PhD Thesis Defense: Gabrielle Rose Moss

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"Structural and Magnetic Characterization of a Magnetite Nanoparticle-Based Aggregation Assay and its Application to In Vitro Cytokine Monitoring"

Abstract

Magnetic particle spectroscopy (MPS) based aggregation assays rely on target induced changes in functionalized magnetic nanoparticle (fMNP) hydrodynamic diameter (DHyd) and/or changes in fMNP aggregation level to enable target detection. fMNP target binding can be modeled via Brownian and Néel relaxation. We are engineering an MPS-based aggregation assay to sense tumor necrosis factor (TNF)-⍺ in a 3D melanoma tissue culture to aid in the development of immunotherapies for cancer treatment. TNF-⍺ is an inflammatory cytokine, whose production can be linked to cell death.  

The first part of this thesis investigates the coercivity (Hc), nearest neighbor distance, and DHyd of fMNPs at various levels of target induced aggregation. We found that Brownian relaxation increased by 3 orders of magnitude as fMNPs transitioned from a fully dispersed state to a fully aggregated state, triggered by target binding. This result agreed with nearest neighbor distance observations from cryogenic transmission electron microscope images. Moreover, an applied field oscillating between -300 and +300 mT decreased dipolar interactions amongst neighboring fMNPs, evident by Hc decreases, suggesting magnetic torque induced aggregate disassembly.

The second part of this thesis investigates the effects of ligand density, off-target proteins, and salts on fMNP targeting efficacy. fMNP aggregation assays can suffer from inaccuracies due to interactions with salts and off-target proteins. Our results suggest that (1) fMNP target affinity was independent of salt concentration for 0.005x to 1.00x phosphate buffered saline solutions, (2) off-target serum proteins triggered an increase in fMNP target affinity, and (3) the aggregation assay’s measurable concentration range can be tuned with surface ligand density.

In the final part of this thesis, we outline several methods to use our assay for TNF-⍺ sensing in a 3D tissue culture. We have developed an approach to localize the fMNPs within the 3D tissue culture to monitor target concentration profiles over time and can calibrate fMNP MPS signals to TNF-⍺ concentrations using a Fickian diffusion model. We also built another Fickian diffusion model to predict protein diffusion and consumption throughout the 3D tissue culture. Lastly, we are developing a site-directed method for MNP anti-TNF-⍺ antibody functionalization to promote optimal targeting efficiency.

Thesis Committee

• Solomon Diamond (chair)
• Jifeng Liu
• Geoffrey Luke
• Kathryn Miller-Jensen (Yale)

Contact

For more information, contact Thayer Registrar at thayer.registrar@dartmouth.edu.