We describe analyses of unusual human alpha-amylase, performed in our laboratory and review available methods for amylase study. Electrophoretic and chromatographic methods provide an effective means for the analysis of amylase isoenzymes and unusual amylase. The recent identification of a selective inhibitor and a monoclonal antibody to amylase isoenzyme contributes to rapid routine clinical assays of amylase isoenzymes. However unusual amylases such as variants, macroamylasemia and sialyl salivary-type amylasemia cannot be detected by those conventional methods. The unusual amylases can only be detected by electrophoresis and can be easily characterized by combination study with chromatographic methods. Electrophoretic and chromatographic methods are universal means to validate unusual amylases found in patient sera. Further basal studies are needed to define the roles of salivary amylase in exocrine fluids using those separation techniques.