The interaction between human heart myoglobin and ten specific monoclonal antibodies was investigated with a new biosensor technology, real time biospecific interaction analysis (RT BIA), using surface plasmon resonance. Analysis of association and dissociation kinetics was monitored in real time, with unlabelled reactants. Antibody isotyping was rapid and simple. Epitope mapping with RT BIA confirmed, with substantial time saving, the sum of results obtained in conventional labelled systems. Monoclonal antibodies with four different epitope specificities and optimal binding function were selected for a myoglobin sandwich assay with enhanced sensitivity. BIAcore can be used directly as a diagnostic tool, or as an analytical tool in immunoassay development.