Ehle H, Horn A
Institute of Biochemistry, Friedrich Schiller University Jena, GDR.
Bioseparation. 1990;1(2):97-110.
Immunoaffinity chromatography of enzymes represents an attractive purification technique suitable for one-step and large-scale purification of enzymes to homogeneity. Monoclonal and polyclonal antibodies can be used equally well. The broad use of the technique is restricted by the harsh elution conditions which are often required. The efforts to overcome these limitations and to optimize the method are reviewed, viz. proenzyme purification, purification of enzymes as part of multienzyme complexes carried out by a mild dissociation step, specific elution by substrates and effectors, enzyme stabilization, electrophoretical desorption and negative elution by adsorbing impurities from the crude extract, and hypotonic elution. The current practice is discussed considering antibody and enzyme selection, optimization of elution conditions, and washing steps using different media. Representative examples are given for various approaches.
酶的免疫亲和色谱法是一种颇具吸引力的纯化技术，适用于将酶一步纯化至均一状态以及大规模纯化。单克隆抗体和多克隆抗体均可同样有效地使用。该技术的广泛应用受到常常所需的严苛洗脱条件的限制。本文综述了为克服这些限制以及优化该方法所做的努力，即酶原纯化、通过温和解离步骤对作为多酶复合物一部分的酶进行纯化、利用底物和效应物进行特异性洗脱、酶的稳定化、电泳解吸以及通过从粗提物中吸附杂质进行负洗脱，还有低渗洗脱。结合抗体和酶的选择、洗脱条件的优化以及使用不同介质的洗涤步骤，对当前的实践进行了讨论。针对各种方法给出了代表性实例。
