Indirect ELISA

Enzyme-linked immunosorbent assay, commonly known as ELISA (or EIA), is similar in principle to RIA but depends on an enzyme rather than a radioactive label. An enzyme conjugated with an antibody reacts with a colorless substrate to generate a colored reaction product. Such a substrate is called a chromogenic substrate.

INDIRECT ELISA

Here, in Indirect ELISA steps are slightly different, though the basic principle remains the same.

Two different antibodies are used.

1. Primary antibody, let's consider this as Ab 1.
2. And a Secondary antibody, Ab 2.

A serum sample or any other sample containing the primary antibody is added to a microtitre plate containing the antigens.

Reaction is allowed between them. and any free Ab 1 is washed off.

Then a secondary antibody(Anti-isotype antibody) linked to an enzyme, specific to the primary antibody is added

Reaction is allowed between them. and any free Ab 2 is washed off.

A substrate specific to the enzyme is added and a chromogenic reactions occurs between the two.

This can be measured by spectrophotometery.

Uses: Generally, serum antibodies to HIV can be detected by indirect ELISA within 6 weeks of infection. 

That's all

Hope it helps

Lots of Love and be Amazing!!

-Staph