Point-of-Need Disposable ELISA System for COVID-19 Serology Testing

Abstract

A disposable enzyme-linked immunosorbent assay (dELISA) device for ate-home or doctor’s office use was developed to detect SARS-CoV-2 antibodies. Serology testing for SARS-CoV-2 antibodies is currently run using well-plate ELISAs in centralized laboratories. However, the scale of serology testing needed for epidemiological and clinical screening studies will overwhelm existing clinical laboratory resources. Instead, a point-of-need device that can be used at home or in doctor’s offices for COVID-19 serology testing must be developed and is one of four target products prioritized by the World Health Organization. Lateral flow assays are common and easy to use, but lack the sensitivity needed to reliably detect SARS-CoV-2 antibodies in clinical samples. This work describes a disposable ELISA device that is as simple to use as a lateral flow assay, but as sensitive as a well-plate ELISA. The device utilizes capillary-driven flow channels made of transparency films and double-sided adhesive combined with paper pumps to drive flow. The geometry of the channels and storage pads enables automated sequential washing and reagent addition steps with two simple end-user steps. An enzyme label is used to produce a colorimetric signal instead of a nanoparticle label in order to amplify signal and increase sensitivity, while the integrated washing steps decrease false positives and increase reproducibility. Naked-eye detection can be used for qualitative results or a smartphone camera for quantitative analysis. The device can detect antibodies at 2.8 ng/mL from whole blood, which was very close the concentration of detectable target in a well-plate ELISA (1.2 ng/mL). In this study the dELISA system was used to detect SARS-CoV-2 antibodies, but we believe that the device represents a fundamental step forward in point-of-care technology that will enable sensitive detection of many other analytes outside of a centralized laboratory.

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

“…This observation is consistent with results published by our group and others. 37 , 42 , 43 Following plasma delivery to the NCM and a subsequent buffer wash ( Figure 2 B), the yellow dye that had dried on the reagent pad to mimic the HRP-antibody was released through the HRP-antibody delivery channel and flowed to the detection zone ( Figure 2 C, 12 min). Simultaneously, buffer flowed through the hydration channel to keep the blood-filtration membrane hydrated, driving the flow to the NCM.…”

“…Future work shall focus on tuning the NFC potentiostat to match its recording range to output signals from samples of clinically low analyte concentration. Whilst there is preliminary evidence showing stability over storage of every active component of the device (the electrode, 41 the N protein modified NCM 37 and the detection antibody reagent pad 40 ), a stability study of the complete device was not conducted yet and would be necessary for the device to access the POC market. Lastly, further developments are needed to increase the reproducibility of our measurements.…”

“…N protein striped NCMs were stored at 4 °C for up to a month before use, as stability over storage was illustrated elsewhere. 37 …”

“…The capillary-flow device was fabricated similarly to a device previously reported, 37 with major differences including the design and number of layers, the materials used and the integrated SPCE. Here, the device was constructed by stacking four layers of a 100 μm thick hydrophilic polyester film (PP2500, 3 M) intercalated with three layers of a 50 μm thick double-sided adhesive (467 MP, 3 M).…”

“…Our novel assay is applied to the detection of IgG antibodies against a SARS-CoV-2 nucleocapsid protein (anti-N antibodies) in human blood samples. An anti-N IgG antibody was chosen as the analytical target of our novel device because of its clinical relevance in COVID-19 diagnosis and patient prognosis. − Based on our previous work, , the device developed here is made of stacked layers of polyester and double-sided adhesive films, which create a capillary-flow microfluidic circuit. The device integrates a blood-filtration membrane for on-board plasma extraction.…”