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Nanoscale Biosensors for Environmental Monitoring and Biological Detection

  • Author(s): Yao, Jingjing
  • Advisor(s): Zhong, Wenwan
  • et al.
Abstract

Biosensors, comprised of a bioreceptor and a signal transduction system, have become one of the most popular analytical methods. In this dissertation, the fabrication of biosensors of high selectivity and sensitivity is accomplished via two means - adapting functional DNAs as bioreceptors and employing innovative nanomaterials as a signal transduction system.

In chapter 2, 3 and 4, functional DNAs of high selectivity are employed to recognize heavy metal ions for environmental monitoring. With the presence of Pb2+, 8-17bp DNAzymes can cleave the substrates at the phosphodiester bond between the ribonucleic adenine (rA) and the deoxyribonucleic guanine (G). This cleavage releases the single stranded-DNAs, which are then adsorbed onto single walled-carbon nanotubes, resulting in fluorescence quenching. The decrease in fluorescence is proportional to the Pb2+ concentration and therefore can be designed into a sensitive Pb2+ biosensor. Another DNA probe was designed to combine T-rich sequence for Hg2+ binding and G-quadruplex for colorimetric signaling. Without sample preparation or analytical instrument involved, Hg2+ in the environmental water can be facilely detected on the blotting membrane.

In chapter 5, 6 and 7, the biomoleuclar recognition event is signaled via cation exchange amplification (CXAmp) in ionic nanoparticles. Zn2+ encapsulated in 5 nm ZnSe nanocrystals can be replaced by free Ag+, rapidly and completely. The freed Zn2+ in turn ignites FluoZin-3, a non-fluorescent Zn-specific dye, resulting in intense fluorescence signal. When ZnSe nanocrystals are attached to the bioreceptors, i.e., the antibodies, the target molecule can be amplified significantly because thousands of Zn2+ ions are freed from one single nanoparticle. The superior labeling efficiency allowed the sensitive detection of IgE at a concentration as low as 1 ng/mL. Further, the size-dependent Zn2+ release property in large porous ZnS nanocrystal clusters (NCCs) is explored. High Zn release efficiency is obtained in 45 nm ZnS NCCs with the assistance of microwave irradiation for only 2 min. By offering an improved labeling efficiency, 5 pg/mL (~33 fM) IgE is detected. Compared with the enzyme-labeled immunosorbent assay, a gold standard for immunoassay, our assay provides sensitivity 1,000 times higher. Besides, the CXAmp is combined with DNA amplification techniques like rolling circle amplification for more challengeable miRNA detection with the detection limit in the attomole range.

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