中文摘要：

利用细胞分层液（比重为1．083）进行养殖牙鲆外周血单个核细胞（包括淋巴细胞和单核细胞）的分离、纯化及培养，并采用膜片钳技术观察了单核细胞膜的电生理特征。结果表明，牙鲆外周血单个核细胞在添加PHA（30μg／ml）的M199（20％FBS）培养液中，于25℃塑料培养皿（Costar，美国）中能够贴壁生长传至6代，存活30天以上。单核细胞呈不规则圆形，细胞长径为（12．30±0．62）μm，短径为（10．29±1．05）μm，并具有较强的吞噬活性，吞噬率（PP）和吞噬指数（PI）分别达到（61．9±3．05）％和1．58±0．51;淋巴细胞多呈圆形或椭圆形，细胞长径为（8．50±0．82）μm，短径为（7．46±1．07）μm，吞噬率和吞噬指数分别达到（42．1±2．33）％和3．17±1．57。膜片钳研究结果表明，牙鲆单核细胞适宜进行膜片钳研究，记录的细胞均表达电压门控钠、钾、钙离子通道。

英文摘要：

Research on cultivation and electrophysiology of mono-nuclear cells from peripheral blood of halibut Paralichthys olivaceus were conducted using patch clamp technique. The results suggest that the cells （including monocyte and lymphocyte） could be isolated and purified by using the mixture of Ficoll-Urogrofin solution. The separated cells increased in number and survived for 6 generations for over 30 days in M199 （20% FBS） medium with PHA （30 μ g/ml） in plastic cell culture plate. Monocytes were abnormal in shape [（12.30±0.62） μm, （10.29± 1.05）μm]. The cultivated cells were more phagocytic to coccus （Staphylococcus aureaus） with PP （phagocytosis percentage） and PI （phagocytosis index） being （61.9±3.05）% and 1.58± 0.51 respectively; lymphocytes were nearly spherical [（8.50± 0.82）μm, （7.46± 1.07） μ m] whose PP = （42.1±2.33）% and PI = 3.17±1.57. Electrophysiological results suggested that most of the cultured monocytes showed readily-formed seals with patch pipettes, which allowed stable, whole-cell patch-clamp recordings. Under a voltage clamp, the net currents include outward and inward current. When bathing solution that suppressed the outward current was used, a tetrodotoxin （TTX）-sensitive Na^＋ current （INa） and a slow Cd^2＋-sensitive Ca^2＋ current （ICa） were dissolved in the presence of 1 μ mol/L TTX and 0.5mmol/L Cd^2＋. The monocytes expressed a 4-aminopyridine （4-AP）-sensitive transient current, analogous to IA, and a slower-rising, tetraethylammonium （TEA）-sensitive current, analogous to IK.