Zweitzig Daniel R, Tibbe Arjan G, Nguyen Ai T, van Rijn Cees J M, Kopnitsky Mark J, Cichonski Kathleen, Terstappen Leon W M M
ZEUS Scientific Inc., 200 Evans Way, Branchburg, NJ 08876, USA.
Department of Research & Development, VyCAP, Hallenweg 23, 7522 NH Enschede, The Netherlands.
J Immunol Methods. 2016 Oct;437:21-7. doi: 10.1016/j.jim.2016.07.002. Epub 2016 Jul 21.
The intrinsic properties of silicon microsieves, such as an optically flat surface, high overall porosity, and low flow resistance have led to an increasing number of biotechnology applications. In this report, the feasibility of creating a microsieve-based immunoassay platform was explored. Microsieves containing 5μm pores were coupled with poly-acrylic acid chains, and then mounted into a plastic holder to enable rapid reagent exchanges via a wicking mechanism. The mounted microsieves were coated with infectious disease-related antigens at [2.5 and 25μg/mL], [20 and 50μg/mL], and [20 and 100μg/mL] to facilitate detection of serum-derived human antibodies against Rubella (3-day measles), B. burgdorferi (Lyme disease), or T. pallidum (syphilis), respectively. The prototype microsieve-based immunoassay platform was able to distinguish positive control sera containing antibodies against Rubella, T. pallidum, and B. burgdorferi from negative control sera with similar qualitative results as FDA-approved ELISA tests. Testing of a WHO IgG syphilitic standard at 0.3, 0.15, 0.075, 0.0375, and 0.01875IU/mL demonstrated that the T. pallidum microsieve assay is able to distinguish disease-specific IgG signal from background signal at similar, and possibly lower, levels than the corresponding ELISA. The T. pallidum microsieve assay prototype also differentiated positive clinical serum samples from negative donor samples, and the results were in good agreement with ELISA (R(2)=0.9046). These feasibility studies demonstrate the potential for utilizing microsieves, along with a reagent wicking device, as a simple diagnostic immunoassay platform.
硅微筛的固有特性，如光学平面表面、高总孔隙率和低流动阻力，已导致其在生物技术领域的应用越来越多。在本报告中，探讨了创建基于微筛的免疫分析平台的可行性。含有5μm孔的微筛与聚丙烯酸链偶联，然后安装到塑料支架中，通过毛细作用机制实现快速试剂交换。将安装好的微筛分别用浓度为[2.5和25μg/mL]、[20和50μg/mL]、[20和100μg/mL]的传染病相关抗原包被，以促进检测血清来源的针对风疹（三日麻疹）、伯氏疏螺旋体（莱姆病）或梅毒螺旋体（梅毒）的人抗体。基于微筛的免疫分析平台原型能够将含有抗风疹、梅毒螺旋体和伯氏疏螺旋体抗体的阳性对照血清与阴性对照血清区分开来，定性结果与FDA批准的ELISA检测相似。对世界卫生组织0.3、0.15、0.075、0.0375和0.01875IU/mL的IgG梅毒标准品进行检测表明，梅毒螺旋体微筛检测能够在与相应ELISA相似甚至可能更低的水平上区分疾病特异性IgG信号与背景信号。梅毒螺旋体微筛检测原型还能区分阳性临床血清样本和阴性供体样本，结果与ELISA高度一致（R(2)=0.9046）。这些可行性研究证明了利用微筛以及试剂毛细作用装置作为简单诊断免疫分析平台的潜力。
