Tags

Type your tag names separated by a space and hit enter

[Preparation of phage-displayed single chain variable fragments of monoclonal antibody MC5 recognizing colorectal carcinoma].
Ai Zheng. 2002 Jun; 21(6):636-9.AZ

Abstract

BACKGROUND & OBJECTIVE

MC5 is a murine monoclonal antibody with a good specificity to human colorectal carcinoma and smaller murine antibody can significantly decrease the possibility of developing human antimouse antibody response in vivo study. The aim of this study was to prepare single chain variable fragments (ScFv) of MC5.

METHODS

mRNA was isolated from the hybridoma cell line producing MC5, and the DNAs encoding variable domains of heavy and light chains(VH and VL DNAs) of the antibody were amplified separately by RT-PCR and assembled into ScFv DNAs with a linker DNA. The ScFv DNAs were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into E. coli TG1. The transformed cells were infected with M13KO7 helper phage to yield recombinant phage antibody ScFvs. After two rounds of panning with cell line SW480 highly expressing MC5-binding antigen, the phage clones displayed ScFv fragments of the antibody were selected by ELISA, and the affinity of the positive phage clones was assayed by competition ELISA.

RESULTS

The VH, VL, and ScFv DNAs were about 340 bp, 320 bp, and 750 bp, respectively. Ten phage clones displayed ScFv of MC5 were selected from 25 enriched phage clones, and 3 of the 10 phage clones had higher affinity of binding to the antigen.

CONCLUSION

The phage-displayed ScFv fragments of monoclonal antibody MC5 are successfully produced by phage display technique, which may provide a way for broadening the application range of the antibody.

Authors+Show Affiliations

Department of Biochemistry and Molecular Biology, Third Military Medical University, Chongqing 400038, P. R. China. hefengtian@163.netNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

English Abstract
Journal Article

Language

chi

PubMed ID

12452065

Citation

He, Feng-tian, et al. "[Preparation of Phage-displayed Single Chain Variable Fragments of Monoclonal Antibody MC5 Recognizing Colorectal Carcinoma]." Ai Zheng = Aizheng = Chinese Journal of Cancer, vol. 21, no. 6, 2002, pp. 636-9.
He FT, Li RF, Zhang Y, et al. [Preparation of phage-displayed single chain variable fragments of monoclonal antibody MC5 recognizing colorectal carcinoma]. Ai Zheng. 2002;21(6):636-9.
He, F. T., Li, R. F., Zhang, Y., Ji, Q., Chen, B. J., Qiao, T. D., & Fan, D. M. (2002). [Preparation of phage-displayed single chain variable fragments of monoclonal antibody MC5 recognizing colorectal carcinoma]. Ai Zheng = Aizheng = Chinese Journal of Cancer, 21(6), 636-9.
He FT, et al. [Preparation of Phage-displayed Single Chain Variable Fragments of Monoclonal Antibody MC5 Recognizing Colorectal Carcinoma]. Ai Zheng. 2002;21(6):636-9. PubMed PMID: 12452065.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - [Preparation of phage-displayed single chain variable fragments of monoclonal antibody MC5 recognizing colorectal carcinoma]. AU - He,Feng-tian, AU - Li,Rong-fen, AU - Zhang,Yan, AU - Ji,Qing, AU - Chen,Bao-jun, AU - Qiao,Tai-dong, AU - Fan,Dai-ming, PY - 2002/11/28/pubmed PY - 2003/1/10/medline PY - 2002/11/28/entrez SP - 636 EP - 9 JF - Ai zheng = Aizheng = Chinese journal of cancer JO - Ai Zheng VL - 21 IS - 6 N2 - BACKGROUND & OBJECTIVE: MC5 is a murine monoclonal antibody with a good specificity to human colorectal carcinoma and smaller murine antibody can significantly decrease the possibility of developing human antimouse antibody response in vivo study. The aim of this study was to prepare single chain variable fragments (ScFv) of MC5. METHODS: mRNA was isolated from the hybridoma cell line producing MC5, and the DNAs encoding variable domains of heavy and light chains(VH and VL DNAs) of the antibody were amplified separately by RT-PCR and assembled into ScFv DNAs with a linker DNA. The ScFv DNAs were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into E. coli TG1. The transformed cells were infected with M13KO7 helper phage to yield recombinant phage antibody ScFvs. After two rounds of panning with cell line SW480 highly expressing MC5-binding antigen, the phage clones displayed ScFv fragments of the antibody were selected by ELISA, and the affinity of the positive phage clones was assayed by competition ELISA. RESULTS: The VH, VL, and ScFv DNAs were about 340 bp, 320 bp, and 750 bp, respectively. Ten phage clones displayed ScFv of MC5 were selected from 25 enriched phage clones, and 3 of the 10 phage clones had higher affinity of binding to the antigen. CONCLUSION: The phage-displayed ScFv fragments of monoclonal antibody MC5 are successfully produced by phage display technique, which may provide a way for broadening the application range of the antibody. UR - https://www.unboundmedicine.com/medline/citation/12452065/[Preparation_of_phage_displayed_single_chain_variable_fragments_of_monoclonal_antibody_MC5_recognizing_colorectal_carcinoma]_ L2 - https://medlineplus.gov/colorectalcancer.html DB - PRIME DP - Unbound Medicine ER -