Wilson Robert, Clavering Charles, Hutchinson Alistair
Department of Chemistry, Liverpool University, Liverpool L69 7ZD, UK.
Anal Chem. 2003 Aug 15;75(16):4244-9. doi: 10.1021/ac034163s.
Electrochemiluminescence enzyme immunoassays for 2,4,6-trinitrotoluene (TNT) and pentaerythritol tetranitrate (PETN) are described. The latter is, to the best of our knowledge, the first report of an immunoassay for PETN. Haptens corresponding to these explosives were covalently attached to high-affinity dextran-coated paramagnetic beads. The beads were mixed with the corresponding Fab fragments and the sample. After adding a second HRP-labeled antispecies-specific antibody, the mixture was pumped into an electrochemiluminometer where beads were concentrated on the working electrode magnetically. The amount of analyte in the sample was determined by measuring light emission when H2O2 was generated electrochemically in the presence of luminol and an enhancer. The detection limits for TNT and PETN were 0.11 and 19.8 ppb, respectively. Details of bead preparation and performance are given. The increase in sensitivity obtained when Fab fragments are used instead of whole antibodies is explained, and the implications of this observation for nanoparticle-based assays are discussed.
本文描述了用于检测2,4,6-三硝基甲苯（TNT）和季戊四醇四硝酸酯（PETN）的电化学发光酶免疫分析方法。据我们所知，后者是关于PETN免疫分析的首次报道。与这些炸药对应的半抗原被共价连接到高亲和力的葡聚糖包被顺磁珠上。将珠子与相应的Fab片段及样品混合。加入第二种辣根过氧化物酶（HRP）标记的抗物种特异性抗体后，将混合物泵入电化学发光仪，珠子在其中通过磁力被浓缩在工作电极上。当在鲁米诺和增强剂存在的情况下通过电化学方法产生过氧化氢时，通过测量发光来确定样品中分析物的量。TNT和PETN的检测限分别为0.11和19.8 ppb。文中给出了珠子制备和性能的详细信息。解释了使用Fab片段而非完整抗体时灵敏度提高的原因，并讨论了这一观察结果对基于纳米颗粒的分析的意义。
