Improved human islet isolation using a new enzyme blend, liberase.
Enzymatic digestion of donor pancreases is a vital step in human and large mammalian islet isolation. The variable enzymatic activities of different batches of commercially available collagenase is a major obstacle in achieving reproducibility in islet isolation procedures. In the present work, the effectiveness of Liberase, a standardized mixture of highly purified enzymes recently developed for the separation of human islets, was compared with that of a traditional collagenase preparation (type P). The results of 50 islet isolations using Liberase enzyme were compared with those of 36 isolations with collagenase, type P. No significant differences in donor age, cold ischemia time, digestion time, or weight of the pancreases were observed between the two groups. Islet yield was significantly higher in the group where the Liberase enzyme was used. All parameters examined (islet number, islet number per gram of tissue, islet equivalent number, and islet equivalent number per gram of tissue) were significantly improved when Liberase enzyme was used. Different lots of Liberase enzyme were tested, and no difference was observed. Islets isolated with Liberase enzyme were also of larger size and were much less fragmented, suggesting a gentler enzymatic action and better preservation of anatomical integrity. Islets isolated with Liberase enzyme, assessed both in vitro and in vivo, revealed a functional profile similar to that of islets separated with collagenase. Liberase enzyme appears, therefore, to represent a new powerful tool for improving the quality of human islet isolation.
Linetsky
E
E
Diabetes Research Institute, University of Miami School of Medicine, and the Miami Veterans Administration Medical Center, Florida 33136, USA.
Bottino
R
R
Lehmann
R
R
Alejandro
R
R
Inverardi
L
L
Ricordi
C
C
D000328:Adult; D000818:Animals; D001786:Blood Glucose / Q000032:analysis / Q000378:metabolism; D017364:Collagenases / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D051379:Mice; D008807:Mice, Inbred BALB C; D008875:Middle Aged; D010179:Pancreas / Q000378:metabolism* / Q000473:pathology; D013820:Thermolysin / Q000378:metabolism*; D013997:Time Factors
D001786:Blood Glucose; D007328:Insulin; D017364:Collagenases; C106635:Liberase; D013820:Thermolysin