Mitoxantrone (MTX) is a clinically used antitumor anthracycline, which is transported to the target tissues by human serum albumin (HSA). Being less toxic unlike other member of this family, its binding characteristics are therefore of immense interest. The protein and the ligand were prepared with the aid of CORINA, protonated with insight II and best conformation was sought by employing Gold V. By docking procedure, site III has been assigned to posses the binding site for MTX with the binding affinity (Ka) = 1.58 x 106 mol-1. Molecular docking calculations placed MTX at digitoxin binding site of HSA. The interaction was found to be thermodynamically favourable (ÄG° = -35.53 KJmol-1). Further analysis of the MTX binding site on to the HSA suggested that the type of interactions that contribute in this binding are hydrophobic contacts, hydrogen bonding and electrostatic interactions. This study presents binding mechanism in a unified way that is simple, yet stringent, more straightforward, more reliable and informative.