镉诱导金属硫蛋白的纯化及其抗体制备

Purification of cadmium induced rat metallothionein and its antiserum preparation

目的：建立简便分离纯化镉诱导金属硫蛋白（ＣｄＭＴ）的方法，并制备抗ＣｄＭＴ抗血清。方法：采用加长ＳｅｐｈａｄｅｘＧ７５柱层析分离纯化由ＣｄＣｌ２诱导的大鼠肝组织ＣｄＭＴ，以此为抗原免疫家兔制得抗ＣｄＭＴ抗血清。结果：纯化的ＣｄＭＴ经ＳＤＳＰＡＧＥ测定相对分子质量及Ｃｄ含量测定与标准ＣｄＭＴ一致，氨基酸组分分析也与文献报道基本一致；获得的抗血清对大鼠ＣｄＭＴ具有特异性反应。结论：本法为一种简便高效的分离纯化ＣｄＭＴ的方法，抗ＣｄＭＴ抗血清的制备可为进一步建立敏感的ＥＬＩＳＡ和免疫组化方法进行组织内ＭＴ含量测定及定位，有利于深入研究Ｃｄ中毒机理和ＭＴ生物学功能。

Objective:To establish a simple method of purification of rat metallothionein(MT) induced by cadmium(Cd), and to prepare polyclonal rabbit antiserum against rat Cd MT. Methods: Cd MT was isolated from rat liver which had been injected with CdCl 2 ,separated by long typical Sephadex G 75 chromatographic column,and then used to immunize rabbit to obtain antiserum against rat Cd MT. Results:The purified Cd MT was verified by SDS PAGE and cadmium contents in per Cd MT molecule,and its molecular weight was 68 000 as same as MT standard.The amino acid composition of purified rat Cd MT also showed only a little difference as compared with the previous reports. The rabbit antiserum was able to react specifically with rat Cd MT.Conclusion:The results suggest that this novel method of purification for rat Cd MT was more simple and effective, and the rabbit antiserum against Cd MT make it possible to establish a sensitive MT ELISA and immunohistochemical analysis, so we can carry out further research on MT biological function .