Isolation of mutants of Aujeszky's disease virus with antigenically altered glycoprotein E by affinity chromatography using monoclonal antibodies

An efficient method for isolation of virus mutants with antigenically alter ed proteins is described. The method is based on the separation of viruses with wild-type and antigenically altered proteins by affinity chromatograph y using monoclonal antibodies (MAbs). A nonessential glycoprotein E (gE) of Aujeszky's disease virus (ADV) was chosen as a model for introducing the a ntigenic changes. The ADV strain Ka mutagenised with 5-bromo-2'-deoxyuridin e was used for the selection of mutants that do not bind to gE-specific MAb conjugated to resin. After three rounds of isolation by affinity chromatog raphy, the resulting viruses that escape the binding to MAb were plaque-pur ified by plating at limiting dilution, and virus isolates were tested by th e gE-specific sandwich ELISA in which the selecting MAb was used as a captu re antibody. About 70% of the ADV isolates tested were not recognised by th e sandwich gE-ELISA. The analysis of some of virus isolates in indirect ELI SA with a panel of 16 gE-specific MAbs revealed that at least several of th e generated virus isolates were mutants expressing gE with alterations in t he epitope of the selecting MAb 75/7, as well as in the majority of other c onformation-dependent epitopes of gE. The method for the production of anti genically altered viruses by affinity chromatography using MAbs is simple a nd convenient, and can be utilised with MAbs irrespective of their virus-ne utralising activity. (C) 1999 Elsevier Science B.V. All rights reserved.