Surface-enhanced Raman scattering immunoassays using a rotated capture substrate
Surface-enhanced Raman scattering immunoassays using a rotated capture substrate
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DOI：
10.1021/ac0701031
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发表时间：
2007-06-01
影响因子：
7.4
通讯作者：
Porter, Marc D.
中科院分区：
文献类型：
--
作者：
Driskell, Jeremy D.;Uhlenkamp, Jill M.;Porter, Marc D.
一种快速、灵敏的免疫吸附分析格式已经开发出来，以满足医疗、兽医和生物恐怖主义预防领域日益增长的性能要求(即减少孵化时间和检测限制)。本文介绍了旋转捕获底物的概念，作为一种简便的方法，增加了抗原和标记到固相表面的通量，从而减少了分析时间。为此，进行了夹心型分析，将抗体-抗原相互作用的特异性与表面增强拉曼散射检测的高灵敏度结合起来。为了研究这一策略，通过硫代偶联剂将抗兔多克隆抗体固定在金捕获底物上。然后将能够控制旋转的捕获底物浸入含有兔免疫球蛋白的样品溶液中，作为模型分析物。底物与目标抗体结合后，浸泡并旋转在外源拉曼标记(ERL)溶液中，该溶液由包被芳香族部分的金纳米颗粒(60 Nm)作为拉曼散射体，抗体作为生物特异性识别元件。研究了底物旋转对抗原结合和ERL标记步骤的影响。优化的旋转条件使分析时间从24小时减少到25分钟，检测限提高了10倍。最后，将所开发的方法应用于山羊血清中悬浮的兔免疫球蛋白的检测，以评估其在生物基质中的性能。
A rapid, sensitive format for immunosorbent assays has been developed to meet the increasing levels of performance (i.e., reduction of incubation times and detection limits) demanded in the medical, veterinary, and bioterrorism prevention arenas. This paper introduces the concept of a rotating capture substrate as a facile means to increase the flux of antigen and label to the solid-phase surface and thereby reduce assay time. To this end, a sandwich-type assay is carried out that couples the specificity of antibody-antigen interactions with the high sensitivity of surface-enhanced Raman scattering detection. To investigate this strategy, polyclonal anti-rabbit IgG was immobilized on a gold capture substrate via a thiolate coupling agent. The capture substrate, capable of controlled rotation, was then immersed in a sample solution containing rabbit IgG, which served as a model analyte. After binding the target IgG, the substrates were immersed and rotated in an extrinsic Raman label (ERL) labeling solution, which is composed of gold nanoparticles (60 nm) coated with an aromatic moiety as the Raman scatterer and an antibody as the biospecific recognition element. The effect of substrate rotation on both the antigen binding and ERL labeling steps was investigated. Implementation of optimized rotation conditions resulted in the reduction of assay times from 24 h to 25 min and a 10-fold improvement in the limit of detection. Finally, the developed protocol was applied to the detection of rabbit IgG suspended in goat serum, which served to assess performance in a biological matrix.