Abstract: Human CD20 is a B lymphocyte surface antigen abnormally high-expressed in B-cell lymphoma, chosen as a cell-therapy target of B-cell malignancies. By molecular cloning, the extracellular domain of human CD20 was successfully constructed into pET28a vector as twins connected with a linker (-GSSGGSSG-）, and with a C-terminal 6His-tag. After confirming by DNA sequencing, the expression vector named pET28a-Bi20 was transformed into E. coli Transetta(DE3). The expression of the recombinant product Bi20 was induced by adding 1 mmol/L IPTG. When the optical density at 600 nm (OD600) of the bacterial suspension reached 0.6-0.8, at 37 ℃ for 5 h. The recombinant Bi20 was expressed as inclusion body. Through washing of inclusion body, purifying by Ni-affinity column under denatured condition, and screening and optimizing the refolding experimental conditions of inclusion body, the target peptide was finally purified as a soluble form of high purity. Western blotting and ELISA demonstrated that the purified Bi20 protein had a desirable immunogenicity.

Key words: human B lymphocyte surface antigen, cell therapy, recombinant expression, linker, inclusion body refolding