Electrochemical Capillary-Flow Immunoassay for Detecting Anti-SARS-CoV-2 Nucleocapsid Protein Antibodies at the Point of Care

Abstract

Rapid and inexpensive serological tests for SARS-CoV-2 antibodies are needed to conduct population-level seroprevalence surveillance studies and can improve diagnostic reliability when used in combination with viral tests. Here, we report a novel low-cost electrochemical capillary-flow device to quantify IgG antibodies targeting SARS-CoV-2 nucleocapsid proteins (anti-N antibody) down to 5 ng/mL in low-volume (10 μL) human whole blood samples in under 20 min. No sample preparation is needed as the device integrates a blood-filtration membrane for on-board plasma extraction. The device is made of stacked layers of a hydrophilic polyester and double-sided adhesive films, which create a passive microfluidic circuit that automates the steps of an enzyme-linked immunosorbent assay (ELISA). The sample and reagents are sequentially delivered to a nitrocellulose membrane that is modified with a recombinant SARS-CoV-2 nucleocapsid protein. When present in the sample, anti-N antibodies are captured on the nitrocellulose membrane and detected via chronoamperometry performed on a screen-printed carbon electrode. As a result of this quantitative electrochemical readout, no result interpretation is required, making the device ideal for point-of-care (POC) use by non-trained users. Moreover, we show that the device can be coupled to a near-field communication potentiostat operated from a smartphone, confirming its true POC potential. The novelty of this work resides in the integration of sensitive electrochemical detection with capillary-flow immunoassay, providing accuracy at the point of care. This novel electrochemical capillary-flow device has the potential to aid the diagnosis of infectious diseases at the point of care.

Electrochemical Immunoassay for the Detection of SARS-CoV-2 Nucleocapsid Protein in Nasopharyngeal Samples

“…Although assay time was greatly reduced compared to traditional lab-based methods such as ELISA, RT-qPCR, and plaque assays, time from the sample to the result remains approximately 70 min. Future work is aimed at automating the assay by integrating it in a fluidic platform amenable to the point of care, 51 further reducing the assay time, and assessing the stability of the functionalized SPCEs over extended time periods under various storage conditions. This will enable the assay to be integrated into a robust, quantitative, and sensitive POC test for the detection of SARS-CoV-2 infection in patients.…”

Emerging Electrochemical Biosensing Trends for Rapid Diagnosis of COVID-19 Biomarkers as Point-of-Care Platforms: A Critical Review

“…This assay meets ASSURED criteria, as it utilizes very low-cost sensing equipment, requires low sample volumes of <10 μL, and allows rapid analysis with high sensitivity. 3 A cotton-tipped electrode was successfully utilized for the detection of SARS-CoV-2 by quantifying nucleocapsid protein with a limit of detection of 0.8 pg/mL. A screen-printed electrode was modified with carbon nanofibers with diazonium by an electrografting method.…”

“…At-home detection methods with smartphone-based readouts are needed for early disease identification. 3 …”

Recent advances of functional nucleic acid-based sensors for point-of-care detection of SARS-CoV-2

“…Analysis of nearly 400 clinical samples yielded 100% sensitivity and 100% specificity. Electrochemical capillary osmosis devices [ 24 ], hematological magnetic immunoassay [ 25 ], and microflow cytometry-based agglutination immunoassay [ 26 ] were also developed for COVID-19 diagnosis, combined with smartphones to obtain faster test results.…”