Deck 28: Enzyme-Linked Immunoabsorbent Assay Elisa Part Fivepublic Health Introduction to Public Health

Question 1

What is the purpose of the chromogen?

Answer 1

The chromogen is a chemical compound produces the colored product, when it get oxidized. In ELISA (Enzyme Linked Immunosorbent Assay), the addition of enzyme labelled antibodies ensures the presence of either test antigen or test antibodies (direct or indirect ELISA), through the final addition of chromogen. The enzyme peroxidase is bounded with the enzyme labelled antibodies, and the Tetramethylbenzidine is used as a chromogen. If, the enzyme labelled antibodies do not washed away means, the enzyme present in the antibodies oxidizes the chromogen and produces color. Moreover, if, the enzyme labelled antibodies are washed away means, the oxidation do not occurs and ends with no color.
Therefore, the purpose of the chromogen in the ELISA is, to produce the color, and confirms the positive reaction.

Question 2

What is attached to the microtiter well in an indirect ELISA?

Answer 2

The indirect ELISA (Enzyme Linked Immunosorbent Assay), refers the detection of antibodies in the test sera. In indirect ELISA, the antigen coated micro-titer plate is used to detect the antibodies. By principle, for the positive reaction, the antigen present in the micro-titer plate got binds to the antibodies present in the test sera. Those, primary antibodies are detected through the enzyme labelled antibodies; and finally the chromogen confirms the positive reaction by the production of color.
Note: In Indirect ELISA, there is no sandwich of antibodies occurs; because, the micro-titer plate is coated with antigen, and the test serum is added to identify the presence of respective antibodies.
Hence, it is clearly understood that, the micro-titer plate in the indirect ELISA is, coated with Antigen.

Question 3

What is the color of a positive ELISA test?

Answer 3

Generally, the Tetramethylbenzidine is used as a chromogen in ELISA (Enzyme Linked Immunosorbent Assay); the oxidation of Tetramethylbenzidine produces the yellow color. During the ELISA, the oxidation of the Tetramethylbenzidine is catalyzed by the peroxidase enzyme present in the enzyme labelled antibodies. If, the enzyme labelled antibodies does not washed away during the ELISA (both direct and indirect) means, the addition of tetramethylbenzidine gives the yellow colored product and confirms the positive reaction.
Therefore, positive ELISA test gives the
.