Amplifying the Macromolecular Crowding Effect Using Nanoparticles

The melting temperature (<i>T</i>_m) of DNA is affected not only by salt but also by the presence of high molecular weight (MW) solutes, such as polyethylene glycol (PEG), acting as a crowding agent. For short DNAs in a solution of low MW PEGs, however, the change of excluded volume upon melting is very small, leading to no increase in <i>T</i>_m. We demonstrate herein that by attaching 12-mer DNAs to gold nanoparticles, the excluded volume change was significantly increased upon melting, leading to increased <i>T</i>_m even with PEG 200. Larger AuNPs, higher MW PEGs, and higher PEG concentrations show even larger effects in stabilizing the DNA. This study reveals a unique and fundamental feature at nanoscale due to geometric effects. It also suggests that weak interactions can be stabilized by a combination of polyvalent binding and the enhanced macromolecular crowding effect using nanoparticles

Aptamer-Functionalized Hydrogel Microparticles for Fast Visual Detection of Mercury(II) and Adenosine

With a low optical background, high loading capacity, and good biocompatibility, hydrogels are ideal materials for immobilization of biopolymers to develop optical biosensors. We recently immobilized mercury and lead binding DNAs within a monolithic gel and demonstrated ultrasensitive visual detection of these heavy metals. The high sensitivity was attributed to the enrichment of the analytes into the gels. The signaling kinetics was slow, however, taking about 1 h to obtain a stable optical signal because of a long diffusion distance. In this work, we aim to understand the analyte enrichment process and improve the signaling kinetics by preparing hydrogel microparticles. DNA-functionalized gel beads were synthesized using an emulsion polymerization technique and most of the beads were between 10 and 50 μm. Acrydite-modified DNA was incorporated by copolymerization. Visual detection of 10 nM Hg²⁺ was still achieved and a stable signal was obtained in just 2 min. The gel beads could be spotted to form a microarray and dried for storage. A new visual sensor for adenosine was designed and immobilized within the gel beads. The adenosine aptamer binds its target about 1000-fold less tightly compared to the mercury binding DNA, allowing a comparison to be made on analyte enrichment by aptamer-functionalized hydrogels

Instantaneous Attachment of an Ultrahigh Density of Nonthiolated DNA to Gold Nanoparticles and Its Applications

The last 16 years have witnessed the landmark development of polyvalent thiolated DNA-functionalized gold nanoparticles (AuNP's) possessing striking properties within the emerging field of nanobiotechnology. Many novel properties of this hybrid nanomaterial are attributed to the dense DNA shell. However, the question of whether nonthiolated polyvalent DNA–AuNP could be fabricated with a high DNA density and properties similar to those of its thiolated counterpart has not been explored in detail. Herein, we report that by simply tuning the pH of the DNA–AuNP mixture an ultrahigh capacity of nonthiolated DNA can be conjugated to AuNP's in a few minutes, resulting in polyvalent DNA–AuNP conjugates with cooperative melting behavior, a typical property of polyvalent thiolated DNA-functionalized AuNP's. With this method, large AuNP's (e.g., 50 nm) can be functionalized to achieve the colorimetric detection of sub-nanometer DNA. Furthermore, this fast, stable DNA loading was employed to separate AuNP's of different sizes. We propose that a large fraction of the attached DNAs are adsorbed via one or a few terminal bases to afford the high loading capacity and the ability to hybridize with the complementary DNA. This discovery not only offers a time- and cost-effective way to functionalize AuNP's with a high density of nonthiolated DNA but also provides new insights into the fundamental understanding of how DNA strands with different sequences interact with AuNP's