Date of Award
Doctor of Philosophy (PhD)
Julia X. Zhao
In the first project, an ultrasensitive assay has been developed for the detection and determination of Hg2+(aq) based on single-particle inductively coupled plasma-mass spectrometry (spICP-MS). In the presence of Hg2+(aq), AuNPs modified with a segment of single-stranded DNA can aggregate due to the formation of well-known thymine (T)-Hg2+-T complex. The spICP-MS can sensitively and quantitatively measure the degree of aggregation by the overall decrease in number of detected AuNPs or NP aggregates. Compared with most other Hg assays that use the same principle of aggregation-dispersion with DNA modified AuNPs, this spICP-MS-based method achieved a much lower detection limit of 0.031 part-per-trillion (155 fM) and a wider (10,000-fold) linear range up to 1 ppb. The method also showed good potential for practical applications with the environment and biomedical samples because it demonstrated minimal interference from the sample matrix. Moreover, this method, with its outstanding sensitivity could be expanded for the detection of other targets by designing the recognition unit on gold nanoparticles.
In the second project, an ultrasensitive assay for biomolecules has been developed using graphene/gold nanoparticles (AuNPs) composites and single-particle inductively coupled plasma-mass spectrometry (spICP-MS). Thrombin was chosen as a model biomolecule for this study. AuNPs modified with thrombin aptamers were first non-selectively adsorbed onto the surface of graphene oxide (GO) to form GO/AuNPs composites. In the presence of thrombin, the AuNPs desorbed from the GO/AuNPs composites due to a conformation change of the thrombin aptamer after binding with thrombin. The desorbed AuNPs were proportional to the concentration of thrombin and
could be quantified by spICP-MS. By counting the individual AuNPs in the spICP-MS measurement, the concentration of thrombin could be determined. This assay achieved an ultralow detection limit of 4.5 fM with a broad linear range from 10 fM to 100 pM. The method also showed excellent selectivity and reproducibility when a complex protein matrix was evaluated. Furthermore, the diversity of ssDNA ligands made this method a promising new technology for ultrasensitive detection of a wide variety of biomarkers in clinical diagnostics.
In the third project, a novel sandwich bioreagent for sensitive and selective detection of E. coli bacteria was developed. The bioreagent combined antibody and aptamer as selective ligands and CuNPs as fluorescent signal labels. To further improve the method sensitivity, a hybridization chain reaction (HCR) amplification strategy was applied. E. coli bacteria in water were first captured by the anti-E. coli antibody modified magnetic beads and used to obtain the primer for HCR based on the aptamer. Ultimately, the fluorescent CuNPs were produced using the long length dsDNA formed during HCR as templates. The measured fluorescence intensity was directly dependent on E. coli concentration. A low detection limit of 5,200 CFU/mL and excellent selectivity were achieved under the optimal conditions. Moreover, the method exhibited great potential for detection of other bacteria in environmental analysis and clinical diagnostics.
In the fourth project, the G-quadruplex/hemin (G/H) complex was applied as an additive to increase the antibacterial efficiency of H2O2 and avoid serious toxicity effect. The G/H complex catalyzes the decomposition of H2O2 and yields a higher hydroxyl radical (·OH) generation during the procedure. The formed ·OH has a higher antibacterial performance than the original H2O2. Using the G/H complex as an additive, the antibacterial activity of H2O2 was vastly enhanced against both Gram-positive and Gram-negative bacteria in vitro experiments. Furthermore, the designed antibacterial system was applied using a mouse wound model in vivo and showed outstanding prevention of wound infection and facilitation of wound healing. The study demonstrated the utility of using the G/H complex to help control both Gram-positive and Gram-negative infections.
Xing, Yuqian, "Development Of DNA-Based Biosensors And Antibacterial Additive" (2019). Theses and Dissertations. 2492.