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VITELLOGENIN ELISA FOR FATHEAD MINNOWS (PIMEPHALES PROMELAS) USING A COMPLETELY HOMOLOGOUS ASSAY SYSTEMS
Citation:
Chow, M., J J. Korte, J. M. Hemming, AND N D. Denslow. VITELLOGENIN ELISA FOR FATHEAD MINNOWS (PIMEPHALES PROMELAS) USING A COMPLETELY HOMOLOGOUS ASSAY SYSTEMS. Presented at 22nd Annual SETAC Meeting, Baltimore, MD, November 11-15, 2001.
Description:
The indication of vitellogenin in fish has been used as a biomarker for estrogen-receptor mediated gene induction pathways resulting from exposure to environmental estrogens. Pimephales promelas (fathead minnows) have been selected as one of the test models to investigate reproductive dysfunction due to exposure. Because this assay will need to be standardized among laboratories, we have prepared a series of monoclonal antibodies that are specific for fathead minnow Vtg that are of high affinity and high sensitivity. We have determined the epitope for one antibody which enabled us to make a synthetic peptide and use it, instead of fathead minnow Vtg, to coat a 96-well plate for a standard competition assay. This removes the requirement for large amounts of purified fathead minnow vitellogenin for coating plates. We are still, however, using a fathead minnow vitellogenin standard curve. The working range of this assay is 5 ng/ml to 100 ng/ml (90% to 20% binding B/Bo) for the standard curve. Since we have to dilute plasma samples by a factor of 10 to avoid interferences, the sensitivity of the assay for plasma samples becomes 50 ng/ml, which compares well to other assays currently available. This assay is useful for determining Vtg induction in fathead minnows arising from environmental exposures.