ELISA is a technique used to detect antibodies or antigens in a sample. It works by first adding antigens to wells which antibodies then bind to. Secondary antibodies with an enzyme are then added, and if the antigens and primary antibodies are present, the enzyme causes a color change reaction. This process allows ELISA to be used to test for conditions like HIV in patients by simulating the immune response in a lab setting.

1. ELISA Workshop
By Ricardo Chiesa
ELISA is a technique used in immunology for the detection of an antibody or
antigen in a sample. An antigen is a molecule or substance that enters the body and is
recognized as a foreign material. As a result, it promotes the production of an antibody that
will bind to the antigen and neutralize it, avoiding any type of infection for the organism.
This process is known as immunization. In the lab, ELISA was done to simulate the test
and diagnose of HIV in patients. First, the antigens were added to the cells of a microplate
strip in order for them bind to the surface. In this case, chicken gamma-globulin (purified
from egg yolks) was used as an antigen representative. After the wells were washed with
detergent, the primary antibodies (polyclonal rabbit antibodies) were added and they were
supposed to bind to the antigens, if there were any. Then, the secondary antibodies are
added, and they were supposed to bind to the primary antibodies. These secondary
antibodies were obtained from goats by injecting rabbit’s antibodies into them. As a result,
rabbit’s antibodies were recognized by the goat’s immune system as antigens, so antibodies
were produced. Finally, the detection of secondary antibodies involves an enzyme-
substrate reaction. These antibodies contained an HRP enzyme bound to them that in the
presence of nitrogen peroxide will catalyze the oxidation of a substrate known as TMB, and
will produce a blue color. In the experiment, the substrate was added to the wells and
oxidation occurred in some of the wells.