HOMOLOGIES BETWEEN GRAIN STORAGE PROTEINS OF DIFFERENT CEREAL SPECIES .2. EFFECTS OF ASSAY FORMAT AND GRAIN EXTRACTANT ON ANTIBODY CROSS-REACTIVITY

A variety of monoclonal antibodies with specificities for different wheat grain storage proteins has been shown to exhibit a number of differing cross-reaction patterns with grain storage proteins from related cereal species (Skerritt and Lew, J. Cereal Sci. 11 (1990) 103-121). This antibody library provides an opportunity to measure the effects of a range of immunoassay methodological variables on the cross-reactivity of these antibodies. Variation of the extractant used for grain proteins from different cereals altered both the amount and composition of protein extracted and thus the cross-reaction of the antibodies. Antibody cross-reactivity was greater when nitrocellulose membrane solid phases compared with polystyrene microwells were used for indirect enzyme linked immunosorbent assay (ELISA). Quite differing cross-reactivity results were obtained when the same grain protein extracts were used in indirect, antigen-competition and sandwich-type assay formats. In the sandwich assay format, alteration of either the solid phase-bound or labelled antibody altered apparent antibody specificity. These results indicate that it is important when reporting immunological homology data, to define cross-reaction of antibodies with respect to the assay format used. Antibody specificity (or cross-reactivity) can be manipulated by variation of sample extractant, solid phase and the format of the assay. © 1990, Academic Press Limited. All rights reserved.