High pressure liquid chromatographic detection of intracellular retinoid binding proteins from cultured cell and tumor cytosols.
High pressure liquid chromatographic detection of intracellular retinoid binding proteins from cultured cell and tumor cytosols.
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高压液相色谱法检测培养细胞和肿瘤细胞质中的细胞内类视黄醇结合蛋白。
DOI：
10.1016/0006-291x(83)90382-0
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发表时间：
1983
影响因子：
3.1
通讯作者：
Haussler,MR
中科院分区：
文献类型：
--
作者：
Allegretto,EA;Kelly,MA;Donaldson,CA;Levine,N;Pike,JW;Haussler,MR
本文报道了高压液相色谱法（HPLC）在细胞维甲酸结合蛋白（CRABP）和细胞视黄醇结合蛋白（CRBP）快速检测中的首次应用。用[3H]维甲酸（30 Ci/mmol）和[3H]视黄醇（43 Ci/mmol）标记培养细胞（3T6和MCF-7）或肿瘤（黑色素瘤和卵巢）的细胞质，并采用60 cm TSK 3000 sw柱进行HPLC分析。在每一种情况下，在22.5 ml的洗脱体积下都可以很容易地检测到CRABP和CRBP，其分子量为14,600。结合蛋白峰的特性是通过饱和性、特异性和有机聚合物选择性抑制结合来确定的。因此，该方法从大多数细胞质蛋白中分离出粗混合物中的CRABP和CRBP，应该是评估维生素a结合蛋白相互作用及其生物学意义的有价值的工具。
We report the first application of high pressure liquid chromatography (HPLC) in the rapid detection of cellular retinoic acid binding protein (CRABP) and cellular retinol binding protein (CRBP). Cytosols from cultured cells (3T6 and MCF-7) or from tumors (melanoma and ovarian) were labeled with [3H]retinoic acid (30 Ci/mmol) and [3H]retinol (43 Ci/mmol) and analyzed via HPLC employing a 60 cm TSK 3000 sw column. In each case CRABP and CRBP were readily detectable at an elution volume of 22.5 ml, consistent with their molecular weights of 14,600. Identity of the binding protein peaks was established by saturability, specificity, and selective inhibition of binding by an organomercurial. Thus, this method, which resolves CRABP and CRBP in crude mixtures from the majority of cytosolic proteins, should be a valuable tool in the evaluation of vitamin A-binding protein interactions and their biological significance.