IMMUNOCHEMICAL CHARACTERIZATION OF TYPE-A BOTULINUM NEUROTOXIN IN ITSPURIFIED AND COMPLEXED FORMS

Immunochemical reactivities of type A botulinum neurotoxin to polyclon al antibodies raised against the neurotoxin complex toroid have been i nvestigated using enzyme-linked immunosorbent assay (ELISA) and with a fiberoptic immunosensor. The complex contains a group of complexing p roteins in addition to the neurotoxin itself. Neurotoxin specific immu noglobulin G (IgG), purified from the IgG raised against the complex u sing an affinity column, showed a two-fold increase in reactivity with purified neurotoxin compared to the neurotoxin in the complex. Antibo dies against the whole complex reacted approximately five times better with the complex than with the neurotoxin, suggesting that the detect ion of the neurotoxin complex may be more sensitive. Considering the f act that the amount of the complexing proteins and neurotoxin appears to be in a 4:1 ratio, a five-fold higher reactivity could suggest a 25 -fold higher detectability of the neurotoxin in the complex. ELISA bin ding curves of complexing proteins and purified neurotoxin with antibo dies raised against the whole complex indicated the complexing protein s to have significantly higher antigenicity. Furthermore, IgG fraction with or without the neurotoxin specific antibodies reacted almost equ ally to the neurotoxin complex, again suggesting higher immunogenicity of the complexing proteins. Increased binding of the complexing prote ins versus the purified neurotoxin to antibodies against the complex a nd thus immunogenicity was also observed in the binding curves generat ed using a fiberoptic immunosensor.