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POLYCLONAL ANTISERA AGAINST ESTUARINE CRUSTACEAN VITELLIN AND VITELLOGENIN: DEVELOPMENT OF AN OOGENESIS DISFUNCTION ASSAY AS AN INDICATOR OF ENDOCRINE DISRUPTION FROM JUVENILE HORMONE AGONIST INSECTICIDES
Citation:
Tuberty, S. R., S. F. Nates, AND C L. McKenney Jr. POLYCLONAL ANTISERA AGAINST ESTUARINE CRUSTACEAN VITELLIN AND VITELLOGENIN: DEVELOPMENT OF AN OOGENESIS DISFUNCTION ASSAY AS AN INDICATOR OF ENDOCRINE DISRUPTION FROM JUVENILE HORMONE AGONIST INSECTICIDES. Presented at Crustacean Society 2000 Summer Meeting, Puerto Vallarta, Mexico, 26-30 June 2000.
Description:
Adult female Lepidophthalmus louisianensis, Palaemonetes pugio, Rhithropanopeus harrisii, Mysidopsis bahia, and Uca panacea were collected from estuarine localities in Santa Rosa Sound, Gulf Breeze, FL during late spring and summer of 1999. Mature ovaries were dissected and homogenized in crustacean extraction buffer, pH 7.3. After centrifugation at 12,000 g for 15 min, crude vitellin extracts were then prepared by gel permeation chromatography followed by anion-exchange chromatography. Eluate was monitored at 280 nm and 474 nm and samples with concomitant peaks were collected. The diluted samples were concentrated and lyophilized. Total protein content of the samples were assayed and PAGE and SDS-PAG electrophoresis was performed to ascertain the purity of the vitellin samples. Rabbits were immunized against each of the purified vitellins and sera collected after suitable blood antibody titers were achieved. Specificity of each antisera was tested against their respective vitellins and a male homogenate as negative controls using SDS-PAGE and Western blotting. The antisera were used to develop enzyme-linked immunoabsorbant assays for determination of hemolymph vitellogenin and egg vitellin concentrations. These assays are used to determine changes in baseline vitellin concentrations of eggs or hemolymph from females exposed to juvenile hormone agonists, which are used as insecticides and contribute to non-point source pollution of estuarine habitats.