Ellipticine (NSC 71795), a plant alkaloid with antitumor activity, is a weakly basic polycyclic molecule with dimensions similar to those of proflavin. Like proflavin, ellipticine exhibits hypocromic and bathochromic changes in absorption spectrum in the presence of DNA. It binds preferentially to helical DNA by intercalation, but the strength of binding is substantially greater than that of proflavin. The evidence for intercalation is based on effects on the sedimentation and viscosity of sheared DNA fragments, removal and reversal of the supercoiling of closed circular DNA, and electric dichroism measurements. The sedimentation and viscosity changes are quantitatively similar to those produced by proflavin. The unwinding angle on binding to supercoiled DNA is estimated to be 7.9 degrees, similar to that of proflavin. Electric dichroism shows the plane of the bound ellipticine molecule to be oriented parallel (plus or minus 7 degrees) to the plane of the bases in helical DNA. Ellipticine differs from proflavin in that it is uncharged at neutral pH and becomes protonated under mildly acid conditions. This feature may influence the intracellular distribution of the drug. Ellipticine bound to DNA is probably in its protonated form.