人糖蛋白130(gp130)ELISA试剂盒

人糖蛋白130(gp130)ELISA试剂盒原理

辣根过氧化物酶 (horse radish peroxidase，简称HRP，EC.1.11.1.7)是植物中研究得最深入的一种过氧化物酶，早在20世纪30年代就有人着手从辣根中分离此酶，以后又制备出结晶。本实验是以辣根为原料，经过水的抽提，硫酸铵和丙酮分级分离，再经锌离子纯化，透析除盐，冰冻干燥，便可得到高纯度的辣根过氧化物酶。

人糖蛋白130(gp130)ELISA试剂盒辣根过氧化物酶是一种含亚铁血红素的蛋白质，HRP广泛分布于植物界，它是由无色的酶蛋白和棕色的铁卟啉结合而成的糖蛋白，糖含量18%。HRP由多个同功酶组成，Mr在40000左右，等电点7.2。溶于水，溶解度为5%(W/V)，溶液呈棕红色，透明。人糖蛋白130(gp130)ELISA试剂盒酶催化的最适PH因供氢体不同而稍有差异，但多在pH5左右。酶溶于水和58%以下的硫酸铵溶液。HRP可溶于0.58饱和度以下的硫酸铵溶液，而0.62饱和度以上则不溶。该酶最适pH7.0(对愈创木酚为氢给体而言)。在室温下，HRP几周内稳定，加热到63℃，在15min内稳定。人糖蛋白130(gp130)ELISA试剂盒氧化还原电势很低，pH6.08时，E 0 =-0.2070V，pH为7.71时，E′0 =-0.2787V。HRP的辅基和酶蛋白最大吸收光谱分别为403nm和275nm，一般以OD403nm/OD275nm的比值RZ(德文Reinheit Zahl)表示酶的纯度。* ?# p. n2 ~1 w. X3 b

酶标记物包括酶标记抗原、酶标记抗体和酶标记SPA等。酶标记物质量的好坏直接关系到免疫酶技术的成功与否，因此被称为关键的试剂。酶标记物中最常用的是酶标记抗体，它是将酶与特异性抗体经适当方法连接而成。酶标记抗体的质量主要取决于纯度好、活性强及亲和力高的酶和抗体，其次要有良好的制备方法。目前，高质量的酶(如辣根过氧化物酶，简称HRP)国内已有商品供应。人糖蛋白130(gp130)ELISA试剂盒高质量的抗体则可通过提取纯化而获得。在制备方法上，宜选用产率高、不影响结合物的活性和不混杂干扰性物质且操作简便易行的方法。

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