RESUMO
3-Succinylaconitine was conjugated with bovine serum albumin (BSA) for use as an immunogen for the preparation of a monoclonal antibody (MAb) against aconitine (Aco). Splenocytes from mice immunized with the Aco-BSA conjugate were fused with an aminopterin-sensitive mouse myeloma cell line, P3-X63-Ag8-653, and a hybridoma secreting a MAb against Aco was successfully obtained. The MAb cross-reacted with mesaconitine, hypaconitine and jesaconitine, which are Aco-type alkaloids, but not with any other compounds examined. The full measurement range of an enzyme-linked immunosorbent assay (ELISA) developed using the new MAb extended from 100ngmL(-1) to 1.5microgmL(-1) of Aco. The concentrations of Aco-type alkaloids in various Aconiti radixes assayed using the new ELISA method showed good agreement with previous reports.
Assuntos
Aconitina/análise , Anticorpos Monoclonais/química , Aconitina/análogos & derivados , Aconitina/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/isolamento & purificação , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Calibragem , Células Cultivadas , Ensaio de Imunoadsorção Enzimática/métodos , Hibridomas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Conformação Molecular , Sensibilidade e Especificidade , Soroalbumina Bovina/química , Soroalbumina Bovina/imunologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Baço/citologia , Baço/imunologia , EstereoisomerismoRESUMO
The purpose of this study was to determine whether larkspur toxins conjugated to protein carriers would promote active immunity in mice. Mice were injected with several larkspur toxin-protein conjugates or adjuvant alone to determine whether the resulting immunological response altered animal susceptibility to methyllycaconitine, the major toxic larkspur alkaloid. Although vaccinations increased the calculated lethal dose 50% (LD50) for intravenous methyllycaconitine toxicity, overlapping confidence intervals did not provide evidence of differences between the vaccinated and control groups. In the lycoctonine conjugate (LYC)-vaccinated group, mouse survival was related (P = 0.001) to serum titers for methyllycaconitine doses up to 4.5 mg/kg of body weight. When mice withlow antibody titers were removed from the vaccinated groups in which titer was related to survival, the recalculated LD50 estimates were 20% greater than the LD50 of the control group. However, the 95% confidence intervals of the recalculated LD50 groups overlapped with the control groups. Overall, these results suggest that vaccination altered methyllycaconitine toxicity in mice and that vaccination may be useful in decreasing the effects of larkspur toxins in animals. Additional studies are warranted to continue development of potential larkspur vaccines for livestock.
