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    Detection of microRNA by electrocatalytically amplified nanoparticle collisions

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    CASTANEDA-DISSERTATION-2017.pdf (4.112Mb)
    Date
    2017-06-16
    Author
    Castaneda, Alma Delia
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    Abstract
    We report a new and general approach that will be useful for adapting the method of electrocatalytic amplification (ECA) to biosensing applications. In ECA, individual collisions of catalytic nanoparticles with a noncatalytic electrode surface lead to bursts of current. In this dissertation, the current arises from catalytic electrooxidation of N₂H₄ at the surface of platinum nanoparticles (PtNPs). As described in Chapter 1, the problem with using ECA for biosensing applications heretofore, is that it is necessary to immobilize a receptor, such as DNA (as in the case here) or an antibody on the PtNP surface. This inactivates the colliding NP, however, and leads to very small collision signatures. In this work, we show that oligonucleotides bound on the PtNP surface can be detected using ECA following enzymatic digestion. Chapter 2 demonstrates the proof-of-concept of this general approach using ssDNA-modified PtNPs and Exonuclease I (Exo I), an enzyme specific to ssDNA. After PtNPs were passivated with ssDNA, we show that the presence of this DNA can be detected by selectively removing a fraction via enzymatic cleavage. About half of the electrocatalytic current is recovered from the PtNPs on both Au and Hg microelectrodes. In Chapter 3, we show the application of this enzyme approach for the specific detection of microRNA (miRNA). The targets are miRNA-203 and miRNA-21, miRNAs of interest for cancer biomarker detection. PtNPs were modified with ssDNA complementary to the target, incubated with the miRNA, and the ssDNA was cleaved by Duplex Specific Nuclease (DSN). This exposes the PtNP surface for ECA, and the signal frequency is correlated to concentration of miRNA. Chapter 4 introduces a technique whereby ECA signals are manipulated via electrostatic interactions by modifying the surface of Au microelectrodes with polyelectrolyte multilayer films (PEMs). We demonstrate that it is possible to control the frequency of the collisions by manipulating the net electrostatic charge on the outer surface of the PEM film, and that electrons are able to tunnel from the PtNPs to the electrode through films of thicknesses up to 5 nm. These results set the stage for future sensing applications
    Department
    Chemistry
    Subject
    Electrochemistry
    Nuclease
    MicroRNA
    Microelectrode
    Nanoparticles
    URI
    https://hdl.handle.net/2152/75134
    http://dx.doi.org/10.26153/tsw/2240
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    • facebook
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