Evaluating suspected CWD feral pig brain samples using RT-QuIC and protocol for the purification and quality control of recombinant monomeric proteins for RT-QuIC assay

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Hepker, Monica
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Anumantha Kanthasamy
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Veterinary Microbiology and Preventive Medicine
Our faculty promote the understanding of causes of infectious disease in animals and the mechanisms by which diseases develop at the organismal, cellular and molecular levels. Veterinary microbiology also includes research on the interaction of pathogenic and symbiotic microbes with their hosts and the host response to infection.
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Veterinary Microbiology and Preventive Medicine

Proteinopathy is a broad term used to describe any disease or condition that arises as

a result of aberrant protein behavior. Most often it is used when discussing any of a wide

range of neurodegenerative diseases that tend to result from an abnormal misfolding and

subsequent aggregation of a normal cellular protein. The pathology of each of these diseases

is largely dependent upon the nature of the protein aggregate that is responsible for or

resulting from the disease. When viewed individually each disease seems only vaguely

similar to others, but when a step back is taken to consider the broader aspects of the disease

it begins to show similarities with other diseases, some of which are better studied and

understood. The advances in other proteinopathies can then be experimentally applied to a

specific disease of interest and the results compared to gain better understand the

mechanisms underlying its pathogenesis and progression.

Two examples of such proteinopathies are transmissible spongiform encephalopathies

(TSE) and Parkinson’s disease (PD). TSEs are a subset of prion misfolding diseases,

specifically identified as being transmissible to other individuals in a manner resembling

viral, bacterial, or toxic exposure, that have been studied in animals for decades. While rare

in humans, prion protein misfolding does occur in a handful of different forms. PD on the

other hand is fairly common in older individuals, affecting approximately one million people

in the United States and an estimated 10 million people worldwide. Both conditions share

such notable traits as characteristic protein aggregates, being neurodegenerative, eventually

fatally so, and having a link to the enteric nervous system. Techniques applied to the study of

TSEs have been successfully applied to the study of PD, and inverse has occurred as well,

leading to leap frog effect of advancement for both diseases.

In this thesis, topics of two proteinopathies are covered. A literature review of chronic

wasting disease (CWD) history, study, and further interests leads the paper. This is followed

by a chapter on analysis of feral pig brainstem by RT-QuIC to determine the presence or

absence of aggregated prion protein. The next chapter is dedicated to describing a protocol

for the efficient and reliable production of recombinant human α-synuclein monomeric

protein for use in research. Each of these techniques has been applied to the study of multiple

diseases, but the refinement of each technique for application to the disease or species of

interest is essential to achieving the best results.

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Wed May 01 00:00:00 UTC 2019