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Truncated CD19 as a selection marker for the isolation of stem cell derived β-cells Huang, Luo Ting (Helen)
Abstract
Stem cell-derived β-cells (SCβ-cell) are a renewable and scalable alternative to cadaveric islet transplants as a cell replacement therapy for type 1 diabetes (T1D). However, heterogeneity within SCβ-cell cultures remains problematic for graft safety and function. Magnetic selection of SCβ-cells expressing a unique cell surface marker may help deplete undesirable cell types and facilitate functional maturation. This is study, I explored CD19 as a potential cell surface marker for the enrichment of insulin-expressing SCβ-cells. Using CRISPR/Cas9 technology, I created a knock-in add-on of CD19-mScarlet downstream of the insulin coding sequence in human embryonic stem cells (hESCs). I established reproducible SCβ-cell surface expression of CD19-mScarlet. Importantly, I developed and optimized a magnetic sorting protocol for CD19-mScarlet-expressing cells, forming enriched SCβ-cell clusters with improved glucose-stimulated c-peptide secretion. This strategy holds promise to facilitate large-scale production of functional SCβ-cells for T1D disease modeling and cell replacement therapy.
Item Metadata
Title |
Truncated CD19 as a selection marker for the isolation of stem cell derived β-cells
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Creator | |
Supervisor | |
Publisher |
University of British Columbia
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Date Issued |
2023
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Description |
Stem cell-derived β-cells (SCβ-cell) are a renewable and scalable alternative to cadaveric islet transplants as a cell replacement therapy for type 1 diabetes (T1D). However, heterogeneity within SCβ-cell cultures remains problematic for graft safety and function. Magnetic selection of SCβ-cells expressing a unique cell surface marker may help deplete undesirable cell types and facilitate functional maturation. This is study, I explored CD19 as a potential cell surface marker for the enrichment of insulin-expressing SCβ-cells. Using CRISPR/Cas9 technology, I created a knock-in add-on of CD19-mScarlet downstream of the insulin coding sequence in human embryonic stem cells (hESCs). I established reproducible SCβ-cell surface expression of CD19-mScarlet. Importantly, I developed and optimized a magnetic sorting protocol for CD19-mScarlet-expressing cells, forming enriched SCβ-cell clusters with improved glucose-stimulated c-peptide secretion. This strategy holds promise to facilitate large-scale production of functional SCβ-cells for T1D disease modeling and cell replacement therapy.
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Genre | |
Type | |
Language |
eng
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Date Available |
2023-07-05
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Provider |
Vancouver : University of British Columbia Library
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Rights |
Attribution-NonCommercial-NoDerivatives 4.0 International
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DOI |
10.14288/1.0434147
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URI | |
Degree | |
Program | |
Affiliation | |
Degree Grantor |
University of British Columbia
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Graduation Date |
2023-11
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Campus | |
Scholarly Level |
Graduate
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Rights URI | |
Aggregated Source Repository |
DSpace
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Rights
Attribution-NonCommercial-NoDerivatives 4.0 International