The DNA interaction of a chiral Troger base derived from proflavine was investigated by DNA melting temperature measurements and complementary biochemical assays. DNase I footprinting experiments demonstrate that the binding of the proflavine-based Troger base is both enantio- and sequence-specific. The (+)-isomer poorly interacts with DNA in a non-sequence-selective fashion. In sharp contrast, the corresponding (-)-isomer recognizes preferentially certain DNA sequences containing both A . T and G . C base pairs, such as the motifs 5'-GTT . AAC and 5'-ATGA . TCAT. This is the first experimental demonstration that acridine-type Troger bases can be used for enantiospecific recognition of DNA sequences,