Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

Production and characterization of anti‐(mucin MUC1) single‐domain antibody in tobacco ( Nicotiana tabacum cultivar Xanthi)

Production and characterization of anti‐(mucin MUC1) single‐domain antibody in tobacco (... Members of the Camelidae (camels, dromedaries, llamas, alpacas, guanacos and vicunas) are known to produce Igs (immunoglobulins) devoid of light chains and CH1s (constant heavy‐chain domains). The antigen‐specific binding fragments of these heavy‐chain antibodies therefore comprise one single domain (the so‐called ‘VHH’) and are of great importance in biotechnological applications. To evaluate the expression and biological activity of sdAbs (single‐domain antibodies) in plants, which, on account of their small size and antigen‐recognition properties, would have a major impact on antibody‐engineering strategies, we constructed a pBI121‐VHH gene encoding the recombinant sdAb fragments with specificity for a cancer‐associated mucin, MUC1. Analysis of transgenic tobacco (Nicotiana tabacum cultivar Xanthi) plants by PCR and Western blotting demonstrated the expression of sdAb, while ELISA results with various MUC1 antigens and immunocytochemistry with cancerous cell lines confirmed that the activity of these molecules compared favourably with that of the parent recombinant antibodies. Protein purification was achieved by using sequential (NH4)2SO4 precipitation, gel filtration and immunoaffinity chromatography. Analysis of the purified VHH by ELISA indicated that the purified antibody fragments were able to react successfully with a MUC1‐related peptide. These results reaffirm that the tobacco plant is a suitable host for the production of correctly folded VHH antibody fragments with diagnostic and therapeutic applications. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology and Applied Biochemistry Wiley

Production and characterization of anti‐(mucin MUC1) single‐domain antibody in tobacco ( Nicotiana tabacum cultivar Xanthi)

Download PDF
9 pages

Loading next page...
 
/lp/wiley/production-and-characterization-of-anti-mucin-muc1-single-domain-ffg5c6rmlj

References (3)

Publisher
Wiley
Copyright
2007 International Union of Biochemistry and Molecular Biology
ISSN
0885-4513
eISSN
1470-8744
DOI
10.1042/BA20060071
pmid
17073735
Publisher site
See Article on Publisher Site

Abstract

Members of the Camelidae (camels, dromedaries, llamas, alpacas, guanacos and vicunas) are known to produce Igs (immunoglobulins) devoid of light chains and CH1s (constant heavy‐chain domains). The antigen‐specific binding fragments of these heavy‐chain antibodies therefore comprise one single domain (the so‐called ‘VHH’) and are of great importance in biotechnological applications. To evaluate the expression and biological activity of sdAbs (single‐domain antibodies) in plants, which, on account of their small size and antigen‐recognition properties, would have a major impact on antibody‐engineering strategies, we constructed a pBI121‐VHH gene encoding the recombinant sdAb fragments with specificity for a cancer‐associated mucin, MUC1. Analysis of transgenic tobacco (Nicotiana tabacum cultivar Xanthi) plants by PCR and Western blotting demonstrated the expression of sdAb, while ELISA results with various MUC1 antigens and immunocytochemistry with cancerous cell lines confirmed that the activity of these molecules compared favourably with that of the parent recombinant antibodies. Protein purification was achieved by using sequential (NH4)2SO4 precipitation, gel filtration and immunoaffinity chromatography. Analysis of the purified VHH by ELISA indicated that the purified antibody fragments were able to react successfully with a MUC1‐related peptide. These results reaffirm that the tobacco plant is a suitable host for the production of correctly folded VHH antibody fragments with diagnostic and therapeutic applications.

Journal

Biotechnology and Applied Biochemistry – Wiley

Published: May 1, 2007

There are no references for this article.