Serology: Enzyme-linked immunosorbent assay (ELISA)
Serology:
Enzyme-linked immunosorbent assay (ELISA)
Authors: Dr. RW Worthington (Retired)
Licensed under a Creative Commons Attribution license.
Antigen capture ELISAs
Antigen capture ELISAs are powerful tools for the demonstration of antigens in samples. They are sensitive, specific, and cheap and results can be obtained in a few hours and can be done with simple equipment in non-specialist laboratories. For these reasons antigen capture ELISA methods are replacing many traditional tissue culture and animal inoculation methods particularly for virus diseases. Antigen capture ELISAs is suitable for exploiting the high specificity of antibodies and particularly MAbs. Capture antibody tests are widely used, for example a test using polyclonal antibodies is used to identify foot and mouth disease virus in tissues and to differentiate between foot and mouth disease virus types. Similarly a test using MAbs is able to distinguish between the closely related rinderpest and peste des petits ruminants (PPR) viruses.
In the case of foot and mouth disease rabbit anti-sera to each of the seven types of foot and mouth disease virus are coated onto rows of wells on a plate. These act as the capture antibody and virus particles bind to them when tissue samples are dispensed into the wells. Guinea pig sera against the seven types of virus are added and finally a rabbit anti-guinea pig conjugate is used to identify the rows in which virus and guinea pig antibody have bound. After each step the plates are thoroughly washed and finally substrate is added and colour allowed developing in the usual way. The test can also be used to identify the virus type in tissue culture amplified virus.
In the case of Rinderpest and PPR a single MAb against an epitope on the N protein, which is common to both viruses, is used as a capture antibody. Virus of either disease will then bind to the plate when infected tissue samples are applied to it. BiotinylatedMAbs with specificity for either rinderpest or PPR are then used to identify the virus that has bound to the plate and an avidin enzyme conjugate is added. Plates are thoroughly washed between each step and finally substrate is added and the colour allowed developing in the normal manner.
Both the above tests could therefore be described as antigen capture, sandwich ELISAs. Many other antigen capture ELISAs are commonly used to identify virus in tissue samples.
1 | Page