RESULTS
Rapid antigen test with phlegm specimens detects SARS-CoV-2 more sensitively than with nasal swab specimens
To know the performance of the rapid antigen test using phlegm specimens in SARS-CoV-2 detection, we compared its sensitivity with that using popular nasal swab specimens. In this test, the “T” band from the phlegm specimen was very strong, but invisible from the nasal swab specimen (Figure 1A, test 1 vs test 2). This result indicated that the patient was infected by SARS-CoV-2, which was detected by the rapid antigen test with the phlegm specimen but not with the nasal swab. The patient continued the antigen test with nasal swab specimens (Figure 1A, test 3), and interestingly found 12 hours later the test with a nasal swab specimen was positive. This result reminded us that the sensitivity in detection of SARS-CoV-2 using rapid antigen tests with phlegm specimens is higher than with nasal swab specimens. Hence, we serially diluted the phlegm specimen 3-fold for more tests (Figure 1B). The 2187-fold dilution showed a weak “T” band (Figure 1B, test 9) compared to the fewer dilutions (Figure 1B, test 2-8), and the 6561-fold dilution had similar levels of SARS-CoV-2 antigen to those of the original nasal swab specimen (Figure 1B, test 10 vs test 1), suggesting the sensitivity in detection of SARS-CoV-2 using an antigen test with phlegm specimens was about 6000-fold higher than that with nasal swab specimens in this case. These results confirmed that rapid SARS-CoV-2 antigen tests with phlegm specimens have higher sensitivity in detection of SARS-CoV-2 than with nasal swab specimens and could be an alternative evaluation technique for COVID-19 diagnosis.
Rapid antigen test with phlegm specimens accurately detects SARS-CoV-2
Among the total of 41 volunteers who volunteered for SARS-CoV-2 tests, 13 patients had a scratchy sensation in the throat when specimens were collected, and their phlegm specimens were positive, but the matched nasal swab specimens were negative; One patient with slight fever had both phlegm and nasal swab specimens tested positive; Seventeen patients with early symptoms of COVID-19 had phlegm specimens tested positive with lack of nasal swab specimens; the rest 10 volunteers without any COVID-19 symptom showed negative phlegm specimens in antigen tests. All 31 patients with detected positive phlegm specimens had typical COVID-19 symptoms in a later stage indicating the accuracy of rapid antigen tests using phlegm specimens was 100%. Considering the collection of phlegm specimens is much milder and easier than that of nasal swab specimens, these results suggested that rapid antigen tests for SARS-CoV-2 with phlegm specimens had high accuracy and can be a good substitute for those with nasal swab specimens.
Rapid antigen test with phlegm specimens earlier detects SARS-CoV-2 infections than with nasal swab specimens
We have demonstrated that the rapid SARS-CoV-2 antigen test with phlegm specimens is more sensitive than that with nasal swab specimens. Hence it is possible that patients can use this test to determine whether they are infected by SARS-CoV-2 earlier than with nasal swab specimens. To test this hypothesis, we chose 3 patients who had positive phlegm in the SARS-CoV-2 antigen test and negative nasal swab to continue antigen tests with nasal swab specimens (Figure 2). Patient 1 had a positive nasal swab specimen 13 hours after SARS-CoV-2 was first detected in the phlegm specimen (Figure 2A, test 4 vs test 2), suggesting the SARS-CoV-2 antigen test with the phlegm specimen detected viral antigen earlier than with the nasal swab specimen for 13 hours. Similarly, Patient 2 and Patient 3 were determined as SARS-CoV-2 infected cases by phlegm specimens earlier than by nasal swab specimens for 24 hours (Figure 2B test 5 vs test 1) and 42 hours (Figure 2C test 5 vs test 1) respectively. Taken together, rapid antigen tests with phlegm specimens detect SARS-CoV-2 12-42 hours earlier than those with nasal swab specimens. Considering the rule of early detection, the better specimen for rapid SARS-CoV-2 antigen tests is from phlegm, but not from nasal swabs.
Rapid antigen test with phlegm specimens earlier notifies patients of SARS-CoV-2 infections than PCR test
Since PCR test is a gold standard in SARS-CoV-2 detection and phlegm specimens in rapid antigen test enhance sensitivity in detection, next we would like to know which of the two methods is better for detection of SARS-CoV-2. We picked a volunteer who was exposed to a positive case in the early stages of COVID-19 for further study. On Day 0, the volunteer had a negative phlegm specimen by the rapid SARS-CoV-2 antigen test (Figure 3, test 1), and the PCR test was notified of a negative result on the next day (Figure 3, 0 h). On Day 1, the volunteer collected phlegm specimens for antigen tests and determined that the sample was positive (Figure 3, 0 h). The SARS-CoV-2 antigen levels increased in a later stage shown by the stronger “T” bands with time increment (Figure 3, test 2, 4, 5, 7). On Day 2, 35 hours later, his antigen test with a nasal swab showed a positive result (Figure 3, test 12), as confirmed our conclusion that phlegm specimens in rapid antigen tests determine SARS-CoV-2 infection earlier than nasal swab specimens for about 12-42 hours. More interestingly, the specimen was collected from the patient for a PCR test on Day 1 when the phlegm specimen was first positive (Figure 3, 0 h), and was notified of a positive result 11 hours (Figure 3, 11 h). Hence, technically rapid antigen tests with phlegm specimens earlier notify patients of a positive result than PCR tests, though the two tests have a similar sensitivity in detection of SARS-CoV-2.
Rapid antigen test with phlegm specimens facilitates monitoring health of COVID-19 patients
Considering rapid antigen tests with phlegm specimens can identify the early SARS-CoV2 infection, it is possible to monitor the health conditions of COVID-19 patients using this test. On Day 1, one volunteer had a positive phlegm specimen in the rapid antigen test (Figure 4, test 1). Due to the weak “T” band in the antigen test with the phlegm specimen and negative nucleic acid sample two days ago, we deduced Day 1 was the earliest time in detection of SARS-CoV-2 in the patient. Simultaneously, the patient had a PCR test. Since the report of PCR tests took time, but the patient was notified of a positive nucleic acid RNA of SARS-CoV-2 on Day 2, 10 hours after the first phlegm specimen was positive (Figure 4, 10 h vs 0 h), as is consistent with the previous result (Figure 3, 11 h vs 0 h). On Day 2, the phlegm specimens remained positive, but the nasal swab specimen tested positive 21 hours after the first phlegm specimen was positive (Figure 4, test 9vs test 1). The patient was then thirsty and had a slight cough and an increased body temperature (37.9 °C) 29 hours after the first phlegm specimen was positive (Figure 4, 21 h and 29 h). Hence, rapid antigen test with phlegm specimens facilitates monitoring health of COVID-19 patients.
Rapid antigen test with phlegm specimens better determines patient recovery from SARS-CoV-2 infection
The sensitivity in detection of SARS-CoV-2 using phlegm specimens in rapid antigen tests is higher than using nasal swab specimens, and comparable to that using oropharyngeal swab specimens in PCR tests. Hence, it is possible that a rapid antigen test with a phlegm specimen could be used to determine whether a patient is recovered from SARS-CoV-2 infection. To this end, we investigated the virus antigen levels using rapid antigen tests with phlegm specimens in 3 volunteers, and the tests with nasal swab specimens were used as a contrast (Figure 5). These volunteers were all infected by SARS-CoV-2 and had common symptoms of COVID-19 by omicron, such as a sore throat, runny nose, and headache. Till Day 9 – Day 11, SARS-CoV-2 was still detectable, but the titer was very low, as shown by the weak “T” band in the rapid antigen test with phlegm specimens (Figure 5A test 1 & 3, 5B test 1, 5C test 3), but not with nasal swab specimens (Figure 5A test 2 & 4, 5B test 2, 5C test 4). Depending on the variation of immunity background, each patient may take a different time to recover. As expected, we found while the phlegm specimen was negative in the rapid antigen test, the nuclei acid sample was simultaneously negative in the PCR test on the same day for both patients (Figure 5B, test 3 and green▽; 5C test 5 and green▽). Taken together, rapid SARS-CoV-2 antigen tests with phlegm specimens better direct patients’ recovery from COVID-19 infection.