ELISA Test for Antigen/Antibody Detection

Enzyme-Linked Immunosorbent Assays (ELISAs) are the most widely used type of assay.

ELISA (Enzyme Linked Immunosorbent Assay) is no named because the test technique involves the use of an enzyme system and immunosorbent.

ELISA method for measuring Antigen Antibody reaction is becoming increasingly used in the detection of antigen (infectious agent) or antibody for its simplicity and sensitivity. It is as sensitive as radioimmunoassay and requires only microlitre quantities of test reagents. It has now been widely applied in detection of a variety of antibody and antigens such as hormones, toxins, and viruses.
Some Salient Features

1. They have high sensitivity and specificity.
2. The result of quantitative ELISA tests can be read visually
3. A large number of tests can be done at one time.
   ELISAs are designed specifically for screening large numbers of specimens at a time, making them suitable for use in surveillance and centralized blood transfusion services.
4. Reagents used for ELISA are stable and can be distributed in district and rural laboratories but as ELISAs require sophisticated equipment and skilled technicians to perform the tests, their use is limited to certain circumstances.

Principle
Most ELISA methods developed for the detection of antigen or antibody consist of use of corresponding antibody or antigen in question which is firmly fixed on solid phase, such as plastic surface of polyvinyl plate or polystyrene tube, inside the well of a microdilution tray or outside of spherical plastic or metal bead. Such systems are also called Solid Phase Immunosorbent Assay.

The enzyme system consists of;

1. An enzyme:  horse radish peroxidase, alkaline phosphatase which is labelled or linked, to a specific antibody.
2.  A specific substrate:

• O-Phenyl-diamine-dihydrochloride for peroxidase
• P Nitrophenyl Phosphate- for Alkaline Phosphatase

Which is added after the antigen-antibody reaction. The enzyme catalyses (usually hydrolyses) the substrate to give a colour end point (yellow compound in case of Alkaline Phosphatase). The intensity of the colour gives an indication of the amount of bound antibody or antigen.

Most Commonly used ELISA are
a. ELISA for Antigen Detection

1. Direct ELISA
2. Indirect ELISA

b. ELISA For Antibody Detection

1. Non Competitive ELISA
2. Competitive ELISA

c. Capture ELISA