Isolation of transporting plasma membrane vesicles from bovine tracheal epithelium.
Isolation of transporting plasma membrane vesicles from bovine tracheal epithelium.
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从牛气管上皮中分离运输质膜囊泡。
DOI：
10.1016/0005-2736(83)90024-x
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
Dubinsky,WP
中科院分区：
文献类型：
--
作者：
Langridge-Smith,JE;Field,M;Dubinsky,WP
介绍了一种从牛气管上皮中分离质膜囊泡的方法。这一过程产生了高度纯化的根尖膜，其标志酶碱性磷酸酶的含量增加了19倍。其他细胞器对这一组分的污染很小。从同一制剂分离的基侧膜与起始材料相比，(Na++K+)-ATPase活性增加了4倍，碱性磷酸酶比活性降低了2倍。顶膜囊泡对Na+摄取的分析表明，它们适合于运输研究。Na+进入囊泡间隙的运输表现为：(1)Na+摄取与介质渗透压呈线性负相关；(2)洗涤剂处理可完全释放累积的Na+；(3)Na+摄取速率显著依赖于温度。Na+转运的其他特征是：(1)被阿米洛利抑制；(2)对速尿不敏感；(3)摄取速率的阴离子依赖性，选择性如下：SCN−>；Cl−>；葡萄糖酸盐−。
A method is described for isolating plasma membrane vesicles from bovine tracheal epithelium. The procedure yields highly purified apical membranes which are enriched 19-fold in the marker enzyme, alkaline phosphatase. Contamination of this fraction by other organelles is minimal. Basolateral membranes isolated from the same preparation have a 4-fold enrichment of (Na++ K+)-ATPase and a 2-fold reduction in alkaline phosphatase specific activity compared to the starting material. Assays of Na+ uptake by the apical membrane vesicles demonstrate their suitability for transport studies. Transport of Na+ into an intravesicular space was demonstrated by (1) a linear inverse correlation between Na+ uptake and medium osmolarity;(2) complete release of accumulated Na+ by treatment with detergent; and (3) a marked temperature-dependence of Na+ uptake rate. Other features of Na+ transport were (1) inhibition by amiloride;(2) insensitivity to furosemide; and (3) anion-dependence of uptake rate with the following selectivity: SCN−> Cl−> gluconate−.