HUMAN HAPTOGLOBINS - ESTIMATION AND PURIFICATION
HUMAN HAPTOGLOBINS - ESTIMATION AND PURIFICATION
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DOI：
10.1042/bj0720115
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发表时间：
1959-01-01
影响因子：
4.1
通讯作者：
SMITHIES, O
中科院分区：
文献类型：
--
作者：
CONNELL, GE;SMITHIES, O
描述了一种基于结合珠蛋白与结合珠蛋白络合物中的血红蛋白的过氧化物酶活性，快速、准确地估计血清和纯化制剂中结合珠蛋白的方法。使用过氧化氢、愈创木酚和高铁血红蛋白试剂，在明确的条件下，将游离血红蛋白的过氧化物酶活性基本上降至零。反应用分光光度法跟踪。对该方法进行了校准，以使溶液中的半球蛋白含量可以用其与血红蛋白的结合能力来表示。用Dowex 2 X-10阴离子交换树脂从pH为4.2的透析血清中大量吸附结合珠蛋白，然后在柱子中洗涤，用0.05M氯化钠溶液洗脱，得到高纯度的结合珠蛋白，产率约为50%。用淀粉凝胶电泳法对3种常见结合珠蛋白类型的血清和改良型HP2-1(Mod.)血清中的纯化结合珠蛋白进行了检测。有证据表明，所获得的结合珠蛋白没有受到纯化的破坏，而且在几种遗传类型中发现的许多单独的结合珠蛋白成分不是人工制品。
A procedure for the rapid, accurate estimation of haptoglobin in serum and purified preparations is described, based on the peroxidase activity of hemoglobin in its complexes with haptoglobin. Hydrogen peroxide, guaiacol and methemoglobin reagents are used, under well-defined conditions which reduce the peroxidase activity of free hemoglobin essentially to zero. The reaction is followed spectrophotometrically. The method is calibrated so that the hapto- globin content of a solution can be expressed in terms of its hemoglobin- binding capacity. Haptoglobin has been prepared in high purity from 1 to 150 ml of serum in approximately 50% yield, by bulk-adsorption from dialyzed serum at pH 4.2 with Dowex 2 X-10 anionic-exchange resin in the chloride form, followed by washing in a column and elution with 0.05 M NaCl solution. Purified haptoglobins from sera of the 3 common haptoglobin types and from serum of the modified type, Hp 2-1 (Mod.), have been examined by starch-gel electrophoresis. Evidence is presented that the haptoglobins obtained are undamaged by the purification, and that the many individual haptoglobin components demon-strated in the several genetic types are not artifacts.