Antibody Phage Display

Methods and Protocols

Philippa M. O’Brien, Robert Aitken (Herausgeber)

Buch | Softcover

401 Seiten

2001
Humana Press Inc. (Verlag)
978-0-89603-711-3 (ISBN)

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The closing years of the 19th century and the start of the 20th century witnessed the emergence of microbiology and immunology as discrete sci- tific disciplines, and in the work of Roux and Yersin, perhaps the first benefits of their synergy—immunotherapy against bacterial infection. As we advance into the new millennium, microbiology and immunology again offer a c- ceptual leap forward as antibody phage display gains increasing acceptance as the definitive technology for monoclonal production and unleashes new - portunities in immunotherapy, drug discovery, and functional genomics. In assembling Antibody Phage Display: Methods and Protocols, we have aimed to produce a resource of real value for scientists who have followed the development of phage display technology over the past decade. The founding principles of phage display have always held an elegant simplicity. We hope that readers will find similar clarity in the technical guidance offered by the book’s contributors. In meeting our objectives, we have tried to cover the broad scope of the technology and the key areas of library construction, scre- ing, antibody modification, and expression. Of course, the technology cont- ues to advance apace, but we trust that readers will be able to gage the potential of phage display from our coverage, that some of its subtleties will emerge, and that our selection of methods will prove appealing. We are indebted to all the contributing authors for sharing their expertise with the wider scientific community.

Overview of Antibody Phage-Display Technology and Its Applications.- Standard Protocols for the Construction of Fab Libraries.- Standard Protocols for the Construction of scFv Libraries.- Broadening the Impact of Antibody Phage Display Technology.- Construction of Large Naïve Fab Libraries.- Construction of Polyclonal Antibody Libraries Using Phage Display.- Antigen-Driven Stimulation of B-Lymphocytes In Vitro.- 8 The Recovery of Immunoglobulin Sequences from Single Human B Cells by Clonal Expansion.- Panning of Antibody Phage-Display Libraries.- Selection of Antibodies Against Biotinylated Antigens.- Isolation of Anti-Hapten Specific Antibody Fragments from Combinatorial Libraries.- Blocking Immunodominant Epitopes by Competitive Deselection.- Rescue of a Broader Range of Antibody Specificities Using an Epitope-Masking Strategy.- Screening of Phage-Expressed Antibody Libraries by Capture Lift.- Antibody-Guided Selection Using Capture-Sandwich ELISA.- Proximity-Guided (ProxiMol) Antibody Selection.- Isolation of Human Monoclonal Antibodies Using Guided Selection with Mouse Monoclonal Antibodies.- Selecting Antibodies to Cell-Surface Antigens Using Magnetic Sorting Techniques.- Isolation of Human Tumor-Associated Cell Surface Antigen-Binding scFvs.- Subtractive Isolation of Single-Chain Antibodies Using Tissue Fragments.- Selection of Antibodies Based on Antibody Kinetic Binding Properties.- Selection of Functional Antibodies on the Basis of Valency.- Two-Step Strategy for Alteration of Immunoglobulin Specificity by In Vitro Mutagenesis.- Targeting Random Mutations to Hotspots in Antibody Variable Domains for Affinity Improvement.- Error-Prone Polymerase Chain Reaction for Modification of scFvs.- Use of Escherichia coli Mutator Cells to Mature Antibodies.- ChainShuffling to Modify Properties of Recombinant Immunoglobulins.- Generation of Bispecific and Tandem Diabodies.- High-Level Periplasmic Expression and Purification of scFvs.- Periplasmic Expression and Purification of Recombinant Fabs.- Expression of Antibody Fragments in Pichia pastoris.- Expression of VHH Antibody Fragments in Saccharomyces cerevisiae.- Intrabodies.- 34 Expression of scFvs and scFv Fusion Proteins in Eukaryotic Cells.- 35 Expression of Antibody Fab Fragments and Whole Immunoglobulin in Mammalian Cells.

Erscheint lt. Verlag	28.12.2001
Reihe/Serie	Methods in Molecular Biology ; 178
Zusatzinfo	XV, 401 p.
Verlagsort	Totowa, NJ
Sprache	englisch
Maße	155 x 235 mm
Themenwelt	Naturwissenschaften ► Biologie ► Biochemie
Themenwelt	Naturwissenschaften ► Biologie ► Mikrobiologie / Immunologie
ISBN-10	0-89603-711-8 / 0896037118
ISBN-13	978-0-89603-711-3 / 9780896037113
Zustand	Neuware