ELISA (AQA AS Biology): Revision Note

ELISA

• ELISA (Enzyme-Linked Immunosorbent Assay) uses antibodies to detect the presence and quantity of a specific antigen or antibody in a sample

• In an ELISA test:

  • An enzyme is attached to antibodies

  • When this enzyme reacts with a certain substrate, a coloured product is formed, causing the solution in the reaction vessel to change colour

  • If a colour change occurs, this shows that the antigen or antibody of interest is present in the sample being tested (e.g. blood plasma)

• There are different types of ELISA test:

  • Direct ELISA tests use a single antibody that is complementary to the antigen being tested for

  • Indirect ELISA tests use two different antibodies (known as primary and secondary antibodies)

Example: indirect ELISA for HIV diagnosis

• An indirect ELISA test can be used to test whether a patient has antibodies to HIV:

  1. HIV antigens are fixed to the bottom of the test well

  2. Patient's blood plasma is added – if HIV-specific antibodies are present, they bind (primary antibodies) to the HIV antigens

  3. The well is washed to remove unbound antibodies

  4. Secondary antibodies with an enzyme attached are added – they bind to the primary antibodies

  5. Another wash step removes unbound secondary antibodies

     • This step avoids a false-positive test

  6. A substrate is added – if the enzyme is present, a colour change occurs

  7. A colour change indicates the patient has HIV-specific antibodies, meaning they are infected with HIV