We succeeded in forming a highly dense and uniform self-assembled monolayer (SAM) composed of thiolated double-stranded oligonucleotide (HS-dsDNA). The HS-dsDNA SAMs were prepared on gold substrates, and were characterized by surface plasmon resonance (SPR), X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). Formation of highly dense and uniform HS-dsDNA SAM was confirmed by the SPR and AFM measurements. XPS measurements bore out existence of the S-Au bond.