ABSTRACT Here, we aimed to investigate the diagnostic value of a serological assay using the nucleocapsid protein developed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection and evaluated its performance using three commercial enzyme-linked immunosorbent assays (ELISAs), namely, Standard E 2019 novel coronavirus disease (COVID-19) total antibody (Ab) ELISA (SD Biosensor), and EDI novel coronavirus COVID-19 IgG and IgM ELISA. A recombinant nucleocapsid protein (rNP) was expressed from plants and Escherichia coli for the detection of serum total Ab. We prospectively collected 141 serum samples from 32 patients with reverse transcription-PCR (RT-PCR)-confirmed COVID-19 and determined the sensitivity and dynamics of their total Ab response. Specificity was evaluated using 158 prepandemic samples. To validate the assays, we evaluated the performance using two different cutoff values. The sensitivity and specificity for each assay were as follows: 92.91% and 94.30% (plant-rNP), 83.69% and 98.73% (SD Biosensor), 75.89% and 98.10% ( E. coli -rNP), 76.47% and 100% (EDI-IgG), and 80.39% and 80% (EDI-IgM). The plant-based rNP showed the highest sensitivity and area under the receiver operating characteristic (ROC) curve (0.980) among all the assays ( P < 0.05). The seroconversion rate for total Ab increased sequentially with disease progression, with a sensitivity of 100% after 10 to 12 days of post-symptom onset (PSO) for both rNP-plant-based and SD Biosensor ELISAs. After 2 weeks of PSO, the seroconversion rates were >80% and 100% for EDI-IgM and EDI-IgG ELISA, respectively. Seroconversion occurred earlier with rNP plant-based ELISA (5 days PSO) compared with E. coli – based (7 days PSO) and SD Biosensor (8 days PSO) ELISA. We determined that rNP produced in plants enables the robust detection of SARS-CoV-2 total Abs. The assay can be used for serosurvey and complementary diagnosis of COVID-19. IMPORTANCE At present, the principal diagnostic methods for COVID-19 comprise the identification of viral nucleic acid by genetic approaches, including PCR-based techniques or next-generation sequencing. However, there is an urgent need for validated serological assays which are crucial for the understanding of immune responses against SARS-CoV-2. In this study, a highly sensitive and specific serological antibody assay was developed for the detection of SARS-CoV-2 with an overall accuracy of 93.56% using a recombinant nucleoprotein expressed from plants.
【저자키워드】 COVID-19, SARS-CoV-2, ELISA, nucleoprotein, total antibody, plant, E. coli, 【초록키워드】 coronavirus disease, coronavirus, immune response, antibody, Genetic, novel coronavirus disease, severe acute respiratory syndrome Coronavirus, RT-PCR, reverse transcription-PCR, ELISA, nucleocapsid protein, Novel coronavirus, Disease progression, 2019 novel coronavirus, nucleic acid, serum, sensitivity, enzyme-linked immunosorbent assay, Recombinant nucleocapsid protein, Seroconversion, Viral, Serological assay, Serosurvey, ROC, plants, Accuracy, Sensitivity and specificity, enzyme-linked immunosorbent assays, immune responses, Next-generation sequencing, Patient, reverse transcription, respiratory, characteristic, serological, disease, ELISAs, Diagnostic method, PSO, Escherichia coli, Standard, complementary, IgG and IgM, receiver operating characteristic, acute respiratory syndrome, E. coli, acute respiratory syndrome coronavirus, serum samples, acute respiratory syndrome coronavirus 2, approaches, diagnostic value, post-symptom onset, viral nucleic acid, Cutoff, seroconversion rates, present, seroconversion rate, SD Biosensor, serum sample, robust, produced, highest, collected, occurred, assays, evaluated, can be used, expressed, diagnosis of COVID-19, 【제목키워드】 recombinant, Total,