1 research outputs found
Development of Polyclonal Antibodies for Detection of Diosbulbin B‑Derived <i>cis</i>-Enedial Protein Adducts
Diosbulbin
B (DSB), a major component of herbal medicine <i>Dioscorea bulbifera</i> L. (DB), can be metabolized to an electrophilic
intermediate, DSB-derived <i>cis</i>-enedial (DDE). DDE
was suggested to contribute to the hepatotoxicity observed in experimental
animals and humans after their exposure to DSB. Our previous work
found that DDE reacted with primary amino and/or sulfhydryl groups
of hepatic protein. The objective of the study was to develop polyclonal
antibodies that can recognize DDE-derived protein adducts. Immunogens
synthesized from DDE and keyhole limpet hemocyanin were employed to
raise polyclonal antibodies in rabbits. An enzyme-linked immunosorbent
assay (ELISA) demonstrated high titers of antisera obtained from immunized
rabbits. Immunoblot analysis showed that DDE-modified bovine serum
albumin (BSA) was recognized by the obtained polyclonal antibodies
in a concentration-dependent manner and without cross-reaction to
native BSA. Competitive ELISA and competitive immunoblot analyses
defined the specificity of the antibodies to recognize BSA modified
by DDE. Immunoblot analysis also detected a multitude of chemiluminescent
bands with a variety of molecular weights in liver homogenates that
were harvested from mice treated with DSB. In summary, we have successfully
raised polyclonal antibodies to detect protein adducts derived from
DDE