Bioscreening and expression of a camel anti-CTGF VHH nanobody and its renaturation by a novel dialysis-dilution method.
The variable regions of the camel heavy chain antibody, also known as nanobody is the smallest antibody with antigen-binding efficiency. CTGF is considered important during extracellular matrix deposition which was involved in the pathogenesis of fibrosis related diseases. There are several anti-CTGF-C nanobody drugs under developing in pharmacy. In this study, we described the screening of a novel anti-CTGF-C nanobody from the peripheral blood of immunized camel by phage display. The screened nanobody was further expressed and purified from E. coli cells. A sophisticated dialysis-dilution method was designed for the in vitro refolding of the nanobody. The results showed that the expressed nanobody was consisted of 135 amino acid and mainly expressed as inclusion body in E. coli cells. The dialysis-dilution method was very effective and the recovery rate of the renaturation was more than 80 %. The ELISA result suggested the nanobody had been well refolded showing a superior CTGF binding activity to the commercial mouse anti-CTGF-C mAb. In conclusion, the anti-CTGF-C nonobody had been successfully screened by phage display. The dialysis-dilution refolding method was very effective and the recovery rate reached over 80 %.
Clinical Biochemistry Department, Medicine School, Southeast University, Nanjing, China.
Xiulei
X
Xue
Clinical Biochemistry Department, Medicine School, Southeast University, Nanjing, China.|Center of Clinical Laboratory Medicine of Zhongda Hospital, Southeast University, Nanjing, China.
Xiaobo
X
Fan
Clinical Biochemistry Department, Medicine School, Southeast University, Nanjing, China.
Qingrong
Q
Qu
Clinical Biochemistry Department, Medicine School, Southeast University, Nanjing, China. guoqiuwu@163.com.|Center of Clinical Laboratory Medicine of Zhongda Hospital, Southeast University, Nanjing, China. guoqiuwu@163.com.
Guoqiu
G
Wu
Connective tissue growth factor; Nanobody; Phage display; Protein renaturation