Dual-detection fluorescent immunochromatographic assay for quantitative detection of SARS-CoV-2 spike RBD-ACE2 blocking neutralizing antibody

Abstract

The global effort against the COVID-19 pandemic dictates that routine quantitative detection of SARS-CoV-2 neutralizing antibodies is vital for assessing immunity following periodic revaccination against new viral variants. Here, we report a dual-detection fluorescent immunochromatographic assay (DFIA), with a built-in self-calibration process, that enables rapid quantitative detection of neutralizing antibodies that block binding between the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein and the angiotensin-converting enzyme 2 (ACE2). Thus, this assay is based on the inhibition of binding between ACE2 and the RBD of the SARS-CoV-2 spike protein by neutralizing antibodies, and the affinity of anti-human immunoglobulins for these neutralizing antibodies. Our self-calibrating DFIA shows improved precision and sensitivity with a wider dynamic linear range, due to the incorporation of a ratiometric algorithm of two-reverse linkage signals responding to an analyte. This was evident by the fact that no positive results (0/14) were observed in verified negative samples, while 22 positives were detected in 23 samples from verified convalescent plasma. A comparative analysis of the ability to detect neutralizing antibodies in 266 clinical serum samples including those from vaccine recipients, indicated that the overall percent agreement between DFIA and the commercial ELISA kit was 90.98%. Thus, the proposed DFIA provides a more reliable and accurate rapid test for detecting SARS-CoV-2 infections and vaccinations in the community. Therefore, the DFIA based strategy for detecting biomarkers, which uses a ratiometric algorithm based on affinity and inhibition reactions, may be applied to improve the performance of immunochromatographic assays.

Ultrabright nanoparticle-labeled lateral flow immunoassay for detection of anti-SARS-CoV-2 neutralizing antibodies in human serum

“…The anti-SARS-CoV-2 NAb level in the body is becoming an indispensable reference to evaluate the acquired protective immunity for SARS-CoV-2 [ 28 ]. Among all the NAb detection methods, LFIA has unique advantages, such as low cost, easy operation, and no need for complex pretreatment and sophisticated instruments [ 9 ]. The marker for signal output is the key component that concerns the performance of LFIA.…”

“…The high-standard conditions cost much and require professional operator, which is unpractical for massive detection of NAbs. Therefore, several convenient methods, such as lateral flow immunoassay (LFIA) [ 9 ], enzyme-linked immunosorbent assay [ 10 ], surface plasmon resonance assay [ 11 ], and some emerging digital microfluidic systems [ [12] , [13] , [14] ], have been developed to detect SARS-CoV-2 NAbs recently. Among all of them, the LFIA method enjoys portability, rapidity, simplicity, and low cost, and it has been widely used in the point-of-care immunosensors.…”

Evaluation of a rapid semiquantitative lateral flow assay for the prediction of serum neutralizing activity against SARS-CoV-2 variants

“…Using a pseudovirus microneutralization of 1:40 as reference, they found that an inhibition in the RBD-ACE2 test higher than 50% yielded a very good prediction [18] . More recently, Duan et al developed a dual-detection fluorescent immunochromatographic assay (DFIA), with a built-in self-calibration process, that enables rapid quantitative detection of NAbs, and observed a good agreement with a commercial ELISA kit [19] .…”

Recent Advances of Representative Optical Biosensors for Rapid and Sensitive Diagnostics of SARS-CoV-2

“…Current clinical diagnostic methods for COVID-19 mainly include molecular (ribonucleic acid (RNA)), antigen, antibody, or imaging tests [ 2 , 11 ]. The reverse transcription polymerase chain reaction (RT-PCR) for the detection of the RNA of SARS-CoV-2 is regarded as the gold standard in epidemic prevention and control because of its high sensitivity and specificity.…”