title: Effect of Hoechst 33342 staining on developmental competence of prepubertal goat oocytes.
abstract: This study was undertaken to evaluate the effects of Hoechst staining on nuclear maturation and fertilisation when used at different stages of in vitro maturation (IVM) in prepubertal goat oocytes. Oocytes were matured in TCM1999 supplemented with 10% fetal bovine serum, 10 microg LH/ml, 10 microg FSH/ml and 1 microM 17beta-estradiol for 27 h. Frozen-thawed sperm cells were prepared by centrifugation in a discontinuous Percoll gradient and resuspended in DMH medium with 20% steer serum. Oocytes were fertilised in DMH medium with 7.75 mM calcium lactate. During IVM oocytes were exposed to 0.5 microg/ml of Hoechst 33342 staining and to ultraviolet light for a mean time of 3 s at 0 h, 8 h, 15 h, 20 h and 27 h. The percentage of metaphase II oocytes decreased significantly when oocytes were stained with Hoechst dye at 0 h, 8 h and 15 h of IVM. There was a decrease in total fertilisation rate and normal fertilisation rate of Hoechest-stained oocytes, independently of the time of Hoechst staining. Hoechst staining produces a significant reduction in oocyte viability when it is used in the early stages of in vitro maturation.
authors.affiliation: Departament de Ciència Animal i dels Aliments, Universitat Autònoma de Barcelona, Ballaterra, Spain.
authors.forename: Esther
authors.initials: E
authors.lastname: Velilla
authors.forename: Manel
authors.initials: M
authors.lastname: López-Béjar
authors.forename: Elisabet
authors.initials: E
authors.lastname: Rodríguez-González
authors.forename: Francesca
authors.initials: F
authors.lastname: Vidal
authors.forename: Maria-Teresa
authors.initials: MT
authors.lastname: Paramio
mesh_terms: D000818:Animals; D001562:Benzimidazoles / Q000633:toxicity*; D002470:Cell Survival / Q000187:drug effects; D005260:Female; D005307:Fertilization in Vitro; D005456:Fluorescent Dyes / Q000633:toxicity*; D006041:Goats; D066298:In Vitro Techniques; D008540:Meiosis / Q000187:drug effects; D009865:Oocytes / Q000166:cytology / Q000187:drug effects* / Q000254:growth & development*; D012741:Sexual Maturation; D013194:Staining and Labeling
chemical_list: D001562:Benzimidazoles; D005456:Fluorescent Dyes; C017807:bisbenzimide ethoxide trihydrochloride