Wang H, Regunathan S, Ruggiero D A, Reis D J
Department of Neurology and Neuroscience, Cornell University Medical College, New York, New York 10021.
Mol Pharmacol. 1993 Apr;43(4):509-15.
Polyclonal antibodies were raised in rabbits against a 70-kDa ligand-binding protein of the imidazoline receptor purified from solubilized bovine adrenal chromaffin cell membranes by ligand affinity chromatography. The antibodies labeled a single protein (approximately 70 kDa) in Western blots of bovine adrenal chromaffin cell membranes, inhibited 40% of specific [3H]idazoxan binding to imidazoline receptors in chromaffin cell membranes, and specifically immunoprecipitated 75% of all imidazoline-binding activity of solubilized chromaffin cell membrane proteins. The antibodies specifically immunostained heterogeneous subsets of cultured bovine chromaffin cells. They stained subpopulations of chromaffin cells of rat adrenal medulla but not the cells of adrenal cortex. We conclude that the antibodies recognize with high specificity and selectivity a approximately 70-kDa binding protein associated with or representing the imidazoline receptor that is expressed in mammalian species. Highly specific antibodies against the imidazoline receptor protein will permit mapping of the distribution of imidazoline receptors in brain and periphery and also may be useful as probes in cloning genes encoding the imidazoline receptors.
通过配体亲和色谱法从溶解的牛肾上腺嗜铬细胞膜中纯化出咪唑啉受体的70 kDa配体结合蛋白，以此免疫家兔制备多克隆抗体。这些抗体在牛肾上腺嗜铬细胞膜的蛋白质免疫印迹中标记了一种单一蛋白质（约70 kDa），抑制了40%的特异性[3H]伊达唑胺与嗜铬细胞膜中咪唑啉受体的结合，并特异性免疫沉淀了溶解的嗜铬细胞膜蛋白中75%的所有咪唑啉结合活性。这些抗体特异性免疫染色培养的牛嗜铬细胞的异质亚群。它们对大鼠肾上腺髓质嗜铬细胞的亚群染色，但不对肾上腺皮质细胞染色。我们得出结论，这些抗体以高特异性和选择性识别一种约70 kDa的结合蛋白，该蛋白与哺乳动物物种中表达的咪唑啉受体相关或代表该受体。针对咪唑啉受体蛋白的高度特异性抗体将有助于绘制咪唑啉受体在脑和外周的分布图谱，也可能作为克隆编码咪唑啉受体基因的探针。
