Strandh M, Ohlin M, Borrebaeck C A, Ohlson S
Department of Natural Sciences, University of Kalmar, Sweden.
J Immunol Methods. 1998 May 1;214(1-2):73-9. doi: 10.1016/s0022-1759(98)00039-8.
IgM antibodies are often of low affinity (dissociation constant (Kd) > 10(-5) M) and therefore they are usually neglected as tools in, e.g., immunoassays. Previous studies have shown that low affinity biological interactions can be studied and exploited in affinity chromatography, biosensor technology and capillary electrophoresis. In this study we have demonstrated that IgM can be a useful ligand for analytical separation of antigens in weak affinity chromatography (WAC). A low affinity human monoclonal IgM antibody, directed at digoxin, was produced in a hybridoma cell culture, purified to homogeneity and immobilized onto an HPLC support. The IgM HPLC column displayed specific weak affinity retention in the 0.01-0.1 mM range as evaluated with digoxin and ouabain. The specificity was not affected when samples of ouabain in a crude environment of diluted serum were separated on the IgM column. These findings suggest an approach in immunoadsorbent technology where biomolecules can be analyzed and separated with weak affinity chromatography using IgM as a general affinity ligand.
IgM抗体通常亲和力较低（解离常数（Kd）> 10^(-5) M），因此在免疫分析等技术中，它们通常不作为工具使用。先前的研究表明，低亲和力的生物相互作用可在亲和色谱、生物传感器技术和毛细管电泳中进行研究和利用。在本研究中，我们证明了IgM在弱亲和力色谱（WAC）中可作为分析分离抗原的有用配体。一种针对地高辛的低亲和力人单克隆IgM抗体在杂交瘤细胞培养物中产生，纯化至同质后固定在HPLC载体上。用地高辛和哇巴因评估时，IgM HPLC柱在0.01 - 0.1 mM范围内显示出特异性的弱亲和力保留。当在稀释血清的粗环境中分离哇巴因样品时，IgM柱上的特异性不受影响。这些发现提示了免疫吸附技术中的一种方法，即使用IgM作为通用亲和配体，通过弱亲和力色谱对生物分子进行分析和分离。
