Phage-displayed Fab fragments against anti-human interleukin-2 receptor alpha. Detection of antigen-bound phages with anti-cpIII monoclonal antibodies.

The genes encoding the VHCH1 and VLCL parts of the mouse anti-human IL-2R alpha antibody 7G7B6 were amplified by PCR and the corresponding antibody fragments displayed on the surface of filamentous phages. The expression of Fab fragments was analysed by immunoblotting using HRP-labelled goat anti-mouse Ig antisera. By traditional hybridoma technology, splenocytes from Balb/c mice, immunized with native phage particles, were fused with P3X63-Ag8.653 myeloma cells in order to yield monoclonal antibodies against filamentous phage proteins. The obtained monoclonal antibody IF8 (mu/kappa) recognized the minor coat protein III as a 65-70 kDa protein band by immunoblotting, whereas the monoclonal antibody IVC8 (mu/kappa), in addition to cpIII, recognized a protein with an approximate molecular weight of 38-43 kDa. Both antibodies were employed to determine the binding specificity of the phage-displayed anti-human IL-2R alpha Fab fragments in an ELISA using recombinant baculovirus-expressed human IL-2R alpha proteins as antigens.