# A Fully Integrated Point-of-Care Test for Ebola

> **NIH NIH R01** · DUKE UNIVERSITY · 2020 · $717,426

## Abstract

PROJECT SUMMARY
The objective of this proposal is to develop a new point-of-care test (POCT)—the D4 assay—for early field-
detection of Ebola virus (EBOV) infection. Currently, EBOV is diagnosed by RNA detection using reverse
transcriptase-polymerase chain reaction (RT-PCR). RT-PCR requires a makeshift BSL-4 grade laboratory in the
field, expensive equipment, and highly trained personnel. Other POCTs, including the user-friendly lateral-flow
assay (LFA), lack the sensitivity for early detection that is critical for timely intervention with the available
antibody cocktail that yields 90% survival rates for patients with low viremia. Our objective is motivated by an
urgent clinical need for a POCT that (1) detects EBOV infection in the field quickly and reliably, (2) requires little
on-field infrastructure, (3) yields results in 30 min, and (4) matches or exceeds the performance of RT-PCR. To
achieve these goals, we have designed an integrated POCT the D4 assay that has four simple steps—dispense,
dissolve, diffuse, and detect that require limited handling and skill to perform. This new-frontier technology takes
advantage of the presence of an unmistakable, viral secreted glycoprotein sGP that is present in the serum of
infected patients very early in infection. We have generated customized monoclonal antibodies (Abs) for sGP to
use in the D4 assay. The current prototype D4 assay that we have designed detects EBOV infection at least one
day earlier than RT-PCR in infected monkeys and at a far lower cost than RT-PCR or LFA. In this proposal,
we plan to advance our development and improve the sensitivity of the D4 assay further as well as reduce the
assay time from 60 min to 30 min. Our strategy is to increase the equilibrium binding constant of our current Ab
pair from ~10-9 M to ~10-11 M with antibody affinity maturation techniques and high-throughput screening of
antibody pairs. The enhanced D4 assay kit will have inkjet-printed capture and detection antibodies on a protein
and cell-resistant polymer brush on a glass plate encased in a passive capillarity microfluidics chip. The assay
output will be fluorescence of microspots on the D4 chip. We have developed a portable handheld fluorescence
detector to capture and image the spots and automatically convert them into the concentration of analytes for
quantitation and uploaded to a secure server. The design will be rigorously tested and validated with samples
from infected human cells and laboratory-challenged non-human primates. At the completion of this project, we
will have a field-ready, user-friendly, and highly sensitive POCT that will allow healthcare workers to detect EBOV
in serum, blood, or other bodily fluids in 30 min. The new design will push the current boundaries of EBOV
detection and facilitate more expedient deployment of infection control and patient support measures that can
yield 90% survival rates or better if implemented early in infection. Because the D4 POCT is multiplexable,...

## Key facts

- **NIH application ID:** 10119782
- **Project number:** 1R01AI150888-01A1
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** Ashutosh Chilkoti
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $717,426
- **Award type:** 1
- **Project period:** 2020-09-23 → 2025-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10119782

## Citation

> US National Institutes of Health, RePORTER application 10119782, A Fully Integrated Point-of-Care Test for Ebola (1R01AI150888-01A1). Retrieved via AI Analytics 2026-05-21 from https://api.ai-analytics.org/grant/nih/10119782. Licensed CC0.

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