Alternative Hapten Design for Zearalenone Immunoreagent Generation

Appropriate hapten design and synthesis have been identified as critical steps to generate high-performance immunoreagents and to develop sensitive and selective immunoanalytical methods. Antibodies and immunoassays for the major mycotoxin zearalenone have been reported and marketed. However, zearalenone haptens have mostly been prepared by the oxime active ester technique, and hapten characterization has generally been poor or non-existent. In the present study, novel haptens of zearalenone with longer linkers and with alternative tethering sites have been designed for immunizing and assay conjugate preparation. All of these molecules were purified and spectroscopically verified, and a structure-activity relationship evaluation was carried out. This approach revealed that the hapten with the linker at the carbonyl group generated antibodies with a higher affinity than the hapten functionalized at the phenyl moiety. Antibodies produced with the latter hapten, on the other hand, showed lower cross-reactivity values to the major zearalenone metabolites. Finally, similar immunoassay sensitivity was achieved with all of the antibodies when heterologous haptens were employed. Furthermore, by altering the structure of the competing antigen, the immunoassay selectivity was modified. These results demonstrate that immunochemical methods for zearalenone rapid analysis can still be improved in terms of sensitivity and selectivity.

Automated summary

Appropriate hapten design and synthesis are critical for generating high-performance immunoreagents and developing sensitive and selective immunoanalytical methods. In this study, novel haptens of zearalenone were designed and evaluated for their structure-activity relationship. The results showed that modifying the hapten structure can affect antibody affinity and cross-reactivity, and changing the competing antigen structure can modify the immunoassay selectivity. These findings suggest that the immunochemical methods for zearalenone rapid analysis can still be improved in terms of sensitivity and selectivity.