The Development and Validation of a Quantitative ELISA for in vivo and in vitro Synthesized Vitellogenin from Mossambicus tilapia (Oreochromis mossambicus)

Abstract

The induction of vitellogenin (VTG), an egg yolk precursor found in most oviparious vertebrates, has been well established as a valuable biomarker for assessing estrogenic endocrine disruption. Oreochromis mossambicus is a tilapiine species indigenous to Southern Africa and, therefore, represents a potential African bio-indicator for the development of in vitro and in vivo screens for estrogenicity using VTG as biomarker. However, few ELISAs have been developed to quantify tilapia VTG protein levels. In the present study, commercially available VTG antiserum that shows cross reactivity with tilapia VTG was used to set up and validate a quantitative competition ELISA for tilapia VTG. This ELISA has a broad detection range between 80 ng/ml–5.4 µg/ml VTG and is able to detect both in vivo and in vitro synthesized VTG. This ELISA is highly repeatable with intra- and inter-assay variations less than 3.4% at the lowest detection limit.

Keywords:

• Biomarker
• ELISA
• Endocrine disruption (EDC)
• Estrogenicity
• Oreochromis mossambicus
• Vitellogenin

ACKNOWLEDGMENTS

Special thanks to Prof. Hannes Van Wyk, University of Stellen Bosch for his valuable inputs. We would like to thank the National Research Foundation (NRF) of South Africa for financially supporting this study.

Notes

^a For routine measurement 1:2000 VTG antigen and 1:1000 antibody dilution was selected.

^b CV = coefficient of variation.