# Simplified Single Molecule Protein Assays with Unprecedented Sensitivity

> **NIH NIH R56** · BRIGHAM AND WOMEN'S HOSPITAL · 2022 · $447,500

## Abstract

PROJECT SUMMARY
The ability to measure extremely low levels of biomolecules accurately and rapidly is essential for diagnosing
and monitoring many diseases. While sufficient for certain biomarkers, the sensitivities of most existing
diagnostic systems are inadequate for measuring many protein biomarkers that exist in easily accessible
biofluids at concentrations much lower than their detection limits, which remain in the picomolar range. In this
application, we propose to develop, design, engineer, and refine a new ultrasensitive single molecule protein
analysis platform that will be able to routinely measure attomolar protein concentrations, which we call
Molecular On-bead Signal Amplification for Individual Counting (MOSAIC). MOSAIC transforms single
molecule measurements into a simplified assay format via on-bead signal localization, which has the potential
to be integrated into a point of care (POC) device. In MOSAIC, a non-diffusible signal is generated on each
bead carrying a target molecule, creating an on-bead signal that remains attached for prolonged periods of
time, thereby enabling alternative detection schemes to be employed that do not require bead loading into
microwells or well sealing to localize signals. A key challenge to be addressed in the proposed work will be to
ensure that this MOSAIC platform can consistently outperform current ultrasensitive protein detection
technologies in sensitivity by two orders of magnitude across many protein analytes, which in turn lays the
foundation for future work in translating this enhanced analytical sensitivity to improved clinical sensitivity and
specificity in diagnostic applications. In Aims 1 and 2, we will optimize signal generation and readout methods
for MOSAIC and expand its multiplexing capabilities. In Aim 3, we plan to integrate MOSAIC into a disposable
cartridge and a portable instrument with automated sample processing, liquid handling, incubation, imaging,
and sample analysis. The resulting sample-to-answer biosensing technology will provide an ultrasensitive
diagnostic platform that can be readily and affordably utilized in the central clinical laboratory, in emergency
departments, and in resource-limited settings, enabling rapid, accurate disease diagnosis and early treatments.

## Key facts

- **NIH application ID:** 10644163
- **Project number:** 1R56EB032826-01
- **Recipient organization:** BRIGHAM AND WOMEN'S HOSPITAL
- **Principal Investigator:** DAVID R. WALT
- **Activity code:** R56 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $447,500
- **Award type:** 1
- **Project period:** 2022-09-01 → 2023-05-04

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10644163

## Citation

> US National Institutes of Health, RePORTER application 10644163, Simplified Single Molecule Protein Assays with Unprecedented Sensitivity (1R56EB032826-01). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10644163. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*