Point-of-Care Antigen Test for SARS-CoV-2 in Asymptomatic College Students

We used the BinaxNOW COVID-19 Ag Card to screen 1,540 asymptomatic college students for severe acute respiratory syndrome coronavirus 2 in a low-prevalence setting. Compared with reverse transcription PCR, BinaxNOW showed 20% overall sensitivity; among participants with culturable virus, sensitivity was 60%. BinaxNOW provides point-of-care screening but misses many infections.

We used the BinaxNOW COVID-19 Ag Card to screen 1,540 asymptomatic college students for severe acute respiratory syndrome coronavirus 2 in a low-prevalence setting. Compared with reverse transcription PCR, Bi-naxNOW showed 20% overall sensitivity; among participants with culturable virus, sensitivity was 60%. Bi-naxNOW provides point-of-care screening but misses many infections.
1 swab immediately using BinaxNOW and sent the other swab to CDC for rRT-PCR.
We conducted BinaxNOW assays on-site in accordance with manufacturer instructions (4). Students received BinaxNOW results after 15-30 minutes. Those who tested positive were counseled to isolate for 10 days and interviewed for contact tracing.
Swab samples collected for rRT-PCR were stored using Remel R12587 viral transport media (Thermo Fisher Scientific, https://www.thermofisher.com) with cold packs; samples were transported daily to CDC and refrigerated at 4°C. We isolated nucleic acid from the specimens using the MagNA Pure 96 Instrument (Roche Molecular Systems, Inc., https:// lifescience.roche.com) within 48 hours of collection, then analyzed the nucleic acid using the CDC Influenza SARS-CoV-2 (Flu SC2) Multiplex Assay (5). Results were reported as SARS-CoV-2-positive (cycle threshold [C t ] <40 for the SARS-CoV-2 target), SARS-CoV-2-negative, or invalid (C t value ≥40 for all viral targets and C t >35 for human RNase P reference gene on repeat testing, according to the manufacturer's guidelines).
We cultured residual frozen SARS-CoV-2-positive samples in 100 µL viral transport media. We limited dilution in Vero CCL-81 cells and monitored 96-well plates daily for cytopathic effects (J. Harcourt, unpub. data, https://www.biorxiv.org/content/10.1 101/2020.03.02.972935v2). We extracted nucleic acid from the wells exhibiting cytopathic effects and confirmed the presence of SARS-CoV-2 by rRT-PCR. We considered a specimen to be culture-positive if the first viral passage had a C t value >2 less than the clinical sample.

Conclusions
In total, 1,540 asymptomatic students provided paired samples (Table). Forty (2.6%) samples tested positive by rRT-PCR; of these, 8 (20%) also tested positive by BinaxNOW. We did not observe any false-positive BinaxNOW results (100% specificity). Concordant samples had a lower median C t value than discordant samples (21.9 vs. 34.9). Students received rRT-PCR results within 72 hours. No specimens tested positive for influenza A or B viruses. All 8 persons who tested positive by BinaxNOW and rRT-PCR later reported symptom onset. Among the 32 students who provided samples that tested negative by BinaxNOW and positive by rRT-PCR, 10 (31.3%) later reported symptom onset (median C t 34.9), 16 (50.0%) later reported no symptoms (median C t 35.1), and 6 (18.8%) did not report information on symptoms (median C t 34.9).
We detected culturable virus in 5 (12.5%) samples that tested positive by rRT-PCR, including 3 (60%) that also tested positive by BinaxNOW (Figure). One person provided a sample (C t 28.9) that tested negative by BinaxNOW but was culture-positive; symptoms later developed in this person, who tested positive by a different antigen test (BD Veritor System; Becton, Dickinson and Company, https:// www.bd.com) the next day. Symptoms did not develop in the other person who provided a sample that tested negative by BinaxNOW and positive by culture (C t 37.3).
BinaxNOW provides rapid, point-of-care results; students received BinaxNOW results 3 days earlier than rRT-PCR results. However, BinaxNOW had low sensitivity, especially among persons with higher C t values, which suggest lower viral load. BinaxNOW did not identify 32 persons who tested positive by rRT-PCR.
Our data are consistent with those of Prince-Guerra et al. (6), which found low overall Bi-naxNOW sensitivity (35.8%; 44/123) compared with RT-PCR among asymptomatic persons. Prince-Guerra et al. (6) found that concordant samples had a lower mean C t value than discordant samples (22.5 vs. 33.9); we observed 88.9% (8/9) sensitivity among samples with C t values <32. However, Prince-Guerra et al. (6) collected samples using disparate methods (nasopharyngeal swab for RT-PCR and anterior nasal swab for BinaxNOW), precluding direct comparison of samples. Our results are Point-of-Care Antigen Test for SARS-CoV-2 inconsistent with those of Pilarowski et al. (7), which showed 81.4% sensitivity among 102 persons who were asymptomatic or had symptom onset >1 week previously. We observed high specificity, consistent with results of both investigations (6,7). Unlike the community investigations of Prince-Guerra et al. (6) and Pilarowski et al. (7), in which testing was offered to persons who might have had specific reasons for seeking testing, our investigation was conducted in a closed, defined population, among persons with no known exposures or symptoms, providing more generalizable performance data for similar institutions. CDC provided guidance on expanded screening testing of asymptomatic individuals to reduce spread of SARS-CoV-2 and for interpretation of antigen tests (8,9). Test performance among asymptomatic persons probably varies for different antigen tests. For example, an assessment of the Sofia SARS Antigen Fluorescent Immunoassay (Quidel Corporation, https://www.quidel.com) reported 41.2% sensitivity and 98.4% specificity among 871 asymptomatic college students (10).
Isolation of SARS-CoV-2 in cell culture demonstrates viral replication. However, because many factors affect the culture performance, lack of culturable virus does not necessarily indicate a lack of infectious virus. The presence of culturable virus in samples that test negative for SARS-CoV-2 antigens suggests that BinaxNOW does not identify some persons with infectious virus. However, the speed of BinaxNOW enabled the immediate identification of 8 SARS-CoV-2-positive persons, thereby limiting transmission that might have occurred during the additional 2 days that students waited for rRT-PCR results.
Although rRT-PCR tests remain standard for SARS-CoV-2 detection, point-of-care antigen tests such as BinaxNOW could increase access to serial screening, enabling the rapid identification and isolation of infectious persons. Because presymptomatic and asymptomatic persons can transmit SARS-CoV-2 (11), screening of asymptomatic persons is a key strategy for interrupting SARS-CoV-2 transmission. Although messaging must clearly communicate the low sensitivity of the test, positive results enable immediate public health action.