Edited by Taekjip Ha, Johns Hopkins University, Baltimore, MD, and approved August 15, 2017 (received for review May 18, 2017)
Basic research and medical diagnostics rely on the ability to detect and quantify specific proteins in biological fluids. While numerous current detection techniques exist, these are often limited by trade-offs between ease of use, sensitivity, and cost. Here, we present the nanoswitch-linked immunosorbent assay (NLISA), an accessible, sensitive, and low-cost detection platform that is based upon nanoscale devices that change confirmation upon binding a target protein. NLISA is surface-free and includes a kinetic-proofreading purification step, enabling both enhanced sensitivity and the ability to accurately distinguish between similar proteins from different strains of the same virus or that differ by only a single mutation. Our method is also readily transferable to point-of-care devices due to an easy readout and few hands-on steps.
Protein detection and quantification play critical roles in both basic research and clinical practice. Current detection platforms range from the widely used ELISA to more sophisticated, and more expensive, approaches such as digital ELISA. Despite advances, there remains a need for a method that combines the simplicity and cost-effectiveness of ELISA with the sensitivity and speed of modern approaches in a format suitable for both laboratory and rapid, point-of-care applications. Building on recent developments in DNA structural nanotechnology, we introduce the nanoswitch-linked immunosorbent assay (NLISA), a detection platform based on easily constructed DNA nanodevices that change conformation upon binding to a target protein with the results read out by gel electrophoresis. NLISA is surface-free and includes a kinetic-proofreading step for purification, enabling both enhanced sensitivity and reduced cross-reactivity. We demonstrate femtomolar-level detection of prostate-specific antigen in biological fluids, as well as reduced cross-reactivity between different serotypes of dengue and also between a single-mutation and wild-type protein. NLISA is less expensive, uses less sample volume, is more rapid, and, with no washes, includes fewer hands-on steps than ELISA, while also achieving superior sensitivity. Our approach also has the potential to enable rapid point-of-care assays, as we demonstrate by performing NLISA with an iPad/iPhone camera for imaging.
↵1Present address: Confer Health, Inc., Charlestown, MA 02129.
Author contributions: C.H.H., D.Y., M.A.K., and W.P.W. designed research; C.H.H. and D.Y. performed research; C.H.H. and D.Y. analyzed data; C.H.H., D.Y., M.A.K., and W.P.W. wrote the paper; and W.P.W. directed research.
Conflict of interest statement: Patent applications have been filed for various aspects of this work by all authors. M.A.K. and W.P.W. are founders of and have equity interest in Confer Health, Inc., a company that is developing point-of-care diagnostics.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1708148114/-/DCSupplemental.
Freely available online through the PNAS open access option.
