[show abstract][hide abstract] ABSTRACT: We studied the interaction of infrared optical traps with controlled-pore glass (CPG) beads in aqueous medium. The lateral optical trapping force and stiffness were experimentally found considerably smaller than those of their solid counterparts. The simulation using an average refractive index revealed significant losses of effective trapping efficiency, which quantitatively agreed well with experimentally fitted curves. This effect was ascribed to the reduced relative refractive index of medium-filled CPG beads with respect to the medium. Combining optical trapping with mechanical confinements, we demonstrated a microfluidic platform allowing for the synthesis of multiple DNA oligonucleotide sequences on individual beads of interest.
[show abstract][hide abstract] ABSTRACT: Light-directed in situ synthesis of DNA microarrays using computer-controlled projection from a digital micromirror device--maskless array synthesis (MAS)--has proved to be successful at both commercial and laboratory scales. The chemical synthetic cycle in MAS is quite similar to that of conventional solid-phase synthesis of oligonucleotides, but the complexity of microarrays and unique synthesis kinetics on the glass substrate require a careful tuning of parameters and unique modifications to the synthesis cycle to obtain optimal deprotection and phosphoramidite coupling. In addition, unintended deprotection due to scattering and diffraction introduce insertion errors that contribute significantly to the overall error rate.
Stepwise phosphoramidite coupling yields have been greatly improved and are now comparable to those obtained in solid phase synthesis of oligonucleotides. Extended chemical exposure in the synthesis of complex, long oligonucleotide arrays result in lower--but still high--final average yields which approach 99%. The new synthesis chemistry includes elimination of the standard oxidation until the final step, and improved coupling and light deprotection. Coupling Insertions due to stray light are the limiting factor in sequence quality for oligonucleotide synthesis for gene assembly. Diffraction and local flare are by far the largest contributors to loss of optical contrast.
Maskless array synthesis is an efficient and versatile method for synthesizing high density arrays of long oligonucleotides for hybridization- and other molecular binding-based experiments. For applications requiring high sequence purity, such as gene assembly, diffraction and flare remain significant obstacles, but can be significantly reduced with straightforward experimental strategies.
Journal of Nanobiotechnology 12/2011; 9:57. · 5.09 Impact Factor
[show abstract][hide abstract] ABSTRACT: An optical tweezers directed parallel DNA oligonucleotide synthesis methodology is described in which controlled pore glass (CPG) beads act as solid substrates in a two-stream microfluidic reactor. The reactor contains two parallel sets of physical confinement features that retain beads in the reagent stream for synthetic reaction but allow the beads to be optically trapped and transferred between the reagent and the inert streams for sequence programming. As a demonstration, we synthesized oligonucleotides of target sequence 25-nt, one deletion and one substitution using dimethoxytrityl (DMT) nucleoside phosphoramidite chemistry. In detecting single-nucleotide mismatches, fluorescence in situ hybridization of the bead-conjugated probes showed high specificity and signal-to-noise ratios. These preliminary results suggest further possibilities of creating a novel type of versatile, sensitive and multifunctional reconfigurable one-bead one-compound (OBOC) bead array.
Lab on a Chip 03/2011; 11(9):1629-37. · 5.70 Impact Factor
[show abstract][hide abstract] ABSTRACT: The last decade has witnessed the development of a variety of single nanoparticle spectroscopy methods. This has facilitated unprecedented growth of knowledge and understanding of fascinating optical properties of individual metal nanoparticles. This has opened up exciting possibilities of single nanoparticles for applications in many areas including advanced photonics, biomedical imaging and sensing. The field of single nanoparticles detection and characterization is still growing. This paper reviews recent advances in single nanoparticles spectroscopy using both near-field and far-field optics. It covers spectroscopy methods for extremely small (approximately 1 nm) to relatively large nanoparticles (approximately 200 nm) and their optical properties. Different optical techniques are described. Finally, a perspective on possible practical applications of single nanoparticle spectroscopy focusing on biomedical fields is given.