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[Cloning of variable region genes of anti-tetanus toxoid antibody and their expression as three kinds of engineered antibodies in E. coli].
Shi Yan Sheng Wu Xue Bao. 1997 Sep; 30(3):285-92.SY

Abstract

The heavy and light chain variable region genes of anti-tetanus toxoid (TT) antibody and the heavy chain Fd genes were amplified and cloned through RT-PCR from mouse hybridoma cells. The sequences of VH and VK were determined. Fd gene fragments were expressed in E. coli. The ELISA results indicated that the expressed Fd showed antigen binding activity but was nonspecific. Furthermore, through SOE and PCR techniques, the VH and VK gene fragments together with ScFv linker were assembled into single chain antibody (ScFv) gene fragment. While together with human heavy chain CH 1 gene fragment and Fab linker, they were assembled into chimeric Fab gene fragment. The two assembled gene fragments were separately inserted into phagemid pHEN 1, which was a fd-based vector containing gene 3 encoding the minor coat protein. In presence of helper phage M 13-VCS the anti-TT phage-ScFv or phage-Fab were displayed on the surface of phage particles respectively. Results from phage-ELISA indicated that both phage antibodies were TT-specific.

Authors+Show Affiliations

Shanghai Institute of Cell Biology, Chinese Academy of Sciences.No affiliation info available

Pub Type(s)

English Abstract
Journal Article
Research Support, Non-U.S. Gov't

Language

chi

PubMed ID

11039023

Citation

Zan, H, and M Yeh. "[Cloning of Variable Region Genes of Anti-tetanus Toxoid Antibody and Their Expression as Three Kinds of Engineered Antibodies in E. Coli]." Shi Yan Sheng Wu Xue Bao, vol. 30, no. 3, 1997, pp. 285-92.
Zan H, Yeh M. [Cloning of variable region genes of anti-tetanus toxoid antibody and their expression as three kinds of engineered antibodies in E. coli]. Shi Yan Sheng Wu Xue Bao. 1997;30(3):285-92.
Zan, H., & Yeh, M. (1997). [Cloning of variable region genes of anti-tetanus toxoid antibody and their expression as three kinds of engineered antibodies in E. coli]. Shi Yan Sheng Wu Xue Bao, 30(3), 285-92.
Zan H, Yeh M. [Cloning of Variable Region Genes of Anti-tetanus Toxoid Antibody and Their Expression as Three Kinds of Engineered Antibodies in E. Coli]. Shi Yan Sheng Wu Xue Bao. 1997;30(3):285-92. PubMed PMID: 11039023.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - [Cloning of variable region genes of anti-tetanus toxoid antibody and their expression as three kinds of engineered antibodies in E. coli]. AU - Zan,H, AU - Yeh,M, PY - 2000/10/20/pubmed PY - 2001/2/28/medline PY - 2000/10/20/entrez SP - 285 EP - 92 JF - Shi yan sheng wu xue bao JO - Shi Yan Sheng Wu Xue Bao VL - 30 IS - 3 N2 - The heavy and light chain variable region genes of anti-tetanus toxoid (TT) antibody and the heavy chain Fd genes were amplified and cloned through RT-PCR from mouse hybridoma cells. The sequences of VH and VK were determined. Fd gene fragments were expressed in E. coli. The ELISA results indicated that the expressed Fd showed antigen binding activity but was nonspecific. Furthermore, through SOE and PCR techniques, the VH and VK gene fragments together with ScFv linker were assembled into single chain antibody (ScFv) gene fragment. While together with human heavy chain CH 1 gene fragment and Fab linker, they were assembled into chimeric Fab gene fragment. The two assembled gene fragments were separately inserted into phagemid pHEN 1, which was a fd-based vector containing gene 3 encoding the minor coat protein. In presence of helper phage M 13-VCS the anti-TT phage-ScFv or phage-Fab were displayed on the surface of phage particles respectively. Results from phage-ELISA indicated that both phage antibodies were TT-specific. SN - 0001-5334 UR - https://www.unboundmedicine.com/medline/citation/11039023/[Cloning_of_variable_region_genes_of_anti_tetanus_toxoid_antibody_and_their_expression_as_three_kinds_of_engineered_antibodies_in_E__coli]_ L2 - http://www.diseaseinfosearch.org/result/7033 DB - PRIME DP - Unbound Medicine ER -