Improved Human Islet Isolation Using a New Enzyme Blend, Liberase

Abstract

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.

Islet isolation for allotransplantation: variables associated with successful islet yield and graft function

“…Pancreas digestion, and in particular the choice of collagenase, are critical for isolation success [7,20]. The use of batches with high collagenase activity per total enzyme was an important determinant for successful isolation.…”

Evaluation of Viable β-Cell Mass is Useful for Selecting Collagenase for Human Islet Isolation: Comparison of Collagenase NB1 and Liberase HI

“…The Liberase HI was introduced in November 1994 as the first dissociation enzyme blend developed to isolate human islets (26, 28, 36). This product contained purified collagenase (both class I and II) from C. histolyticum and a purified bacterial neutral protease (thermolysin or Dispase ® )(28). The development of Liberase HI has dramatically changed human islet isolation (28), contributing to obtaining reproducibly high numbers of islets from large animal pancreata, including human, and establishing human islet transplantation as an option of therapies for patients with type 1 diabetes (41).…”

“…Collagenase plays a crucial role in dissociating the pancreas during enzymatic digestion phase. Liberase HI collagenase (Liberase; Boeringer-Mannheim/Roche) has been utilized as a standard blend for clinical islet transplantation since 1994 (28). More recently, the collagenase NB1 blend (Serva) has been rendered available and utilized for clinical trials in human islet isolation.…”

IL-10 Gene Modified Dendritic Cells Inhibit T Helper Type 1-Mediated Alloimmune Responses and Promote Immunological Tolerance in Diabetes

“…Pancreatic islets were isolated from the male BALB/c mice by collagenase technique (13). After washed twice, the islets were cultured in 1 ml Krebs-Ringer bicarbonate buffer with HEPES and its insulin secretory function was assayed by immunoradio assay (14).…”