An understanding of DNA-ligand, interactions at the molecular level is important for the design of new drugs and probes that can recognize specific DNA sequences and structural motifs. Interestingly, determining the mode-of-binding of a DNA ligand is not always straightforward due to uncertainties inherent in traditional assays. We have recently reported an exciting new assay utilizing scanning force microscopy (SFM) that can discern whether a ligand binds to DNA by intercalative or nonintercalative modes [Coury et al, Proc. Natl. Acad. Sc; U.S.A. 1996, 93, 12283-12286]. Visualization of individual DNA molecules by SFM and observation of ligand-induced lengthening provides direct evidence for intercalation. Metal complexes of polypyridyl ligands have been extensively studied as new probes of DNA structure and function because they exist tie chiral molecules with the potential of enantioselective recognition of DNA, The binding mode of even the most. widely studied of the members of this group, tris(o-phenanthroline)ruthenium(II) (Ru(phen)(3)(2+)), remains somewhat controversial due in large part to its low binding affinity. We report here the use of Ru(phen)(3)(2+) as a test of our new assay toward the studies of weakly-binding ligands and to resolve the ambiguity surrounding the mode-of-binding of Delta and Lambda-Ru(phen)(3)(2+). Experiments reported here reveal that the experimental conditions of our assay do not preclude the binding of Ru(phen)(3)(2+) to DNA and that NO lengthening occurs. Our findings are consistent with the view that Ru(phen)(3)(2+) binds to duplex nucleic acids through nonintercalative modes.