Publication Date

2019-05-05

Availability

Open access

Embargo Period

2019-05-05

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Chemistry (Arts and Sciences)

Date of Defense

2019-04-04

First Committee Member

Angel E. Kaifer

Second Committee Member

Sapna K. Deo

Third Committee Member

Roger M. Leblanc

Fourth Committee Member

James N. Wilson

Abstract

Bioconjugation is one of the common ways of modifying biomolecules with reporters and other moieties for applications in the development of various sensing systems. However, achieving specific and efficient bioconjugation is challenging. Bioorthogonal chemistry can overcome these challenges as these reactions are designed to work under physiological conditions and target a specific functionality on the target biomolecule. Different functional groups can be introduced genetically or chemically on biomolecules to have a unique reactive group in order to perform bioorthogonal reactions. Herein, work relating to the development of bioluminescent biosensing systems by applying bioorthogonal chemistry between the bioluminescent proteins and other biomolecules is discussed. In particular, antibodies for the development of immunoassays that are labeled bioorthogonally with a bioluminescent protein, Gaussia luciferase were prepared and an immunoassay for the detection of interferon-γ was developed. Also, bioluminescent molecular aptamer beacons utilizing Gaussia luciferase as a reporter is described. In this work, an inhibitor molecule was attached on the beacon to reduce the signal-to-noise ratio and improve the sensitivity. Finally, the prospective research is highlighted as new biosensors can be designed with different bioorthogonal reactions and techniques.

Keywords

bioorthogonal chemistry; immunoassay; molecular aptamer beacon; bioluminescence

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