Novel Point-of-Care Disposable Device and Cell Culture Bioprocessing Technique

• Main Author: Okarski, Kevin M.
• Other Authors: Powers, Linda S.
• Language: en_US
• Published: The University of Arizona. 2016
• Subjects: Diagnostic; Pathogen Detection; Rapid; Regenerative Medicine; Stem Cell; Biomedical Engineering; Bioprocessing
• Online Access: http://hdl.handle.net/10150/620668; http://arizona.openrepository.com/arizona/handle/10150/620668

This dissertation is composed of two projects dedicated to the development of techniques and technologies for improving the quality of life for patients in both clinical and resource-limited settings. The purpose of the first project was to design a rapid diagnostic device to screen whole blood samples for the presence of infectious agents. Point-of-care (PoC) technologies are becoming increasingly important for the detection of infectious agents in resource-limited settings (RSLs) where state-of-the-art blood screening practices are not feasible for implementation. For this project, a rapid diagnostic device was developed to directly detect pathogen content within freshly drawn whole blood samples using a ligand-binding assay format. The assay is completely self-contained within a hermetically sealed device to minimize operational complexity and ensure operator safety. The diagnostic device is capable of processing complex sample matrices by selectively capturing, concentrating, and labeling infectious agents upon functionalized surfaces. Following sample processing, the assay is optically interrogated with a fluorescence-based reader to provide rapid feedback regarding sample purity. Designs of the rapid diagnostic platform evolved over several prototype generations corresponding to project milestones emphasizing ergonomic performance, military specification testing for environmental resilience, and manufacture to yield production-grade devices for future diagnostic performance data collection. The goal of the regenerative therapy-based portion of this research was to develop a novel technique for the selective enrichment of cells demonstrating enhanced regenerative capacity in tissue-extracted cell samples. Adherent cell cultures of stromal vascular fractions (SVFs) extracted from adipose tissues were exposed to nutrient deficient conditions' eliciting a bimodal cellular response between two dissimilar cell culture subpopulations. The regenerative capacity of these two distinct subpopulations was evaluated by assessing their characteristic morphology, metabolic activity, and ability to undergo multilineage differentiation. The SVF subpopulation which demonstrated sensitivity to the nutrient deficient conditions expressed typical morphological expression of adherent cell cultures, elevated metabolic activity, and the ability to differentiate along adipogenic, chondrogenic, and osteogenic lineages. The SVF subpopulation which demonstrated resistance to the nutrient deficient conditions, however, expressed atypical morphologies, impaired metabolic activity, and did not survive culture with differentiation growth media. Based on the data, the 'treatment-sensitive' SVF subpopulation demonstrated a greater regenerative capacity than the‘treatment-resistant' subpopulation. Furthermore, the treatment-resistant subpopulation of the SVF may be representative of the damaged, senescent, and otherwise less-functional cells that comprise a significant portion of tissue-extracted cell samples and pose a significant risk to therapeutic efficacy and reproducibility. Ultimately, this expedient and inexpensive bioprocessing technique may serve to improve cell-based regenerative therapies by eliminating undesirable cells from culture.