Evaluation of Liberase, a purified enzyme blend, for the isolation of human primordial and primary ovarian follicles.
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.
Dolmans
Marie-Madeleine
MM
Department of Gynecology, Université Catholique de Louvain, Avenue Hippocrate 10, 1200 Brussels, Belgium.
Michaux
Nathalie
N
Camboni
Alessandra
A
Martinez-Madrid
Belen
B
Van Langendonckt
Anne
A
Nottola
Stefania Annarita
SA
Donnez
Jacques
J
D017364:Collagenases; D005260:Female; D006107:Granulosa Cells / Q000166:cytology / Q000502:physiology; D006801:Humans; D046529:Microscopy, Electron, Transmission; D009865:Oocytes / Q000166:cytology / Q000502:physiology; D006080:Ovarian Follicle / Q000033:anatomy & histology* / Q000254:growth & development / Q000648:ultrastructure; D013820:Thermolysin; D020858:Tissue and Organ Harvesting / Q000379:methods*
D017364:Collagenases; C106635:Liberase; D013820:Thermolysin