Design and characterization of a "click" chemistry-based electrochemical aptasensor for detection of vascular endothelial growth factor
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Vascular endothelial growth factor (VEGF) is an inflammatory cytokine that has been linked to diseases such as cancer, rheumatoid arthritis, and age-related macular degeneration. A recent study also suggests that chronic methamphetamine (METH) exposure may induce an up-regulation in VEGF expression. Motivated by these studies, researchers have actively pursued the discovery of therapeutic ligands to inhibit VEGF from binding to its receptors, thereby preventing or attenuating the effect of overexpression of VEGF. Similar to antibodies, aptamers, a new class of target-binding ligands, have demonstrated the ability to bind to their targets with comparably high affinity. While a recently developed VEGF aptamer can potentially be used therapeutically, the same aptamer can also be employed as the sensing element in the fabrication of a VEGF biosensor.
The focus of this thesis is the design and characterization of an electrochemical aptamer-based (E-AB) sensor using a DNA aptamer for direct detection of VEGF in blood serum. The goal is to evaluate the feasibility of employing a relatively new surface conjugation method, namely Sharpless “click” chemistry, for the fabrication of this sensor. The performance of the “click”-based sensor is compared to that observed with a sensor fabricated via a conventional two-step method. The results show that both sensors perform comparably when interrogated with VEGF in a serum-containing buffer. Despite the differences in the total signal gain upon target binding, the “click”-based sensor exhibits similar selectivity, regenerability and reusability to the conventional sensor. This work for the first time confirms the feasibility of using “click” chemistry in the fabrication of an E-AB sensor. More importantly, this versatile sensor fabrication approach could potentially be used in the fabrication of a multi-pixel electrode array for direct detection of a wide range of inflammatory cytokines relevant to METH exposure.
Adviser: Rebecca Y. Lai
A THESIS Presented to the Faculty of The Graduate College at the University of Nebraska In Partial Fulfillment of Requirements For the Degree of Master of Science, Major: Chemistry, Under the Supervision of Rebecca Y. Lai. Lincoln, Nebraska: July 2011
Copyright 2011 Cody Schaefer