Journal
NATURE PROTOCOLS
Volume 3, Issue 11, Pages 1766-1777Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nprot.2008.176
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Here we describe a protocol for the selection of full-length IgG antibodies from repertoires displayed on Escherichia coli. In the method described here, full-length heavy and light chains are assembled in the periplasm into aglycosylated IgGs that are fully functional for antigen binding. Expression of an inner membrane-tethered Fc-binding protein is used to capture the IgG molecules and anchor them to the cell. Following outer-membrane permeabilization, fluorescently labeled ligand-binding library clones are selected by multiple rounds of fluorescence-activated cell sorting. Selection of a comprehensive set of IgG clones can typically be obtained within 3-4 weeks, a timescale that is comparable with most prevalent antibody display technologies. The isolated antibodies are well expressed in bacteria and exhibit affinities per binding site in the nanomolar range.
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