Immobilization and hybridization of oligonucleotides on maleimido-terminated self-assembled monolayers.

Maleimido-terminated self-assembled monolayers were prepared via a one-step reaction of a maleimido-functionalized trichlorosilane with an oxide-covered surface. Through conjugate addition, thiol-tagged DNA was immobilized on these maleimido-functionalized self-assembled monolayers, and these immobilized oligonucleotides were further hybridized with complementary strands. The surface concentrations of oligonucleotides were quantified with radioactivity measurements. The surface concentrations of immobilized oligonucleotides increase in a nonlinear manner with increasing molar fraction of maleimido moieties on the surface and achieve high values of 1.6 (+/-0.6) x 10(13) oligonucleotides/cm2 if monolayers having full coverage of maleimido groups are used. High hybridization efficiencies of complementary strands (approximately 75%) are obtained even if the surface concentration of immobilized strands exceeds a value of 5.0 x 10(12) oligonucleotides/cm2. Furthermore, five cycles of denaturation and rehybridization result in only a 4% loss of the immobilized DNA after each run.