Entrapment of Fluorescent Signaling DNA Aptamers in Sol−Gel-Derived Silica

Abstract

We report on the first successful immobilization of a DNA aptamer, in particular, a fluorescence-signaling DNA aptamer, within a sol-gel-derived matrix. The specific aptamer examined in this study undergoes a structural switch in the presence of adenosine triphosphate (ATP) to release a dabcyl-labeled nucleotide strand (QDNA), which in turn relieves the quenching of a fluorescein label that is also present in the aptamer structure. It was demonstrated that aptamers containing a complementary QDNA strand along with either a short complimentary strand bearing fluorescein (tripartite structure) or a directly bound fluorescein moiety (bipartite structure) remained intact upon entrapment within biocompatible sol-gel derived materials and retained binding activity, structure-switching capabilities, and fluorescence signal generation that was selective and sensitive to ATP concentration. Studies were undertaken to evaluate the properties of the immobilized aptamers that were either in their native state or bound to streptavidin using a terminal biotin group on the aptamer, including response time, accessibility, and leaching. Furthermore, signaling abilities were optimized through evaluation of different QDNA constructs. These studies indicated that the aptamers remained in a state that was similar to solution, with moderate leaching, only minor decreases in accessibility to ATP, and an expected reduction in response time due to diffusional barriers to mass transport of the analyte through the silica matrix. Entrapment of the aptamer also resulted in protection of the DNA against degradation from nucleases, improving the potential for use of the aptamer for in vivo sensing. This work demonstrates that sol-gel-derived materials can be used to successfully immobilize and protect DNA-based biorecognition elements and, in particular, DNA aptamers, opening new possibilities for the development of DNA aptamer-based devices, such as affinity columns, microarrays, and fiber-optic sensors.

Visual optical biosensors based on DNA-functionalized polyacrylamide hydrogels

“…These features are not readily available for most other types of substrates commonly used for biomolecular immobilization. We only reviewed polyacrylamide gels in this work but there are many other types of gel forming materials such as silica-based gels which are also very popular for interfacing with DNA sensors [87][88][89]. DNA itself is a polymer and it is actually possible to prepare hydrogels made of pure DNA.…”

Aptamer-Based Hydrogels and Their Applications

“…In addition to being immobilized on the polymer networks, aptamer sensors can be entrapped within the hydrogel [131,132]. Brennan and co-authors developed a method that entrapped ATP aptamer sensor inside a sol-gel-derived silica hydrogel [131].…”

Oligonucleotide-functionalized hydrogels as stimuli responsive materials and biosensors

“…Two types of sensor designs were tested as shown in Figure 10A and B, where the sensors respectively contained either three or two strands of DNA. 101 In both cases, the aptamer (in green)…”