Abstract: Objective To optimize the conditions for enhancing the refolding of a novel recombinant human (rh) endostatin and test the biological activities of refolded endostatin. Methods The partial purified inclusion bodies of rh-endostatin were dissolved with 6mol/L Guanidine-HCl followed by combination of dilution and dialysis of the dissolved endostatin. The refolded endostatin was then purified by cation-exchange chromatography. The biological activities of purified rh-endostatin were assessed by using endostatin-specific monoclonal antibody and chick embryo chorioallantoic membrane assay. Results The 63% refolding yield was achieved after optimizing the refolding conditions. The purified endostatin was reacted with specific anti-endostatin monoclonal antibody and showed significant inhibition of angiogenesis in chick embryo chorioallantoic membrane assay. Conclusion The method of highest refolding yield of human endostatin so far was developed. This optimized method will greatly increase the application of this novel human endostatin to preclinical and clinical studies.

Key words: recombinant human endostatin, refolding, purification, chick embryo chorioallantoic membrane assay