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A sensitive enzyme-linked immunosorbent assay for IgA protease activity

, : A sensitive enzyme-linked immunosorbent assay for IgA protease activity. Journal of Immunological Methods 63(3): 367-376

IgA proteases are enzymes of bacterial origin capable of cleaving human IgA1 into Fab alpha and Fc alpha fragments. This article describes a solid phase assay employing microamounts of protease as well as substrate for the quantitation of IgA protease activity. IgA substrate (IgA paraprotein, colostrum S-IgA, or simply diluted saliva) is bound to the surface of a polystyrene microtitration plate coated with anti-light chain antibody in order to assure binding of substrate molecules through Fab alpha. Incubation of such bound substrate with IgA protease, either prepared or as protease-producing whole bacteria, results in release of Fc alpha whereas Fab alpha is still retained after wash. Loss of Fc alpha is detected through a reduced capacity for binding of peroxidase-conjugated anti-alpha-chain antibody, the binding of which is detected using standard ELISA techniques. Simplicity and extreme sensitivity make this assay useful for quantitation of IgA protease activity, for kinetic studies of the enzyme, and for detection of IgA protease activity in single agar plate colonies of bacteria.


PMID: 6355301

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