A new type of connective tissue growth factor in human soluble single-chain antibody expression and purification.docVIP

 PAGE \* MERGEFORMAT 20 A new type of connective tissue growth factor in human soluble single-chain antibody expression and purification Study: Fan Xiaobo, Gou Li-Xia, Liu Zhao, Chen Zilong, REMEDIES, Wu ball, Xi Tao [Abstract] Objective: To study the single-chain antibody (scFv) in different expression vectors and strains and the immunological activity of expression product evaluation. Methods: The humanized single-chain antibody gene by restriction enzyme digestion, the connection method such as biotechnology cloned into pET28a, pET32a and pET EI: X 3 different vector and the expression of the different strains used to express, observed the expression of single chain antibodies and soluble expression of antibody to optimize the use of affinity chromatography and gel layer Analysis of the purified protein, and in vitro biological activity of the target protein. Results: The anti-connective tissue growth factor single-chain antibody pET28a/BL21 (DE3) can not be expressed in pET32a/BL21 (DE3) and pET EI: X / BLR (DE3) was successfully expressed in both, but in the pET EI: X / BLR (DE3) containing the scFv antibody from a peptide could not (intein) with cleavage. scFv in pET32a/BL21 (DE3) in the total protein expression about 30%, after two steps of chromatography, the purity of 85% or more. vitro immunoassay showed that the target protein with good immunogenicity. Conclusion: The humanized anti-connective tissue growth factor single chain antibody scFv in Escherichia coli need for molecular chaperone expression, the expression of policy by low temperature in pET32a/BL21 (DE3) was expressed successfully. [Keywords:] Human single chain antibody; connective tissue growth factor; elastin; separation and purification [Abstract] Objective: To investigate the expression of a noval anti CTGF single chian Fv (scFv) antibody within different plasmid and / or host cell and the in vitro immunoreactivity of this scFv antibody. Methods: In this study, a scFv antib