A thermodynamic and kinetic study on the mode of binding of 9-amino-6-chloro-2-methoxi-acridine (ACMA) to poly(dA–dT)·poly(dA–dT) and poly(dG–dC)·poly(dG–dC) has been undertaken at pH = 7.0 and I = 0.1 M. The spectrophotometric, kinetic (T-jump), circular dichroism, viscometric and calorimetric information gathered point to formation of a fully intercalated ACMA complex with poly(dA–dT)·poly(dA–dT) and another one only partially intercalated (7%) with poly(dG–dC)·poly(dG–dC). The ACMA affinity with the A–T bases was higher than with the G–C bases. The two polynucleotide sequences give rise to external complexes when the ACMA concentration is raised, namely, the electrostatic complex poly(dA–dT)·poly(dA–dT)–ACMA and the major groove binding complex poly(dG–dC)·poly(dG–dC)–ACMA. A considerable quenching effect of the ACMA fluorescence is observed with poly(dA–dT)·poly(dA–dT), ascribable to face-to-face location in the intercalated A–T–ACMA base-pairs. The even stronger effect observed in the presence of poly(dG–dC)·poly(dG–dC) is related to the guanine residue from on- and off-slot ACMA positions.