Lab 7. Enzyme-Linked Immunosorbent
Assay (ELISA)
Goals for this lab:
1. Understand how antigen-antibody interactions are utilized in ELISA.
2. Compare indirect and sandwich ELISA.
3. Describe the purpose of the controls and reagents in the ELISA protocol.
4. Apply the ELISA method by performing an anti-nuclear antibody (ANA) test.
5. Learn how to use a standard curve generated from samples with known concentrations to
determine unknown concentrations of anti-nuclear antibody.
The enzyme-linked immunosorbent assay (ELISA) technique is an extremely sensitive method for
detecting antigens or antibodies, and can detect an antigen or antibody at a concentration as low as 1
ng/ml. The ELISA is commonly used to quantify antigens or antibodies, but is not good for detecting
the presence of antigens or antibodies. The ELISA procedure has been widely applied for medical
diagnosis and research. For example, ELISA can be used to determine the serum concentration of
antibodies specific for Salmonella typhi for diagnosis of the infection, determine the serum
concentration of the thyroid hormone for diagnosis of Graves disease (an autoimmune disease
causing hyperthyroidism), and determine the serum antibody titers of immunized animals or humans
to evaluate the efficacy of a vaccine.
Antibodies conjugated to enzymes are used in ELISA. Horse radish peroxidase (HRP) and alkaline
phosphatase (AP) are two enzymes that are commonly conjugated to antibodies. These types of
antibody conjugates also can be used for Western blot and immunohistochemistry