Chan B, Madras B K, Davis A, Seeman P
Eur J Pharmacol. 1981 Aug 27;74(1):53-9. doi: 10.1016/0014-2999(81)90322-8.
Receptors with high affinity for neuroleptics (D2-type dopamine receptors) were solubilized from dog striatum using 1% digitonin. Soluble receptors were labelled using [3H]spiperone, and the bound 3H-ligand was separated from the free [3H]spiperone by molecular sieving on the Sephadex G-50 columns. This paper presents a polyethylene glycol (PEG) precipitation method designed to replace this column method. IC50 values for both neuroleptics and dopamine agonists, as well as dissociation constants for [3H]spiperone were similar for both methods. The precipitation method yielded a receptor density of 64 fmol/mg protein whereas the molecular sieving technique yielded a value of 194 fmol/mg protein. The qualitative similarities between the methods validates using the rapid precipitation method for monitoring the receptor during various stages of its purification.
使用1%的洋地黄皂苷从犬纹状体中溶解出对神经安定药具有高亲和力的受体（D2型多巴胺受体）。用[3H]螺哌隆标记可溶性受体，并通过在Sephadex G - 50柱上进行分子筛将结合的3H配体与游离的[3H]螺哌隆分离。本文介绍了一种旨在替代这种柱法的聚乙二醇（PEG）沉淀法。两种方法测定的神经安定药和多巴胺激动剂的IC50值以及[3H]螺哌隆的解离常数相似。沉淀法得到的受体密度为64 fmol/mg蛋白质，而分子筛技术得到的值为194 fmol/mg蛋白质。这些方法之间的定性相似性验证了在受体纯化的各个阶段使用快速沉淀法监测受体的可行性。
