Multiplexed and High-Throughput Label-Free Detection of RNA/Spike Protein/IgG/IgM Biomarkers of SARS-CoV-2 Infection Utilizing Nanoplasmonic Biosensors

Abstract

To tackle the COVID-19 outbreak, which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), there is an unmet need for highly accurate diagnostic tests at all stages of infection with rapid results and high specificity. Here, we present a label-free nanoplasmonic biosensor-based, multiplex screening test for COVID-19 that can quantitatively detect 10 different biomarkers (6 viral nucleic acid genes, 2 spike protein subunits, and 2 antibodies) with a limit of detection in the aM range, all within one biosensor platform. Our newly developed nanoplasmonic biosensors demonstrate high specificity, which is of the upmost importance to avoid false responses. As a proof of concept, we show that our detection approach has the potential to quantify both IgG and IgM antibodies directly from COVID-19-positive patient plasma samples in a single instrument run, demonstrating the high-throughput capability of our detection approach. Most importantly, our assay provides receiving operating characteristics, areas under the curve of 0.997 and 0.999 for IgG and IgM, respectively. The calculated p -value determined through the Mann–Whitney nonparametric test is <0.0001 for both antibodies when the test of COVID-19-positive patients ( n = 80) is compared with that of healthy individuals ( n = 72). Additionally, the screening test provides a calculated sensitivity (true positive rate) of 100% (80/80), a specificity (true negative rate) >96% (77/80), a positive predictive value of 98% at 5% prevalence, and a negative predictive value of 100% at 5% prevalence. We believe that our very sensitive, multiplex, high-throughput testing approach has potential applications in COVID-19 diagnostics, particularly in determining virus progression and infection severity for clinicians for an appropriate treatment, and will also prove to be a very effective diagnostic test when applied to diseases beyond the COVID-19 pandemic.

Silver nanotriangle array based LSPR sensor for rapid coronavirus detection

“…These sensors are compact, durable, repeatable, and more reliable than traditional sensors, offering real-time and label-free chemical and biological detection. Label-free LSPR biosensors for SARS-CoV-2 screening have been reported for nucleic acids [10] , [11] , surface protein subunits (spike, envelope, and membrane) [12] , and SARS-CoV-2 virus [13] , [14] , as well as antibodies (IgG, IgM) [15] , [16] , [17] . For example, Ventura et al proposed a colorimetric sensor using gold nanoparticles for SARS-CoV-2 surface spike protein detection [12] .…”

A Study of the Detection of SARS-CoV-2 ORF1ab Gene by the Use of Electrochemiluminescent Biosensor Based on Dual-Probe Hybridization

“…The method can meet the requirements for analyzing simulated samples such as saliva and urine, with the advantages of high sensitivity, stable reproducibility, and anti-interference solid abilities. Compared with the existing reported detection method for the SARS-CoV-2 gene by ECL biosensor based on the amplification technique [ 24 , 25 ] and other biosensors [ 26 , 27 , 28 , 29 , 30 , 31 ], the ECL biosensor based on dual-probe hybridization detection of SARS-CoV-2 has improved sensitivity, as shown in Table 4 . It also has the advantages of simplifying the operation steps without amplifying and shortening the detection time, achieving rapid and highly sensitive detection within 30 min.…”

The Effects of Three-Dimensional Ligand Immobilization on Kinetic Measurements in Biosensors

“…The label-free sensors currently available on the market enable the investigation of hundreds of probes at a time against a single antigen [5], as well as a single probe against multiple targets [6]. The dynamic range of the available technology is incredibly large, both in terms of analyte molecular weight (MW) and concentration.…”