Hybridization of 12-mers peptide nucleic acid (PNA) to complementary DNA was investigated in solution and on gold surfaces. The oligomers were designed to improve mismatch discrimination and minimize formation of secondary structures. Thermal denaturation experiments indicate high thermal stabilities for PNA-DNA hybrid with T_m values close to calculated values. Hybridization of PNA-DNA at 45°C and room temperature showed no difference. Hybridization on gold surface was also investigated with complementary and noncomplementary DNAs. The results show that 12-mer PNA and DNA hybridization at room temperature retained high specificity within ∼5 ng.