Microbiology Ph.D., School of Life Science, Arizona State University, Tempe, AZ, 2017. Thesis: Quantitating the Functional TCRab Repertoire Using DNA Origami Nanostructures. (3.71)
Advisor: Dr. Joseph Blattman
Coastal Marine and Wetland Studies, Dept. of Coastal Marine and Wetland Studies, Coastal Carolina University, Conway, SC. Thesis: The Effects of Dissolved Inorganic Nutrient Concentrations and Ratios on Microbial Stoichiometry. (3.88)
Advisor: Dr. Vladislav Gulis
B.S. Biology, The Ohio State University, Columbus, OH. 2009 (3.63)
T cells are important for the control of many infectious diseases. In order to recognize and combat a diverse array of invading pathogens, T cells express a large repertoire of clonotypic ab dimeric T cell receptors (TCRs), with few cells on average expressing TCR specific for any given antigen but an enormous number of specificities at the population level within an individual. Quantitatively defining this balance between diversity and protection has been problematic, in large part due to the lack of methods for quantitating total TCR diversity; despite the quantitation of sequence diversity independently for TCRa and TCRb chains in the naive T cell repertoire for both humans and mice, a systematic accurate measurement of TCR combinatorial diversity, due to pairing of different TCR a and b chains, has not been attempted. The brute force method of sequencing both TCR chains at the single cell level is financially unfeasible for large naive T cell populations; each sequencing reaction costs ~$2 so for a single naive mouse with ~10000000 total T cells this would be a $20,000,000 experiment. Molecular strategies for linking TCRa and TCRb mRNA have not been adequately developed to generate suitable input material for standard multiplex deep sequencing of TCR sequences that would provide information on both TCR chains from a single cell. One major limitation to such approaches is that hybrid structures, generated by transfection with oligonucleotides, results in activation of nucleases that destroy the template and therefore, preclude TCR sequence analysis. In order to address these issues, we have developed novel DNA origami nanostructures (in collaboration with Dr. Hao Yan, a leading pioneer in the field of DNA origami) that can be transfected into T cells to ligate and protect both TCRa and TCRb mRNA, and after re-isolation, can be used as a template for ligation of a dual-primed reverse transcription reaction, and following with PCR amplification will provide input material for deep sequencing to obtain linked sequence information for both TCR chains from individual cells within polyclonal T cell populations, and thereby provide a first estimate of total TCR diversity. Using the developed DNA origami nanostructures to quantitatively define TCR diversity in individuals undergoing immune reconstitution after lymphodepletion prior to and after challenge with viral pathogens, we will be able to begin to understand the balance between immune diversity and protection.
Louie was recruited to ASU in 2012, where he began working for Dr. Joseph Blattman pursuing his PhD in Immunology. Louie has accrued nearly $50,000 in grants/fellowships. Louie was a 3-year Roche/ARCS Fellowship recipient and an American Association of Immunologists Careers In Immunology Fellowship recipient. During his PhD he helped develop a spin-out company, Gemneo Biosceince where he is currently the Vice President of Research and Production.