The exploration of a quantum dot fluorescence resonance energy transfer (QD-FRET) based bioassay for label-free target nucleic acid detection is reported herein. This work explores the potential for developing a displacement assay for detection of nucleic acid sequences of various lengths, including one of 484 bases. Short probe oligonucleotides conjugated to QDs were allowed to hybridize to short partially mismatched dye-labelled oligonucleotide targets. The non-labelled target of interest, a 484-base segment of heat shock protein 70 (HSP 70), contained a portion that was fully complementary to the probe. Thermodynamic parameters suggested that HSP 70 would displace dye-labelled targets; however, detection was not observed. Modifications were made to this assay to reduce sterics and increase the stability of hybrids. The results obtained using this modified assay indicated that detection of non-labelled, long oligonucleotide sequences was possible using a displacement assay that relied on a short probe oligonucleotide.

The exploration of a quantum dot fluorescence resonance energy transfer (QD-FRET) based bioassay for label-free target nucleic acid detection is reported herein. This work explores the potential for developing a displacement assay for detection of nucleic acid sequences of various lengths, including one of 484 bases. Short probe oligonucleotides conjugated to QDs were allowed to hybridize to short partially mismatched dye-labelled oligonucleotide targets. The non-labelled target of interest, a 484-base segment of heat shock protein 70 (HSP 70), contained a portion that was fully complementary to the probe. Thermodynamic parameters suggested that HSP 70 would displace dye-labelled targets; however, detection was not observed. Modifications were made to this assay to reduce sterics and increase the stability of hybrids. The results obtained using this modified assay indicated that detection of non-labelled, long oligonucleotide sequences was possible using a displacement assay that relied on a short probe oligonucleotide.

Yes / Dye-functionalized polymers have been extensively studied to understand polymer chain dynamics, intra or inter-molecular association and conformational changes as well as in practical applications such as signal amplification in diagnostic tests and light-harvesting antennas. In this work, the Förster resonance energy transfer (FRET) of dye-functionalized poly(2-ethyl-2-oxazoline) (PEtOx) was studied to evaluate the effect of dye positioning and polymer chain length on the FRET efficiency. Therefore, both α (initiating terminus)- or ω (terminal chain end)-fluorophore single labeled and dual α,ω-fluorescent dye labeled PEtOx were prepared via cationic ring opening polymerization (CROP) using 1-(bromomethyl)pyrene as the initiator and/or 1-pyrenebutyric acid or coumarin 343 as the terminating agent, yielding well-defined PEtOx with high labeling efficiency (over 91%). Fluorescence studies revealed that intramolecular FRET is most efficient for heterotelechelic PEtOx containing both pyrene and coumarin 343 fluorophores as chain ends, as expected. A strong dependence of the energy transfer on the chain length was found for these dual labeled polymers. The polymers were tested in both dilute organic (chloroform) and aqueous media revealing a higher FRET efficiency in water due to the enhanced emissive properties of pyrene. The application of dual labeled polymers as fluorescent probes for temperature sensing was demonstrated based on the lower critical solution temperature behavior of the PEtOx. Furthermore, these polymers could be successfully processed into fibers and thin films. Importantly, the fluorescence properties were retained in the solid state without decreasing the FRET efficiency, thus opening future possibilities for application of these materials in solar cells and/or sensors.

Thesis advisor: Abhishek Chatterjee / Förster Resonance Energy Transfer (FRET) offers a powerful approach to study biomolecular dynamics in vitro as well as in vivo. The ability to apply FRET imaging to proteins in living cells provides an excellent tool to monitor important dynamic events such as protein conformational changes, protein-protein interactions, and proteolysis reactions. However, selectively incorporating two distinct fluorophores into the target protein(s) that are capable of FRET interaction within the complex cellular milieu is challenging. Consequently, terminal fusion to genetically encoded fluorescent proteins has emerged as the predominant labeling strategy for FRET studies in vivo. However, a major limitation of this strategy stems from the large size of the fluorescent proteins, which may perturb the native properties of the target, and restricted attachment only to the termini of the target. We reasoned that using genetically encoded fluorescent unnatural amino acids would overcome several of these challenges associated with currently available labeling strategies owing to their small size and the ability to introduce them site- specifically and co-translationally. Here, we report the use of the fluorescent unnatural amino acid “Anap” as a FRET donor with green and yellow fluorescent protein acceptors. We demonstrate the utility of this labeling strategy using proteolysis and conformational change models, and step towards in vivo studies by further developing a proteolysis system in cell lysates. / Thesis (MS) — Boston College, 2016. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Chemistry.

Molecular assemblies capable of harvesting light and using the absorbed energy have attracted great interest in recent years because of their applicability in such domains as light emitting diodes, fluorescent labelling of biological molecules, and photonic devices. Nature has also developed in plants and photosynthetic bacteria several examples of photonic nanostructures which guide light over small distances and harvest light energy, using resonance energy transfer (RET). For some time, researchers have tried to mimic the spatial arrangements of high energy transfer efficiency found in Nature. Recent progress in the application, creation and manipulation of individual or small groups of molecules are opening new perspectives for further developments in this field. These recent advances are commonly considered to lie at the root of what is being called "Nanotechnology". Although the definitions of nanotechnology are diverse, it is commonly admitted that this new domain of Science draws ideas and concepts from disciplines including engineering, physics, chemistry, biology, mathematics and computer science. The central dogma of the “bottom up” version of nanotechnology is the notion of self-assembly, which is the spontaneous assembly of materials into predetermined ordered structures or complexes. Presented here is an example from a field of nanotechnology that utilizes self-assembly onto nano-patterned surfaces to generate nano-structured systems and devices. More precisely, in the present case we target photo-active devices based on Fluorescence Resonance Energy Transfer (FRET), taking inspiration from photosynthetic light harvesting systems, where concentric nanometric rings of chromophores funnel light energy to a reaction center. Here, we synthesize nano-patterned chromophore surfaces which are able to collect light energy over a large surface and funnel it in regions of ~100 nm size. Our results indicate that an efficient collection and transfer of light energy can be performed by properly nano-designed surfaces, which may have practical consequences for the fabrication of light-powered active nano-devices.

This thesis reports the use of both dc and ac electric fi eld induced resonant energy transfer, RET, between cold Rydberg atoms as a useful tool for enhancement of interatomic interactions. A general technique for laser frequency stabilization and its suitability for Rydberg atom excitation is also demonstrated. RET between cold Rydberg atoms was used to determine Rydberg atom energy levels. The ⁸⁵Rb atoms are laser cooled and trapped in a magneto-optical trap. For energy level determination experiment, atoms were optically excited to 32d₅/₂ Rydberg states. The two-atom process 32d₅/₂ + 32d₅/₂ → 34p₃/₂+30g is resonant at an electric fi eld of approximately 0.3 V/cm through dipole dipole interaction. The experimentally observed resonant fi eld, together with the Stark map calculation is used to make a determination of the ⁸⁵Rb ng-series quantum defect to be ⵒg(n = 30) = 0.00405(6). The ac Stark eff ect was also used to induce RET between cold Rydberg atoms. When a 28.5 GHz dressing field was set at speci fic fi eld strengths, the two-atom dipole-dipole process 43d₅/₂ + 43d₅/₂ → 45p₃/₂ + 41f was dramatically enhanced, due to induced degeneracy of the initial and final states. This method for enhancing interactions is complementary to dc electric- field-induced RET, but has more flexibility due to the possibility of varying the applied frequency. At a dressing field of 28.5 GHz all of the participating levels (43d₅/₂, 45p₃/₂ and 41f) show signi cant shifts and these give a complicated series of resonances. An oscillating electric fi eld at 1.356 GHz was also used to promote the above RET process where the atoms are initially excited to the 43d₅/₂ Rydberg states. The ac fi eld strength was scanned to collect RET spectra. Di fferent resonances were observed for diff erent magnetic sublevels involved in the process. Compared to the higher dressing field frequency of 28.5 GHz, the choice of dressing frequency of 1.356 GHz, which is slightly blue detuned from the 41f - 41g transition, and structure of the spectra may be understood, by analogy with the dc field case.

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 71-79).

This thesis reports the use of both dc and ac electric fi eld induced resonant energy transfer, RET, between cold Rydberg atoms as a useful tool for enhancement of interatomic interactions. A general technique for laser frequency stabilization and its suitability for Rydberg atom excitation is also demonstrated. RET between cold Rydberg atoms was used to determine Rydberg atom energy levels. The ⁸⁵Rb atoms are laser cooled and trapped in a magneto-optical trap. For energy level determination experiment, atoms were optically excited to 32d₅/₂ Rydberg states. The two-atom process 32d₅/₂ + 32d₅/₂ → 34p₃/₂+30g is resonant at an electric fi eld of approximately 0.3 V/cm through dipole dipole interaction. The experimentally observed resonant fi eld, together with the Stark map calculation is used to make a determination of the ⁸⁵Rb ng-series quantum defect to be ⵒg(n = 30) = 0.00405(6). The ac Stark eff ect was also used to induce RET between cold Rydberg atoms. When a 28.5 GHz dressing field was set at speci fic fi eld strengths, the two-atom dipole-dipole process 43d₅/₂ + 43d₅/₂ → 45p₃/₂ + 41f was dramatically enhanced, due to induced degeneracy of the initial and final states. This method for enhancing interactions is complementary to dc electric- field-induced RET, but has more flexibility due to the possibility of varying the applied frequency. At a dressing field of 28.5 GHz all of the participating levels (43d₅/₂, 45p₃/₂ and 41f) show signi cant shifts and these give a complicated series of resonances. An oscillating electric fi eld at 1.356 GHz was also used to promote the above RET process where the atoms are initially excited to the 43d₅/₂ Rydberg states. The ac fi eld strength was scanned to collect RET spectra. Di fferent resonances were observed for diff erent magnetic sublevels involved in the process. Compared to the higher dressing field frequency of 28.5 GHz, the choice of dressing frequency of 1.356 GHz, which is slightly blue detuned from the 41f - 41g transition, and structure of the spectra may be understood, by analogy with the dc field case.

We have calculated the structures, the heats of hydrogenation and the resonance energies of benzene isomers. All structures and energies were calculated by using the MMX force fields. Using PC model, the calculated structure parameters of benzene's 8 isomers are generally in good agreements with the experimental data. The heats of hydrogenation and resonance energy of benzene isomers are parallel to those experimental data and need a. systematic adjustment of 4.5 kJ/mol.

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