A fluorescence-based method for determining the surface coverage and hybridization efficiency of thiol-capped oligonucleotides bound to gold thin films and nanoparticles.

Using a fluorescence-based method, we have determined the number of thiol-derivatized single-stranded oligonucleotides bound to gold nanoparticles and their extent of hybridization with complementary oligonucleotides in solution. Oligonucleotide surface coverages of hexanethiol 12-mer oligonucleotides on gold nanoparticles (34 +/- 1 pmol/cm2) were significantly higher than on planar gold thin films (18 +/- 3 pmol/cm2), while the percentage of hybridizable strands on the gold nanoparticles (1.3 +/- 0.3 pmol/cm2, 4%) was lower than for gold thin films (6 +/- 2 pmol/cm2, 33%). A gradual increase in electrolyte concentration over the course of oligonucleotide deposition significantly increases surface coverage and consequently particle stability. In addition, oligonucleotide spacer sequences improve the hybridization efficiency of oligonucleotide-modified nanoparticles from approximately 4 to 44%. The surface coverage of recognition strands can be tailored using coadsorbed diluent oligonucleotides. This provides a means of indirectly controlling the average number of hybridized strands per nanoparticle. The work presented here has important implications with regard to understanding interactions between modified oligonucleotides and metal nanoparticles, as well as optimizing the sensitivity of gold nanoparticle-based oligonucleotide detection methods.

Selective colorimetric detection of polynucleotides based on the distance-dependent optical properties of gold nanoparticles.

A highly selective, colorimetric polynucleotide detection method based on mercaptoalkyloligonucleotide-modified gold nanoparticle probes is reported. Introduction of a single-stranded target oligonucleotide (30 bases) into a solution containing the appropriate probes resulted in the formation of a polymeric network of nanoparticles with a concomitant red-to-pinkish/purple color change. Hybridization was facilitated by freezing and thawing of the solutions, and the denaturation of these hybrid materials showed transition temperatures over a narrow range that allowed differentiation of a variety of imperfect targets. Transfer of the hybridization mixture to a reverse-phase silica plate resulted in a blue color upon drying that could be detected visually. The unoptimized system can detect about 10 femtomoles of an oligonucleotide.

A DNA-based method for rationally assembling nanoparticles into macroscopic materials.

Colloidal particles of metals and semiconductors have potentially useful optical, optoelectronic and material properties that derive from their small (nanoscopic) size. These properties might lead to applications including chemical sensors, spectroscopic enhancers, quantum dot and nanostructure fabrication, and microimaging methods. A great deal of control can now be exercised over the chemical composition, size and polydispersity of colloidal particles, and many methods have been developed for assembling them into useful aggregates and materials. Here we describe a method for assembling colloidal gold nanoparticles rationally and reversibly into macroscopic aggregates. The method involves attaching to the surfaces of two batches of 13-nm gold particles non-complementary DNA oligonucleotides capped with thiol groups, which bind to gold. When we add to the solution an oligonucleotide duplex with 'sticky ends' that are complementary to the two grafted sequences, the nanoparticles self-assemble into aggregates. This assembly process can be reversed by thermal denaturation. This strategy should now make it possible to tailor the optical, electronic and structural properties of the colloidal aggregates by using the specificity of DNA interactions to direct the interactions between particles of different size and composition.