Daniele A, Di Natale P
Hum Genet. 1987 Mar;75(3):234-8. doi: 10.1007/BF00281065.
Polyclonal antibodies were obtained from rabbits by injection of iduronate sulfatase purified 35,000-fold from human placenta, after elution of the enzyme from sodium dodecyl sulfate (SDS) polyacrylamide gels. The specificity of these antibodies towards iduronate sulfatase was demonstrated by immunoprecipitation of enzyme activity; the level of other lysosomal hydrolases and sulfatases remained constant. Immunoblot of iduronate sulfatase from various human sources showed that the antibody recognises a polypeptide of mol. wt. 72,000 daltons in placenta and serum, and a form of mol. wt. 60,000 daltons in fibroblasts. No immunoprecipitable peptide was found in urine or in the culture medium of fibroblasts. Polypeptides of the same molecular weight were recognised in serum and in fibroblasts of Hunter patients. The presence of altered proteins in these patients was also shown by competition experiments. The addition of Hunter proteins alters the binding of normal enzyme to the antibody.
通过从人胎盘中纯化35000倍的艾杜糖醛酸硫酸酯酶注入兔子体内来获取多克隆抗体,该酶是从十二烷基硫酸钠(SDS)聚丙烯酰胺凝胶上洗脱下来的。这些抗体对艾杜糖醛酸硫酸酯酶的特异性通过酶活性的免疫沉淀得以证明;其他溶酶体水解酶和硫酸酯酶的水平保持不变。对来自各种人类来源的艾杜糖醛酸硫酸酯酶进行免疫印迹分析表明,该抗体识别胎盘和血清中分子量为72000道尔顿的一种多肽,以及成纤维细胞中分子量为60000道尔顿的一种形式。在尿液或成纤维细胞培养基中未发现可免疫沉淀的肽。在亨特氏病患者的血清和成纤维细胞中识别出相同分子量的多肽。竞争实验也表明这些患者中存在改变的蛋白质。添加亨特氏病患者的蛋白质会改变正常酶与抗体的结合。