[mono[125I]iodo-Tyr10,MetO17]-Vasoactive intestinal polypeptide. Preparation, characterization, and use for radioimmunoassay and receptor binding

J. L. Martin, K. Rose, G. J. Hughes, Pierre Magistretti

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52 Scopus citations

Abstract

Vasoactive intestinal polypeptide (VIP) was labeled with sodium [125I]iodide using the chloramine-T method and subsequently purified by reverse-phase high performance liquid chromatography. Three main 125I-labeled peaks designated A, B, and C resulted from the radioiodination and purification procedures. They were characterized by: (a) electrophoresis of tryptic fragments; (b) Edman degradation (for Peaks A and C); (c) enzymatic digestion to amino acids by leucine aminopeptidase, carboxypeptidase Y and Pronase; and (d) treatment with cyanogen bromide. Peak A corresponds to VIP monoiodinated on Tyr10 and with the Met17 residue oxidized to methionine sulfoxide. This [mono[125I]iodo-Tyr10,MetO17]VIP displays the following characteristics. 1) It constitutes quantitatively the major product of the iodination procedure (62.5%); 2) it is well resolved from other labeled and unlabeled products; 3) it is stable (2 months at -20°C); 4) it possesses a high specific activity (2050 Ci/mmol); 5) it maintains the biological activity of native VIP; and 6) it binds to antibody and membrane recognition sites in a specific, saturable, and reversible manner. Reduction of [mono[125I]iodo-Tyr10,MetO17]VIP to [mono[125I]iodo-Tyr10]VIP does not improve the performance of the tracer in a radioimmunoassay. The method described in this article is simple and rapid and yields a molecular form of 125I-labeled VIP that has been fully characterized and is suitable for use in biological studies.

Original languageEnglish (US)
Pages (from-to)5320-5327
Number of pages8
JournalJournal of Biological Chemistry
Volume261
Issue number12
StatePublished - Jan 1 1986

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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