Tertiary structure changes in plasmid DNA, induced by ethidium bromide intercalation, have been observed in aqueous solutions by the use of an atomic force microscope. A relaxed closed circular pBR322 molecule became a positively supercoiled complex on the drug binding. The supercoiling always resulted in an interwound (or a plectonemic) form, but never a solenoidal (or a toroidal) form. A quantitative analysis of the compactness of such supercoiled complexes has been carried out.