Development and characterization of polyclonal antibodies against subtype specific vitellogenin of the dojo loach, Misgurnus anguillicaudatus

Fish vitellogenin (Vtg) has become an important biomarker for assessing the estrogenic potency of chemicals and the exposure of animals to estrogenic contaminants present in aquatic environments. In the current study, the polyclonal antibodies against subtype-specific Vtgs of the dojo loach (Misgurnus anguillicaudatus) were developed and identified by gene recombination and expression. Immuno-biochemical analyses revealed that VtgAo1 protein appeared to be the major Vtg type in this species. Enhanced chemiluminescent Western blotting was developed using the antiserum against VtgAo1. Exposure of male loach to 17 alpha-ethinylestradiol (EE2) and 17 beta-estradiol (E2) via water induced the VtgAo1 protein induction with a lowest-observed-effect concentration (LOEC) by 100 ng L-1 and 1000 ng L(-1)group at 7 day post-initiation (dpi), respectively. The results indicated that this method provided a valuable tool for detecting estrogenic activities in aquatic environments and excluded any difficulties expected in purification procedures to separate a highly pure Vtg subtype from circulating proteins, and the other Vtg subtypes.

Automated summary

The study developed a method for detecting estrogenic activities in aquatic environments by developing polyclonal antibodies against subtype-specific Vtgs of the dojo loach. It was found that VtgAo1 protein was the major Vtg type in this species and successfully used for Western blotting analysis, indicating induction of VtgAo1 protein after exposure to EE2 and E2.