Kneba M, Krieger G, Kehl A, Bause I, Nagel G A
J Immunol Methods. 1983 Jul 15;61(2):233-43. doi: 10.1016/0022-1759(83)90167-9.
A sensitive method which permits analysis of IgG containing circulating immune complexes without detailed knowledge of the nature of the antigens and the specificity of the antibodies involved is described. Soluble BSA: anti-BSA were used as model immune complexes and isolated from serum. The procedure involves the use of gel chromatography for the separation of the high molecular weight fraction containing the immune complexes as measured by binding to 125I-labeled Clq, followed by absorption of the immune complex fraction to immobilized protein A-Sepharose CL-4B. After desorption from protein A-Sepharose the complexes were dissociated and separated into free antigen and antibody by chromatofocusing in the presence of urea. The isolated free antigen and antibody retained their immunological activity as shown by immunodiffusion, binding after their recombination to 125I-labeled Clq, and by recombining antigen and antibody with much enhanced sensitivity using a microplate ELISA system. By means of the ELISA recombination technique it is possible to analyze less than 1 microgram of BSA:anti-BSA model complexes. Application of this technique may provide more information about the nature of immune complex like material associated with diseases.
本文描述了一种灵敏的方法，该方法无需详细了解所涉及抗原的性质和抗体的特异性，即可分析含IgG的循环免疫复合物。可溶性牛血清白蛋白：抗牛血清白蛋白用作模型免疫复合物，并从血清中分离出来。该程序包括使用凝胶色谱法分离含有免疫复合物的高分子量部分，通过与125I标记的Clq结合来测定，随后将免疫复合物部分吸附到固定化蛋白A-琼脂糖CL-4B上。从蛋白A-琼脂糖上解吸后，复合物被解离，并在尿素存在下通过色谱聚焦分离成游离抗原和抗体。通过免疫扩散、重组后与125I标记的Clq结合以及使用微孔板ELISA系统以大大提高的灵敏度重组抗原和抗体，所分离的游离抗原和抗体保留了它们的免疫活性。借助ELISA重组技术，可以分析不到1微克的牛血清白蛋白：抗牛血清白蛋白模型复合物。该技术的应用可能会提供更多关于与疾病相关的免疫复合物样物质性质的信息。
