ELISA for Detection of Viruses, Fungi, Bacteria, MLOs, Mycotoxins and Hormones.
ELISA is based on the ability of low molecular weight antibodies to couple with enzymes, to produce enzymatically active immunological conjugates. This allows the detection of immune reaction with histochemical staining techniques, because the antibody component is involved in immune reaction and the conjugated enzyme can be used for staining reaction utilizing appropriate substrate. Using this principle, ELISA was initially developed in 1971 independently by two groups (Engvall and Perlmann; Weeman and Schuurs). Later, Vollar et. Al. (1976) and Clark and Adams (1977) utilized ELISA for detection of virus infection. (In India, initially Usha m. Joshi of Bombay, and later several other workers also utilized it for detection of viruses). Several modification of original ELISA are now know and are widely used for detection fo viruses, fungi, bacteria, MLOs (mycoplasma like organisms), etc.

In all the three techniques, a polystyrene or polyvinylchloride microtitre plate is used, which has wells to provide for a solid phase for the immune reaction.
(i) DAS-ELISA. In this double antibody sandwich (DAS) ELISA, the wells are coated with antibody (or immunoglobulins = Ig) prepared from antisera. The test sample is then added to allow trapping of virus antigen by the coated antibody in the well. This is followed by the addition of enzyme labelled antivirus IG, which will attach to the trapped antigen. A substrate for the enzyme is now added for producing colour reaction. Peroxidase (OD 450) or alkaline phosphatase (OD 405) is used, which allows quantitative measurement through a spectrophotometric device.