Clinical validation of automated and rapid mariPOC SARS-CoV-2 antigen test

Abstract

COVID-19 diagnostics was quickly ramped up worldwide early 2020 based on the detection of viral RNA. However, based on the scientific knowledge for pre-existing coronaviruses, it was expected that the SARS-CoV-2 RNA will be detected from symptomatic and at significant rates also from asymptomatic individuals due to persistence of non-infectious RNA. To increase the efficacy of diagnostics, surveillance, screening and pandemic control, rapid methods, such as antigen tests, are needed for decentralized testing and to assess infectiousness. A novel automated mariPOC SARS-CoV-2 test was developed for the detection of conserved structural viral nucleocapsid proteins. The test utilizes sophisticated optical laser technology for two-photon excitation and individual detection of immunoassay solid-phase particles. We validated the new method against qRT-PCR. Sensitivity of the test was 100.0% (13/13) directly from nasopharyngeal swab specimens and 84.4% (38/45) from swab specimens in undefined transport mediums. Specificity of the test was 100.0% (201/201). The test's limit of detection was 2.7 TCID₅₀/test. It showed no cross-reactions. Our study shows that the new test can detect infectious individuals already in 20 min with clinical sensitivity close to qRT-PCR. The mariPOC is a versatile platform for syndromic testing and for high capacity infection control screening of infectious individuals.

Figure 1

(a) Diagnostic workflow in the mariPOC SARS-CoV-2 test and schematic sample dispensing into the test cartridge. Nasopharyngeal sample is suspended into the sample buffer by vortexing before placing the sample tube into the analyzer for automated analysis. The sample is automatically dispensed into the test plate reaction well(s) through resealing multilayer cover, which upper and lower pierceable layers are aluminum foil and cross-cut sheet, respectively. Immunometric reactions start when the sample dissolves the dried reagents. Test result is reported objectively as positive or negative. (b) Schematic principle of the SARS-CoV-2 assay where nucleocapsid proteins are detected based on sandwich immunoassay and two-photon excitation fluorescent measurement of individual microparticles by confocal microscopy. Grey hourglass-shaped area designates excitation light bath. Reddish oval-shaped area designates the focal volume where two-photon excitation of fluorescence takes place.

Figure 2

Ct values of qRT-PCR for mariPOC test positive (green dots) and negative (red dots) samples in the validation sample cohort 2. Dashed line is at Ct 33.24, which was the lowest detected Ct.