ingest cdrApp 2018-08-23T19:16:18.487Z d39a25df-af15-48e9-aec2-c9af81a997a2 modifyDatastreamByValue RELS-EXT fedoraAdmin 2018-08-23T19:17:10.001Z Setting exclusive relation addDatastream MD_TECHNICAL fedoraAdmin 2018-08-23T19:17:21.249Z Adding technical metadata derived by FITS addDatastream MD_FULL_TEXT fedoraAdmin 2018-08-23T19:17:44.539Z Adding full text metadata extracted by Apache Tika modifyDatastreamByValue RELS-EXT fedoraAdmin 2018-08-23T19:18:06.741Z Setting exclusive relation modifyDatastreamByValue MD_DESCRIPTIVE cdrApp 2018-09-26T22:50:22.195Z modifyDatastreamByValue MD_DESCRIPTIVE cdrApp 2019-03-20T17:04:01.362Z Matthew DiSalvo Author UNC/NCSU Joint Department of Biomedical Engineering School of Medicine Automation of Microraft Arrays for Stem Cell Analysis and Sorting Induced pluripotent stem cells (iPSCs) are reprogrammed somatic cells with the potential to revolutionize personalized medicine, disease modeling, and tissue engineering. Emerging therapies based upon human iPSCs (hiPSCs) are already under development for degenerative diseases such as age-related macular degeneration (AMD). Despite the ready availability of hiPSCs, their enormous clinical and research potential is limited by the need to purify the cells during generation, genetic editing, and differentiation using tedious manual methodologies. This dissertation describes the automation and further development of microraft arrays to perform the isolation and splitting of hiPSCs colonies, which is the primary bottleneck in hiPSC purification pipelines. Microraft arrays are elastomeric microwell arrays with releasable magnetic cell culture and transfer elements, or “microrafts,” held within each microwell. Microraft arrays enable the identification of cells by imaging cytometry and the isolation of cells and their associated microrafts by dislodgement from the microarray, followed by magnetic manipulation into secondary vessels. The microraft array platform has been previously shown to be automatable and able to sort cells with exceptional viability and efficiencies. However, previous platforms have lacked the speed and robustness to perform large-scale microraft releases. Furthermore, previous microraft array designs were not designed to culture isolated microcolonies of hiPSCs. In this dissertation, microraft arrays were redesigned to isolate hundreds of microcolonies of cells, each within a nested grid of microrafts. Novel microarray microfabrication and computational modeling methods were developed to enable automated and robust imaging of microraft arrays. Image acquisition and analysis software was created to perform label-free detection of hiPSC microcolonies on microraft arrays and, in a separate application, to monitor colonic organoids. Additionally, a high-throughput automated microraft release and collection platform was developed that, for the first time, made used of real-time imaging to intelligently maximize the robustness and speed of microraft releases. This platform was utilized to isolate, culture, monitor, and successfully split hundreds of hiPSC microcolonies, thus demonstrating its utility for hiPSC purification. Summer 2018 2018 Biomedical engineering Automation, Cell Sorting, Image Analysis, Microarrays, Microraft Arrays, Stem Cells eng Doctor of Philosophy Dissertation University of North Carolina at Chapel Hill Graduate School Degree granting institution Biomedical Engineering John Ramsey Thesis advisor Nancy Allbritton Thesis advisor Dale Cowley Thesis advisor Frances Ligler Thesis advisor David Lalush Thesis advisor text Matthew DiSalvo Creator UNC/NCSU Joint Department of Biomedical Engineering School of Medicine Automation of Microraft Arrays for Stem Cell Analysis and Sorting Induced pluripotent stem cells (iPSCs) are reprogrammed somatic cells with the potential to revolutionize personalized medicine, disease modeling, and tissue engineering. Emerging therapies based upon human iPSCs (hiPSCs) are already under development for degenerative diseases such as age-related macular degeneration (AMD). Despite the ready availability of hiPSCs, their enormous clinical and research potential is limited by the need to purify the cells during generation, genetic editing, and differentiation using tedious manual methodologies. This dissertation describes the automation and further development of microraft arrays to perform the isolation and splitting of hiPSCs colonies, which is the primary bottleneck in hiPSC purification pipelines. Microraft arrays are elastomeric microwell arrays with releasable magnetic cell culture and transfer elements, or “microrafts,” held within each microwell. Microraft arrays enable the identification of cells by imaging cytometry and the isolation of cells and their associated microrafts by dislodgement from the microarray, followed by magnetic manipulation into secondary vessels. The microraft array platform has been previously shown to be automatable and able to sort cells with exceptional viability and efficiencies. However, previous platforms have lacked the speed and robustness to perform large-scale microraft releases. Furthermore, previous microraft array designs were not designed to culture isolated microcolonies of hiPSCs. In this dissertation, microraft arrays were redesigned to isolate hundreds of microcolonies of cells, each within a nested grid of microrafts. Novel microarray microfabrication and computational modeling methods were developed to enable automated and robust imaging of microraft arrays. Image acquisition and analysis software was created to perform label-free detection of hiPSC microcolonies on microraft arrays and, in a separate application, to monitor colonic organoids. Additionally, a high-throughput automated microraft release and collection platform was developed that, for the first time, made used of real-time imaging to intelligently maximize the robustness and speed of microraft releases. This platform was utilized to isolate, culture, monitor, and successfully split hundreds of hiPSC microcolonies, thus demonstrating its utility for hiPSC purification. Biomedical engineering Automation; Cell Sorting; Image Analysis; Microarrays; Microraft Arrays; Stem Cells Doctor of Philosophy Dissertation University of North Carolina at Chapel Hill Graduate School Degree granting institution Biomedical Engineering John Ramsey Thesis advisor Nancy Allbritton Thesis advisor Dale Cowley Thesis advisor Frances Ligler Thesis advisor David Lalush Thesis advisor 2018 2018-08 eng text Matthew DiSalvo Creator UNC/NCSU Joint Department of Biomedical Engineering School of Medicine Induced pluripotent stem cells (iPSCs) are reprogrammed somatic cells with the potential to revolutionize personalized medicine, disease modeling, and tissue engineering. Emerging therapies based upon human iPSCs (hiPSCs) are already under development for degenerative diseases such as age-related macular degeneration (AMD). Despite the ready availability of hiPSCs, their enormous clinical and research potential is limited by the need to purify the cells during generation, genetic editing, and differentiation using tedious manual methodologies. This dissertation describes the automation and further development of microraft arrays to perform the isolation and splitting of hiPSCs colonies, which is the primary bottleneck in hiPSC purification pipelines. Microraft arrays are elastomeric microwell arrays with releasable magnetic cell culture and transfer elements, or “microrafts,” held within each microwell. Microraft arrays enable the identification of cells by imaging cytometry and the isolation of cells and their associated microrafts by dislodgement from the microarray, followed by magnetic manipulation into secondary vessels. The microraft array platform has been previously shown to be automatable and able to sort cells with exceptional viability and efficiencies. However, previous platforms have lacked the speed and robustness to perform large-scale microraft releases. Furthermore, previous microraft array designs were not designed to culture isolated microcolonies of hiPSCs. In this dissertation, microraft arrays were redesigned to isolate hundreds of microcolonies of cells, each within a nested grid of microrafts. Novel microarray microfabrication and computational modeling methods were developed to enable automated and robust imaging of microraft arrays. Image acquisition and analysis software was created to perform label-free detection of hiPSC microcolonies on microraft arrays and, in a separate application, to monitor colonic organoids. Additionally, a high-throughput automated microraft release and collection platform was developed that, for the first time, made used of real-time imaging to intelligently maximize the robustness and speed of microraft releases. This platform was utilized to isolate, culture, monitor, and successfully split hundreds of hiPSC microcolonies, thus demonstrating its utility for hiPSC purification. Biomedical engineering Automation; Cell Sorting; Image Analysis; Microarrays; Microraft Arrays; Stem Cells Doctor of Philosophy Dissertation University of North Carolina at Chapel Hill Graduate School Degree granting institution John Ramsey Thesis advisor Nancy Allbritton Thesis advisor Dale Cowley Thesis advisor Frances Ligler Thesis advisor David Lalush Thesis advisor 2018 2018-08 eng text DiSalvo_unc_0153D_18033.pdf uuid:949246f3-51cd-4ffe-b039-c340389715c4 2020-08-23T00:00:00 2018-07-13T16:01:18Z proquest application/pdf 5748581