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Determining the mode-of-binding of a DNA ligand is not always straightforward. Here, we establish a scanning force microscopic assay for mode-of-binding that is (i) direct: lengths of individual DNA-ligand complexes are directly measured; (ii) rapid: there are no requirements for staining or elaborate sample preparation; and (iii) unambiguous: an observed increase in DNA length upon addition of a ligand is definitive evidence for an intercalative mode-ofbinding. Mode-of-binding, binding affinity, and site-exclusion number are readily determined from scanning force microscopy measurements of the changes in length of individual drug-DNA complexes as a function of drug concentration. With this assay, we resolve the ambiguity surrounding the mode of binding of 2,5-bis(4-amidinophenyl)furan (APF) to DNA and show that it binds to DNA by nonintercalative modes. APF is a member of an important class of aromatic dicationic drugs that show significant activity in the treatment of Pneumocystis carinii pneumonia, an opportunistic infection that is the leading cause of death in AIDS patients.Nucleic acid ligands have been discovered and successfully engineered to act as anticancer drugs (1), probes of nucleic acid damage and structure (2), and as sequence-specific binding and cleavage agents (3, 4). Nucleic acid ligands are used in the treatment of genetic, oncogenic, and viral diseases. A molecular level understanding of nucleic acid interactions is a minimum first step in the development of useful new ligands.Ligands bind to DNA (i) by intercalation, (ii) within the major or minor grooves, (iii) by "nonclassical" modes (5), or (iv) by a combination of these. Definitive assays for mode-ofbinding are three-dimensional structure determination by x-ray diffraction or NMR spectroscopy. However, these structural techniques are labor-intensive, are often precluded by lack of site-specificity, rapid exchange, or multiple binding modes, and are limited to short DNA fragments. Assays suitable to long DNA fragments involve viscometry, sedimentation, and linear and circular dichroism. These methods are reliable when ligands bind by conventional intercalative or minor groove modes (6), but can be confounded by mixed and nonclassical modes. Difficulties arise in part because these assays are indirect and inferential.Here we establish scanning force microscopy (SFM) as a direct, rapid, and unambiguous assay for mode-of-binding conventional and nonclassical ligands to DNA. SFM has been shown previously to be a high-resolution method for imaging nucleic acids (7-10). We have demonstrated that the site of binding of intercalators can be pinpointed with SFM (11).Intercalators stack between base pairs, lengthening DNA by an amount equivalent to the van der Waals thickness of each intercalating moiety (3.4 A) (12). A ligand that does not lengthen DNA does not intercalate. Our SFM assay directly measures the lengths of individual DNA molecules. Increases in DNA length upon ligand binding provides direct evidence for intercalation. Bindi...

This work presents a model to predict the effect of surface stresses on the ith-mode bending resonant frequency of microcantilevers and its experimental validation. With this model, one can calculate the surface stress acting upon the microcantilever solely by measuring resonant frequencies whereas previously one needed to measure the deflection. Resonant frequency measurement has distinct advantages in terms of ease and accuracy of measurement.

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The first, systematic study of the interacting effects of solvent and axially coordinated monovalent anions on the electroreduction mechanisms and redox potentials of iron porphyrins is presented. Five different anions were coordinated to (5,10,15,20tetraphenylporphinato)iron(III), and their respective redox reactions were investigated in 12 nonaqueous solvents. Potential shifts with changes in solvent were directly related to solvent donicity. Potential shifts with changes in counterion were related to the degree of iron(III)-counterion interaction and were also found to be solvent dependent. The counterion is the predominant factor governing the type of electron-transfer mechanism observed for the electrode reactions of iron porphyrins.With the results obtained, the apparent independence of redox potentials of the iron-center spin state is established.