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[劳雷尔电免疫扩散法。免疫亲和色谱洗脱液研究中各种技术的问题与局限]

[Laurell electroimmunodiffusion. Concerns and limits of various technics for the study of immunoaffinity chromatographic eluants].

作者信息

Biou D, Durand G, Feger J

出版信息

Clin Biochem. 1977 Aug;10(4):141-7. doi: 10.1016/s0009-9120(77)92088-4.

Abstract

Monodimensional and bidimensional electroimmunodifusion techniques were applied to the study of immunochromatography eluates. They were obtained with human sera as samples, and rabbit anti human sera alpha1AGP antibodies coupled to CNBr Sepharose 4B packed in a chromatography column. By monodimensional electroimmunodiffusion alpha1AGP could be quantitated. By tandem bidimensional immunoelectrophoresis the eluate purity could be shown, as well as its identity to alpha1AGP. Besides, this last technique was used to the critical study of elution conditions, and to put forward the influence of some parameters (such as pH ionic strength) on alpha1AGP denaturation and its transformation into a protein which is still antigenic but has lost a part of its electrophoretic mobility at pH 8.6. This phenomenon could explain the alpha1AGP loss observed after column elution. The precautions in the interpretation, the interest and the sensitiveness of the electroimmunodiffusion techniques are exposed.

摘要

采用单向和双向电免疫扩散技术对免疫色谱洗脱液进行研究。以人血清为样本,将兔抗人血清α1AGP抗体偶联到装填于色谱柱中的溴化氰活化琼脂糖4B上,以此获得洗脱液。通过单向电免疫扩散可对α1AGP进行定量。通过串联双向免疫电泳可显示洗脱液的纯度及其与α1AGP的一致性。此外,最后这项技术用于洗脱条件的关键研究,并提出一些参数(如pH值、离子强度)对α1AGP变性及其在pH 8.6条件下转变为一种仍具有抗原性但部分电泳迁移率丧失的蛋白质的影响。这种现象可以解释柱洗脱后观察到的α1AGP损失。阐述了电免疫扩散技术在解释过程中的注意事项、优势及灵敏度。

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