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Use of antibody gene library for the isolation of specific single chain antibodies. by ampicillin-antigen conjugates

  • Isolation of recombinant antibodies from antibody libraries is commonly performed by different molecular display formats including phage display and ribosome display or different cell-surface display formats. We describe a new method which allows the selection of Escherichia coil cells producing the required single chain antibody by cultivation in presence of ampicillin conjugated to the antigen of interest. The method utilizes the neutralization of the conjugate by the produced single chain antibody which is secreted to the periplasm. Therefore, a new expression system based on the pET26b vector was designed and a library was constructed. The method was successfully established first for the selection of E. coli BL21 Star (DE3) cells expressing a model single chain antibody (anti-fluorescein) by a simple selection assay on LB-agar plates. Using this selection assay, we could identify a new single chain antibody binding biotin by growing E. coil BL21 Star (DE3) containing the library in presence of a biotin-ampicillin conjugate. InIsolation of recombinant antibodies from antibody libraries is commonly performed by different molecular display formats including phage display and ribosome display or different cell-surface display formats. We describe a new method which allows the selection of Escherichia coil cells producing the required single chain antibody by cultivation in presence of ampicillin conjugated to the antigen of interest. The method utilizes the neutralization of the conjugate by the produced single chain antibody which is secreted to the periplasm. Therefore, a new expression system based on the pET26b vector was designed and a library was constructed. The method was successfully established first for the selection of E. coli BL21 Star (DE3) cells expressing a model single chain antibody (anti-fluorescein) by a simple selection assay on LB-agar plates. Using this selection assay, we could identify a new single chain antibody binding biotin by growing E. coil BL21 Star (DE3) containing the library in presence of a biotin-ampicillin conjugate. In contrast to methods as molecular or cell surface display our selection system applies the soluble single chain antibody molecule and thereby avoids undesired effects, e.g. by the phage particle or the yeast fusion protein. By selecting directly in an expression strain, production and characterization of the selected single chain antibody is possible without any further cloning or transformation steps.show moreshow less

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Metadaten
Author:Meina Neumann-Schaal, Katrin Messerschmidt, Nicole Grenz, Katja Heilmann
DOI:https://doi.org/10.1016/j.imlet.2013.02.005
ISSN:0165-2478 (print)
Parent Title (English):Immunology letters : an international journal providing for the rapid publication of short reports in immunology
Publisher:Elsevier
Place of publication:Amsterdam
Document Type:Article
Language:English
Year of first Publication:2013
Year of Completion:2013
Release Date:2017/03/26
Tag:Anti-biotin antibody; Naive single chain library; Selection method; Single chain antibody
Volume:151
Issue:1-2
Pagenumber:5
First Page:39
Last Page:43
Funder:Federal Ministry of Education and Research [InnoProfile 703]
Organizational units:Mathematisch-Naturwissenschaftliche Fakultät / Institut für Biochemie und Biologie
Peer Review:Referiert