Mouse whole embryo culture: Evaluating the requirement for rat serum as culture medium.
birth defects
development
embryo culture
neural tube
organogenesis
postimplantation
teratogen
yolk sac
Journal
Birth defects research
ISSN: 2472-1727
Titre abrégé: Birth Defects Res
Pays: United States
ID NLM: 101701004
Informations de publication
Date de publication:
01 10 2019
01 10 2019
Historique:
received:
27
04
2019
revised:
31
05
2019
accepted:
06
06
2019
pubmed:
27
6
2019
medline:
9
7
2020
entrez:
26
6
2019
Statut:
ppublish
Résumé
Whole embryo culture is a valuable research method in mammalian developmental biology and birth defects research, enabling longitudinal studies of explanted organogenesis-stage rodent embryos. Rat serum is the primary culture medium, and can sustain growth and development over limited periods as in utero. However, the cost, labor, and time to produce culture serum are factors limiting the uptake of the methodology. The goal of replacing or at least reducing rat serum usage in culture would be in accordance with the principles of "replacement, reduction, and refinement" of animals in research (the 3Rs). We performed cultures of mouse embryos for 24 hr from embryonic day 8.5 in serum-free media or in rat serum diluted with defined media, compared with 100% rat serum. Developmental parameters scored after culture included yolk sac circulation, dorsal axial length, somite number, protein content, and completion of cranial neural tube closure. A literature review revealed use of both serum-free and diluted rat serum-based media in whole embryo culture studies, but with almost no formal comparisons of culture success against 100% rat serum. Two serum-free media were tested, but neither could sustain development as in 100% rat serum. Dilution of rat serum 1:1 with Glasgow Minimum Essential Medium plus defined supplements supported growth and development as well as whole rat serum, whereas other diluent media yielded substandard outcomes. Rat serum usage cannot be avoided, to achieve high quality mouse embryo cultures, but rat usage can be reduced using medium containing diluted serum.
Sections du résumé
BACKGROUND
Whole embryo culture is a valuable research method in mammalian developmental biology and birth defects research, enabling longitudinal studies of explanted organogenesis-stage rodent embryos. Rat serum is the primary culture medium, and can sustain growth and development over limited periods as in utero. However, the cost, labor, and time to produce culture serum are factors limiting the uptake of the methodology. The goal of replacing or at least reducing rat serum usage in culture would be in accordance with the principles of "replacement, reduction, and refinement" of animals in research (the 3Rs).
METHODS
We performed cultures of mouse embryos for 24 hr from embryonic day 8.5 in serum-free media or in rat serum diluted with defined media, compared with 100% rat serum. Developmental parameters scored after culture included yolk sac circulation, dorsal axial length, somite number, protein content, and completion of cranial neural tube closure.
RESULTS
A literature review revealed use of both serum-free and diluted rat serum-based media in whole embryo culture studies, but with almost no formal comparisons of culture success against 100% rat serum. Two serum-free media were tested, but neither could sustain development as in 100% rat serum. Dilution of rat serum 1:1 with Glasgow Minimum Essential Medium plus defined supplements supported growth and development as well as whole rat serum, whereas other diluent media yielded substandard outcomes.
CONCLUSION
Rat serum usage cannot be avoided, to achieve high quality mouse embryo cultures, but rat usage can be reduced using medium containing diluted serum.
Identifiants
pubmed: 31237114
doi: 10.1002/bdr2.1538
pmc: PMC6778057
mid: EMS83735
doi:
Substances chimiques
Culture Media
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
1165-1177Subventions
Organisme : Wellcome Trust
Pays : United Kingdom
Organisme : Wellcome Trust
ID : 087525
Pays : United Kingdom
Organisme : National Centre for the Replacement, Refinement and Reduction of Animals in Research
ID : NC/M001555/1
Pays : United Kingdom
Informations de copyright
© 2019 The Authors. Birth Defects Research published by Wiley Periodicals, Inc.
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Birth Defects Res. 2019 Oct 1;111(16):1165-1177
pubmed: 31237114