Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.

Recombinant single-chain fragments (scFv) of the murine anti-MUC1 monoclonal antibody C595 have been produced using the original hybridoma cells as a source of variable heavy (V(H))- and variable light (V(L))-chain-encoding antibody genes. The use of the polymerase chain reaction (PCR), bacteriophag...

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Autores principales: Denton, G., Sekowski, M., Spencer, D. I., Hughes, O. D., Murray, A., Denley, H., Tendler, S. J., Price, M. R.
Formato: Texto
Lenguaje:English
Publicado: Nature Publishing Group 1997
Materias:
Research Article
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2228010/
https://www.ncbi.nlm.nih.gov/pubmed/9303360
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author Denton, G.
Sekowski, M.
Spencer, D. I.
Hughes, O. D.
Murray, A.
Denley, H.
Tendler, S. J.
Price, M. R.
author_facet Denton, G.
Sekowski, M.
Spencer, D. I.
Hughes, O. D.
Murray, A.
Denley, H.
Tendler, S. J.
Price, M. R.
author_sort Denton, G.
collection PubMed
description Recombinant single-chain fragments (scFv) of the murine anti-MUC1 monoclonal antibody C595 have been produced using the original hybridoma cells as a source of variable heavy (V(H))- and variable light (V(L))-chain-encoding antibody genes. The use of the polymerase chain reaction (PCR), bacteriophage (phage) display technology and gene expression systems in E. coli has led to the production of soluble C595 scFv. The scFv has been purified from the bacterial supernatant by peptide epitope affinity chromatography, leading to the recovery of immunoreactive C595 scFv, which was similar in activity to the C595 parent antibody. Analysis by DNA sequencing, SDS-PAGE and Western blotting has demonstrated the integrity of the scFv, while ELISA, FACScan analysis, fluorescence quenching, quantitative immunoreactivity experiments and immunohistochemistry confirm that the activity of the scFv compares favourably with that of the parent antibody. The retention of binding activity to MUC1 antigen on human bladder and breast carcinoma tissue specimens illustrates the potential application of this novel product as an immunodiagnostic and immunotherapeutic reagent. IMAGES:
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spelling pubmed-22280102009-09-10 Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas. Denton, G. Sekowski, M. Spencer, D. I. Hughes, O. D. Murray, A. Denley, H. Tendler, S. J. Price, M. R. Br J Cancer Research Article Recombinant single-chain fragments (scFv) of the murine anti-MUC1 monoclonal antibody C595 have been produced using the original hybridoma cells as a source of variable heavy (V(H))- and variable light (V(L))-chain-encoding antibody genes. The use of the polymerase chain reaction (PCR), bacteriophage (phage) display technology and gene expression systems in E. coli has led to the production of soluble C595 scFv. The scFv has been purified from the bacterial supernatant by peptide epitope affinity chromatography, leading to the recovery of immunoreactive C595 scFv, which was similar in activity to the C595 parent antibody. Analysis by DNA sequencing, SDS-PAGE and Western blotting has demonstrated the integrity of the scFv, while ELISA, FACScan analysis, fluorescence quenching, quantitative immunoreactivity experiments and immunohistochemistry confirm that the activity of the scFv compares favourably with that of the parent antibody. The retention of binding activity to MUC1 antigen on human bladder and breast carcinoma tissue specimens illustrates the potential application of this novel product as an immunodiagnostic and immunotherapeutic reagent. IMAGES: Nature Publishing Group 1997 /pmc/articles/PMC2228010/ /pubmed/9303360 Text en https://creativecommons.org/licenses/by/4.0/This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit https://creativecommons.org/licenses/by/4.0/.
spellingShingle Research Article
Denton, G.
Sekowski, M.
Spencer, D. I.
Hughes, O. D.
Murray, A.
Denley, H.
Tendler, S. J.
Price, M. R.
Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title_full Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title_fullStr Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title_full_unstemmed Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title_short Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.
title_sort production and characterization of a recombinant anti-muc1 scfv reactive with human carcinomas.
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2228010/
https://www.ncbi.nlm.nih.gov/pubmed/9303360
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