High-Throughput, High-Multiplex Digital Protein Detection with Attomolar Sensitivity

多路复用 分析物 多路复用 仪表(计算机编程) 微流控 生物分子 吞吐量 灵敏度(控制系统) 计算机科学 纳米技术 计算生物学 材料科学 化学 生物信息学 色谱法 电子工程 生物 工程类 电信 操作系统 无线
作者
Connie Wu,Tyler Dougan,David R. Walt
出处
期刊:ACS Nano [American Chemical Society]
卷期号:16 (1): 1025-1035 被引量:94
标识
DOI:10.1021/acsnano.1c08675
摘要

A major challenge in many clinical diagnostic applications is the measurement of low-abundance proteins and other biomolecules in biological fluids. Digital technologies such as the digital enzyme-linked immunosorbent assay (ELISA) have enabled 1000-fold increases in sensitivity over conventional protein detection methods. However, current digital ELISA technologies still possess insufficient sensitivities for many rare protein biomarkers and require specialized instrumentation or time-consuming workflows that have limited their widespread implementation. To address these challenges, we have developed a more sensitive and streamlined digital ELISA platform, Molecular On-bead Signal Amplification for Individual Counting (MOSAIC), which attains low attomolar limits of detection, with an order of magnitude enhancement in sensitivity over these other methods. MOSAIC uses a rapid, automatable flow cytometric readout that vastly increases throughput and is easily integrated into existing laboratory infrastructure. As MOSAIC provides high sampling efficiencies for rare target molecules, assay bead number can readily be tuned to enhance signal-to-background with high measurement precision. Furthermore, the solution-based signal readout of MOSAIC expands the number of analytes that can simultaneously be measured for higher-order multiplexing with femtomolar sensitivities or below, compared with microwell- or droplet-based digital methods. As a proof of principle, we apply MOSAIC toward improving the detectability of low-abundance cytokines in saliva and ultrasensitive multiplexed measurements of eight protein analytes in plasma and saliva. The attomolar sensitivity, high throughput, and broad multiplexing abilities of MOSAIC provide highly accessible and versatile ultrasensitive capabilities that can potentially accelerate protein biomarker discovery and diagnostic testing for diverse disease applications.
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