Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica

Acta Academiae Medicinae Sinica ›› 2011, Vol. 33 ›› Issue (5): 543-548.doi: 10.3881/j.issn.1000-503X.2011.05.013

• Original Articles • Previous Articles     Next Articles

Prokaryotic Expression of Recombinant Human Bone Morphogenetic Protein-2 and Preparation of Monoclonal Antibodies

WANG Jing1, ZHANG Jie2, ZHANG Ying2, MA Ning-ning1, XIE Liang-zhi1   

  1. 1Cell Culture and Center, CAMS and PUMC, Beijing 100005, China; 2Sinocelltech Ltd, Beijing 100176, China
  • Received:2011-02-16 Revised:2011-10-28 Online:2011-10-28 Published:2011-10-28
  • Contact: XIE Liang-zhi
  • Supported by:
    Supported by the Key Project of the “Eleventh Fiveyear Plan” for Medical Science Development of China(2009ZX090306-004)

Abstract: Objective To express and purify the recombinant human bone morphogenetic protein-2 mature peptide (rhBMP-2m) in prokaryotic system and to develop highly-specific monoclonal antibodies. Method An engineered E.coli strain expressing rhBMP-2m was fermented. The bacterial cells were firstly lysed and then the rhBMP-2m inclusion bodies were isolated by centrifugation. After the inclusion bodies had been solubilized by high-concentration denaturing agents, denatured rhBMP-2m was purified by cantion ion-exchange chromatography. Biologically active rhBMP-2m was obtained by refolding of purified denatured rhBMP-2m through direct dilution. The refolded rhBMP-2m was used to immunize Balb/c mice to develop anti-rhBMP-2m monoclonal antibodies using classic hybridoma technique. Results rhBMP-2m with a purity greater than 95% was obtained on reduced SDS-PAGE. The refolded rhBMP-2m was measured to be bioactive by the induction of alkaline phosphatase activity in MC3T3-E1 cells. Two hybridoma cell lines that stably secreted anti-rhBMP-2m antibody were developed from the immunized mice. Conclusion Bioactive rhBMP-2m protein and its monocloncal antibodies were successfully prepared, which will provides a solid base for future studies on rhBMP-2.

Key words: recombinant human bone morphogenetic protein-2 mature peptide, protein refolding, monoclonal antibody

CLC Number: