Abstract
The detection of proteins is of central importance to biomolecular analysis and diagnostics, yet fundamental limitations due to the surface-based nature of most sensing approaches persist, and limited improvements have been designed to integrate multimodal information beyond concentration measurements. Here we present a single-molecule microfluidic sensing platform for digital protein biomarker detection in solution, termed digital immunosensor assay (DigitISA). DigitISA is based on microchip electrophoretic separation combined with single-molecule detection and enables absolute number–concentrations quantification of proteins in a single, solution-phase step. Applying DigitISA to a range of targets including amyloid aggregates, exosomes, and biomolecular condensates, we demonstrate that the assay provides information beyond stoichiometric interactions, and enables characterization of immunochemistry, binding affinity, and protein biomarker abundance. Together, DigitISA constitutes a new experimental paradigm for the digital sensing of protein biomarkers, and enables analyses of targets that would otherwise be hard or impossible to address by conventional immuno-sensing techniques.
Competing Interest Statement
G.K., K.L.S., W.E.A., and T.P.J.K. declare the following competing interests. Parts of this work have been the subject of a patent application filed by Cambridge Enterprise Limited, a fully owned subsidiary of the University of Cambridge. Inventors: Krainer, G.; Saar, K.L.; Arter, W.E., Knowles, T.P.J.; Applicant: Cambridge Enterprise Ltd.; Title: Highly sensitive biomolecule detection and quantification. Publication Number: WO/2021/176065; Publication Date: 10.09.2021; International Application No.: PCT/EP2021/055614; International Filing Date: 05.03.2021. The remaining authors declare no competing interests.
Footnotes
n/a