Syversen U, Waldum H L, O'Connor D T
Institute of Cancer Research, University of Trondheim, Norway.
Neuropeptides. 1992 Aug;22(4):235-40. doi: 10.1016/0143-4179(92)90052-x.
Chromogranin A (CgA) is a useful probe of human neuroendocrine neoplasia and exocytotic sympathoadrenal activity, but the application of CgA immunoassays has not been widespread because of limited availability of purified human CgA. Here we describe a rapid, high yield isolation of human CgA. After obtaining and lysing pheochromocytoma chromaffin granules, the soluble core proteins (chromogranins) were depleted of dopamine-beta-hydroxylase by passage over a concanavalin A-Sepharose affinity column, then lyophilized, resuspended in volatile buffer, and gel filtered on Sephacryl S-300. SDS-PAGE-analyzed column fractions contained homogeneous human CgA, which was verified structurally (N-terminal amino acid sequence) and immunologically (radioimmunoassay and immunoblot). The overall 22.6 mg yield of purified CgA represented 5.7% of the starting vesicle core protein. This preparation will be useful in evaluating the sympathoadrenal system and endocrine neoplasia in man.
嗜铬粒蛋白A（CgA）是一种用于检测人类神经内分泌肿瘤和胞吐性交感肾上腺活动的有用探针，但由于纯化的人CgA可用性有限，CgA免疫测定的应用并不广泛。在此，我们描述了一种快速、高产率分离人CgA的方法。获取并裂解嗜铬细胞瘤嗜铬粒后，通过伴刀豆球蛋白A-琼脂糖亲和柱去除可溶性核心蛋白（嗜铬粒蛋白）中的多巴胺-β-羟化酶，然后冻干，重悬于挥发性缓冲液中，并在Sephacryl S-300上进行凝胶过滤。SDS-PAGE分析的柱级分含有均一的人CgA，通过结构（N端氨基酸序列）和免疫（放射免疫测定和免疫印迹）进行了验证。纯化的CgA总产率为22.6 mg，占起始囊泡核心蛋白的5.7%。该制剂将有助于评估人类的交感肾上腺系统和内分泌肿瘤。
