Induction of vitellogenesis by estradiol-17β and development of enzyme-linked immunosorbant assays to quantify plasma vitellogenin levels in green turtles (Chelonia mydas)

Lawrence H. Herbst, Linda Siconolfi-Baez, John H. Torelli, Paul A. Klein, Marci J. Kerben, Isabella M. Schumacher

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

Treatment of juvenile green turtles (Chelonia mydas) with estradiol-17β resulted in the induction of a 200 kDa plasma protein, consistent with vitellogenin (Vtg). The N-terminal 15 amino acids of the anion exchange purified protein shared sequence homologies with vitellogenins of several vertebrate species. Rabbit antiserum raised against purified Vtg recognized the plasma protein as well as several yolk proteins. Monoclonal antibody (Mab) HL1248, produced by inoculating mice with turtle yolk granules, showed specificity for plasma Vtg as well as a set of yolk proteins 120, 82, 43 and 32 kDa in size. The N-terminal 22 amino acids of the 43 kDa yolk protein was similar to the lipovitellin I subunit of Vtg of several vertebrate species. The peptide mass map of the 82 kDa yolk protein shared enough ions with that of purified plasma Vtg to support the conclusion that this protein was derived from plasma Vtg. Taken together, these results validate the specificity of Mab HL1248 for Vtg. Using purified Vtg concentration standards, competition and antigen capture enzyme-linked immunosorbant assays (ELISAs) were shown to quantitatively detect Vtg in green turtle plasma. Pre-induced plasma of juvenile turtles had Vtg levels of 2-4 μg/ml whereas post-estradiol exposure samples had 38-40 mg/ml. The plasma Vtg concentration of a nesting female turtle was 4.6 mg/ml, approximately 20-fold higher than that of a non-nesting adult female. The antigen capture ELISA will be useful in population studies of this endangered species, to detect vitellogenesis in females that will nest in a given year and to detect inappropriate Vtg levels in turtles exposed to xenoestrogens.

Original languageEnglish (US)
Pages (from-to)551-563
Number of pages13
JournalComparative Biochemistry and Physiology - B Biochemistry and Molecular Biology
Volume135
Issue number3
DOIs
StatePublished - Jul 1 2003

Fingerprint

Vitellogenesis
Vitellogenins
Turtles
Estradiol
Assays
Plasmas
Enzymes
Egg Proteins
Vertebrates
Blood Proteins
Monoclonal Antibodies
Antiporters
Antigens
Amino Acids
Amino Acid Sequence Homology
Endangered Species

Keywords

  • Chelonia mydas
  • Enzyme-linked immunosorbant assay
  • Estrogen
  • Green turtle
  • Monoclonal antibodies
  • Reptiles
  • Vitellogenin
  • Yolk

ASJC Scopus subject areas

  • Biochemistry
  • Physiology

Cite this

Induction of vitellogenesis by estradiol-17β and development of enzyme-linked immunosorbant assays to quantify plasma vitellogenin levels in green turtles (Chelonia mydas). / Herbst, Lawrence H.; Siconolfi-Baez, Linda; Torelli, John H.; Klein, Paul A.; Kerben, Marci J.; Schumacher, Isabella M.

In: Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology, Vol. 135, No. 3, 01.07.2003, p. 551-563.

Research output: Contribution to journalArticle

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abstract = "Treatment of juvenile green turtles (Chelonia mydas) with estradiol-17β resulted in the induction of a 200 kDa plasma protein, consistent with vitellogenin (Vtg). The N-terminal 15 amino acids of the anion exchange purified protein shared sequence homologies with vitellogenins of several vertebrate species. Rabbit antiserum raised against purified Vtg recognized the plasma protein as well as several yolk proteins. Monoclonal antibody (Mab) HL1248, produced by inoculating mice with turtle yolk granules, showed specificity for plasma Vtg as well as a set of yolk proteins 120, 82, 43 and 32 kDa in size. The N-terminal 22 amino acids of the 43 kDa yolk protein was similar to the lipovitellin I subunit of Vtg of several vertebrate species. The peptide mass map of the 82 kDa yolk protein shared enough ions with that of purified plasma Vtg to support the conclusion that this protein was derived from plasma Vtg. Taken together, these results validate the specificity of Mab HL1248 for Vtg. Using purified Vtg concentration standards, competition and antigen capture enzyme-linked immunosorbant assays (ELISAs) were shown to quantitatively detect Vtg in green turtle plasma. Pre-induced plasma of juvenile turtles had Vtg levels of 2-4 μg/ml whereas post-estradiol exposure samples had 38-40 mg/ml. The plasma Vtg concentration of a nesting female turtle was 4.6 mg/ml, approximately 20-fold higher than that of a non-nesting adult female. The antigen capture ELISA will be useful in population studies of this endangered species, to detect vitellogenesis in females that will nest in a given year and to detect inappropriate Vtg levels in turtles exposed to xenoestrogens.",
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