Development of an Enzyme-linked Immunosorbent Assay Using Vitellin for Vitellogenin Measurement in the Pale Chub, <i>Zacco platypus</i>

Abstract

ObjectivesFish vitellogenin (VTG) is produced in the female liver during oogenesis through the estradiol cycle and produced in the male liver by endocrine disrupting chemicals (EDCs) such as alkylphenols. In this study, we propose that the VTG concentration in the pale chub could be detected using monoclonal antibodies and polyclonal antibodies against vitellin (Vn) in a VTG enzyme-linked immunosorbent assay (ELISA) system.MethodsMonoclonal antibodies and polyclonal antibodies were produced using the Vn extracted from the matured ovum of the ovary. The VTG was extracted from the plasma of the male pale chub. The Vn and VTG were confirmed by measuring the molecular weight of their proteins using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and the specificity of the antibodies was checked through western blotting methods. The assay system was validated with respect to optimal assay concentrations, specificity, recovery, and intra- and inter-assay variations.ResultsThe Vn consisted of two protein bands with apparent molecular weights of 64 and 37 kDa. The SDS-PAGE indicated protein weights of 146 and 77 kDa in the VTG. The assay range was 15.6 ng/mL to 2,000 ng/mL, and the value of the intra- and inter-assay variations were within 10.0% and 14.7%, respectively. The recovery rate was 99.5±5.5%.ConclusionsA sandwich ELISA was developed that could be used to qualify the VTG of pale chub in screening for EDCs. Pale chub is an ideal species for observing estrogen activity in the environment because of its extensive habitat and extensive food chain. The ELISA developed here would be more favorable than those for other species for determining the effect of long-term food chain accumulation of EDCs in aquatic environments.

<p>Toxicopathological and immunological studies on different concentrations of chitosan-coated silver nanoparticles in rats</p>

“…The toxicities that come from the different administration routes regularly fluctuated because of the distribution patterns. For instance, in a single-dose oral administration study,13 the tissue distribution of Ag in the liver, kidneys, and lungs was higher when Ag + was administered compared with AgNPs. However, intravenously administered AgNPs predominantly aggregated in the liver and spleen, and the free Ag + was in this way released and excreted, a large amount of which remained in the kidneys, lungs, and brain 14.…”