Refinement of the Automated Method for Human Islet Isolation and Presentation of a Closed System for In Vitro Islet Culture

Abstract

A standardized technique of islet isolation is presented applying novel means to improve enzymatic digestion and to meet cGMP standards.

cAMP Induces Stromal Interaction Molecule 1 (STIM1) Puncta but neither Orai1 Protein Clustering nor Store-operated Ca2+ Entry (SOCE) in Islet Cells

“…Human pancreatic islets from five normoglycemic cadaveric donors (two female, three male; aged 39 -57 years) were generously provided by the Nordic Network for Clinical Islet Transplantation. The islets were isolated with semiautomated digestion filtration (27) and purified on a continuous density gradient in a refrigerated cell processor (COBE 2191; COBE Blood Component Technology, Lakewood, CO). After purification, the islets were kept for 2-5 days at 37 ºC in an atmosphere of 5% CO 2 in CMRL 1066 culture medium (Mediatech, Herndon, VA) containing 5.5 mM glucose and supplemented with 10 mM nicotinamide, 10 mM HEPES, 0.25 g/ml Fungizone, 50 g/ml gentamicin, 2 mM glutamine, 10 g/ml ciprofloxacin, and 10% fetal calf serum.…”

Detection of a Low-Grade Enteroviral Infection in the Islets of Langerhans of Living Patients Newly Diagnosed With Type 1 Diabetes

“…Pancreatic islets were isolated in Uppsala using methods previously developed for clinical islet isolation (13). Isolated islets were cultured for a few days, and aliquots of the culture medium were collected 1, 3, and 6 days postisolation for enterovirus PCR analyses.…”

Anchoring of Vascular Endothelial Growth Factor to Surface-Immobilized Heparin on Pancreatic Islets: Implications for Stimulating Islet Angiogenesis

“…Insulin secretion in response to glucose stimulation in a dynamic perifusion system was assessed as previously described 39 at 24 h after heparinization. Islets were initially perifused with 1.67 mmol=L glucose, thereafter with 16.7 mmol=L, and finally again with 1.67 mmol=L glucose.…”