title: Procedures for the reconstruction, primary culture and experimental use of rainbow trout gill epithelia.
abstract: This protocol describes how to reconstruct and culture the freshwater rainbow trout gill epithelium on flat permeable membrane supports within cell culture inserts. The protocol describes gill cell isolation, cultured gill epithelium formation, maintenance, monitoring and preparation for use in experimental procedures. To produce a heterogeneous gill epithelium, as seen in vivo, seeding of isolated gill cells twice over a 2-d period is required. As a consequence, this is termed the double-seeded insert technique. Approximately 5-12 d after cell isolation and seeding, preparations develop electrically tight gill epithelia that can withstand freshwater on the apical cell surface. The system can be used to study freshwater gill physiology, and it is a humane alternative for toxicity testing, bioaccumulation studies and environmental water quality monitoring.
authors: Division of Diabetes and Nutritional Sciences, Metal Metabolism Group, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Sabine
authors: S
authors: Schnell
authors: Division of Diabetes and Nutritional Sciences, Metal Metabolism Group, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Lucy C
authors: LC
authors: Stott
authors: Division of Diabetes and Nutritional Sciences, Metal Metabolism Group, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Christer
authors: C
authors: Hogstrand
authors: Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada.
authors: Chris M
authors: CM
authors: Wood
authors: Department of Biology, York University, Toronto, Canada.
authors: Scott P
authors: SP
authors: Kelly
authors: European Commission, Joint Research Centre, Institute of Environment and Sustainability, Ispra (Varese), Italy.
authors: Peter
authors: P
authors: Pärt
authors: Global Safety Health and Environment, AstraZeneca, Alderley Park, Macclesfield, Cheshire, UK.
authors: Stewart F
authors: SF
authors: Owen
authors: Division of Diabetes and Nutritional Sciences, Metal Metabolism Group, Faculty of Life Sciences and Medicine, King's College London, London, UK.
authors: Nic R
authors: NR
authors: Bury
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D004784:Environmental Monitoring; D004847:Epithelial Cells / Q000166:cytology*; D005880:Gills / Q000166:cytology*; D017686:Oncorhynchus mykiss / Q000033:anatomy & histology