title: Development and characterization of primary cell cultures from the hematopoietic tissues of the Dublin Bay prawn, Nephrops norvegicus.
abstract: Improved maintenance in vitro of the hematopoietic tissue of the Dublin Bay prawn Nephrops norvegicus (L.) resulted by using 10% (v/v) 2x Leibovitz's medium prepared in seawater (salinity = 25 per thousand), and supplemented with 10% (v/v) heat inactivated fetal bovine serum plus 5% (v/v) Nephrops serum or 5% (v/v) Nephrops muscle extract, and 0.06 g/l of L-proline and 1 g/l of glucose. Pronase at 100 microg/ml improved tissue dissociation and subsequent spreading of hematopoietic cell cultures. The addition of epithelial growth factor (EGF), based fibroblast growth factor (bFGF) or insulin growth factor 1 (IGF-I) did not enhance cell growth. Cell culture contained several types of maturing hemocytes, in the size range of 6-24 microm diameter. Acid phosphatase, alpha-naphthyl butyrate esterase, alpha-naphthyl acetate esterase, naphthyl AS-D chloroacetate esterase activity and phenoloxidase activity was demonstrated, but not so alkaline phosphatase or peroxidase. Small PAS (= Periodic acid Schiff) positive granules, unsaturated lipids and phospholipids were observed. Cultures remained functional for over two weeks. Mitosis was noticed occasionally; however, cell proliferation was not recorded by use of nuclear proliferation markers.
authors.affiliation: Department of Biological Sciences, Heriot-Watt University, Riccarton, Edinburgh, Scotland, UK.
authors.forename: A L
authors.initials: AL
authors.lastname: Mulford
authors.forename: F
authors.initials: F
authors.lastname: Lyng
authors.forename: C
authors.initials: C
authors.lastname: Mothersill
authors.forename: B
authors.initials: B
authors.lastname: Austin
mesh_terms: D000135:Acid Phosphatase / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002467:Cell Nucleus / Q000648:ultrastructure; D048429:Cell Size / Q000502:physiology; D002478:Cells, Cultured; D003470:Culture Media / Q000737:chemistry; D003593:Cytoplasm / Q000648:ultrastructure; D033364:Decapoda / Q000166:cytology* / Q000201:enzymology; D005260:Female; D006413:Hematopoietic System / Q000166:cytology*; D006434:Hemocytes / Q000166:cytology* / Q000502:physiology; D008297:Male; D008938:Mitosis / Q000502:physiology*; D014442:Monophenol Monooxygenase / Q000032:analysis / Q000378:metabolism; D009283:Naphthol AS D Esterase / Q000378:metabolism; D012623:Seawater / Q000737:chemistry
chemical_list: D003470:Culture Media; D014442:Monophenol Monooxygenase; D009283:Naphthol AS D Esterase; D000135:Acid Phosphatase