Department of Chemical Engineering and Centre for Biosensors, Biodevices and Bioelectronics (C3Bio), University of Bath, Claverton Down, Bath BA2 7AY, United Kingdom.
Reading School of Pharmacy, University of Reading, Whiteknights Campus, Reading, RG6 6AD United Kingdom.
ACS Sens. 2021 Dec 24;6(12):4338-4348. doi: 10.1021/acssensors.1c01524. Epub 2021 Dec 2.
A range of biosensing techniques including immunoassays are routinely used for quantitation of analytes in biological samples and available in a range of formats, from centralized lab testing (e.g., microplate enzyme-linked immunosorbent assay (ELISA)) to automated point-of-care (POC) and lateral flow immunochromatographic tests. High analytical performance is intrinsically linked to the use of a sequence of reagent and washing steps, yet this is extremely challenging to deliver at the POC without a high level of fluidic control involving, e.g., automation, fluidic pumping, or manual fluid handling/pipetting. Here we introduce a microfluidic siphon concept that conceptualizes a multistep ″dipstick″ for quantitative, enzymatically amplified immunoassays using a strip of microporous or microbored material. We demonstrated that gravity-driven siphon flow can be realized in single-bore glass capillaries, a multibored microcapillary film, and a glass fiber porous membrane. In contrast to other POC devices proposed to date, the operation of the siphon is only dependent on the hydrostatic liquid pressure (gravity) and not capillary forces, and the unique stepwise approach to the delivery of the sample and immunoassay reagents results in zero dead volume in the device, no reagent overlap or carryover, and full start/stop fluid control. We demonstrated applications of a 10-bore microfluidic siphon as a portable ELISA system without compromised quantitative capabilities in two global diagnostic applications: (1) a four-plex sandwich ELISA for rapid smartphone dengue serotype identification by serotype-specific dengue virus NS1 antigen detection, relevant for acute dengue fever diagnosis, and (2) quantitation of anti-SARS-CoV-2 IgG and IgM titers in spiked serum samples. Diagnostic siphons provide the opportunity for high-performance immunoassay testing outside sophisticated laboratories, meeting the rapidly changing global clinical and public health needs.
一系列生物传感技术，包括免疫测定法，通常用于定量分析生物样本中的分析物，并且有多种格式可供选择，从集中式实验室测试（例如微孔板酶联免疫吸附测定（ELISA））到自动化即时检测（POC）和侧向流动免疫层析测试。高分析性能与试剂和洗涤步骤的顺序使用密切相关，但如果没有高水平的流体控制（例如自动化、流体泵送或手动流体处理/移液），在 POC 中实现这一点极具挑战性。在这里，我们介绍了一种微流虹吸概念，该概念将多步“比色条”概念化为使用微孔或微孔材料条进行定量、酶放大免疫测定。我们证明，重力驱动的虹吸流可以在单孔玻璃毛细管、多孔微毛细管膜和玻璃纤维多孔膜中实现。与迄今为止提出的其他 POC 设备不同，虹吸的操作仅取决于静水液体压力（重力）而不是毛细作用力，并且独特的逐步输送样品和免疫测定试剂的方法导致设备中无死体积、无试剂重叠或残留，以及完全的起始/停止流体控制。我们展示了 10 孔微流虹吸作为无牺牲定量能力的便携式 ELISA 系统的应用，在两个全球诊断应用中：(1) 用于通过血清型特异性登革热病毒 NS1 抗原检测快速智能手机登革热血清型鉴定的四聚体夹心 ELISA，这与急性登革热诊断相关，以及 (2) 定量测定 Spike 血清样本中的抗 SARS-CoV-2 IgG 和 IgM 滴度。诊断虹吸为在复杂实验室之外进行高性能免疫测定测试提供了机会，满足了快速变化的全球临床和公共卫生需求。
