USE OF AN ALBUMIN-BINDING DOMAIN FOR THE SELECTIVE IMMOBILIZATION OF RECOMBINANT CAPTURE ANTIBODY FRAGMENTS ON ELISA PLATES

A small albumin-binding domain (ABD) of 46 amino acids derived from st reptococcal protein G was employed for the directed attachment of reco mbinant immunoglobulin (Ig) fragments to microtitre plates that had be en coated with human serum albumin (HSA). Generic vectors were constru cted in order to produce the Fv or Fab fragments fused with the ABD in Escherichia coli. Using the anti-lysozyme antibody D1.3 as the captur e antibody fragment it was possible to quantify the non-radioactively labelled antigen with high sensitivity in a sandwich ELISA. The new st rategy avoids denaturation or an unfavourable orientation of the Ig fr agment, which can occur during direct adsorption to the microtitre pla te. The HSA that serves to complex the ABD ensures efficient saturatio n of reactive binding sites on the plastic surface as well so that no additional blocking steps are necessary and the assay can be quickly p erformed. (C) 1998 Elsevier Science B.V. All rights reserved.