibody can be calculated from the intensity of the colour change

Question 4

How does a direct ELISA work?

Answer 4

• A direct ELISA uses a single antibody that is 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 is 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 well and the detection antibody will bind to it
• The well is washed 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
• This is a positive result for the presence of the antigen

Question 5

How does an indirect ELISA work?

Answer 5

• Indirect ELISA uses two different antibodies
• An indirect ELISA test can be used to see if a patient has the antibodies to HIV
• HIV antigen is bound to the bottom of a well plate
• Sample of the patient’s blood plasma -which might contain different antibodies- is added to the well
• If there are any HIV-specific antibodies in the plasma these will bind to the HIV antigen stuck to the bottom of the well - the well is then washed out to remove any unbound antibodies
• A secondary antibody, that has a specific enzyme attached, is added to the well
• This secondary antibody can bind to the HIV-specific antibody (aka the primary antibody)
• The well is washed again to remove any secondary antibody - if there’s no primary antibody, all of the secondary antibody will be washed away because there will be nothing to bind to
• A solution is added to well
• Solution contains substrate - can react with enzyme attached to secondary antibody to make a coloured product
• If soluion changes colour, it shows the patients has HIV-specific antibodies in their blood and is infected with HIV