ELISA test

  1. Carried out using a plastic plate with lots of wells

  2. apply sample (antigen) to well

  3. material at bottom of the well is designed so antigen molecules stick

  4. the well is then washed many times to remove antigen molecules that didn’t stick

  5. antibody specific to antigen is added

  6. wait for antibodies to bind to antigens

  7. we wash again to remove excess antibody molecules

  8. Second antibody with an enzyme attached to it is then added

  9. wait for the second antibody to bind to the first antibody

  10. wash again to remove any unbound second antibody

  11. we add a substrate to the enzyme which is usually colourless

  12. the enzyme converts the substrate into a product which is coloured

  13. normally the ELISA is read after a fixed amount of time

  14. the intensity of the colour produced depends on the number of enzyme molecules which depends on the number of antigens present

  15. the intensity of the colour can then be used to determine the amount of antigen

this makes the ELISA a quantitative test

-When doing an ELISA we use a number of different dilutions to ensure all the antigens stick to the plate

-if the sample contains too much antigen the well may be overloaded and some of the antigen might not stick which would mean we wouldn’t get an accurate value for the quantity of antigen

very effective for testing for infections such as HIV