Further automation in pancreatic islet isolations

Abstract

For patients with severe cases of type 1 diabetes mellitus, the standard treatment of exogenous insulin administration is not sufficient, and, therefore, are eligible for β-cell replacement therapies, such as pancreatic islet transplantations, to regain glycemic control. In order to transplant pancreatic islets, they first need to be separated from the rest of the pancreatic tissue through pancreatic islet isolation procedure. Researchers from the Leiden University medical Center (LUMC) are working on a system, called PancReatic Islet Separation Method (PRISM), that automates large parts of this isolation procedure. During validation studies of the prototype device, PRISM 1.0, two main areas have been identified that can be further improved upon during the development of a version two PRISM device. One of these areas is the need for the ability to identify pancreatic islets from exocrine tissue in a label free, non-destructive an in- or on-line manner, as an alternative for the use of Dithizone (DTZ), a cytotoxic zinc chelating agent which stains the pancreatic islets red. The other area for improvement is to find a solution for the crowding effects, resulting in a disturbed density gradient, from a large volume of tissue in the limited size Latham bowl. In an effort to explore alternative options to identify pancreatic islets from exocrine tissue, a research has been conducted in which optical and electrical properties of both tissues were investigated. To this end, experiments have been performed in which the optical absorption and electrical impedance spectra have been investigated. While some differences between pancreatic islets and exocrine tissue have been measured, the results are within the margin of error, and, therefore, no definitive conclusion has been reached on the usability of these methods for pancreatic islet identification. To overcome the difficulties of a large volume of tissue in the continuous flow centrifuge bowl during the pancreatic islet isolation procedure, research has been done into the optimization of the controllable variables during the creation of sequential and simultaneous density gradients. This research showed that the used tubing could gain more control after applying a treatment protocol and that the developed pump system is linearly progressive and accurate. These findings were then used to re-visit the simultaneous density gradient process, and evaluate the accuracy and repeatability of this process. Ultimately, this process proved to be a robust and reliable method to create simultaneous density gradients, and would, therefore, be able to solve the Latham bowl size limitations.