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Effect of Hoechst 33342 staining on developmental competence of prepubertal goat oocytes

Published online by Cambridge University Press:  19 August 2002

Esther Velilla
Affiliation:
Departament de Ciència Animal i dels Aliments, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain
Manel López-Béjar
Affiliation:
Departament de Sanitat i Anatomia Veterinaria, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain
Elisabet Rodríguez-González
Affiliation:
Departament de Ciència Animal i dels Aliments, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain
Francesca Vidal
Affiliation:
Departament de Biologia Cellular, Fisiología i Immunologia, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain
Maria-Teresa Paramio
Affiliation:
Departament de Ciència Animal i dels Aliments, Universitat Autònoma de Barcelona, Bellaterra, Barcelona, Spain

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 μg LH/ml, 10 μg FSH/ml and 1 μM 17β-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 μg/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.

Type
Research Article
Copyright
2002 Cambridge University Press

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