Palacios Oscar, Ruiz Encinar Jorge, Schaumlöffel Dirk, Lobinski Ryszard
Equipe de Chimie Analytique Bio-inorganique, CNRS UMR 5034, Hélioparc, 2, av. Pr. Angot, 64053, Pau, France.
Anal Bioanal Chem. 2006 Mar;384(6):1276-83. doi: 10.1007/s00216-005-0286-0. Epub 2006 Feb 22.
Immunoaffinity chromatography has been investigated for fractionation of serum into selenoalbumin and true selenoproteins. Among several albumin-depletion kits tested, a multiaffinity column specifically binding albumin and five other major serum proteins provided the best results. It extracted ca 95% of both albumin and selenoalbumin, which enabled interference-free determination of glutathione peroxidase, selenoprotein P, and selenoalbumin by size-exclusion chromatography combined with inductively coupled plasma mass spectrometry (SEC-ICPMS). The efficiency of the multiaffinity column did not vary over a period of 18 months. The purity of fractions separated by immunoaffinity LC was confirmed by elution-volume matching with standards in SEC-ICPMS and by selenopeptide mapping in capillary HPLC-ICPMS. Quantification of the selenium distribution among the different proteins in human serum from a control group and from a person on a selenium-rich diet revealed that 67% of the supplemented selenium was incorporated into albumin, 30% into glutathione peroxidase, and 3% into selenoprotein P.
已对免疫亲和色谱法进行了研究，用于将血清分离为硒白蛋白和真正的硒蛋白。在测试的几种白蛋白去除试剂盒中，一种特异性结合白蛋白和其他五种主要血清蛋白的多亲和柱效果最佳。它提取了约95%的白蛋白和硒白蛋白，这使得通过尺寸排阻色谱结合电感耦合等离子体质谱法（SEC-ICPMS）能够无干扰地测定谷胱甘肽过氧化物酶、硒蛋白P和硒白蛋白。多亲和柱的效率在18个月内没有变化。通过在SEC-ICPMS中与标准品的洗脱体积匹配以及在毛细管HPLC-ICPMS中的硒肽图谱分析，证实了通过免疫亲和液相色谱分离的组分的纯度。对对照组和富含硒饮食者的人血清中不同蛋白质之间硒分布的定量分析表明，补充的硒中有67%掺入白蛋白，30%掺入谷胱甘肽过氧化物酶，3%掺入硒蛋白P。
