View source for COVID-19 Testing, Reporting, and Information Management in the Laboratory/Adding COVID-19 and other virus testing to your laboratory/What methodologies will you use?

==3. Adding COVID-19 and other virus testing to your laboratory==
Maybe you've been running an environmental health laboratory and want to expand into clinical health testing. Perhaps you're in charge of an academic research lab but want to expand to the clinical diagnostic side. Or maybe you're running a [[physician office laboratory]] (POL) and are wondering if it's even possible to expand your waived testing efforts to COVID-19. Where the previous chapter discussed the "what" of COVID-19 and viral testing, this chapter aims to help you with the "how" of adding it to your laboratory offerings.

Naturally, many questions come with the "how":

* Does using one method make the most sense, or will your lab turn to multiple methods for virus testing? This may be determined by current equipment, space considerations, and budget.
* What type of lab are you running? A POL is going to have fewer options available than a CLIA moderate- or high-complexity lab.
* How interoperable are you existing laboratory and clinical informatics solutions? Research laboratories face more challenges in integrating their systems with EHRs and other clinical systems.
* What vendors and consultants are out there to help get equipped? Some vendors have very specific solutions, whereas others may have a broader range of offerings.

These questions and more are addressed in this chapter.

===3.1 What methodologies will you use?===
====3.1.1 PCR====
[[File:Test PCR dépistage SARS-CoV-2 prélèvement nasal Strasbourg 21 août 2020.jpg|right|thumb|400px|SARS-CoV-2 PCR screening test by nasal swab in Strasbourg on August 21, 2020.]]In the previous chapter, the most common testing methodologies for [[COVID-19]] and other [[Coronavirus|coronaviruses]] were discussed in detail. The prevailing method (often called the "gold standard") among them all is real-time reverse-transcription [[polymerase chain reaction]] (rRT-PCR) assays for testing. Broadly speaking, PCR is useful in pharmaceutical, biotechnology, and genetic engineering endeavors, as well as clinical diagnostics. As such, labs in those industries that already have PCR infrastructure in place have a theoretical step-up over labs that do not.

PCR technology has advanced to the point where it is more efficient and user-friendly than prior, yet "the high cost of the instruments, servicing contracts, and reagents pose major challenges for the market, especially to the price-sensitive academics."<ref name="MWPoly20">{{cite web |url=https://www.marketwatch.com/press-release/polymerase-chain-reaction-market-sector-analysis-report-regional-outlook-competitive-share-forecast---2023-2020-06-23 |archiveurl=https://web.archive.org/web/20200809215548/https://www.marketwatch.com/press-release/polymerase-chain-reaction-market-sector-analysis-report-regional-outlook-competitive-share-forecast---2023-2020-06-23 |title=Polymerase Chain Reaction Market Sector Analysis Report, Regional Outlook & Competitive Share & Forecast - 2023 |author=Kenneth Research |work=MarketWatch |date=23 June 2020 |archivedate=09 August 2020 |accessdate=08 September 2021}}</ref> Writing about the thirty-fifth anniversary of PCR in 2018, science writer Alan Dove not only highlighted these cost issues but also the size and energy requirements for running the equipment. "As a result, one of the defining techniques of modern molecular biology has remained stubbornly inaccessible to educators and unusable in many remote locations."<ref name="DovePCR18">{{cite journal |title=PCR: Thirty-five years and counting |journal=Science |author=Dove, A. |volume=360 |issue=6389 |pages=670–672 |year=2018 |doi=10.1126/science.360.6389.673-c}}</ref> Various efforts have been made over the years to bring costs down by modifying how heating and temperature control are performed<ref name="WongARapid15">{{cite journal |title=A Rapid and Low-Cost PCR Thermal Cycler for Low Resource Settings |journal=PLoS One |author=Wong, G.; Wong, I. Chan, K. et al. |volume=10 |issue=7 |at=e0131701 |year=2015 |doi=10.1371/journal.pone.0131701}}</ref><ref name="KuznetsovDIYbio15">{{cite journal |title=DIYbio Things: Open Source Biology Tools as Platforms for Hybrid Knowledge Production and Scientific Participation |journal=Proceedings of the 33rd Annual ACM Conference on Human Factors in Computing Systems |author=Kuznetsov, S.; Doonan, C.; Wilson, N. et al. |pages=4065–68 |year=2015 |doi=10.1145/2702123.2702235}}</ref><ref name="NortonPhila16">{{cite web |url=https://www.bizjournals.com/philadelphia/news/2016/05/11/government-contract-biomeme-hiring-med-tech.html |title=Phila. med tech startup working on multimillion dollar government contract |author=Norton, D. |work=Philadelphia Business Journal |date=11 May 2016 |accessdate=06 August 2020}}</ref><ref name="AnLow20">{{cite journal |title=Low-Cost Battery-Powered and User-Friendly Real-Time Quantitative PCR System for the Detection of Multigene |journal=Micromachines |author=An, J.; Jiang, Y.; Shi, B. et al. |volume=11 |at=435 |year=2020 |doi=10.3390/mi11040435}}</ref>, but many of those system aren't typically optimal during a pandemic when turnaround time is critical.

Amidst the pandemic, additional challenges also exist to those wanting to conduct PCR testing for COVID-19 and other viruses. As was discussed at the end of the previous chapter, supplies of reagents and consumables are not particularly robust mid-pandemic, with various shortages being reported off and on since the start of the pandemic.<ref name="HerperShortage20">{{cite web |url=https://www.statnews.com/2020/03/10/shortage-crucial-chemicals-us-coronavirus-testing/ |title=Shortage of crucial chemicals creates new obstacle to U.S. coronavirus testing |work=STAT |author=Herper, M.; Branswell, H. |date=10 March 2020 |accessdate=10 April 2020}}</ref><ref name="HaleQiagen20">{{cite web |url=https://www.fiercebiotech.com/medtech/qiagen-aims-to-more-than-quadruple-its-covid-19-reagent-production-6-weeks |title=Qiagen aims to more than quadruple its COVID-19 reagent production in 6 weeks |work=Fierce Biotech |author=Hale, C. |date=18 March 2020 |accessdate=10 April 2020}}</ref><ref name="MehtaMystery20">{{cite web |url=https://www.chemistryworld.com/news/mystery-surrounds-uk-claim-of-covid-19-test-reagent-shortage/4011457.article |title=Mystery surrounds UK claim of Covid-19 test reagent ‘shortage’ |work=Chemistry World |author=Mehta, A. |date=03 April 2020 |accessdate=07 September 2021}}</ref><ref name="RocheIrish20">{{cite web |url=https://www.irishtimes.com/news/science/irish-scientists-develop-reagent-in-effort-to-ease-covid-19-testing-delays-1.4223897 |title=Irish scientists develop reagent in effort to ease Covid-19 testing delays |work=The Irish Times |author=Roche, B. |date=08 April 2020 |accessdate=10 April 2020}}</ref><ref name="PadmaEfforts20">{{cite web |url=https://www.chemistryworld.com/news/efforts-to-combat-covid-19-in-india-hit-by-imported-reagent-shortages/4011718.article#/ |title=Efforts to combat Covid-19 in India hit by imported reagent shortages |author=Padma, T.V. |work=Chemistry World |date=13 May 2020 |accessdate=19 May 2020}}</ref><ref name="DavidSurvey20">{{cite web |url=https://abcnews.go.com/Health/survey-shows-resources-covid-19-diagnostic-testing-limited/story?id=71341885 |title=Survey shows resources for COVID-19 diagnostic testing still limited months later |author=David, E.; Farber, S.E. |work=ABC News |date=20 June 2020 |accessdate=08 July 2020}}</ref><ref name="JohnsonNCLabs20">{{cite web |url=https://patch.com/north-carolina/charlotte/nc-labs-facing-shortages-covid-19-testing-chemicals |title=NC Labs Facing Shortages In COVID-19 Testing Chemicals |author=Johnson, K. |work=Patch |date=02 July 2020 |accessdate=08 July 2020}}</ref><ref name="MervoshItsLike20">{{cite web |url=https://www.nytimes.com/2020/08/04/us/virus-testing-delays.html |title=‘It’s Like Having No Testing’: Coronavirus Test Results Are Still Delayed |author=Mervosh, S.; Fernandez, M. |work=The New York Times |date=04 August 2020 |accessdate=05 August 2020}}</ref><ref name="CourageCovid20">{{cite web |url=https://www.vox.com/2020/7/31/21336212/covid-19-test-results-delays |title=Should we be testing fewer people to stop the spread of Covid-19? |author=Courage, K.H. |work=Vox |date=31 July 2020 |accessdate=05 August 2020}}</ref><ref name="ASMSupply20">{{cite web |url=https://asm.org/Articles/2020/September/Clinical-Microbiology-Supply-Shortage-Collecti-1 |title=Supply Shortages Impacting COVID-19 and Non-COVID Testing |author=American Society for Microbiology |publisher=American Society for Microbiology |date=09 November 2020 |accessdate=18 November 2020}}</ref><ref name="AbbottCovid20">{{cite web |url=https://www.wsj.com/articles/covid-19-testing-saps-supplies-needed-for-other-medical-tests-11604926800 |title=Covid-19 Testing Saps Supplies Needed for Other Medical Tests |author=Abbott, B.; Krouse, S. |work=The Wall Street Journal |date=09 November 2020 |accessdate=18 November 2020}}</ref><ref name="WilliamsSupply21">{{cite web |url=https://www.the-scientist.com/news-opinion/supply-shortages-hit-life-science-labs-hard-68695 |title=Supply Shortages Hit Life Science Labs Hard |author=Williams, S. |work=The Scientist |date=21 April 2021 |accessdate=07 September 2021}}</ref> Some of these shortages have gradually worked themselves out over time, but they highlight the need for other varying methods that don't necessarily depend on the same reagents and consumables that are in short supply.

For those labs wishing to adopt PCR testing of viruses—particularly COVID-19—into their workflow while providing reasonable turnaround times, all is not lost. However, careful planning is required. For example, you'll want to keep in mind that some PCR machines require vendor-specific reagents. If you're going to acquire a particular instrument, you'll want to do due diligence by verifying not only the supported reagents but also those reagents' overall availability (real and projected). You'll also want to consider factors such as anticipated workload (tests per day), what your workflow will look like, and how to balance overall investment with the need for reasonable turnaround times. 

An increasing body of research is being produced suggesting ways to improve turnaround times with PCR testing for COVID-19, with many research efforts focusing on cutting out RNA extraction steps entirely. Alcoba-Florez ''et al.'' [[Journal:Fast SARS-CoV-2 detection by RT-qPCR in preheated nasopharyngeal swab samples|propose]] direct heating of the sample-containing nasopharyngeal swab at 70 °C for 10 minutes in place of RNA extraction.<ref name="Alcoba-FlorezFast20">{{cite journal |title=Fast SARS-CoV-2 detection by RT-qPCR in preheated nasopharyngeal swab samples |journal=International Journal of Infectious Diseases |author=Alcoba-Florez, J.; González-Montelongo, R.; Íñigo-Campos, A.; García-Martínezde Artola, D.; Gil-Campesino, H.;<br />The Microbiology Technical Support Team; Ciuffreda, L.; Valenzuela-Fernández, A.; Flores, C. |volume=97 |pages=66–68 |year=2020 |doi=10.1016/j.ijid.2020.05.099}}</ref> Adams ''et al.'' have proposed an "adaptive PCR" method using a non-standard reagent mix that skips RNA extraction and can act "as a contingency for resource‐limited settings around the globe."<ref name="ShapiroStream20">{{cite web |url=https://news.vanderbilt.edu/2020/07/29/streamlined-diagnostic-approach-to-covid-19-can-avoid-potential-testing-logjam/ |title=Streamlined diagnostic approach to COVID-19 can avoid potential testing logjam |author=Shapiro, M. |work=Research News @ Vanderbilt |date=29 July 2020 |accessdate=06 August 2020}}</ref><ref name="AdamsCOVID20">{{cite journal |title=COVID‐19 diagnostics for resource‐limited settings: Evaluation of “unextracted” qRT‐PCR |journal=Journal of Medical Virology |author=Adams, N.M.; Leelawong, M.; Benton, A. et al. |year=2020 |doi=10.1002/jmv.26328}}</ref> Wee ''et al.'' skip RNA extraction and nucleic acid purification by using a single-tube homogeneous reaction method run on a lightweight, portable thermocycler.<ref name="MeharImproving20">{{cite web |url=https://www.techexplorist.com/improving-speed-gold-standard-covid-19-diagnostic-test/34069/ |title=Improving the speed of gold-standard COVID-19 diagnostic test |author=Mehar, P. |work=Tech Explorist |date=27 July 2020 |accessdate=06 August 2020}}</ref><ref name="WeeRapid20">{{cite journal |title=Rapid Direct Nucleic Acid Amplification Test without RNA Extraction for SARS-CoV-2 Using a Portable PCR Thermocycler |journal=Genes |author=Wee, S.K.; Sivalingam, S.P.; Yap, E.P.H. |volume=11 |issue=6 |at=664 |year=2020 |doi=10.3390/genes11060664}}</ref> Other innovations include tweaking reagents and enzymes to work with one step, skipping the reverse transcription step,<ref name="CSIRFaster20">{{cite web |url=https://www.engineeringnews.co.za/article/faster-local-covid-19-test-kits-could-be-ready-by-year-end-2020-07-30/ |title=Faster, local COVID-19 test kits could be ready by year-end |author=Council for Scientific and Industrial Research |work=Engineering News |publisher=Creamer Media |date=30 July 2020 |accessdate=07 August 2020}}</ref> and using saliva-based molecular testing that skips RNA extraction.<ref name="RanoaSaliva20">{{cite journal |title=Saliva-Based Molecular Testing for SARS-CoV-2 that Bypasses RNA Extraction |journal=bioRxiv |author=Ranoa, D.R.E.; Holland, R.L.; Alnaji, F.G. et al. |year=2020 |doi=10.1101/2020.06.18.159434}}</ref><ref name="ThomasFast20">{{cite web |url=https://www.news-medical.net/news/20200806/Fast-cheap-and-easy-COVID-19-test-from-Yale.aspx |title=Fast, cheap and easy COVID-19 test from Yale |author=Thomas, L. |work=News Medical - Life Sciences |date=06 August 2020 |accessdate=16 August 2020}}</ref>

'''Saliva as a specimen'''

The saliva molecular tests in particular are intriguing. Talk of the potential utility of using saliva as a specimen for COVID-19 was occurring as early as April 2020<ref name="XuSaliva20">{{cite journal |title=Saliva: Potential diagnostic value and transmission of 2019-nCoV |journal=International Journal of Oral Science |author=Xu, R.; Cui, B.; Duan, X. et al. |volume=12 |at=11 |year=2020 |doi=10.1038/s41368-020-0080-z}}</ref><ref name="GreenwoodSaliva20">{{cite web |url=https://news.yale.edu/2020/04/24/saliva-samples-preferable-deep-nasal-swabs-testing-covid-19 |title=Saliva samples preferable to deep nasal swabs for testing COVID-19 |author=Greenwood, M. |work=YaleNews |date=24 April 2020 |accessdate=01 May 2020}}</ref>, and the first saliva-based COVID-19 test, produced by Spectrum Solutions in cooperation with RUCDR Infinite Biologics Laboratory<ref name="SSFirst20">{{cite web |url=https://spectrumsolution.com/fda-authorized-covid-19-updates/ |title=First saliva collection device FDA EUA authorized for COVID-19 testing |publisher=Spectrum Solutions |date=2020 |accessdate=16 August 2020}}</ref> and Vault Health<ref name="VHVault20">{{cite web |url=https://www.prnewswire.com/news-releases/vault-health-launches-first-of-its-kind-saliva-based-fda-eua-approved-test-for-covid-19-301039633.html |title=Vault Health Launches First-of-its-Kind Saliva-based FDA EUA Approved Test for COVID-19 |author=Vault Health |work=PR Newswire |date=14 April 2020 |accessdate=01 May 2020}}</ref>, was given an FDA EUA in April 2020. On August 15, 2020, Yale School of Public Health was given an EUA for it SalivaDirect molecular test. Although still PCR-based (and a CLIA high-complexity test), SalivaDirect is being touted as a means to improve specimen collection safety, consume fewer reagents, prove compatible with high-throughput workflow, and cut overall turnaround time. Not only is saliva easier to collect and safer for healthcare staff, the test is essentially "open sourced," not requiring proprietary equipment from Yale, making the test more flexible by being validated to reliably function with a wider array of reagents and instruments.<ref name="GallagherFDA20">{{cite web |url=https://www.contagionlive.com/view/fda-grants-emergency-covid19-authorization-yale-open-source-method-saliva-testing |title=FDA Grants Emergency COVID-19 Authorization to Yale's Open Source Method of Saliva Testing |author=Gallagher, G.M. |work=ContagionLive |date=15 August 2020 |accessdate=16 August 2020}}</ref><ref name="ZillgittFDA20">{{cite web |url=https://www.usatoday.com/story/sports/nba/2020/08/15/fda-approves-covid-19-saliva-test-developed-yale-nba-nbpa-aid/5590452002/ |title=FDA approves COVID-19 saliva test developed at Yale in partnership with the NBA, NBPA |author=Zillgitt, J. |work=USA Today |date=15 August 2020 |accessdate=16 August 2020}}</ref> When compared to using a nasopharyngeal swab specimen using the ThermoFisher Scientific TaqPath COVID-19 combo kit, results were comparable 94.1% of the time.<ref name="ThomasFast20" /> While sensitivity and specificity may be slightly less comparable to other PCR options<ref name="WeisslederCOVID20">{{cite web |url=https://csb.mgh.harvard.edu/covid |title=COVID-19 Diagnostics in Context |author=Weissleder, R.; Lee, H.; Ko, J. et al. |publisher=Harvard Center for Systems Biology |date=15 August 2020 |accessdate=16 August 2020}}</ref>, the overall advantages during reagent shortages and a definitive need for broader testing likely outweigh the slightly lesser sensitivity and specificity. In November 2020, public health agencies in Arizona and Minnesota reportedly began running trials of free saliva-based molecular testing.<ref name="ParsonsPlaces20">{{cite web |url=https://www.azfamily.com/news/continuing_coverage/coronavirus_coverage/places-with-saliva-based-covid-testing-expecting-influx-of-people/article_76ac95c4-26b5-11eb-b34e-3728b1308927.html |title=Places with saliva-based COVID testing expecting influx of people |author=Parsons, J. |work=AZFamily |date=14 November 2020 |accessdate=19 November 2020}}</ref><ref name="MDoHState20">{{cite web |url=https://www.health.state.mn.us/news/pressrel/2020/covid102220.html |title=State launches pilot of COVID-19 test at home saliva program |author=Minnesota Department of Health |publisher=Minnesota Department of Health |date=22 October 2020}}</ref> 

As the pandemic has progressed into 2021, saliva testing has become even more attractive, in particular for at-home over-the-counter testing.<ref name="PugleNininv21">{{cite web |url=https://www.healthline.com/health-news/noninvasive-saliva-tests-for-covid-19-as-effective-as-nose-throat-swabs |title=Noninvasive Saliva Tests for COVID-19 as Effective as Nose, Throat Swabs |author=Pugle, M. |work=Healthline |date=20 January 2021 |accessdate=08 September 2021}}</ref><ref name="KarkusDiff21">{{cite web |url=https://www.pharmacytimes.com/view/differences-between-saliva-covid-19-tests-nasal-swab-covid-19-tests |title=Differences Between Saliva COVID-19 Tests, Nasal Swab COVID-19 Tests |author=Karkus, T. |work=Pharmacy Times |date=05 April 2021 |accessdate=08 September 2021}}</ref> In August 2021, Spectrum Solutions received an EUA for its Spectrum Solutions SDNA-1000 saliva collection system, specifically designed "to avoid user collection errors" and eliminate "the requirement for any bio-sample temperature-controlled storage or transport,"<ref name="NSMedStaffSpect21">{{cite web |url=https://www.nsmedicaldevices.com/news/spectrum-solutions-covid-19-testing/ |title=Spectrum Solutions’ device gets FDA EUA for unsupervised saliva collection for Covid-19 testing |author=NS Medical Staff Writer |work=NS Medical Devices |date=18 August 2021 |accessdate=08 September 2021}}</ref> arguably upping the game for new saliva-based test kits going forward. Additionally, as variants of COVID-19 continue to crop up, additional saliva-based at-home tests are coming into development. For example, researchers at the Wyss Institute, the Massachusetts Institute of Technology, and Boston-area hospitals have been working on a laboratory-developed test called Minimally Instrumented SHERLOCK (miSHERLOCK) based on CRISPR (clustered regularly interspaced short palindromic repeats) technology. The researchers claim that the test, able to be used with typical off-the-shelf components, "works as well as the gold standard PCR tests and could cost as little as $3 per test."<ref name="HealthDayAtHome21">{{cite web |url=https://www.webmd.com/lung/news/20210807/at-home-saliva-test-can-spot-covid-variants#1 |title=At-Home Saliva Test Can Spot COVID Variants |author=HealthDay News |work=WebMD |date=06 August 2021 |accessdate=08 September 2021}}</ref><ref name="DePuigMinim21">{{cite journal |title=Minimally instrumented SHERLOCK (miSHERLOCK) for CRISPR-based point-of-care diagnosis of SARS-CoV-2 and emerging variants |journal=Science Advances |author=De Puig, H.; Lee, R.A.; Najjar, D. et al. |volume=7 |issue=32 |at=eabh2944 |year=2021 |doi=10.1126/sciadv.abh2944 |pmid=34362739 |pmc=PMC8346217}}</ref>

====3.1.2 Pooled testing====
Another method some labs are taking to speed up turnaround time is using pooled testing. The general concept involves placing two or more test specimens together and testing the pool as one specimen. The most obvious advantage to this is that the process saves on reagents and other supplies, particularly when supply chains are disrupted, and it reduces the amount of time required to analyze large quantities of specimens.<ref name="CDCInterimPool21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/lab/pooling-procedures.html |title=Interim Guidance for Use of Pooling Procedures in SARS-CoV-2 Diagnostic and Screening Testing |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=30 June 2021 |accessdate=19 September 2021}}</ref> This methodology is best used "in situations where disease prevalence is low, since each negative pool test eliminates the need to individually test those specimens and maximizes the number of individuals who can be tested over a given amount of time."<ref name="RohdeCOVID20">{{cite web |url=https://asm.org/Articles/2020/July/COVID-19-Pool-Testing-Is-It-Time-to-Jump-In |title=COVID-19 Pool Testing: Is It Time to Jump In? |author=Rohde, R. |publisher=American Society for Microbiology |date=20 July 2020 |accessdate=06 August 2020}}</ref> However, it's best left to situations where expectations are that less than 10 percent of the population being tested is affected by what's being tested for.<ref name="RohdeCOVID20" /><ref name="MashaPooled20">{{cite web |url=https://asiatimes.com/2020/08/pooled-virus-tests-help-stretched-health-services/ |title=Pooled virus tests help stretched health services |author=Masha, M.; Chau, S. |work=Asia Times |date=04 August 2020 |accessdate=06 August 2020}}</ref><ref name="CitronerHowPool20">{{cite web |url=https://www.healthline.com/health-news/how-pooled-testing-can-help-us-fight-spread-of-covid-19 |title=How Pooled Testing Can Help Us Fight Spread of COVID-19 |author=Citroner, G. |work=Healthline |date=03 August 2020 |accessdate=06 August 2020}}</ref> 

The downside of pooled testing comes with the issues of dilution, contamination, and populations with 10 or more percent infected. A target-positive specimen that commingles with other target-free specimens is itself diluted and in some cases may cause issues with the limit of detection for the assay. Additionally, if the pool tests positive, target-free specimens may become contaminated by a target-positive specimen. This may cause issues with any individual specimen assays that get ran. And the workflows involving pooling must be precise, as a technician working with multiple specimens at the same time increases the chance of lab errors.<ref name="RohdeCOVID20" /><ref name="MashaPooled20" /><ref name="CitronerHowPool20" /> Finally, at least in the U.S., a Food and Drug Administration (FDA) emergency use authorization (EUA) for a validated pooled testing method is required.<ref name="RohdeCOVID20" /> (Validation of pooled methods may differ in other countries.<ref name="MashaPooled20" />) The U.S. Centers for Disease Control and Prevention (CDC) has published [https://www.cdc.gov/coronavirus/2019-ncov/lab/pooling-procedures.html interim guidance] on pooled testing strategies for [[SARS-CoV-2]].

On April 20, 2021, the FDA updated its policies to allow for pooled testing to be added to the use case scenarios for several existing test kits. "This means that tests with EUAs that are amended by this authorization may be used with pooled anterior nasal specimens from individuals without known or suspected COVID-19 when such individuals are tested as part of a testing program that includes testing at regular intervals, at least once per week."<ref name="FDAInVitroEUAs21">{{cite web |url=https://www.fda.gov/medical-devices/coronavirus-disease-2019-covid-19-emergency-use-authorizations-medical-devices/in-vitro-diagnostics-euas-molecular-diagnostic-tests-sars-cov-2 |title=In Vitro Diagnostics EUAs - Molecular Diagnostic Tests for SARS-CoV-2 |publisher=U.S. Food and Drug Administration |date=07 September 2021 |accessdate=07 September 2021}}</ref> However, affected kits can only be used in high-complexity CLIA labs, though "tests authorized for use in specific named or designated high-complexity laboratories can only be used in such laboratories."<ref name="FDAInVitroEUAs21" /> As of September 2021, four PCR test kit EUAs were amended to allow for pooled testing<ref name="FDAInVitroEUAs21" />:

* Biomeme SARS-CoV-2 Real-Time RT-PCR Test
* Clinical Enterprise SARS-SoV-2-RT-PCR Assay
* CRSP SARS-CoV-2 Real-time Reverse Transcriptase (RT)-PCR Diagnostic Assay (Version 3)
* Viracor SARS-CoV-2 assay

====3.1.3 Rapid antigen testing====
As mentioned in the previous chapter, the benefits of antigen testing For COVID-19 and other viral infections are 1. specimen collection can typically be done with a simple nasal swab rather than a more invasive [[nasopharyngeal swab]], 2. testing is more rapid and convenient, and 3. it takes some pressure off the PCR supply chain. However, antigen testing only tests what's there, rather than amplifying the amount, resulting in generally lower sensitivities.<ref name="ServiceRadical2020">{{cite journal |title=Radical shift in COVID-19 testing needed to reopen schools and businesses, researchers say |journal=Science |author=Service, R.F. |year=2020 |doi=10.1126/science.abe1546}}</ref><ref name="GuglielmiTheExp20">{{cite journal |title=The explosion of new coronavirus tests that could help to end the pandemic |journal=Nature |author=Guglielmi, G. |volume=583 |pages=506–09 |year=2020 |doi=10.1038/d41586-020-02140-8}}</ref> As such, the real utility of antigen testing, despite its lower sensitivity, appears to be surveillance situations where a large group of individuals who are at risk can be screened at regularly scheduled intervals of two to four days. If your lab is able to support this sort of testing, then this type of testing may be an option. As of September 2021, thirty-four FDA EUAs for antigen tests have been issues; 28 of those 34 include allowances for CLIA-waived testing, and 10 were authorized for home use.<ref name="FDAInVitroAntigen21">{{cite web |url=https://www.fda.gov/medical-devices/coronavirus-disease-2019-covid-19-emergency-use-authorizations-medical-devices/in-vitro-diagnostics-euas-antigen-diagnostic-tests-sars-cov-2 |title=In Vitro Diagnostics EUAs - Antigen Diagnostic Tests for SARS-CoV-2 |publisher=U.S. Food and Drug Administration |date=07 September 2021 |accessdate=07 September 2021}}</ref> Review [https://www.fda.gov/medical-devices/coronavirus-disease-2019-covid-19-emergency-use-authorizations-medical-devices/in-vitro-diagnostics-euas-antigen-diagnostic-tests-sars-cov-2 the FDA list] to further examine your options.

The CDC emphasizes that molecular testing remains the "gold standard" for detecting SARS-CoV-2 in a sample, and it "may be necessary to confirm an antigen test result with a laboratory-based NAAT, especially if the result of the antigen test is inconsistent with the clinical context." However, some molecular tests designed for point-of-care testing may not be sufficiently designed for confirmatory testing; consult the instructions for use for any confirmatory test.<ref name="CDCInterim21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/lab/resources/antigen-tests-guidelines.html |title=Interim Guidance for Antigen Testing for SARS-CoV-2 |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=09 September 2021 |accessdate=18 September 2021}}</ref> The CDC [https://www.cdc.gov/coronavirus/2019-ncov/lab/resources/antigen-tests-guidelines.html#anchor_1631295114633 makes available] two antigen testing algorithms for determining when confirmatory testing is actually recommended.<ref name="CDCInterim21" />

====3.1.4 LAMP and CRISPR====
Early on in the pandemic, while PCR was getting most of the attention, reverse transcription loop-mediated isothermal amplification (RT-LAMP), an isothermal [[Nucleic acid test|nucleic acid amplification]] technique that allows for RNA amplification, was also quietly being discussed<ref name="LambRapid20">{{cite journal |title=Rapid Detection of Novel Coronavirus (COVID-19) by Reverse Transcription-Loop-Mediated Isothermal Amplification |journal=medRxiv |author=Lamb, L.E.; Barolone, S.N.; Ward, E. et al. |year=2020 |doi=10.1101/2020.02.19.20025155}}</ref><ref name="Schmid-BurgkLAMP20">{{cite journal |title=LAMP-Seq: Population-Scale COVID-19 Diagnostics Using Combinatorial Barcoding |journal=bioRxiv |author=Schmid-Burgk, J.L.; Li, D.; Feldman, D. et al. |year=2020 |url=https://www.biorxiv.org/content/10.1101/2020.04.06.025635v2.article-info |doi=10.1101/2020.04.06.025635}}</ref>, and it has since gained more attention.<ref name="YuRapid20">{{cite journal |title=Rapid Detection of COVID-19 Coronavirus Using a Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) Diagnostic Platform |journal=Clinical Chemistry |author=Yu, L.; Wu, S.; Hao, X. et al. |volume=66 |issue=7 |pages=975–77 |year=2020 |doi=10.1093/clinchem/hvaa102 |pmid=32315390 |pmc=PMC7188121}}</ref><ref name="ParkDevelop20">{{cite journal |title=Development of Reverse Transcription Loop-Mediated Isothermal Amplification Assays Targeting Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) |journal=Journal of Molecular Diagnostics |author=Park, G.-S.; Ku, K.; Baek, S.-H. et al. |volume=22 |issue=6 |pages=729–35 |year=2020 |doi=10.1016/j.jmoldx.2020.03.006 |pmid=32276051 |pmc=PMC7144851}}</ref><ref name="KellnerARapid20">{{cite journal |title=A rapid, highly sensitive and open-access SARS-CoV-2 detection assay for laboratory and home testing |journal=bioRxiv |author=Kellner, M.J.; Ross, J.J.; Schnabl, J. et al. |year=2020 |doi=10.1101/2020.06.23.166397}}</ref><ref name="ThiAColor20">{{cite journal |title=A colorimetric RT-LAMP assay and LAMP-sequencing for detecting SARS-CoV-2 RNA in clinical samples |journal=Science Translational Medicine |author=Thi, V.L.D.; Herbst, K.; Boerner, K. et al. |at=eabc7075 |year=2020 |doi=10.1126/scitranslmed.abc7075 |pmid=32719001}}</ref><ref name="EsbinOver20">{{cite journal |title=Overcoming the bottleneck to widespread testing: a rapid review of nucleic acid testing approaches for COVID-19 detection |journal=RNA |author=Esbin, M.N.; Whitney, O.N.; Chong, S. et al. |volume=26 |issue=7 |pages=771–83 |year=2020 |doi=10.1261/rna.076232.120 |pmid=32358057 |pmc=PMC7297120}}</ref><ref name="HaleOxford20">{{cite web |url=https://www.fiercebiotech.com/medtech/oxford-researchers-develop-portable-covid-19-test-costing-less-than-25 |title=Oxford researchers develop portable COVID-19 test costing less than $25 |author=Hale, C. |work=Fierce Biotech |date=09 July 2020 |accessdate=07 August 2020}}</ref> In July 2020, the University of Oxford was in the process of getting a rapid, affordable, clinically-validated RT-LAMP test approved for the European market. Oxford also notes that "[a]n advantage of using LAMP technology is that it uses different reagents to most laboratory-based PCR tests."<ref name="HaleOxford20" /> Thi ''et al.'' have tested a two-color RT-LAMP assay with an N gene primer set and diagnostic validation using LAMP-sequencing, concluding that the pairing of the two "could offer scalable testing that would be difficult to achieve with conventional qRT-PCR based tests."<ref name="ThiAColor20" /> And California-based Color Genomics set up their own proprietary RT-LAMP system in the summer of 2020, capable of handling up to 10,000 tests per day.<ref name="SheridanThisCal20">{{cite web |url=https://www.statnews.com/2020/08/06/better-simpler-faster-covid-19-test/ |title=This California company has a better version of a simpler, faster Covid-19 test |author=Sheridan, K. |work=STAT |date=06 August 2020 |accessdate=08 August 2020}}</ref>

In most cases, LAMP-based testing is much simpler than PCR, lacking the requirement of specialized instruments. Despite LAMP generally being thought of as less sensitive than PCR<ref name="SheridanThisCal20" /><ref name="GuglielmiTheExp20">{{cite journal |title=The explosion of new coronavirus tests that could help to end the pandemic |journal=Nature |author=Guglielmi, G. |volume=583 |pages=506–09 |year=2020 |doi=10.1038/d41586-020-02140-8}}</ref><ref name="HeidtSaliva20">{{cite web |url=https://www.the-scientist.com/news-opinion/saliva-tests-how-they-work-and-what-they-bring-to-covid-19-67720 |title=Saliva Tests: How They Work and What They Bring to COVID-19 |author=Heidt, A. |work=The Scientist |date=09 July 2020 |accessdate=08 August 2020}}</ref>, the explosion of research into RT-LAMP methods for testing for the presence of SARS-CoV-2 continues to indicate that "under optimized conditions," RT-LAMP methods may actually be able to rival the sensitivity and specificity of many RT-PCR COVID-19 tests.<ref name="EsbinOver20" /> Esbin ''et al.'' add<ref name="EsbinOver20" />:

<blockquote>These methods allow for faster amplification, less specialized equipment, and easy readout. LAMP methods also benefit from the ability to multiplex targets in a single reaction and can be combined with other isothermal methods, like &#91;[[recombinase polymerase amplification]]&#93; in the RAMP technique, to increase test accuracy even more. These techniques may be particularly useful for rapid, point-of-care diagnoses or for remote clinical testing without the need for laboratory equipment.</blockquote>

CRISPR methods are also being used in conjunction with RT-LAMP.<ref name="GuglielmiTheExp20" /><ref name="EsbinOver20" /><ref name="BroughtonCRISPR20">{{cite journal |title=CRISPR–Cas12-based detection of SARS-CoV-2 |journal=Nature Biotechnology |author=Broughton, J.P.; Deng, X.; Yu, G. et al. |volume=38 |pages=870–74 |year=2020 |doi=10.1038/s41587-020-0513-4 |pmid=32300245}}</ref> RT-LAMP creates complementary double-stranded DNA (cDNA) from specimen RNA and then copies (amplifies) it. Then CRISPR methods are used to detect a predefined coronavirus sequence (from a cleaved molecular marker) in the resulting amplified specimen. Though as of September 2021 approved assays using CRISPR-based detection of SARS-CoV-2 are limited to a handful of companies<ref name="FDAInVitroEUAs21" /><ref name="GuglielmiTheExp20" /><ref name="GDHCRISPR20">{{cite web |url=https://www.medicaldevice-network.com/comment/crispr-biotechnology-disrupt-covid-19-testing-market/ |title=CRISPR biotechnology set to disrupt Covid-19 testing market |author=GlobalData Healthcare |work=Verdict Medical Devices |date=14 July 2020}}</ref>, the technology has some promise as an alternative testing method. CRISPR has the additional advantage of being readily coupled with lateral flow assay technology to be deployed in the point-of-care (POC) setting<ref name="EsbinOver20" /><ref name="GDHCRISPR20" />, though it's worth noting the currently EUAed RT-LAMP-based CRISPR kits are only approved for high-complexity CLIA labs. (The current molecular diagnostic test kits running CRISPR technology are Sherlock BioSciences' Sherlock CRISPR SARS-CoV-2 Kit and Mammoth Biosciences' SARS-CoV-2 DETECTR Reagent Kit, both high-complexity.<ref name="FDAInVitroEUAs21" />)

====3.1.5 Point-of-care and other alternative testing====
[[File:Microfluidic chip for point-of-care medical devices.jpg|right|thumb|400px|Example of a microfluidic chip used in point-of-care medical devices]]On September 28, 2020, the WHO published its blueprint for what they call Target Product Profiles (TPP), which "describe the desirable and minimally acceptable profiles" for four different COVID-19 test categories.<ref name="WHOCOVIDTarget20">{{cite web |url=https://www.who.int/publications/m/item/covid-19-target-product-profiles-for-priority-diagnostics-to-support-response-to-the-covid-19-pandemic-v.0.1 |title=COVID-19 Target product profiles for priority diagnostics to support response to the COVID-19 pandemic v.1.0 |author=World Health Organization |publisher=World Health Organization |date=28 September 2020 |accessdate=08 September 2021}}</ref> Addressing POC testing, the WHO recommends that such assays<ref name="WHOCOVIDTarget20" /><ref name="PeplowRapid20">{{cite web |url=https://cen.acs.org/analytical-chemistry/diagnostics/Rapid-COVID-19-testing-breaks/98/web/2020/08 |title=Rapid COVID-19 testing breaks free from the lab |author=Peplow, M. |work=Chemical & Engineering News |date=10 August 2020 |accessdate=12 August 2020}}</ref>:

* have a sensitivity (true positive rate) of at least 80 percent, with 90 percent or better being desirable;
* have a specificity (true negative rate) of at least 97 percent, with greater than 99 percent being desirable;
* provide results in less than 40 minutes, with less 20 minutes or less being desirable;
* have "a cost that allows broad use, including in low- and middle-income countries";
* be simple enough that only a half day to, optimally, a few hours of training are required to run the test; and
* operate reliably outside a clean laboratory environment.

Though at the time of the announcement few of the available test systems could likely meet all these requirements, it's clear this and other urgencies have put pressure on manufacturers to expand COVID-19 testing to the point of care setting.<ref name="PeplowRapid20" /><ref name="KriegerCorona20">{{cite web |url=https://www.mercurynews.com/2020/08/10/coronavirus-how-to-test-everyone-all-the-time/ |title=Coronavirus: How to test everyone, all the time |author=Krieger, L.M. |work=The Mercury News |date=10 August 2020 |accessdate=12 August 2020}}</ref><ref name="BrownPoint20">{{cite web |url=https://www.mcknights.com/news/point-of-care-testing-could-be-biggest-advance-in-covid-19-fight/ |title=Point-of-care testing could be ‘biggest advance’ in COVID-19 fight |author=Brown, D. |work=McKnight's |date=10 August 2020 |accessdate=12 August 2020}}</ref><ref name="WissonCOVID20">{{cite web |url=https://www.healtheuropa.eu/covid-19-and-effective-cohorting-rapid-point-of-care-triage-testing/101696/ |title=COVID-19 and effective cohorting: Rapid point of care triage testing |author=Wisson, J. |work=Health Europa |date=28 July 2020 |accessdate=12 August 2020}}</ref> Additional incentives were offered by the U.S. National Institutes of Health's Rapid Acceleration of Diagnostics (RADx) funding program, which sought to speed up innovation in COVID-19 testing and promote "truly nontraditional approaches for testing that have a slightly longer horizon."<ref name="TrombergRapid20">{{cite journal |title=Rapid Scaling Up of Covid-19 Diagnostic Testing in the United States — The NIH RADx Initiative |journal=New England Journal of Medicine |author=Tromberg, B.J.; Schwetz, T.A.; Pérez-Stable, E.J. et al. |year=2020 |doi=10.1056/NEJMsr2022263}}</ref> In August 2020, RADx had chosen to fund seven biomedical diagnostic companies making new lab-based and POC tests that could significantly ramp up overall testing in the U.S. into September 2020. Four of those offerings were lab-based (from Ginkgo Bioworks, Helix OpCo, Fluidigm, and Mammoth Biosciences) and three were POC tests (from Mesa Biotech, Quidel, and Talis Biomedical), all using varying technologies and methods such as next-generation sequencing, CRISPR, microfluidic chips, nucleic acid testing, antigen testing, and saliva testing.<ref name="NIHDelivering20">{{cite web |url=https://www.nih.gov/news-events/news-releases/nih-delivering-new-covid-19-testing-technologies-meet-us-demand |title=NIH delivering new COVID-19 testing technologies to meet U.S. demand |author=National Institutes of Health |publisher=National Institutes of Health |work=News Releases |date=31 July 2020 |accessdate=12 August 2020}}</ref> On October 28, 2020, RADx added an additional 15 biomedical diagnostics projects for funding, for a total of 22.<ref name="NIHRADxTech20">{{cite web |url=https://www.nih.gov/research-training/medical-research-initiatives/radx/funding#radx-tech-atp-funded |title=Funded Projects - RADx Tech/ATP |publisher=National Institutes of Health |date=28 October 2020 |accessdate=19 November 2020}}</ref> As of September 2021, some of those 22 programs have come to fruition, garnering FDA EUAs, including Mesa Biotech's rapid cartridge-based RT-PCR Accula System, Quidel's rapid Sofia SARS Antigen FIA test, Mammoth Bioscience's SARS-CoV-2 DETECTR Reagent Kit, and Visby Medical's COVID-19 Point of Care Test.<ref name="FDAInVitroEUAs21" />

Outside the RADx program, enterprising researchers in other parts of the world are also attempting non-traditional approaches to improving COVID-19 testing options. Examples include<ref name="EsbinOver20" /><ref name="WissonCOVID20" /><ref name="Leichman10Ways20">{{cite web |url=https://www.israel21c.org/how-israeli-scientists-are-improving-corona-testing/ |title=10 ways Israeli scientists are improving corona testing |author=Leichman, A.K. |work=Isael21c |date=27 July 2020 |accessdate=11 August 2020}}</ref><ref name="UNRCOVID20">{{cite web |url=https://www.sciencedaily.com/releases/2020/10/201014141032.htm |title=COVID-19 rapid test has successful lab results, research moves to next stages: Engineers and virologists team up for novel approach |author=University of Nevada, Reno |work=ScienceDaily |date=14 October 2020 |accessdate=19 November 2020}}</ref>:

* a method of DNA nanoswitch detection of virus particles;
* a dual biomarker-based finger-stick test for acute respiratory infections;
* a rapid breath test to detect volatile organic chemicals from the lungs;
* an affordable, hand-held spectral imaging device to detect virus in blood or saliva in seconds;
* an ultrahigh frequency spectroscopic scanning device to see virus particles resonating;
* a method that combines optical devices and magnetic particles to detect virus RNA;
* an RNA extraction protocol that uses magnetic bead-based kits;
* a nanotube-based electrochemical biosensor for detecting biomarkers in a sample in less than a minute;
* the additional use of an [[artificial intelligence]] (AI) application to better scrutinize test results; and
* the miniaturization of PCR technology to make it more portable and user-friendly.

Of course, most of these are largely experimental technologies, and realistically getting them into the lab may be far out. But they represent out-of-the-box ideas that have some kind of chance at playing a greater role in the clinical laboratory or in point-of-care settings in the future.

====3.1.6 Multiplex testing====
As the pandemic has progressed and test manufacturers have become more experienced with SARS-CoV-2 test development, multiplex testing has become an option. The multiplex assay—an immunoassay test able to measure multiple analytes in a single test—is certainly not new in itself. In 1989, R. Ekins developed the ambient analyte theory, which stated that miniaturizing an immunoassay can lead to an improved limit of detection (LOD). That work influenced the future development of microarray multiplex technology principles.<ref name="TigheELISA15">{{Cite journal |last=Tighe |first=Patrick J. |last2=Ryder |first2=Richard R. |last3=Todd |first3=Ian |last4=Fairclough |first4=Lucy C. |date=2015-04 |title=ELISA in the multiplex era: Potentials and pitfalls |url=https://onlinelibrary.wiley.com/doi/10.1002/prca.201400130 |journal=PROTEOMICS – Clinical Applications |language=en |volume=9 |issue=3-4 |pages=406–422 |doi=10.1002/prca.201400130 |issn=1862-8346 |pmc=PMC6680274 |pmid=25644123}}</ref> By 2013, development of multiplex protein immunoassays was becoming increasingly prominent.<ref name="TigheELISA15" />

As of September 2021, eighteen "multi-analyte" ''in vitro'' molecular diagnostic tests are shown as receiving EUAs by the FDA, four of them even authorized for CLIA waived testing.<ref name="FDAInVitroEUAs21">{{cite web |url=https://www.fda.gov/medical-devices/coronavirus-disease-2019-covid-19-emergency-use-authorizations-medical-devices/in-vitro-diagnostics-euas-molecular-diagnostic-tests-sars-cov-2 |title=In Vitro Diagnostics EUAs - Molecular Diagnostic Tests for SARS-CoV-2 |publisher=U.S. Food and Drug Administration |date=07 September 2021 |accessdate=07 September 2021}}</ref> Common additional targets for analysis among the various kits include influenza A, influenza B, and respiratory syncytial virus (RSV).<ref name="FDAInVitroEUAs21" /> However, several multiplex test kits cover an even broader array of respiratory-affecting organism types and subtypes such as adenovirus and a few other coronavirus types, to name a few. Kits include the ePlex Respiratory Pathogen Panel 2<ref name="HintonePlex20">{{cite web |url=https://www.fda.gov/media/142902/download |format=PDF |title=ePlex Respiratory Pathogen Panel 2 (ePlex RP2 Panel) |author=Hinton, D.M. |publisher=U.S. Food and Drug Administration |date=07 October 2020 |accessdate=19 September 2021}}</ref>, the NxTAG Respiratory Pathogen Panel + SARS-CoV-2<ref name="HintonNxTAG21">{{cite web |url=https://www.fda.gov/media/146492/download |format=PDF |title=NxTAG Respiratory Pathogen Panel + SARS-CoV-2 |author=Hinton, D.M. |publisher=U.S. Food and Drug Administration |date=03 March 2021 |accessdate=19 September 2021}}</ref>, the QIAstat-Dx Respiratory SARS-CoV-2 Panel<ref name="HintonQIAstat21">{{cite web |url=https://www.fda.gov/media/136569/download |format=PDF |title=QIAstat-Dx Respiratory SARS-CoV-2 Panel |author=Hinton, D.M. |publisher=U.S. Food and Drug Administration |date=29 July 2021 |accessdate=19 September 2021}}</ref>, and the BioFire Respiratory Panel 2.1-EZ.<ref name="HintonBioFireRes21">{{cite web |url=https://www.fda.gov/media/142693/download |format=PDF |title=BioFire Respiratory Panel 2.1-EZ (RP2.1-EZ) |author=Hinton, D.M. |publisher=U.S. Food and Drug Administration |date=30 August 2021 |accessdate=19 September 2021}}</ref> (Of the four, the BioFire panel is approved for CLIA waived testing.<ref name="FDAInVitroEUAs21" />) Adding multiplex testing of SARS-CoV-2 plus other organisms to your laboratory will largely revolve around your lab's CLIA status and assessment of the available options.

Multiplexing provides a variety of benefits for laboratories and patients. In their 2015 paper on ELISA and multiplex technologies, Tighe ''et al'' find that multiplexed immunoassays have the potential to decrease diagnosis times and reduce assay costs. "At the same time, such multiplexing offers more comprehensive analysis whether for research purposes, differential diagnoses, or monitoring of therapeutic interventions."<ref name="TigheELISA15" /> They also note the potential for improved health surveillance of patients, catching early-onset diseases by looking for informative biomarkers.<ref name="TigheELISA15" /> From the perspective of diagnosing infections of SARS-CoV-2 or influenza, the CDC adds that multiplexing helps preserve testing supplies that may be in short supply, conduct more tests in a given time period, and paint a clearer picture of both viruses and their prevalence in a given population.<ref name="CDCInflu21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/lab/multiplex.html |title=CDC’s Influenza SARS-CoV-2 Multiplex Assay and Required Supplies |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=13 July 2021 |accessdate=19 September 2021}}</ref>

====3.1.7 Variant testing====
As the pandemic has progressed, you may have heard talk of a "delta" variant of SARS-CoV-2, which is reportedly more contagious and virulent than the initial strain that kicked off the pandemic.<ref name="CDCDelta21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/variants/delta-variant.html |title=Delta Variant: What We Know About the Science |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=26 August 2021 |accessdate=18 September 2021}}</ref> One or more variants of a virus are expected as time progresses, and some of those variants can cause significantly more problems than the source virus. As such, analytical testing of the virus over time is vital to public health.

The purpose of variant testing can be described in two ways, one for public health reasons and another for clinical care reasons. On the public health side, analysis of SARS-CoV-2 variants provides an unbiased, population-level view "of the specific viral strains in circulation and monitors changes in the viral genome over time."<ref name="BuchanSARS21">{{cite web |url=https://www.amp.org/AMP/assets/File/clinical-practice/COVID/AMP_RC_VariantTestingforSARSCOV2_4_28_21.pdf |format=PDF |title=SARS-CoV-2 Variant Testing |work=Rapid Communication |author=Buchan, B.W.; Wolk, D.M.; Yao, J.D. |publisher=Association for Molecular Pathology |date=28 April 2021 |accessdate=18 September 2021}}</ref> With enough public health laboratories conducting this type of analysis—typically whole-genome sequencing (WGS) using [[next-generation sequencing]] (NGS) techniques—a clearer picture of how an outbreak spreads is gained, as well as what variants are taking hold and further threatening human populations (even those that are vaccinated). This information is typically shared through the public health system for surveillance and reporting purposes, though the affected patients themselves may never see the data.<ref name="BuchanSARS21" />

On the clinical care side, analysis of SARS-CoV-2 variants provides further insights into improving COVID-19 patient outcomes. Buchan ''et al.'' identify three potential insights that clinicians may gain, noting that variant testing allows the clinician<ref name="BuchanSARS21" />:

* to distinguish between an existing, persistent infection caused by one viral strain vs. re-infection by a different viral strain;
* to determine whether a patient not responding to a treatment is affected by a specific viral spike protein (S) gene mutation that is "potentially resistant or less susceptible to neutralizing antibodies or monoclonal antibodies"; and
* to detect in the serum or plasma of a patient post-vaccination "viral S gene substitutions in specific variants that are potentially resistant or less susceptible" to the antibodies the vaccine generates.

If, for example, a patient is diagnosed with a variant that is tied to heightened disease severity, the clinician can opt for additional treatments early on to counteract the variant's effects on the patient. This testing is done in a hospital or reference lab by WGS or by targeting a portion of the genome (e.g., a spike protein) or a specific mutation (using RT-PCR). However, according to Buchan ''et al.'', the contributions a mutation makes to a "variant's attributes is not entirely understood, and there is no definitive evidence that directly links a given mutation to poor outcomes, significantly reduced efficacy of SARS-CoV-2 therapies, or vaccine coverage."<ref name="BuchanSARS21" />

That said, and leaving the public health element to the side, if you are a laboratory conducting clinical analyses of SARS-CoV-2 specimens, the likelihood of including viral sequencing and sequence analysis for variant testing may be low for your facility. Such testing is a multi-step process requiring a non-trivial set of resources, often available to large commercial diagnostic laboratories.<ref name="BuchanSARS21" /><ref name="WilliamsEnhanc21">{{cite web |url=https://health.mo.gov/emergencies/ert/alertsadvisories/pdf/update21921.pdf |format=PDF |title=Enhancing Public Health Surveillance for Variant SARSCoV-2 Viruses in Missouri |author=Williams, R.W. |publisher=Missouri Department of Health & Senior Services |date=19 February 2021 |accessdate=18 September 2021}}</ref> The CDC represents one possible workflow for genomic sequencing as such<ref name="CDCRole21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/variants/cdc-role-surveillance.html |title=CDC’s Role in Tracking Variants |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=08 September 2021 |accessdate=18 September 2021}}</ref>:

# A specimen containing the SARS-CoV-2 virus is received by the lab and promptly entered into the [[laboratory information system]] (LIS).
# The RNA of the SARS-CoV-2 virus is extracted from the sample and then converted to complimentary DNA. It is then enriched and loaded into the appropriate NGS instrument.
# The instrument runs the sequencing and raw data is collected, with the lab maintaining quality control steps. The raw data is turned into actionable sequence data.
# The sequence data is verified for suitability, with a resequencing occurring if found to be inadequate. Otherwise, the data is then analyzed and interpreted.
# The final approved sequencing results are reported to the appropriate state, local, tribal, or territorial public health department.<ref name="CDCGuidanceSeq21">{{cite web |url=https://www.cdc.gov/coronavirus/2019-ncov/lab/resources/reporting-sequencing-guidance.html |title=Guidance for Reporting SARS-CoV-2 Sequencing Result |author=Centers for Disease Control and Prevention |publisher=Centers for Disease Control and Prevention |date=23 June 2021 |accessdate=19 September 2021}}</ref>

If your diagnostic lab has or is planning on adding sequencing tools to supplement clinical diagnostics, it may make sense to consider adding variant testing to your available options. But in reality, this sort of testing may largely be left to large institutions, such as the University of Rochester Vaccine Treatment Evaluation Unit or the Yale School of Public Health.<ref name="DupuyCOVID21">{{cite web |url=https://apnews.com/article/fact-checking-549965482111 |title=COVID-19 variants tested through genome sequencing |author=Dupuy, B. |work=Reuters Fact-Checking |date=28 July 2021 |accessdate=18 September 2021}}</ref>