Synthesis of IGFBP-3 fragments in a baculovirus system and characterization of monoclonal anti-IGFBP-3 antibodies

Peter Vorwerk, Youngman Oh, Phillip D.K. Lee, Aruna Khare, Ron G. Rosenfeld

Research output: Contribution to journalArticlepeer-review

24 Scopus citations


IGFBPs play an important role in IGF biological actions by modulating IGF binding to its receptors. The major IGFBP in serum is IGFBP-3, which transports 70-90% of the circulating IGFs. In target cell systems, it sequesters IGFs and inhibits their hormonal actions, but may potentiate IGF activity or exert IGF-independent effects under specific conditions. IGFBP-3 can be modified by IGFBP-3 proteases, which degrade it into smaller fragments. IGFBP-3 fragments generated by proteolysis have reduced affinity for IGFs, thereby modifying IGF action. To study IGFBP-3 fragments in vivo and in vitro, we constructed six different IGFBP-3 fragments by use of a baculovirus expression system and generated 8 different monoclonal IGFBP-3 antibodies. Based on the known cleavage sitos of IGFBP-3 for PSA, MMPs, and the predicted plasmin cleavage sites, we expressed a N-terminal IGFBP-31- 97 fragment and a C-terminal IGFBP-398-264 fragment. By stepwise truncation from the C- lerminal end, we created IGFBP-398-232, IGFBP- 398-206, IGFBP-398-179, and IGFBP-398-159. A strong recognition of the C- terminus and the intermediate parts of IGFBP-3 by six antibodies was found. Four of these mAbs were able to recognize the intermediate fragment alone. Two mAbs were found to immunorcact only with the N-terminal IGFBP-3 fragment and two additional mAbs recognized the N- as well as the C-terminal parts and lacked immunoreactivity for the intermediate part of IGFBP-3. The 15 kDa IGFBP-3 fragment resulting from plasmin digestion was found to only react with N-terminal antibodies, while the 29 kDa fragment in pregnancy serum reacted with both N- and C-terminal antibodies. Thus, these mAbs will be useful tools to determine whether IGFBP-3 fragments found in vivo derive from either the N- or C-terminal domains of IGFBP-3.

Original languageEnglish (US)
Pages (from-to)2368-2370
Number of pages3
JournalJournal of Clinical Endocrinology and Metabolism
Issue number7
StatePublished - 1997
Externally publishedYes

ASJC Scopus subject areas

  • Endocrinology, Diabetes and Metabolism
  • Biochemistry
  • Endocrinology
  • Clinical Biochemistry
  • Biochemistry, medical


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