WP2: Generation of protein-binding affinity reagents

The central objective of the programme is to generate a resource of affinity reagents to all protein targets in the five related categories. Binder types will be: affinity-purified polyclonal rabbit antibodies (KTH); mouse monoclonal antibodies (EMBL-MACF, HMGU); recombinant human single chain antibodies (BBT,ULUND,TUBS) and camelid VH nanobodies (VIB); and two recombinant protein scaffolds, DARPins (ankyrin repeats, UZH) and Affibody binders (protein A scaffolds, KTH). Recombinant systems include selection of antibody fragments (scFv, VH/K, VH) from synthetic, naive, or immune (camelid) libraries by phage display (ULUND, TUBS,VIB) and ribosome display (BBT), the latter of which will also be used for binder improvement by in vitro molecular evolution. Alternative binders built on the DARPin and Affibody molecular scaffolds will be selected by ribosome display (UZH) or phage display (KTH) respectively. The producing groups will carry out initial validations using ELISA and other assays, preparatory to QC in WP3. Efficiency of recombinant selection processes will be increased in WP4 and the improved methods employed in the later part of WP2.