Subcellular fractionation of pig platelets.
Subcellular fractionation of pig platelets.
复制标题
猪血小板的亚细胞分离。
DOI：
10.1016/0304-4165(75)90369-4
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发表时间：
1975
期刊：
影响因子：
--
通讯作者：
M. Fukami
中科院分区：
文献类型：
--
作者：
L. Salganicoff;P. A. Hebda;J. Yandrasitz;M. Fukami
从猪血小板中获得亚细胞组分，通过French press破碎并分离成4个主要部分。对颗粒级分（10000 g）进行蔗糖梯度分级。通过电镜和生化方法观察了初级组分和颗粒亚组分的膜、α-颗粒溶酶体、线粒体和致密颗粒标记物的分布。采用这种血小板均质化技术，发现80%的5-羟色胺和93%的β-N-乙酰氨基葡萄糖苷酶为颗粒状。在梯度中，线粒体在比活性比梯度的其它级分高10-100倍的级分（密度1.16-1.17）中明显条带化。在密度大于1.27的小球中发现含5-羟色胺的颗粒，并且含有颗粒级分的60%的5-羟色胺和腺嘌呤核苷酸。所监测的溶酶体标志物酸性磷酸酶和β-N-乙酰氨基葡萄糖苷酶表现出不同的分布模式。酸性磷酸酶在微粒体组分中显示出最高的比活性，在颗粒组分中仅为2.8%，并且在进一步分级分离后，后者的量似乎也与膜相关。β-N-乙酰氨基葡萄糖苷酶在颗粒组分和微粒体组分中均存在，具有几乎相同的比活性。然而，颗粒部分中存在的物质显然与在蔗糖梯度上分布在宽密度范围内的颗粒有关。随后的钙分布，试图确定其亚细胞位置; 19%被发现在相同的亚组分作为含胡萝卜素的颗粒，但至少50%的颗粒钙与颗粒明显分开的存储颗粒。
Subcellular components were obtained from pig platelets, disrupted by means of a French press and separated into 4 primary fractions. The granule fraction (10 000 g) was subjected to a sucrose gradient fractionation. Primary fractions and the granule subfractions were studied electron microscopically and biochemically by following the distribution of markers of membranes, lysosomes of α-granules, mitochondria and dense granules. With this technique of platelet homogenization, 80% of the serotonin and 93% of theβ-N-acetylglucosaminidase were found to be particulate. In the gradient, mitochondria were sharply banded in a fraction (density 1.16–1.17) having a specific activity 10–100 times higher than the other fractions of the gradient. Serotonin-containing granules were found in a pellet of density greater than 1.27 and contained 60% of the serotonin and adenine nucleotides of the granule fraction. The lysosome markers that were monitored, acid phosphatase andβ-N-acetylglucosaminidase, exhibited different distribution patterns. Acid phosphatase showed the highest specific activity in the microsomal fraction with only 2.8% in the granule fraction, and this latter amount also appeared to be associated with membranes upon further fractionation.β-N-Acetylglucosaminidase was present in both the granule fraction and in the microsomal fraction with nearly the same specific activity. However, that present in the granule fraction was clearly associated with granules that distributed over a wide range of densities on a sucrose gradient. The calcium distribution was followed to attempt to determine its subcellular location; 19% was found in the same subfraction as the serotonin-containing granules, but at least 50% of the particulate calcium was associated with granules distinctly separate from the storage granules.