Comparison of automated SARS-CoV-2 antigen test for COVID-19 infection with quantitative RT-PCR using 313 nasopharyngeal swabs, including from seven serially followed patients

Abstract

In routine clinical practice, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is determined by reverse-transcription PCR (RT-PCR). In the current pandemic, a more rapid and high-throughput method is in growing demand. Here, we validated the performance of a new antigen test (LUMIPULSE) based on chemiluminescence enzyme immunoassay. A total of 313 nasopharyngeal swabs (82 serial samples from 7 infected patients and 231 individual samples from 4 infected patients and 215 uninfected individuals) were analyzed for SARS-CoV-2 with quantitative RT-PCR (RT-qPCR) and then subjected to LUMIPULSE. We determined the cutoff value for antigen detection using receiver operating characteristic curve analysis and compared the performance of the antigen test with that of RT-qPCR. We also compared the viral loads and antigen levels in serial samples from seven infected patients. Using RT-qPCR as the reference, the antigen test exhibited 55.2% sensitivity and 99.6% specificity, with a 91.4% overall agreement rate (286/313). In specimens with > 100 viral copies and between 10 and 100 copies, the antigen test showed 100% and 85% concordance with RT-qPCR, respectively. This concordance declined with lower viral loads. In the serially followed patients, the antigen levels showed a steady decline, along with viral clearance. This gradual decline was in contrast with the abrupt positive-to-negative and negative-to-positive status changes observed with RT-qPCR, particularly in the late phase of infection. In summary, the LUMIPULSE antigen test can rapidly identify SARS-CoV-2-infected individuals with moderate to high viral loads and may be helpful for monitoring viral clearance in hospitalized patients.

Keywords: Antigen; COVID-19; Immunoassay; Infection; RT-qPCR; SARS-CoV-2.

Fig. 1

Comparison of the results between the SARS-CoV-2 antigen (Ag) test and RT-qPCR. (A) A total of 313 nasopharyngeal samples were subjected to RT-qPCR (58 positive and 255 negative). The same samples were also subjected to the Ag test. The beeswarm plot shows the SARS-CoV-2 Ag levels in the RT-qPCR-positive (light blue) and -negative samples (red). (B) Receiver operating characteristic (ROC) curve analysis. The Ag test achieved an area under the ROC curve (AUC) value of 0.848 ± 0.044. (C) Comparison of data obtained with the Ag test and RT-qPCR. An overall agreement of 91.4% was achieved between the two tests, with 55.2% sensitivity and 99.6% specificity obtained with the Ag test.

Fig. 2

Correlation between antigen levels and viral loads. A positive correlation (R² = 0.768) was observed between the SARS-CoV-2 antigen (Ag) level (log₁₀ pg/mL) and the viral titer (log₁₀ copies/test).

Fig. 3

Dynamic changes in antigen levels and viral loads in hospitalized patients. A series of 82 nasopharyngeal swabs was collected from seven hospitalized patients. The data shown are from three representative hospitalized patients. Longitudinal antigen levels (pink) and viral loads (light blue) were measured in these samples after the onset of symptoms. The labels on the right of the diagram indicate the SARS-CoV-2 viral loads (log₁₀ copies/test) and the labels on the left show the SARS-CoV-2 Ag levels (log₁₀ pg/mL). The dashed light-blue line indicates the cutoff level below which SARS-CoV-2 was not detectable (ND) by RT-qPCR. The dashed pink line indicates the cutoff level for the antigen test.