Gerdes M, Meusel M, Spener F
Institut für Chemo- und Biosensorik, Münster, Germany.
J Immunol Methods. 1999 Mar 4;223(2):217-26. doi: 10.1016/s0022-1759(98)00211-7.
The influence of antibody valency in a displacement immunoassay was investigated by comparing the whole antibody molecule with the corresponding Fab-fragment. The displacement immunoassay for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) takes advantage of the cross-reactivity of monoclonal anti-2,4-D antibodies and the Fab-fragments toward immobilized 2-methyl-4-chlorophenoxyacetic acid (MCPA). Due to the low affinity of the antibodies toward MCPA (cross-reactivity of approximately 30%), the addition of 2,4-D resulted in a displacement of the antibodies or the fragments. The detection limits obtained with whole anti-2,4-D antibodies and Fab-fragments were 0.1 microg/l and 0.01 microg/l 2,4-D, respectively. The whole antibodies and the Fab-fragments show similarities, such as the cross-reactivity toward MCPA (26% and 33%), and some characteristics of the calibration curve, for example the large detection range and the sensitivity. In contrast to the bivalent antibodies, however, increasing the hapten/protein ratios of the immobilized MCPA-BSA conjugates did not affect the detection limit when using the Fab-fragments. Moreover, kinetic experiments reveal a faster displacement reaction with the Fab-fragments. A disadvantage of using the Fab-fragments is the generation of lower absorbance values in the ELISA.
通过将完整抗体分子与相应的Fab片段进行比较，研究了置换免疫分析中抗体价数的影响。除草剂2,4-二氯苯氧乙酸（2,4-D）的置换免疫分析利用了单克隆抗2,4-D抗体和Fab片段对固定化2-甲基-4-氯苯氧乙酸（MCPA）的交叉反应性。由于抗体对MCPA的亲和力较低（交叉反应性约为30%），加入2,4-D会导致抗体或片段的置换。用完整抗2,4-D抗体和Fab片段获得的检测限分别为0.1μg/l和0.01μg/l 2,4-D。完整抗体和Fab片段表现出相似性，如对MCPA的交叉反应性（分别为26%和33%）以及校准曲线的一些特征，例如检测范围大、灵敏度高。然而，与二价抗体不同的是，使用Fab片段时，固定化MCPA-BSA偶联物的半抗原/蛋白质比率增加不会影响检测限。此外，动力学实验表明，Fab片段的置换反应更快。使用Fab片段的一个缺点是在酶联免疫吸附测定（ELISA）中产生较低的吸光度值。
