The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform

Bernard E Tuch - One of the best experts on this subject based on the ideXlab platform.

  • comparison of size viability and function of fetal pig islet like cell clusters after digestion using collagenase or Liberase
    Cell Transplantation, 2002
    Co-Authors: Pauline Georges, Roslyn P Muirhead, Lindy Williams, Sara Holman, Muhammad T Tabiin, Sophia K Dean, Bernard E Tuch
    Abstract:

    Liberase is a highly purified blend of collagenases that has been specifically developed to eliminate the numerous problems associated with the conventional use of crude collagenase when isolating islet-like cell clusters (ICCs) from pancreases of different species. The influence of Liberase on yield, size, viability, and function of ICCs has been documented when this enzyme was used to digest adult but not fetal pancreases. In this study, we compared the effects of collagenase and Liberase on fetal pig ICCs. A total of eight fetal pig pancreas digestions were analyzed. Fetuses were obtained from Large White Landrace pigs of gestational age 80 ± 2.1 days. The pancreases were digested with either 3 mg/ml collagenase P or 1.2 mg/ml Liberase HI. The time taken to digest the pancreas was shorter for collagenase when compared with Liberase (22 ± 2 vs. 31 ± 2 min). The size of ICCs was similar for both collagenase (83 ± 0.5 µm) and Liberase (79 ± 0.4 µm) as was the number of ICCs produced per pancreas (7653 ± 1297 vs. 8101 ± 1177). Viability, as assessed using fluorescent markers, was slightly greater for Liberase (79 ± 1% vs. 76 ± 1%, p < 0.05). Responsiveness to β-cell stimulus (20 mM KCl) was similar for both methods of isolation, as was the insulin content of the ICCs, both in vitro and at 1 month after transplantation of 1500 ICCs beneath the renal capsule of immunoincompetent mice. Despite the high content of endotoxins in collagenase, the above results show that this enzyme was equally as efficient as Liberase in isolating functional ICCs from fetal pig pancreas.

  • comparison of size viability and function of fetal pig islet like cell clusters after digestion using collagenase or Liberase
    Cell Transplantation, 2002
    Co-Authors: Pauline Georges, Roslyn P Muirhead, Lindy Williams, Sara Holman, Muhammad T Tabiin, Sophia K Dean, Bernard E Tuch
    Abstract:

    Liberase is a highly purified blend of collagenases that has been specifically developed to eliminate the numerous problems associated with the conventional use of crude collagenase when isolating islet-like cell clusters (ICCs) from pancreases of different species. The influence of Liberase on yield, size, viability, and function of ICCs has been documented when this enzyme was used to digest adult but not fetal pancreases. In this study, we compared the effects of collagenase and Liberase on fetal pig ICCs. A total of eight fetal pig pancreas digestions were analyzed. Fetuses were obtained from Large White Landrace pigs of gestational age 80 +/- 2.1 days. The pancreases were digested with either 3 mg/ml collagenase P or 1.2 mg/ml Liberase HI. The time taken to digest the pancreas was shorter for collagenase when compared with Liberase (22 +/- 2 vs. 31 +/- 2 min). The size of ICCs was similar for both collagenase (83 +/- 0.5 microm) and Liberase (79 +/- 0.4 microm) as was the number of ICCs produced per pancreas (7,653 +/- 1,297 vs. 8,101 +/- 1,177). Viability, as assessed using fluorescent markers, was slightly greater for Liberase (79 +/- 1% vs. 76 +/- 1%, p < 0.05). Responsiveness to beta-cell stimulus (20 mM KCl) was similar for both methods of isolation, as was the insulin content of the ICCs, both in vitro and at I month after transplantation of 1,500 ICCs beneath the renal capsule of immunoincompetent mice. Despite the high content of endotoxins in collagenase, the above results show that this enzyme was equally as efficient as Liberase in isolating functional ICCs from fetal pig pancreas.

Jacques Donnez - One of the best experts on this subject based on the ideXlab platform.

  • enzymatic isolation of human primordial and primary ovarian follicles with Liberase dh protocol for application in a clinical setting
    Fertility and Sterility, 2011
    Co-Authors: Julie Vanacker, Alessandra Camboni, Jacques Donnez, Marie-madeleine Dolmans, Catherine Dath, Anne Van Langendonckt, Christiani Andrade Amorim
    Abstract:

    To set up a protocol to isolate human preantral follicles with an enzyme produced in good manufacturing practice conditions for use in a clinical setting.

  • evaluation of Liberase a purified enzyme blend for the isolation of human primordial and primary ovarian follicles
    Human Reproduction, 2006
    Co-Authors: Marie-madeleine Dolmans, Nathalie Michaux, Alessandra Camboni, Belen Martinezmadrid, Anne Van Langendonckt, Stefania A Nottola, Jacques Donnez
    Abstract:

    BACKGROUND: The purpose of this study is to evaluate the effectiveness of a standardized mixture of purified enzymes (Liberase), for the isolation of human ovarian follicles. METHODS: This is an experimental prospective study. Ovarian biopsies were obtained from eight young women undergoing laparoscopy for benign gynaecological disease. Follicles were isolated by Liberase or collagenase enzymatic digestion. Follicle quality was assessed by evaluating their general morphology and viability after fluorescent staining, and the ultrastructure by electron microscopy. RESULTS: The number of fully isolated follicles recovered from the Liberase-treated group was lower than from the collagenase group (156 versus 263) despite equal-sized biopsies being taken. A high proportion of follicles (98.6%, 70/71) were viable after Liberase isolation and most follicles were of good morphology with a complete granulosa cell layer (70.4%, 31/44). Ultrastructural studies indicated that Liberase-isolated follicles showed signs of atresia only occasionally and that the oolemma-follicular cell interface was well preserved. CONCLUSIONS: Liberase treatment allows the isolation of highly viable follicles from human ovarian tissue, with an unaltered morphology and ultrastructure. This purified endotoxin-free enzyme preparation is a promising alternative to impure collagenase preparations for the reproducible isolation of intact primordial and primary follicles for culture and grafting purposes.

Pauline Georges - One of the best experts on this subject based on the ideXlab platform.

  • comparison of size viability and function of fetal pig islet like cell clusters after digestion using collagenase or Liberase
    Cell Transplantation, 2002
    Co-Authors: Pauline Georges, Roslyn P Muirhead, Lindy Williams, Sara Holman, Muhammad T Tabiin, Sophia K Dean, Bernard E Tuch
    Abstract:

    Liberase is a highly purified blend of collagenases that has been specifically developed to eliminate the numerous problems associated with the conventional use of crude collagenase when isolating islet-like cell clusters (ICCs) from pancreases of different species. The influence of Liberase on yield, size, viability, and function of ICCs has been documented when this enzyme was used to digest adult but not fetal pancreases. In this study, we compared the effects of collagenase and Liberase on fetal pig ICCs. A total of eight fetal pig pancreas digestions were analyzed. Fetuses were obtained from Large White Landrace pigs of gestational age 80 ± 2.1 days. The pancreases were digested with either 3 mg/ml collagenase P or 1.2 mg/ml Liberase HI. The time taken to digest the pancreas was shorter for collagenase when compared with Liberase (22 ± 2 vs. 31 ± 2 min). The size of ICCs was similar for both collagenase (83 ± 0.5 µm) and Liberase (79 ± 0.4 µm) as was the number of ICCs produced per pancreas (7653 ± 1297 vs. 8101 ± 1177). Viability, as assessed using fluorescent markers, was slightly greater for Liberase (79 ± 1% vs. 76 ± 1%, p < 0.05). Responsiveness to β-cell stimulus (20 mM KCl) was similar for both methods of isolation, as was the insulin content of the ICCs, both in vitro and at 1 month after transplantation of 1500 ICCs beneath the renal capsule of immunoincompetent mice. Despite the high content of endotoxins in collagenase, the above results show that this enzyme was equally as efficient as Liberase in isolating functional ICCs from fetal pig pancreas.

  • comparison of size viability and function of fetal pig islet like cell clusters after digestion using collagenase or Liberase
    Cell Transplantation, 2002
    Co-Authors: Pauline Georges, Roslyn P Muirhead, Lindy Williams, Sara Holman, Muhammad T Tabiin, Sophia K Dean, Bernard E Tuch
    Abstract:

    Liberase is a highly purified blend of collagenases that has been specifically developed to eliminate the numerous problems associated with the conventional use of crude collagenase when isolating islet-like cell clusters (ICCs) from pancreases of different species. The influence of Liberase on yield, size, viability, and function of ICCs has been documented when this enzyme was used to digest adult but not fetal pancreases. In this study, we compared the effects of collagenase and Liberase on fetal pig ICCs. A total of eight fetal pig pancreas digestions were analyzed. Fetuses were obtained from Large White Landrace pigs of gestational age 80 +/- 2.1 days. The pancreases were digested with either 3 mg/ml collagenase P or 1.2 mg/ml Liberase HI. The time taken to digest the pancreas was shorter for collagenase when compared with Liberase (22 +/- 2 vs. 31 +/- 2 min). The size of ICCs was similar for both collagenase (83 +/- 0.5 microm) and Liberase (79 +/- 0.4 microm) as was the number of ICCs produced per pancreas (7,653 +/- 1,297 vs. 8,101 +/- 1,177). Viability, as assessed using fluorescent markers, was slightly greater for Liberase (79 +/- 1% vs. 76 +/- 1%, p < 0.05). Responsiveness to beta-cell stimulus (20 mM KCl) was similar for both methods of isolation, as was the insulin content of the ICCs, both in vitro and at I month after transplantation of 1,500 ICCs beneath the renal capsule of immunoincompetent mice. Despite the high content of endotoxins in collagenase, the above results show that this enzyme was equally as efficient as Liberase in isolating functional ICCs from fetal pig pancreas.

Camillo Ricordi - One of the best experts on this subject based on the ideXlab platform.

  • improved human islet isolation using a new enzyme blend Liberase
    Diabetes, 1997
    Co-Authors: Elina Linetsky, Rita Bottino, Roger Lehmann, Rodolfo Alejandro, Luca Inverardi, Camillo Ricordi
    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.

Evgenia Isachenko - One of the best experts on this subject based on the ideXlab platform.

  • construction of human artificial ovary from cryopreserved ovarian tissue appearance of apoptosis and necrosis after enzymatic isolation of follicles
    Cryobiology, 2018
    Co-Authors: Viola Maria Schmidt, Evgenia Isachenko, Gunter Rappl, Gohar Rahimi, Bettina Hanstein, Bernd Morgenstern, P Mallmann, Vladimir Isachenko
    Abstract:

    Abstract Earlier it was shown that number of retrieved follicles was significantly higher in Tumor Dissociation Enzyme (TDE)-treatment group compare to standard Liberase TM-group. The aim of our present investigations was to examine the effect of TDE on appearance of apoptosis and necrosis in follicles and stromal cells after digesting of cryopreserved ovarian cortex. Fresh and frozen ovarian cortex fragments (OCF) from 14 patients (29 ± 6 years old), sized 20–210 mm3 were randomly distributed into four treatment groups and digested with 16% TDE or 0.05 mg/ml Liberase TM: Group 1 frozen OCF digested with TDE; Group 2 frozen OCF digested with LiberaseTM; Group 3 fresh OCF digested with TDE; Group 4 fresh OCF digested with Liberase TM. To differentiate the live, early apoptotic, late apoptotic and necrotic cells in digested ovarian cortex suspension, a flow cytometric apoptosis/necrosis assay with FITC Annexin V Apoptosis Detection Kit and with 7-AAD was performed. Most of fresh (not frozen) cells digested with TDE or Liberase TM (95 ± 2.4% vs. 90.4 ± 3.1%, respectively) as well as in frozen ovarian cortex digested with TDE or Liberase TM (93.1 ± 3.4% vs. 89.7 ± 4.4%, respectively) has located in Q3 quadrant and these cells both negative to 7-AAD and Annexin V were considered as viable. It was established that both types of enzymatic treatment applying to fresh as well as to frozen ovarian cortex resulted to high rate of viable cells (Group 1: 93.8 ± 3.4%; Group 2: 91.8 ± 6.0%; Group 3: 90.5 ± 6.9%; Group 4: 87.3 ± 2.3%) and are non significantly different (P > 0.1) between all treatment groups. The amount of early apoptotic (Group 1: 3.5 ± 1.6%; Group 2: 4.4 ± 1.6%; Group 3: 1.6 ± 1.1%; Group 4: 2.4 ± 1.5%), late apoptotic (Group 1: 2.7 ± 2.4%; Group 2: 44.0 ± 1.9%; Group 3: 3.1 ± 1.1%; Group 4: 2.8 ± 0.7% and necrotic (Group 1: 0.9% ± 0.1%; Group 2: 2.9 ± 0.8%; Group 3: 3.4 ± 4.5%; Group 4: 1.1 ± 0.6%) cells was low and was not significantly different in all treatment groups (P > 0.1). It was concluded that the use of Tumor Dissociation Enzyme, effectiveness of which is higher than Liberase TM, does not lead to increasing of apoptosis and necrosis in follicles and stromal cells after enzymatic digesting of cryopreserved ovarian cortex.

  • Comparison of the enzymatic efficiency of Liberase TM and tumor dissociation enzyme: effect on the viability of cells digested from fresh and cryopreserved human ovarian cortex
    Reproductive Biology and Endocrinology, 2018
    Co-Authors: Viola Maria Schmidt, Gunter Rappl, Gohar Rahimi, Bettina Hanstein, Bernd Morgenstern, P Mallmann, Vladimir Isachenko, Evgenia Isachenko
    Abstract:

    Background The aim of this study was to examine the effectiveness of Tumor Dissociation Enzyme (TDE) on the viability of follicles after digestion of fresh and cryopreserved ovarian cortex fragments (OCFs). Methods Fresh and thawed OCF from 14 patients (29 ± 6 years), sized 20 to 210 mm^3 were randomly distributed into four treatment groups and digested with 16% TDE or 0.05 mg/ml Liberase TM: Group 1, frozen OCF digested with TDE; Group 2, frozen OCF digested with LiberaseTM; Group 3, fresh OCF digested with TDE; and Group 4, fresh OCF digested with Liberase TM. Evaluation of follicle viability was performed under light microscope after staining with Neutral red. For visualization of viable and dead cells under a confocal laser scanning microscope, the follicles were stained with Calcein AM and ethidium homodimer-1. Results The results showed that the number of retrieved follicles was significantly higher (990 vs 487; P