Pancreatic Islet Purification from Large Mammals and Humans Using a COBE 2991 Cell Processor versus Large Plastic Bottles

Abstract

The islet purification step in clinical islet isolation is important for minimizing the risks associated with intraportal infusion. Continuous density gradient with a COBE 2991 cell processor is commonly used for clinical islet purification. However, the high shear force involved in the purification method using the COBE 2991 cell processor causes mechanical damage to the islets. We and other groups have shown human/porcine islet purification using large cylindrical plastic bottles. Shear stress can be minimized or eliminated using large cylindrical plastic bottles because the bottles do not have a narrow segment and no centrifugation is required during tissue loading and the collection processes of islet purification. This review describes current advances in islet purification from large mammals and humans using a COBE 2991 cell processor versus large cylindrical plastic bottles.

Pancreas Preservation with a Neutrophil Elastase Inhibitor, Alvelestat, Contributes to Improvement of Porcine Islet Isolation and Transplantation

“…To isolate the islets, the ducts were perfused in a controlled fashion with a cold enzyme blend of Liberase T-Flex (1.0 mg/mL) with thermolysin (0.075 mg/mL) (Roche Diagnostics Corporation, Indianapolis, IN, USA). The islets were then separated by gentle mechanical dissociation [ 5 , 35 , 36 , 37 , 38 , 39 ] and purified using a continuous gradient of purification solution [ 40 ]. To generate purification solutions, iodixanol was combined with preservation solution.…”

“…To generate purification solutions, iodixanol was combined with preservation solution. We adopted bottle purification (size 500 mL; Nalgene, Rochester, NY, USA) in this step [ 35 , 40 ]. The digested tissue was divided in half so that equal amounts of tissue were used for each bottle.…”

“…A gradient was generated using a gradient marker (Biorep Technologies, Miami Lakes, FL, USA), and candy-cane-shaped stainless steel pipes (length, 30 cm; UMIHIRA, Kyoto, Japan) to enable loading from the low-density solution to high-density solution, leaving the stainless steel pipe in place. After generating a continuous gradient, the digested tissue was loaded as the top layer [ 40 ]. The bottles were centrifuged at 1000 rpm (235× g ) for 5 min at 4 °C.…”

Engineering a thixotropic and biochemically tunable hyaluronan and collagen bioink for biofabrication of multiple tissue construct types

“…Over the past 5 years, we have optimized the environmental conditions that cells and cellular spheroids or organoids experience during the phases of the bioprinting process to the point that viability of printed tissue (and tumor) constructs has largely been a non-issue, demonstrating that our bioinks are sufficiently supporting the cells from a biological, biochemical, and mechanical perspective (11,18,22,27,44,45). However, pancreatic islets, which are crucial for any diabetes-related studies, have been notably difficult to handle ex vivo/in vitro for biofabrication applications due to their inherent fragility during processing (46). To address this hurdle, we specifically assessed the viability of human pancreatic islets after having been bioprinted in 4 different bioinks (Fig.…”

Clinical Islet Transplantation Covered by Health Insurance in Japan

“…The high success rate of CIT with not only DBD (94%) but also DCD (81%) is due to our modifications of the Ricordi/Edmonton islet isolation methods. These modifications included pancreatic ductal injection of a preservation solution [ 16 , 17 ], pancreas preservation with MK solution [ 18 ], and the use of an iodixanol-based purification solution [ 17 , 19 , 20 , 21 ] and islet culture/preservation [ 22 , 23 , 24 ]. The islet isolation technique also enabled us to perform successful single-donor CITs with DBD from 2007 to 2010 in the United States [ 25 ].…”