Abstract The reaction of anthramycin with DNA was studied with respect to the nature of the binding and the selectivity of the reaction for various forms of nucleic acid. The binding appears to involve the formation of a labile covalent bond between anthramycin and DNA. The following observations are consistent with this conclusion: 1. (1) The complex resists dissociation by dialysis, gel filtration, alcohol precipitation, sodium lauryl sulfate or silver ion. 2. (2) The native DNA complex can be denatured by alkali, and the anthramycin remains bound to the separated strands (under conditions which would not allow direct reaction between anthramycin and denatured DNA). 3. (3) The reaction is relatively slow, and the rate is proportional to hydrogen ion concentration. 4. (4) The complex dissociates at a measurable rate below pH 3. 5. (5) Bound anthramycin has enhanced stability with respect to decomposition by heat or alkali. The reaction is highly selective for native DNA. RNA and deoxymononucleotides do not react. The helical RNA-like polymer, rG:rC, failed to react, whereas dG:dC reacted very well. In the case of DNA altered by heat-denaturation or by extensive alkylation, the rates of reaction with anthramycin were markedly reduced, but only minor reductions in numbers of binding sites were detected. The dependence of the reaction rates on ionic strength indicated that the reaction with denatured DNA is sensitive to conformation. Selectivity was also noted with regard to base sequence.