Abstract: The synthesis of 10,10'-substituted-9,9'-biacridine molecules and their derivatives is disclosed. These molecules are shown to catalyze the production of light by chemiluminescence in the presence of a signal solution having at a pH from about 10.0 to about 14.0, at a concentration effective for producing a chemiluminescent signal, a chelating agent, a sulfoxide, a reducing sugar, an oxidant or combination of oxidants, an alcohol and aqueous sodium tetraborate. These 10,10'-substituted-9,9'-biacridines are used alone or attached to haptens or macromolecules and are utilized as labels in the preparation of chemiluminescent, homogeneous or heterogeneous assays. They are also used in conjunction with other chemiluminescent label molecules to produce multiple analyte chemiluminescent assays.