title: Use of micro-emulsion technology for the directed evolution of antibodies.
abstract: Affinity reagents, such as antibodies, are needed to study protein expression patterns, sub-cellular localization, and post-translational modifications in complex mixtures and tissues. Phage Emulsion, Secretion, and Capture (ESCape) is a novel micro-emulsion technology that utilizes water-in-oil (W/O) emulsions for the identification and isolation of cells secreting phage particles that display desirable antibodies. Using this method, a large library of antibody-displaying phage will bind to beads in individual compartments. Rather than using biopanning on a large mixed population, phage micro-emulsion technology allows us to individually query clonal populations of amplified phage against the antigen. The use of emulsions to generate microdroplets has the promise of accelerating phage selection experiments by permitting fine discrimination of kinetic parameters for binding to targets. In this study, we demonstrate the ability of phage micro-emulsion technology to distinguish two scFvs with a 300-fold difference in binding affinities (100nM and 300pM, respectively). In addition, we describe the application of phage micro-emulsion technology for the selection of scFvs that are resistant to elevated temperatures.
authors: AxioMx, Inc., 688 E. Main St., Branford, CT 06405, United States.
authors: Diane L
authors: DL
authors: Buhr
authors: Felicity E
authors: FE
authors: Acca
authors: Erika G
authors: EG
authors: Holland
authors: Katie
authors: K
authors: Johnson
authors: Gail M
authors: GM
authors: Maksymiuk
authors: Ada
authors: A
authors: Vaill
authors: Brian K
authors: BK
authors: Kay
authors: David A
authors: DA
authors: Weitz
authors: Michael P
authors: MP
authors: Weiner
authors: Margaret M
authors: MM
authors: Kiss
mesh_terms: D000915:Antibody Affinity; D017104:Bacteriophage M13 / Q000235:genetics; D061505:Cell Surface Display Techniques; D019020:Directed Molecular Evolution; D004655:Emulsions; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007700:Kinetics; D016296:Mutagenesis; D019151:Peptide Library; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D015202:Protein Engineering; D055550:Protein Stability; D057127:Single-Chain Antibodies / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*
chemical_list: D004655:Emulsions; D019151:Peptide Library; D057127:Single-Chain Antibodies