Phage antibodies

Sblattero, D., and Bradbury, A. (2000). Exploiting recombination in single bacteria to make large phage antibody libraries. Nat. Biotechnol. 18, 75-80. 

Model selections, in which phage antibodies with defined specificity are mixed with nonspecific phage and the enrichment and yield for a selection procedure is measured, provides a rapid experimental approach for studying such complex selection procedures. In order to compare different selection approaches on complex preparations, and to determine the best selection parameters for each approach, an extensive study of various in vitro and in vivo model selections was recently carried out by our group [73]. 

In immunocytochemistry, signals are usually amplified by means of catalyzed reporter deposition. In ProxiMol, catalyzed reporter deposition has been modified to permit the isolation of phage antibodies binding to near neighbor biotinylated guide molecule. Deposition of biotin molecule can be detected by electron microscopy. 

Phage antibodies are directly used to label tissue sections. Interpretation of the data is performed by a team of experts including histochemists and must be subjected to cross-examination. Images are captured using microscopes with a digital camera attachment and archived in databases for further analysis of pathologists. 

Waterhouse, P, Griffiths, A D., Johnson, K S., and Winter, G. (1993) Combinatorial infection and in vivo recombination, a strategy for making large phage antibody repertoires. Nucleic Acids Res 21,2265,2266. 

Hawkins, R. E., Russell, S. J., and Winter, G. (1992) Selection of phage antibodies by binding affinity mimicking affinity maturation J Mol Biol. 226, 889-896. 

McCafferty, J, Griffiths, A. D, Winter, G., and Chiswell, D. J (1990) Phage antibodies filamentous phage displaying antibody variable domains. Nature 348, 552-554... 

In this approach, the phage antibodies react with biotinylated antigen in solution, and the complex is then captured using Streptavidin coupled to Dynabeads. 

Polyclonal or monoclonal phage antibodies can be used as reagents to detect antigens on Western blots (see Chapter 20). Here we describe the detection of purified antigen, but the same protocol can be used for antigen present in complex protein mixtures. 

De Kruif, J., Terstappen, L, Boel, E, and Logtenberg, T (1995) Rapid selection of cell subpopulation-specific human monoclonal antibodies from a synthetic phage antibody library Proc Natl Acad Sci. USA 92, 3938—3942... 

Kretzschmar, T., Zimmermann, C, and Geiser, M (1995) Selection procedures for nonmatured phage antibodies—a quantitative comparison and optimization strategies. A nal Biochem 224,413-419. 

Phage antibodies filamentous phage displaying antibody variable domain. Nature 348, 552-554. 

Sheets, M. D., Amersdorfer, P., Finnem, R., Sargent, P., Lindqvist, E., Schier, R., et al. (1998) Efficient construction of a large nonimmune phage antibody library the production of high-affinity human single-chain antibodies to protein antigens. Proc. Natl. Acad. Sci. USA 95, 6157-6162. 

Chiswell DJ, McCafferty I, Phage antibodies Will new coliclonal antibodies replace monoclonal antibodies Trends Biotechnol., 10 80-84, 1992. 

McCafferty, Griffiths AD, Winter G, Chiswell DJ, Phage antibodies Filamentous phage displaying antibody variable domains, Nature, 348 552-554, 1990. 

Making and Selecting from Phage Antibody Libraries... 

Although a number of different phage (mid) antibody libraries have been published (see Table 18.1), the number which are naive is relatively small. To date, all phage antibody libraries have been made on the filamentous phage, Ff (which comprises fd, fl, and Ml3). In the creation of such libraries a number of different choices can be made ... 

The choice of which form of antibody fragment to use in a phage antibody library (Figure 18.5) is on the whole dictated by practical considerations (Table 18.4). 

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