Scanning electrochemical microscopic (SECM) imaging of oligonucleotides and polynucleotides (poly[G] and calf thymus DNA) immobilized onto aldehyde-modified glass substrates was achieved through oxidation of guanine residues on the DNA molecules by SECM tip-generated Ru(bpy)(3)(3+). Several parameters affecting the tip current associated with the guanine oxidation reaction (e.g. tip/substrate separation, the amount of guanine residues, and the voltammetric parameters) were investigated. The relationship between the tip current and the DNA concentration used for immobilization was also studied. The potential of utilizing the tip-generated Ru(bpy)(3)(3+) to image DNA hybridization was demonstrated. This study demonstrates that SECM imaging based on guanine oxidation is straightforward and sensitive and obviates the necessity of labeling the DNA targets with an electroactive marker.