Particle Motion Analysis in Pancreatic Islets Isolation using a Quadrupole Magnetic Flow Sorter
Sajja, Venkata Sunil Kumar
Type of Degreedissertation
MetadataShow full item record
Pancreatic islet transplantation offers a viable option to achieve permanent metabolic control in Type 1 diabetes patients. However, large quantities of pure viable donor islet cells are necessary for transplantation. Using currently available islet isolation methods multiple donor organs are required to achieve successful transplantation, and there is a demand for an isolation method with high islet yield and viability. Additionally, with porcine xeno-islet cell transplantation providing much hope, improving the porcine islet isolation process has become a worthwhile endeavor. This dissertation is the summery of the work aimed to develop a Quadrupole Magnetic Sorter to isolate pancreatic islets from exocrine tissue. Computational Fluid Dynamics (CFD) simulations were used (Chapter 2) to predict the flow patterns, pressure drop and nonspecific crossover in a newly designed QMS flow channel for the isolation of pancreatic islets of Langerhans. Simulation results were compared with the theoretically and experimentally determined results to validate the CFD model. CFD simulations were employed to compare performance of two models of QMS flow channels with differing splitter positions. Results of the simulations were used to show that one design gives up to 10% less nonspecific crossover than another and this model can be used to optimize the flow channel design to achieve maximum purity of magnetic particles. Magnetic isolation is a promising method for separating and concentrating pancreatic islets for transplantation in Type 1 Diabetes patients. Continuous magnetic islet sorter was designed to overcome the restrictions of current purification methods that result in limited yield, viability and purity of the isolated islets. The performance of the islet sorter depends on the resulting speed of the islets in an applied magnetic field, a property known as magnetophoretic mobility. Essential to the design and operation of the magnetic sorter is a method to measure the magnetophoretic mobilities of magnetically infused islets. Magnetic particle tracking velocimeter (MPTV) was developed to measure (Chapter 3) the magnetophoretic mobility of particles up to 1000 microns in diameter. Velocity measurements are performed in a well-characterized isokinetic magnetic energy gradient using video imaging followed by analysis of the video images using a computer algorithm that produces histogram of absolute mobilities. Mobility distributions obtained indicated that magnetized islets have sufficient mobility to be captured by the proposed sorting method, with this result confirmed in test isolations of magnetized islets. To achieve islet isolation with high purity and yield Quadrupole Magnetic Sorting (QMS), a single cell separation method, is being modified for the isolation of pancreatic islets (Chapter 4). Islets are infused with 4.6μm Dynabeads® and separated continuously with QMS. Results from 10 porcine pancreas isolations indicated possibility of infusing islets with magnetic beads and isolating them continuously by reducing the exposure time of islets to enzymes. QMS isolated islets showed good morphology compared to standard COBE isolated islets and the Oxygen Consumption Rate (OCR) per DNA measurements confirmed the viability of the islets after isolation. QMS isolation can save the culture time and help to eliminate the mechanical stress due to centrifugation on the islets. Nude mice transplantation results confirmed Dynabeads do not affect the functionality of the islets.