Nilson B, Björck L, Akerström B
Department of Physiological Chemistry, University of Lund, Sweden.
J Immunoassay. 1988;9(2):207-25. doi: 10.1080/15321818808057041.
Protein G, an IgG-binding protein, purified from the surface of group G streptococci, was coupled to alkaline phosphatase. The conjugate was used for detection of polyclonal goat and rabbit antibodies and monoclonal mouse IgG1, IgG2a and IgG2b in an enzyme-linked immunosorbent assay. A two-step coupling procedure was used, in which glutaraldehyde was allowed to react with the enzyme, excess glutaraldehyde was then removed by dialysis, and finally protein G added to the glutaraldehyde-activated and polymerized alkaline phosphatase. The activity and yield of the conjugates were then tested in an enzyme-linked immunosorbent assay. Coupling of 25 micrograms protein G to 5 mg alkaline phosphatase gave a conjugate which could be used for more than 10,000 determinations with maximal antibody binding giving an absorbance of 2.0. Under these conditions, there was no need for separation of the reactants before using the protein G-alkaline phosphatase complex.
从G组链球菌表面纯化得到的IgG结合蛋白G与碱性磷酸酶偶联。该偶联物用于酶联免疫吸附测定中检测多克隆山羊和兔抗体以及单克隆小鼠IgG1、IgG2a和IgG2b。采用两步偶联程序，其中戊二醛先与酶反应，然后通过透析去除过量的戊二醛，最后将蛋白G加入到经戊二醛活化和聚合的碱性磷酸酶中。然后在酶联免疫吸附测定中测试偶联物的活性和产率。将25微克蛋白G与5毫克碱性磷酸酶偶联得到的偶联物可用于10000多次测定，最大抗体结合时吸光度为2.0。在这些条件下，使用蛋白G-碱性磷酸酶复合物前无需分离反应物。
