Immunological detection of viruses and their components by monoclonal antibodies is a powerful method for studying the structure and function of viral molecules. Here we describe detailed methods for establishing monoclonal antibodies against severe acute respiratory syndrome coronavirus (SARS-CoV). B cell hybridomas are generated from mice that are hyperimmunized with inactivated SARS-CoV virions. The hybridomas produce monoclonal antibodies that recognize viral component molecules, including the spike protein (S) and the nucleocapsid protein (N), enabling the immunological detection of SARS-CoV by immunofluorescence staining, immunoblot, or an antigen-capture ELISA system. In addition, several S protein-specific antibodies are shown to have in vitro neutralization activity. Thus the monoclonal antibody approach provides useful tools for rapid and specific diagnosis of SARS, as well as for possible antibody-based treatment of the disease.