Microfluidic sieve and detector for rapid ultrasensitive assays with single-molecule sensitivity

Abstract

The advent of ultrasensitive assays enabled many new research and clinical applications, but the time-to-result is hours and has not significantly improved over time, limiting their potential. We introduce the microfluidic sieve and detector (MFSD) that can sieve 200 microlitres in just one minute, and quantify captured analytes in four minutes, down to zeptomolar (10^-19M) concentration. The MFSD comprises half a million 8-m-diameter pores that bind analytes upon Brownian motion-induced wall collision. Conventional ensemble models and a stochastic Brownian agent dynamics introduced here reveal rapid, predictable and tuneable analyte capture under flow. Digital sandwich assays are completed by sequentially flowing the sample and reagents, partitioning the pores, and digitally revealing single trapped analytes by enzymatic amplification. The MFSD tests are specific and reproducible, exhibit a large dynamic range, and are simple, tuneable and versatile, enabling measurement of different proteins including influenza A nucleoprotein in clinical samples, and even nucleic acids while being both faster and more sensitive than PCR. As such, the MFSD could open a new chapter for analysis and diagnostics.