Because the selection process takes place in vitro, as opposed to animal immunization where the immune response is not under control of the experimentator, the panning conditions can be adjusted to drive the selection toward desired antibody properties. Typical examples for guided selections include the way of presenting the antigen to the phage library (native, denatured, captured, masked), the stringency of washing (which determines the affinity range of enriched antibodies), the method of phage elution (e.g., by competition with free antigen), the simulation of assay conditions during panning (e.g., by adding denaturants to select for highly stable antibodies, see ref. 9 as an example), the selection of cross-reactive antibody by alteration of two or more antigens during the panning rounds, and last but not least the selection of epi-tope-specific antibodies by blocking or subtracting the library with a related antigen. The latter can be used, for instance, to select phospho-specific antibodies (see Fig. 2). Such methods have been shown to reliably enrich desired antibody properties, thereby dramatically reducing the subsequent screening effort.
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