April 21, 2020 – ARUP Laboratories
The CDC recommends use of nasopharyngeal swabs to collect specimens for COVID-19 molecular diagnostic tests. Optimal specimen collection is vital for accurate test results.
How accurate are the laboratory tests used for COVID-19?
Clinicians and patients always want to know they can trust the accuracy of lab test results. This has never been more true than it is now, given the attention on COVID-19 testing and its role in helping to halt the spread of COVID-19. This article explains what we know so far about both nucleic acid tests and antibody detection tests for the SARS-CoV-2 virus, and what factors can affect the reliability of an individual test result. As with all lab tests, a number of factors determine the accuracy of a COVID-19 test result. These include not only the instrument and chemical reagents used to perform the test, but also the timing and quality of specimen collection and the biology of the individual patient.
Laboratory tests are characterized by their ability to detect a positive case (sensitivity) and their ability to determine a negative case (specificity). So a sensitive test is less likely to provide a false-negative result and a specific test is less likely to provide a false-positive result. Test manufacturers and laboratories often report the “analytic” sensitivity and specificity of a test, which are based on the analysis of a set of known positive and negative samples. These numbers, however, represent the accuracy of a test under ideal conditions in which specimens have been collected from patients with either high viral loads or a complete absence of exposure. Sensitivity and specificity under real-world conditions, in which patients are more variable and specimen collection may not be ideal, can often be lower than reported numbers.
Nucleic Acid Amplification Testing (NAAT)
There are two main types of tests for COVID-19. The first detects viral RNA using molecular methods such as polymerase chain reaction (PCR). You can read more about this test in ARUP Consult’s COVID-19 topic. These tests are highly specific because they are based on the unique genetic sequence of SARS-CoV-2. If a test comes back positive, you can be confident that there was SARS-CoV-2 viral RNA in the specimen collected from the patient. However, the sensitivity of these tests varies based on both the timing and the way the sample is collected.
The CDC recommends use of nasopharyngeal (NP) swabs for molecular testing because in most patients, the nasopharynx, or the space above the soft palate at the back of the nose, appears to have the highest concentration of virus. NP swab samples are technically challenging to obtain, and a suboptimal collection may reduce test sensitivity and increase the likelihood of obtaining a false-negative result in a patient with the virus.
Testing a swab from the oropharynx or nose is also likely to reduce sensitivity.1,2 Other sample types such as saliva or blood likely result in even lower sensitivity.3 For patients with frank pneumonia, on the other hand, specimens such as bronchoalveolar lavage collected from the lower respiratory tract may have sensitivity equal to or better than an NP swab, although collection of these types of samples increases the biosafety risk to healthcare workers.4
The timing of sample collection is also important because the amount of virus present in the nasopharynx varies over the course of infection. Ideally, samples should be collected near the time of symptom onset to achieve the highest test sensitivity.1 Patients who are infected but not yet symptomatic may have false-negative test results, as may those whose symptoms are waning.
Antibody Testing
In contrast to nucleic acid testing, which directly detects the virus, antibody or serology testing is used to detect an immune response in the patient. You can read more about the role of antibody testing in COVID-19 in this article. Antibody tests include both traditional enzyme immunoassays and rapid lateral flow immunoassays.5 There are not yet any published data on whether samples drawn from a vein result in better sensitivity or specificity than fingerstick specimens.
Most patients have detectable IgG antibodies by day 14 following symptom onset, and the likelihood of detection increases over time. In studies, antibody tests that detected both IgG and IgM were positive in 90% of symptomatic individuals by days 11-24.6,7
SARS-CoV-2 hasn’t been around long enough for investigators to know whether detectable antibodies may decline or even disappear over time. We do know that immunity to other coronaviruses responsible for colds can wane after ONE year,8 whereas immunity to the more closely related SARS-CoV-1 lasts closer to three years.9 Finally, there is early but inconclusive evidence that children and some individuals with mild or asymptomatic SARS-CoV-2 infections may be less likely to develop detectable antibodies.10
Validation studies of a number of currently available antibody tests, using serum from uninfected individuals, suggest that at least some COVID-19 antibody tests have high specificity, i.e., the probability of a false-positive test is low. However, this specificity could vary by the type of assay. Different assays use antigens from different parts of SARS-CoV-2, and some combine IgM and IgG, and so different levels of cross-reactivity with other coronavirus antibodies are possible.
Conclusion
Because they detect molecules that are specific to SARS-CoV-2, the specificity of nucleic acid tests for COVID-19 is very high, meaning that a positive result can generally be trusted. Specificity of available antibody tests may vary by assay; it is important to check the validation data provided by the manufacturer and/or performing laboratory. Sensitivity of both nucleic acid tests and antibody tests is affected by number of variables. The likelihood of a false-negative result depends on both the timing of sample collection and the type of specimen collected (in the case of the molecular test).
Kirsten Meek, PhD, Medical Writer and Editor
References
- Zou L, Ruan F, Huang M, et al. SARS-CoV-2 Viral Load in Upper Respiratory Specimens of Infected Patients. N Engl J Med. 2020;382(12):1177–1179. doi:10.1056/NEJMc2001737. PMID: 32074444
- Carver C, Jones N. Comparative accuracy of oropharyngeal and nasopharyngeal swabs for diagnosis of COVID-19. Oxford COVID-19 evidence service team centre for evidence based medicine. 2020.
- Wang W, Xu Y, Gao R, et al. Detection of SARS-CoV-2 in Different Types of Clinical Specimens. JAMA. Published online March 11, 2020. doi:10.1001/jama.2020.3786
- Loeffelholz MJ, Tang YW. Laboratory diagnosis of emerging human coronavirus infections – the state of the art. Emerg Microbes Infect. 2020;9(1):747–756. doi:10.1080/22221751.2020.1745095
- Sheridan C. Fast, portable tests come online to curb coronavirus pandemic. Nat Biotechnol2020. [Epub ahead of print] doi:10.1038/d41587-020-00010-2 pmid:32203294
- Li Z, Yi Y, Luo X, et al. Development and clinical application of a rapid IgM-IgG combined antibody test for SARS-CoV-2 infection diagnosis. J Med Virol2020. [Epub ahead of print]. . doi:10.1002/jmv.25727 pmid:32104917
- Zhao J, Yuan Q, Wang H, et al. Antibody responses to SARS-CoV-2 in patients of novel coronavirus disease 2019. Clin Infect Dis2020:ciaa344; [Epub ahead of print]. doi:10.1093/cid/ciaa344 pmid:32221519
- Callow KA, Parry HF, Sergeant M, Tyrrell DA. The time course of the immune response to experimental coronavirus infection of man. Epidemiol Infect. 1990;105(2):435–446. doi:10.1017/s0950268800048019
- Wu LP, Wang NC, Chang YH, et al. Duration of antibody responses after severe acute respiratory syndrome. Emerg Infect Dis. 2007;13(10):1562–1564. doi:10.3201/eid1310.070576
- Wu F, Wang A, Wang Q, et al. Neutralizing antibody responses to SARS-CoV-2 in a COVID-19 recovered patient cohort and their implications. medRxiv. [Epub ahead of print].
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