The development of a highly sensitive and quantitative SARS-CoV-2 rapid antigen test applying newly developed monoclonal antibodies to an automated chemiluminescent flow-through membrane immunoassay device

Abstract

Abstract Background The rapid and accurate diagnosis of individuals with SARS-CoV-2 infection is an effective way to prevent and control the spread of COVID-19. Although the detection of SARS-CoV‐2 viral RNA by RT‐qPCR is the gold standard for COVID‐19 testing, the use of antigen-detecting rapid diagnostic tests (Ag-RDTs) is emerging as a complementary surveillance tool as Omicron case numbers skyrocket worldwide. However, the results from Ag-RDTs are less accurate for individuals with low viral loads. Methods To develop a more sensitive and accurate Ag-RDT, we screened a total of 90 candidate monoclonal antibodies (mAbs) obtained from guinea pigs immunized with SARS-CoV-2 nucleocapsid protein (CoV-2-NP), and a highly specific epitope-characterized mAb set suitable for detecting the antigen was selected. By applying the mAb set to an automated chemiluminescence flow-through membrane immunoassay device, we developed a highly sensitive and quantitative Ag-RDT, CoV-2-POCube. Results CoV-2-POCube exclusively recognizes a variety of CoV-2-NP variants but not the nucleocapsid proteins of SARS-CoV and other human coronaviruses. CoV-2-POCube achieved a limit of detection sensitivity of 0.20 ~ 0.66 pg/mL of a variety of CoV-2-NP variants, showing over 100 times greater sensitivity than commercially airable SARS-CoV-2 Ag-RDTs. Conclusion CoV-2-POCube is a promising alternative to currently available diagnostic devices for faster clinical decision-making in individuals with suspected COVID-19 in limited-resource settings.