Structure–immunogenicity relationship of kresoxim-methyl regioisomeric haptens

Abstract

Kresoxim-methyl was one of the two first strobilurins to be discovered, and nowadays it is widely used as an antifungal agent in crop protection. Because of its low molecular weight and negligible structural complexity, the generation of antibodies to kresoxim-methyl noticeably requires the preparation of functionalized haptens. In this study, the introduction of a hydrocarbon spacer arm at the aromatic moieties of the target molecule was carried out by a convergent strategy based on the Sonogashira cross-coupling reaction, and functionalized linkers of the same length were also tethered to the aliphatic toxophore group by the O-alkylation reaction. Evaluation of the immune response, in terms of antibody affinity, showed a differential behavior among five synthesized haptens whose sole dissimilarity was the derivatization site. The characteristic (methoxyimino)acetate moiety of strobilurins was revealed as the optimum linker position for high-affinity polyclonal and monoclonal antibody production. However, good monoclonal antibodies were isolated from mice immunized with a hapten carrying the linker at an opposite site, which otherwise generated a poor polyclonal response in rabbits. Site-heterology was confirmed as a feasible approach for the improvement of the apparent affinity, particularly with polyclonal antibodies. Several of the monoclonal antibodies generated in the context of this project could be proper binders for kresoxim-methyl immunosensing over different analytical platforms.

Combined heterologies for monoclonal antibody-based immunoanalysis of fluxapyroxad

“…[23][24][25] In summary, the antibodies obtained with hapten FXn showed superior affinity and specificity to fluxapyroxad than those derived from hapten FXb, thus evidencing the relevance of the linker position in the binding properties of the generated antibodies, as also demonstrated for other low molecular weight compounds. [26][27][28] For further cELISA development, two combinations of immunoreagents were selected according to the lowest IC 50 value and moderate slope of the inhibition curve, one for the direct format and the other for the indirect format. The optimum concentration of each immunoreagent was determined in order to obtain A max values between 1 and 2 absorbance units.…”

High-affinity Antibodies from a Full Penthiopyrad-mimicking Hapten and Heterologous Immunoassay Development for Fruit Juice Analysis

“…Immunizing haptens should resemble as much as possible the target compound, being the linker tethering site one of the major determinants for a correct display of the characteristic chemical moieties of the molecule (López-Moreno et al 2013). Conjugate BSA-PPm, in which the aliphatic moiety of the molecule was used as linker, was chosen for antibody generation since it carried the hapten that was the best mimic of penthiopyrad .…”

Rationally designed haptens for highly sensitive monoclonal antibody-based immunoanalysis of fenhexamid

“…Moreover, CAMM-based structure−acƟvity relaƟonship analyses can contribute to better understand affinity and specificity characteristics of the produced biomolecules. [17][18] Structurally, the fenhexamid framework is made up by an amido bridge that links an aliphatic moiety to an aromatic group (Fig. 1).…”