Development of a sandwich enzyme-linked immunosorbant assay for the quantification of vitellogeinin in Bullfrog ( Rana catesbeiana)

• Published in: Chemosphere (Oxford) 2006-06, Vol.63 (9), p.1485-1490
• Main Authors: Li, Chun-Ri, Jung, Kwun-Soon, Lee, Sang-Hoon, Kim, Andre, Park, Jang-Su
• Format: Article
• Language: English
• Subjects: Animal, plant and microbial ecology; Animals; Antibodies, Monoclonal; Applied ecology; Biological and medical sciences; Bullfrog-VTG; Ecotoxicology, biological effects of pollution; ELISA; Enzyme-Linked Immunosorbent Assay - methods; Estradiol - pharmacology; Estrogenic activity; Female; Freshwater; Fundamental and applied biological sciences. Psychology; General aspects; Male; Rana catesbeiana; Rana catesbeiana - metabolism; Seminiferous Tubules - drug effects; Seminiferous Tubules - pathology; Sensitivity and Specificity; Testis - drug effects; Testis - pathology; Vitellogenins - analysis; Vitellogenins - drug effects; Vitellogenins - immunology
• ISSN: 0045-6535; 1879-1298
• DOI: 10.1016/j.chemosphere.2005.09.041

A sandwich enzyme-linked immunosorbant assay (ELISA) was developed to quantitatively detect Bullfrog ( Rana catesbeiana) vitellogenin (VTG) levels. This procedure involved a sandwich ELISA using monoclonal antibodies of BVmA1 and BVmA2 against Bullfrog-VTG, and BVmA1 conjugated to horseradish peroxidase as the detection antibody. The assay range was between 9.4 ng/ml and 1200 ng/ml and the recovery of the VTG added to Bullfrog control male serum was 92.0–108.8%. Male Bullfrog was induced by injection 17β-estradiol (E 2) for four weeks and Bullfrog-VTG levels were measured each week. Histological analysis was performed for investigating the correlation of the effect to male reproduction and Bullfrog-VTG level variation depending on E 2 dose. After two weeks of E 2 exposure, the induced Bullfrog-VTG level was significantly higher than Bullfrog control female ( p < 0.05). After four weeks of E 2 exposure, the rupture and fusion of seminiferous tubule in the testes of male Bullfrog were shown and provided direct evidence that the reproduction of male Bullfrog was affected by estrogenic compounds. Bullfrog-VTG bioassay, using the sandwich ELISA, could be a sensitive and useful tool for quantification of estrogenic principles in aquatic environments.