Development and characterization of primary cell cultures from the hematopoietic tissues of the Dublin Bay prawn, Nephrops norvegicus.
Development and characterization of primary cell cultures from the hematopoietic tissues of the Dublin Bay prawn, Nephrops norvegicus.
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DOI：
10.1023/a:1017971618398
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发表时间：
2000
期刊：
影响因子：
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通讯作者：
A. Mulford;F. Lyng;C. Mothersill;B. Austin
中科院分区：
文献类型：
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作者：
A. Mulford;F. Lyng;C. Mothersill;B. Austin
改进了都柏林海湾对虾Nephrops norveicus(L.)在海水(盐度=25‰)中配制10%(v/v)2×Leibovitz‘s培养液，添加10%(v/v)热灭活胎牛血清+5%(v/v)肾血清或5%(v/v)肾小球提取物，添加0.06g/L的L-脯氨酸和1g/L的葡萄糖。100g/ml的Pronase可改善造血细胞培养的组织解离和随后的扩散。加入上皮生长因子(EGF)、成纤维细胞生长因子(BFGF)或胰岛素生长因子-1(IGF-I)对细胞生长无促进作用。细胞培养中含有几种类型的成熟血细胞，直径在6-24微米范围内。表现为酸性磷酸酶、α-丁酸萘酯酶、α-乙酸萘酯酶、氯乙酸萘酯酶和酚氧化酶活性，而碱性磷酸酶和过氧化物酶活性较弱。PAS阳性颗粒、不饱和脂类和磷脂颗粒较小。培养物在两周多的时间里仍能正常工作。偶尔也能观察到有丝分裂；然而，用核增殖标记物不能记录细胞的增殖。
Improved maintenancein vitroof the hematopoietic tissue of the Dublin Bay prawnNephrops norvegicus(L.) resulted by using 10% (v/v) 2× Leibovitz's medium prepared in seawater (salinity = 25‰), and supplemented with 10% (v/v) heat inactivated fetal bovine serum plus 5% (v/v)Nephropsserum or 5% (v/v)Nephropsmuscle extract, and 0.06 g/l of L-proline and 1 g/l of glucose. Pronase at 100 g/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 µm diameter. Acid phosphatase, α-naphthyl butyrate esterase, α-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.