The SARS-CoV-2 spreading rapidly has aroused catastrophic public healthcare issues and economy crisis worldwide. It plays predominant role to rapidly and accurately diagnose the virus for effective prevention and treatment. As an abundant transmembrane protein, spike protein (SP) is one of the most valuable antigenic biomarkers for diagnosis of COVID-19. Herein a phage expression of WNLDLSQWLPPM peptide specific to SARS-CoV-2 SP was screened. Molecular docking revealed that the isolated peptide binds to major antigenic epitope locating at S2 subunit with hydrogen bonding. Taking the specific peptide as antigen sensing probe and tyramine signal amplification (TSA), an ultrasensitive "peptide-antigen-antibody" ELISA (p-ELISA) was explored, by which the limit of detection (LOD) was 14 fM and 2.8 fM SARS-CoV-2 SP antigen for first TSA and secondary TSA, respectively. Compared with the LOD by the p-ELISA by direct mode, the sensitivity with 2nd TSA enhanced 100 times. Further, the proposed p-ELISA method can detect SARS-CoV-2 pseudoviruses down to 10 and 3 TCID50/mL spiked in healthy nasal swab sample with 1st TSA and 2nd TSA, separately. Thus, the proposed p-ELISA method with TSA is expected to be a promising ultrasensitive tool for rapidly detecting SARS-CoV-2 antigen to help control the infectious disease.
Keywords: ELISA; Peptide; SARS-CoV-2 S antigen; SARS-CoV-2 pseudoviruses; Tyramine signal amplification.
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