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The most suitable primary screening assay for soluble antigen is the enzyme-linked immuno-absorbant assay (ELISA). Soluble antigen is bound to the wells of a 96-well microtitre plate. Tissue culture supernatants are then incubated with this bound antigen and any antibodies present that are antigen specific will bind to the well in the plate. An enzyme linked secondary antibody with specificity for the monoclonal immunoglobulins is used to detect wells where the primary antibody has bound. This binding is then visualised by the addition of an appropriate enzyme substrate that, after reaction with the enzyme, yields a coloured product.

Schematic diagram of a completed ELISA.
The purple wells represent antigen/antibody complexes formed by the binding of specific antibodies to antigen present in the well, C+ marks the position of the positive control well.


Preparation of ELISA plate
Performing the ELISA