Difficulties in the quantification of asialoglycoprotein receptors on the rat hepatocyte.
Difficulties in the quantification of asialoglycoprotein receptors on the rat hepatocyte.
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DOI：
10.1016/s0021-9258(19)70522-5
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发表时间：
1980-10
期刊：
影响因子：
--
通讯作者：
A. Schwartz;D. Rup;H. Lodish
中科院分区：
文献类型：
--
作者：
A. Schwartz;D. Rup;H. Lodish
通过灌注不含 Ca2+ 的溶液，然后灌注含有 Ca2+ 和胶原酶的缓冲液来分离大鼠肝细胞。它们特异性且强烈地结合 125I-asialo-orosumucoid。 4°C 时，细胞在基本必需培养基中，在饱和浓度的 125I-脱唾液酸类粘蛋白存在下孵育，每个细胞结合 500,000 个分子。在结合反应中添加 10% 马血清，结合数量减少至 200,000。用 1% 牛血清白蛋白预洗涤细胞可使结合分子的数量减少 60%。在所有情况下，Kd 值约为 7 x 10(-9) M。在 37°C 时，125I-asialo-orosumucoid 的摄取率（约 0.1 pmol/min/10(6) 个细胞）因血清或白蛋白制剂的存在而成比例降低。因此，看来在血清或白蛋白存在的情况下，大鼠肝细胞表面上脱唾液酸糖蛋白受体的绝对数量不容易测定，并且约为500,000/细胞。
Rat hepatocytes were isolated by perfusion with a Ca2+ free solution, followed by a buffer containing Ca2+ and collagenase. They specifically and avidly bind 125I-asialo-orosomucoid. At 4 degrees C, cells in minimal essential medium, incubated in the presence of saturating concentrations of 125I-asialo-orosomucoid, bound 500,000 molecules/cell. Addition of 10% horse serum to the binding reaction reduced the number bound to 200,000. Prewashing of the cells in 1% bovine serum albumin reduced the number of bound molecules by 60%. In all cases, the Kd value was approximately 7 x 10(-9) M. At 37 degrees C, the rate of uptake of 125I-asialo-orosomucoid (approximately 0.1 pmol/min/10(6) cells) is proportionately reduced by the presence of serum or albumin preparatons. Thus, it appears that the absolute number of asialoglycoprotein receptors on the surface of rat hepatocytes is not readily determined in the presence of serum or albumin and is about 500,000/cell.