title: Follicles of various maturation stages react differently to enzymatic isolation: a comparison of different isolation protocols.
abstract: Isolation of human follicles is based on digestion of the tissue by combinations of enzymes. Follicle vitality and morphology are often based on the analysis of pooled follicles of different maturation stages. Information is therefore lacking on the effect of the isolation protocol to individual follicles of different maturation stages. A study was conducted using five protocols combining different enzymes and varying concentrations. Isolated follicles were classified according to their maturation stages, counted and characterized for vitality, morphology, early apoptosis and organization of transzonal projections. No statistical differences were found between the protocols when outcome parameters were analysed on a pool of follicles regardless of their maturation status. Differences were observed in quality when the follicles were analysed separately according to their maturation status. Combining morphologic characteristics and vitality, both Liberase DH and Liberase TM combined with collagenase IV were better at isolating high-quality primordial follicles, compared with collagenase IV. No statistical difference between the isolation protocols was found for primary follicles. If only high-quality isolated secondary follicles are needed, collagenase IV is found to be most advantageous. Follicles of different maturation stages react differently when enzymatic isolation protocols are compared.
authors.affiliation: Department of Reproductive Medicine, Ghent University Hospital, 9000 Ghent, Belgium. Electronic address: sylvie.lierman@ugent.be.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Lierman
authors.affiliation: Department of Reproductive Medicine, Ghent University Hospital, 9000 Ghent, Belgium.
authors.forename: Kelly
authors.initials: K
authors.lastname: Tilleman
authors.affiliation: Department of Basic Medical Science, Faculty of Medicine and Health Science, Ghent University, 9000 Ghent, Belgium.
authors.forename: Maria
authors.initials: M
authors.lastname: Cornelissen
authors.affiliation: Department of Veterinary Sciences, Faculty of Biochemical, Pharmaceutical and Veterinary Sciences, Antwerp University, 2020 Antwerp, Belgium; Department of Molecular Biotechnology, Faculty of Bioscience Engineering, Ghent University, 9000 Ghent, Belgium.
authors.forename: Winnok H
authors.initials: WH
authors.lastname: De Vos
authors.affiliation: Department of Gynaecology, Ghent University Hospital, 9000 Ghent, Belgium.
authors.forename: Steven
authors.initials: S
authors.lastname: Weyers
authors.affiliation: Department of Endocrinology, Ghent University Hospital, 9000 Ghent, Belgium.
authors.forename: Guy
authors.initials: G
authors.lastname: T'Sjoen
authors.affiliation: Department of Pathology, Ghent University Hospital, 9000 Ghent, Belgium.
authors.forename: Claude A
authors.initials: CA
authors.lastname: Cuvelier
authors.affiliation: Department of Reproductive Medicine, Ghent University Hospital, 9000 Ghent, Belgium.
authors.forename: Petra
authors.initials: P
authors.lastname: De Sutter
mesh_terms: D000199:Actins / Q000737:chemistry; D000328:Adult; D017209:Apoptosis; D002470:Cell Survival; D017364:Collagenases / Q000737:chemistry; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D006801:Humans; D020287:In Situ Nick-End Labeling; D008297:Male; D054315:Oocyte Retrieval / Q000379:methods*; D009865:Oocytes / Q000166:cytology; D006080:Ovarian Follicle / Q000187:drug effects* / Q000473:pathology; D013739:Testosterone / Q000627:therapeutic use; D013820:Thermolysin / Q000737:chemistry; D063106:Transgender Persons
chemical_list: D000199:Actins; D013739:Testosterone; D017364:Collagenases; C106635:Liberase; D013820:Thermolysin
keywords: apoptosis; enzyme; follicle isolation; human; ovarian tissue; vitality