Department of Physics, University of Ottawa, Ottawa, Canada.
Nat Commun. 2021 Sep 9;12(1):5348. doi: 10.1038/s41467-021-25566-8.
Single-molecule counting is the most accurate and precise method for determining the concentration of a biomarker in solution and is leading to the emergence of digital diagnostic platforms enabling precision medicine. In principle, solid-state nanopores-fully electronic sensors with single-molecule sensitivity-are well suited to the task. Here we present a digital immunoassay scheme capable of reliably quantifying the concentration of a target protein in complex biofluids that overcomes specificity, sensitivity, and consistency challenges associated with the use of solid-state nanopores for protein sensing. This is achieved by employing easily-identifiable DNA nanostructures as proxies for the presence ("1") or absence ("0") of the target protein captured via a magnetic bead-based sandwich immunoassay. As a proof-of-concept, we demonstrate quantification of the concentration of thyroid-stimulating hormone from human serum samples down to the high femtomolar range. Further optimization to the method will push sensitivity and dynamic range, allowing for development of precision diagnostic tools compatible with point-of-care format.
单分子计数是确定溶液中生物标志物浓度的最准确和最精确的方法，它正在引领数字诊断平台的出现，从而实现精准医疗。原则上，具有单分子灵敏度的固态纳米孔——全电子传感器非常适合这项任务。在这里，我们提出了一种数字免疫测定方案，能够可靠地定量检测复杂生物流体中目标蛋白的浓度，该方案克服了使用固态纳米孔进行蛋白传感时与特异性、灵敏度和一致性相关的挑战。这是通过使用易于识别的 DNA 纳米结构作为存在（“1”）或不存在（“0”）的替代物来实现的，这些结构是通过基于磁珠的三明治免疫测定捕获目标蛋白的。作为概念验证，我们证明了可以从人血清样本中定量检测到促甲状腺激素的浓度，低至高飞摩尔范围。进一步优化该方法将提高灵敏度和动态范围，从而开发出与即时诊断格式兼容的精密诊断工具。
