Optimization of human pancreatic islet isolation with a newly designed cooling system for COBE 2991.
OBJECTIVE
Islet purification is performed using the cell separator COBE 2991, which allows the purification of large amounts of islets through a continuous density gradient. However, the original cell separator COBE 2991 was not refrigerated, and islets were exposed to inappropriately high temperatures during the purification step of the isolation process. Our aim was to design a cooling system for the purification of human pancreatic islets using COBE 2991, to increase the viability and quality of the preparations.


METHODS
We designed and adapted a cooling system for the COBE 2991 based on a cooling bath connected to a coil containing a recirculation solution with a temperature below 0 degrees C which was placed around the rotor of the COBE 2991. Cell viability was determined by acridine orange/propidium iodide staining, and islet cell function by measuring glucose-stimulated insulin secretion expressed as the insulin stimulation index.


RESULTS
Pancreata from 10 consecutive organ donors (mean age, 42.8 +/- 4.3 years) were digested and purified using the newly generated cooling system for COBE 2991. At the end of the purification process, the temperature of the density gradient that contained the islet preparation was reduced by 8 degrees C to 10 degrees C compared with that of a system with no refrigeration. Islet viability increased to 83% +/- 4%, and the insulin stimulation index increased to 11.4 +/- 1.6 (average +/- SEM).


CONCLUSIONS
This innovative cooling system for COBE 2991 achieved substantial reductions in temperature and improved the quality of human pancreatic islet preparations that were suitable for transplantation.
Nacher
M
M
Servei d'Endocrinologia, Hospital Universitari de Bellvitge, Institute d'Investigació Biomèdica de Bellvitge, Barcelona, Spain. mnacher@bellvitgehospital.cat
Barceló
V
V
Escoriza
J
J
Joanny
G
G
Núñez-Ollé
M
M
Montanya
E
E
D000328:Adult; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods; D004867:Equipment Design; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D009926:Organ Preservation / Q000295:instrumentation / Q000379:methods*; D013696:Temperature
D007328:Insulin; D005947:Glucose