| Abstract: | Two related agglutinins are present in the seeds of Ricinus communis (castor): ricin, a dichain ribosome-inactivating protein and Ricinus communis agglutinin-1, a much less toxic tetrameric hemagglutinin. The immunochemical analysis of these agglutinins is of special interest because ricin toxicity has resulted in both accidental and intentional poisonings, while it has also provided a potential cancer chemotherapeutic in the form of an immunoconjugate. We previously characterized a panel of monoclonal antibodies (mAbs) for the analysis of potential contamination with ricin in several food matrices. In this study, an optical sensing technique, biolayer interferometry (BLI), was used to study the binding of two mAbs to the agglutinins. MAbs were immobilized on sensors with amine-reactive, Fc-binding, and streptavidin-coated tips to study the interactions with the agglutinins and with ricin A- and B-chains in solution. The kinetically determined equilibrium dissociation constants generally agreed with the relative binding observed in ELISA, although binding was less predictable for the isolated ricin chains. BLI analysis of kinetic constants for mAb 1797 was not affected by nonfat milk (0.5% by volume). BLI provides a useful method to characterize the binding of antibodies, with the potential for immunodiagnostic applications in food matrices. |
