Expression, identification and bioactivity characterization of an anti-CD3/anti-CD20 bispecific single-chain antibody.
- Author:
Rui YU
1
;
Shi-Chong LI
;
Ben-Chuan WU
;
Hong LIU
;
Ling-Ling YE
;
Zhao-Lie CHEN
Author Information
1. Institute of Biotechnology, Academy of Military Medical Sciences, Beijing 100071, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Antibodies, Bispecific;
biosynthesis;
genetics;
immunology;
Antibodies, Monoclonal;
immunology;
Antigens, CD20;
immunology;
CD3 Complex;
immunology;
CHO Cells;
Cricetinae;
Cricetulus;
Humans;
Immunoglobulin Variable Region;
biosynthesis;
genetics;
immunology;
Lymphoma, B-Cell;
metabolism;
Recombinant Proteins;
Transfection;
Tumor Cells, Cultured
- From:
Chinese Journal of Biotechnology
2005;21(2):289-293
- CountryChina
- Language:Chinese
-
Abstract:
The synthetic gene with 1640bp encoding for the anti-CD3/anti-CD20 bispecific single-chain antibody was designed and obtained by SOE (splicing by overlap extension) PCR. The cDNA was cloned into Flp-In expression vector pcDNA5/FRT and transfeced into Flp-In CHO cells to generate a stable expression cell line with a capacity for expressing anti-CD3/anti-CD20 bispecific single-chain antibody at 300 microg/L. The protein, which had a molecular weight of about 70 kD,was purified by Ni-NTA affinity chromatography and identified by SDS-PAGE and Western-blot analysis. Immunofluorescence assay and cellular rosetting showed that it can react specifically on Jurkat (CD3+) and Ramous (CD20+) cells. The lysis of human PBL against CD20-positive lymphoma Ramous cells in the presence of the anti-CD3/anti-CD20 bispecific single-chain antibody can observed by microscope. All these results would lighten the further study of its biological functions in vitro and in vivo.
