The Xpert Xpress CoV-2 plus test, performed on the GeneXpert Infinity Systems, is a rapid real-time RT-PCR test intended for the qualitative detection of SARS-CoV-2 RNA in nasopharyngeal and anterior nasal swab specimens collected from individuals with signs and symptoms of respiratory tract infection.

The Xpert Xpress CoV-2 plus test is intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical, epidemiologic, and laboratory findings. Positive results are indicative of the presence of SARS-CoV-2 RNA. Positive results do not rule out bacterial infection or co-infection with other pathogens.

Negative results do not preclude SARS-CoV-2 infection. The results of this test should not be used as for diagnosis and patient management decisions.

Device Description

The Xpert Xpress CoV-2 plus test is a rapid, automated in vitro diagnostic test for the qualitative detection of viral RNA from SARS-CoV-2 in nasopharyngeal swab (NPS) and anterior nasal swab (NS) specimens obtained from individuals with signs and symptoms of respiratory tract infection.

The Xpert Xpress CoV-2 plus test is performed on the Cepheid GeneXpert® Instrument Systems (GeneXpert Dx, GeneXpert Infinity-48s and GeneXpert Infinity-80 systems), which consist of an instrument, computer and preloaded software for running tests and viewing the results. The GeneXpert Instrument Systems automate and integrate sample preparation, nucleic acid extraction and amplification, and detection of the target sequences in simple or complex samples using real-time reverse transcription (RT)-polymerase chain reaction (PCR) and PCR assays. Depending on the instrument, the GeneXpert Instrument Systems can have from 1 and up to 80 randomly accessible modules, each capable of performing separate sample preparation and real-time RT-PCR and PCR tests. Each module contains a syringe drive for dispensing fluids (i.e., the syringe drive activates the plunger that works in concert with the rotary valve in the cartridge to move fluids between chambers), an ultrasonic horn for lysing cells or spores, and a proprietary I-CORE® thermocycler for performing real-time RT-PCR and PCR as well as detection. The systems require the use of single-use disposable cartridges that hold the RT-PCR reagents and host sample purification, nucleic acid amplification, and detection of the target sequences. Because the cartridges are selfcontained, cross-contamination between cartridges during the testing process is minimized.

The Xpert Xpress CoV-2 plus test includes reagents for the detection of viral RNA from SARS-CoV-2 in NPS and NS specimens. The primers and probes in the Xpert Xpress CoV-2 plus test are designed to amplify and detect sequences in the genes that encode the following SARS-CoV-2 proteins: nucleocapsid (N2), envelope (E), and RNA-dependent RNA polymerase (RdRP). A Sample Processing Control (SPC) and a Probe Check Control (PCC) are also included in the cartridge utilized by the GeneXpert instrument. The SPC is present to control for adequate processing of the sample and to monitor for the presence of potential inhibitor(s) in the RT-PCR reaction. The SPC also ensures that the RT-PCR reaction conditions (temperature and time) are appropriate for the amplification reaction and that the RT-PCR reagents are functional. The PCC verifies reagent rehydration. PCR tube filling, and confirms that all reaction components are present in the cartridge including monitoring for probe integrity and dve stability.

AI/ML Overview

Here's a breakdown of the acceptance criteria and the study details for the Cepheid Xpert Xpress CoV-2 plus, based on the provided document:

Acceptance Criteria and Device Performance

Note: The document explicitly states the "claimed LoD" which can be interpreted as an acceptance criterion for analytical sensitivity. For clinical performance, the reported PPA and NPA values serve as the acceptance criteria.

Acceptance Criterion (Test Type)	Reported Device Performance
Analytical Sensitivity (LoD) - Clinical NPS-UTM/VTM Matrix	403 copies/mL (N2 target)
Analytical Sensitivity (LoD) - Clinical NS-UTM/VTM Matrix	462 copies/mL (N2 target)
Analytical Sensitivity (LoD) - WHO First International Standard SARS-CoV-2 RNA in clinical NS-UTM/VTM Matrix	1000 IU/mL
Analytical Inclusivity (Primers E, N2, RdRP amplicons)	E: 100%, N2: 99.95%, RdRP: 100% (Predicted)
Analytical Inclusivity (Probes E, N2, RdRP)	E: 100%, N2: 100%, RdRP: 99.6% (Predicted)
Analytical Inclusivity (Wet-Testing, 61 strains)	100% detection of all 61 SARS-CoV-2 strains tested (3/3 replicates positive for all)
Analytical Exclusivity (In silico)	No potential unintended cross-reactivity with listed organisms (E gene cross-reactivity with SARS-CoV, Human and Bat SARS-coronavirus expected)
Analytical Exclusivity (Wet-Testing)	No cross-reactivity with 62 non-SARS-CoV-2 microorganisms (except expected E gene cross-reactivity with SARS-coronavirus Urbani)
Microbial Interference	No interference from 18 commensal microorganisms at tested concentrations (8/8 correct results)
Potentially Interfering Substances	No interference from 21 of 23 substances at tested concentrations. Fluticasone propionate nasal spray and mucin type I-S interfered at higher concentrations, but not at half the concentration.
Carryover Contamination	0% carryover contamination (40/40 positive, 42/42 negative correctly identified)
Reproducibility (Qualitative Agreement - Negative)	99.3% (142/143) [96.1% - 99.9% CI]
Reproducibility (Qualitative Agreement - Low Positive)	100% (144/144) [97.4% - 100% CI]
Reproducibility (Qualitative Agreement - Moderate Positive)	100% (144/144) [97.4% - 100% CI]
Single-Site Precision (Qualitative Agreement - Negative)	100% (80/80) [95.4%-100.0% CI]
Single-Site Precision (Qualitative Agreement - Low Positive)	100% (80/80) [95.4%-100.0% CI]
Single-Site Precision (Qualitative Agreement - Moderate Positive)	100% (80/80) [95.4%-100.0% CI]
Clinical Performance (Overall PPA)	98.1% (95% CI: 96.7% - 98.9%)
Clinical Performance (Overall NPA)	98.3% (95% CI: 97.7% - 98.7%)
Clinical Performance (NPS PPA)	97.0% (95% CI: 94.4% - 98.4%)
Clinical Performance (NPS NPA)	98.2% (95% CI: 97.4% - 98.8%)
Clinical Performance (NS PPA)	99.3% (95% CI: 97.5% - 99.8%)
Clinical Performance (NS NPA)	98.3% (95% CI: 97.5% - 98.8%)

Study Details

2. Sample size used for the test set and the data provenance (e.g. country of origin of the data, retrospective or prospective)

Clinical Performance Test Set:
- Initial Specimens: 4047 specimens (2029 Nasopharyngeal Swab (NPS) and 2018 Anterior Nasal Swab (NS)).
- Valid Specimens (after exclusions): 3750 specimens (1879 NPS and 1871 NS).
- Data Provenance: United States (32 geographically diverse sites), prospectively collected, fresh (98.6%) and frozen (1.4%) clinical specimens collected in 2022.
Analytical Performance Test Sets:
- LoD (NPS & NS): Replicates of 20 per reagent lot (2 lots).
- LoD (WHO Standard): 20 replicates.
- Analytical Inclusivity (Wet Testing): 61 SARS-CoV-2 strains, 3 replicates each.
- Analytical Exclusivity (Wet Testing): 62 microorganisms, 3 replicates each.
- Microbial Interference: 18 commensal microorganisms, 8 replicates each with SARS-CoV-2.
- Potentially Interfering Substances: 23 substances, 8 replicates each for negative and positive samples.
- Carryover Contamination: 40 positive samples and 42 negative samples.
- Reproducibility: 144 observations per panel member (3 sites x 2 operators x 3 lots x 2 days/lot x 2 runs x 2 replicates).
- Single-Site Precision: 80 observations per panel member (1 site x 1 operator x 1 lot x 20 days x 2 runs x 2 replicates).
- Data Provenance: Not explicitly stated for analytical studies, but typically performed in-house or by contracted labs. The substances and strains used are clearly defined.

3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts (e.g. radiologist with 10 years of experience)

The clinical performance study used a U.S. FDA-cleared molecular respiratory panel that includes SARS-CoV-2 as the comparator (reference method). This type of comparator assay is itself considered a "ground truth" established by its own regulatory clearance process, relying on its validated accuracy rather than individual expert adjudication for each case in this study.
Qualifications of experts: Not applicable as ground truth was established by a cleared molecular diagnostic test.

4. Adjudication method (e.g. 2+1, 3+1, none) for the test set

Clinical Performance: The comparator was a U.S. FDA-cleared molecular respiratory panel. For discrepant results with this comparator, additional testing was performed using a U.S. FDA EUA SARS-CoV-2 molecular test. The text indicates that these discrepant test results were used to resolve the discrepancies (e.g., 7/9 SARS-CoV-2 negative; 8/28 SARS-CoV-2 positive from the specific footnotes), implying a third party or a consensus reference method was used for discordant samples. This suggests a form of discrepant analysis, where a third, highly accurate method or a re-test with a known method, is used to resolve differences between the investigational device and the primary comparator.

5. If a multi reader multi case (MRMC) comparative effectiveness study was done, If so, what was the effect size of how much human readers improve with AI vs without AI assistance

Not applicable. This submission describes a molecular diagnostic test (in vitro diagnostic device), not an imaging-based AI diagnostic device requiring human reader interpretation in an MRMC study. The device provides a direct qualitative result (POSITIVE/NEGATIVE).

6. If a standalone (i.e. algorithm only without human-in-the-loop performance) was done

Yes, this is a standalone device. The Xpert Xpress CoV-2 plus test is an automated, real-time RT-PCR test performed on GeneXpert Instrument Systems. Its output is a qualitative result (SARS-CoV-2 POSITIVE or NEGATIVE) based on the algorithm within the instrument, without human interpretation of raw data for diagnosis. Although human operators load samples and review results, the diagnostic determination itself is made by the system.

7. The type of ground truth used (expert consensus, pathology, outcomes data, etc)

Clinical Performance: The ground truth for clinical performance was established by concordance with a U.S. FDA-cleared molecular respiratory panel and, for discrepant cases, a U.S. FDA EUA SARS-CoV-2 molecular test. This represents a highly accurate molecular diagnostic reference method.
Analytical Performance: Ground truth for analytical studies was established using known concentrations of inactivated SARS-CoV-2 virus, WHO International Standard for SARS-CoV-2 RNA, genomic RNA/DNA of various microorganisms, and in silico analyses against sequence databases.

8. The sample size for the training set

Not explicitly stated in terms of a "training set". For molecular diagnostic tests, "training" typically refers to the development and optimization of the primer and probe design, and algorithmic thresholds. This development process often involves internal studies and iterations rather than a distinct "training set" in the machine learning sense. The extensive analytical performance studies (LoD, inclusivity, exclusivity) serve to validate the developed algorithm's performance against diverse known samples. In silico analyses involved millions of SARS-CoV-2 sequences.

9. How the ground truth for the training set was established

As noted above, a distinct "training set" in the machine learning context is not explicitly described. However, the ground truth for optimizing the assay's primers, probes, and detection thresholds would have been established through:
- Known concentrations of synthetic SARS-CoV-2 genetic material or inactivated virus.
- In silico analysis against extensive public sequence databases (e.g., GISAID, NCBI) to identify target regions and ensure inclusivity of variants while maintaining exclusivity against other pathogens.
- Laboratory-prepared samples with known microbial content and concentrations used to optimize and validate analytical performance characteristics like sensitivity and specificity.

Summary

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October 13, 2023

Image /page/0/Picture/1 description: The image shows the logo of the U.S. Food and Drug Administration (FDA). The logo consists of two parts: the Department of Health & Human Services logo on the left and the FDA logo on the right. The FDA logo includes the letters "FDA" in a blue square, followed by the words "U.S. FOOD & DRUG" in blue, with the word "ADMINISTRATION" underneath.

Cepheid Jennifer Motto Principal Regulatory Affairs Specialist 904 Caribbean Drive Sunnyvale, California 94089

Re: K230440

Trade/Device Name: Xpert Xpress CoV-2 plus Regulation Number: 21 CFR 866.3981 Regulation Name: Device To Detect And Identify Nucleic Acid Targets In Respiratory Specimens From Microbial Agents That Cause The SARS-Cov-2 Respiratory Infection And Other Microbial Agents When In A Multi-Target Test Regulatory Class: Class II Product Code: QQX, OOI Dated: September 1, 2023 Received: September 12, 2023

Dear Jennifer Motto:

We have reviewed your section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (the Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database available at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device"

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(https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).

Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30. Design controls; 21 CFR 820.90. Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review. the OS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safetyreporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely. Himani Bisht -S

Himani Bisht, Ph.D. Assistant Director Viral Respiratory and HPV Branch Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Quality

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Enclosure

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Indications for Use

510(k) Number (if known) K230440

Device Name Xpert Xpress CoV-2 plus

Indications for Use (Describe)

Negative results do not preclude SARS-CoV-2 infection. The results of this test should not be used as for diagnosis and patient management decisions.

Type of Use (Select one or both, as applicable)

☑ Prescription Use (Part 21 CFR 801 Subpart D)
☐ Over-The-Counter Use (21 CFR 801 Subpart C)

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Image /page/4/Picture/0 description: The image contains the logo for Cepheid, a molecular diagnostics company. The logo features a stylized blue wing-like graphic above the company name. Below the company name, the text "Xpert® Xpress CoV-2 plus" is displayed, indicating a product related to COVID-19 testing.

5. 510(k) Summary

As required by 21 CFR Section 807.92(c).

Submitted by:	Cepheid904 Caribbean DriveSunnyvale, CA 90489Phone number: (408) 541-4191
Contact:	Jennifer Motto, M.S., RAC
Date of Preparation:	February 17, 2023
Device:
Trade name:Common name:Type of Test:	Xpert® Xpress CoV-2 plusXpert Xpress CoV-2 plusQualitative real-time reverse transcription polymerase chainreaction (RT-PCR) and detection test
Regulation number:Classification name:	21 CFR 866.3981Multi-target respiratory specimen nucleic acid test includingSARS-CoV-2 and other microbial agents,Respiratory Specimen Nucleic Acid SARS-CoV-2 Test,
Primary productcode:	QQX
Secondary productcode	OOI
ClassificationAdvisory Panel	Microbiology (83)
Prescription Use	Yes
Predicate DeviceAssay:	BioFire COVID-19 Test 2 (K211079)

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5.1. Device Description

The Xpert Xpress CoV-2 plus test is designed for use with NPS or NS specimen collected with nylon flocked swabs and placed into a viral transport medium (VTM), Universal Transport Medium (UTM) or eNAT®. Examples of the ancillary specimen collection kits that are compatible for use with the Xpert Xpress CoV-2 plus test include:

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. Nasopharyngeal Sample Collection Kit for Viruses

Copan UTM® 3C057N (Flexible Minitip Flocked Swab with UTM® Medium O w/o Beads)
Becton Dickinson Universal Viral Transport Kit P/N 220531 (Flexible Minitip O Flocked Swab with UVT Medium)
Copan eNAT® Molecular Collection and Preservation Medium P/N 6U074S01 O (Flexible Minitip Flocked Swab with eNAT® Medium)

● Nasal Sample Collection Kit for Viruses

Copan UTM® 3C064N (Regular Flocked Swab with UTM® Medium w/o O Beads)
Copan eNAT® Molecular Collection and Preservation Medium P/N o 6U073S01 (Regular Flocked Swab with eNAT® Medium)
Alternatively, swabs and transport media can be obtained separately:
- Nylon flocked swab (Copan P/N 502CS01, 503CS01) O
- Viral transport medium, 3 mL (Copan P/N 3C047N, BD Universal o Transport Medium, Remel M4RT, Remel M5) or equivalent

These sample collection kits for viruses allow NPS and NS specimens from patients to be collected, preserved and transported to laboratory prior to analysis with the Xpert Xpress CoV-2 plus test.

5.2. Device Intended Use

The Xpert Xpress CoV-2 plus test, performed on the GeneXpert Dx and GeneXpert Infinity Systems, is a rapid real-time RT-PCR test intended for the qualitative detection of SARS-CoV-2 RNA in nasopharyngeal and anterior nasal swab specimens collected from individuals with signs and symptoms of respiratory tract infection.

Negative results do not preclude SARS-CoV-2 infection. The results of this test should not be used as the sole basis for diagnosis and patient management decisions.

5.3. Substantial Equivalence

Table 5-1 shows the similarities and differences between Xpert Xpress CoV-2 plus and the predicate device, BioFire COVID-19 Test 2 [K211079].

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Table 5-1. Comparison of Similarities and Differences Between Xpert Xpress CoV-2 plus and
the Predicate Device

Attribute	New DeviceCepheidXpert® Xpress CoV-2 plus	Predicate Device – K211079BioFire Defense, LLCBioFire COVID-19 Test 2
Regulation	Same	21 CFR 866.3981Devices to detect and identify nucleic acid targets in respiratory samples from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-analyte test
Product Code	Same	QQXRespiratory Specimen Nucleic Acid SARS-CoV-2 Test
Device Class	Same	II (Special Controls)
Technology/Detection	Real-time reverse transcription polymerase chain reaction (RT-qPCR)	Nested multiplex RT-PCR followed by high resolution melting analysis to confirm identity of amplified product
Intended Use	The Xpert Xpress CoV-2 plus test, performed on the GeneXpert Dx and GeneXpert Infinity Systems, is a rapid real-time RT-PCR test intended for the qualitative detection of SARS-CoV-2 RNA in nasopharyngeal and anterior nasal swab specimens collected from individuals with signs and symptoms of respiratory tract infection.The Xpert Xpress CoV-2 plus test is intended for use as an aid in the diagnosis of COVID-19 if used in conjunction with other clinical, epidemiologic, and laboratory findings. Positive results are indicative of the presence of SARS-CoV-2 RNA. Positive results do not rule out bacterial infection or co-infection with other pathogens.Negative results do not preclude SARS-CoV-2 infection. The results of this test should not be used as the sole basis for diagnosis and patient management decisions.	The BioFire COVID-19 Test 2 is a qualitative nested multiplexed RT-PCR in vitro diagnostic test intended for use with the BioFire FilmArray 2.0 and BioFire FilmArray Torch Systems. The BioFire COVID-19 Test 2 detects nucleic acids from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in nasopharyngeal swabs (NPS) from symptomatic individuals suspected of COVID-19 by their healthcare provider.Results are for the identification of SARS-CoV-2 RNA. The SARS-CoV-2 RNA is generally detectable in NPS specimens during the acute phase of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA; clinical correlation with patient history and other diagnostic information is necessary to determine patient infection status. Positive results do not rule out co-infection with other pathogens.Results are meant to be used in conjunction with other clinical, epidemiologic, and laboratory data, in accordance with the guidelines provided by the relevant public health authorities. The BioFire COVID-19 Test 2 is intended for use by trained medical and laboratory professionals in a
	New Device	Predicate Device - K211079
Attribute	CepheidXpert® Xpress CoV-2 plus	BioFire Defense, LLCBioFire COVID-19 Test 2laboratory setting or under the supervisionof a trained laboratory professional.
Assay Targets	SARS-CoV-2(E, N2, RdRP)	SARS-CoV-2(Multiple Targets)
Specimen Type	• Nasopharyngeal swab (NPS)• Anterior nasal swab (NS)	Nasopharyngeal swab (NPS)
TransportMedia	• Universal Transport Medium (UTM) /Viral Transport Medium (VTM)• eNAT	• Transport Medium• Saline
Test Format	Same	Single Use
Automation	Same	Automated Nucleic Acid Extraction,Detection and Results Interpretation
Assay Results	Same	Qualitative
InternalControl	• Sample Processing Control (SPC)• Probe Check Control (PCC)	• Sample Processing Control• PCR and Melt Analysis Control
InstrumentSystems	Cepheid GeneXpert® Instrument Systems	BioFire® FilmArray® 2.0 orBioFire® FilmArray® Torch Systems
Time to Result	~30 minutes	~ 45 minutes

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Image /page/8/Picture/0 description: The image features the Cepheid logo, which includes a stylized blue wing-like design above the word "Cepheid" in a serif font. Below the logo, the text "Xpert® Xpress CoV-2 plus" is displayed, with "Xpert" followed by a registered trademark symbol. The text is in a smaller, sans-serif font, indicating a product or service related to COVID-19 testing.

Traditional 510(k) Submission

The following performance data (analytical and clinical) were provided in support of the substantial equivalence determination.

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5.4. Performance Studies

5.4.1. Analytical Performance

Analytical Sensitivity/Limit of Detection (LoD) - Clinical NPS Matrix

The analytical sensitivity of the Xpert Xpress CoV-2 plus test was first estimated by testing limiting dilutions of a NATtrol inactivated SARS-CoV-2 virus in pooled negative clinical NPS-UTM/VTM matrix, using two reagent lots and following the guidance in Clinical and Laboratory Standards Institute (CLSI) document EP17-A2. LoD was estimated by considering each target gene (E, N2, and RdRP) in addition to the overall positivity rate for the Xpert Xpress CoV-2 plus test. The estimated LoD value as determined by Probit regression analysis was based on the weakest target gene (N2) and verified using two lots of Xpert Xpress CoV-2 plus reagents in replicates of 20 per lot for two clinical NPS matrices (UTM/VTM and eNAT). The concentration levels with observed hit rates greater than or equal to 95% in the LoD determination study were 403, 200 and 70 copies/mL for the N2 target, RdRP target and E target, respectively. The claimed LoD is 403 copies/mL (Tables 5-2 and 5-3).

Table 5-2. Xpert Xpress CoV-2 plus Limit of Detection -- Verification of NATtrol SARS-CoV-2 LoD in NPS-UTM/VTM Matrix

Virus Strain	Reagent Lot	LoD(copies/mL)	PositiveResults outof # ValidReplicates	%Positive	MeanE Ct	MeanN2 Ct	MeanRdRP Ct
NATtrol™SARS-CoV-2	Reagent Lot 1	403	20/20	100%	34.3	38.3	36.6
	Reagent Lot 2	403	20/20	100%	34.1	37.7	36.5

Table 5-3. Xpert Xpress CoV-2 plus Limit of Detection - Verification of NATtrol SARS-CoV-2 LoD in NPS-eNAT Matrix

Virus Strain	Reagent Lot	LoD(copies/mL)	Positivesout of #ValidReplicates	%Positive	MeanE Ct	MeanN2 Ct	MeanRdRP Ct
NATtrol™	Reagent Lot 1	403	20/20ª	100%	33.4	36.8	35.5
SARS-CoV-2	Reagent Lot 2	403	20/20	100%	33.5	36.8	35.5

a. One of 20 replicates tested reported INVALID. The run was successfully repeated to obtain 20 valid replicates.

Analytical Sensitivity/Limit of Detection (LoD) – Clinical NS-UTM/VTM Matrix

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Image /page/10/Picture/0 description: The image contains the logo for Cepheid, a molecular diagnostics company. The logo features a stylized blue wing-like shape above the company name. Below the company name is the text "Xpert® Xpress CoV-2 plus", which refers to one of Cepheid's diagnostic tests.

NS-UTM/VTM matrix were determined using Probit regression analysis and the point estimated LoD values were 97, 462 and 157 copies/mL for the E target, N2 target and RdRP target, respectively. The estimated LoD value of the weakest target gene (N2) was verified using two lots of Xpert Xpress CoV-2 plus reagents in replicates of 20 per lot for clinical NS-UTM/VTM matrix. The claimed LoD is 462 copies/mL (Table 5-4).

Table 5-4. Xpert Xpress CoV-2 plus Limit of Detection - Verification of NATtrol SARS-CoV-2 LoD in NS-UTM/VTM Matrix

Virus Strain	Reagent Lot	LoD(copies/mL)	PositiveResults outof # ofValidReplicates	%Positive	MeanE Ct	MeanN2 Ct	MeanRdRP Ct
NATtrol™SARS-CoV-2	Reagent Lot 1	462	20/20	100%	34.4	38.9	36.8
	Reagent Lot 2	462	20/20	100%	34.1	37.6	36.1

Analytical Sensitivity (Limit of Detection) - WHO First International Standard SARS-CoV-2 RNA in clinical NS-UTM/VTM Matrix

The analytical sensitivity of the Xpert Xpress CoV-2 plus test was estimated by testing limiting dilutions of WHO First International Standard for SARS-CoV-2 RNA in pooled negative clinical NS-UTM/VTM matrix, using one reagent lot. The study was performed in a random and blinded fashion.

The lowest concentration level of the First WHO International Standard for SARS-CoV-2 RNA that generated >95% positive results (19/20 positive results) for the weakest target gene (N2) by the Xpert Xpress CoV-2 plus test was used to confirm be the analytical sensitivity or Limit of Detection (LoD). The verified LoD for the First WHO International Standard for SARS-CoV-2 RNA tested in NS-UTM/VTM matrix with the Xpert Xpress CoV-2 plus test was 1000 IU/mL. The positivity rates for the overall SARS-CoV-2 results and the analyte targets, including the mean Ct values for E, N2 and RdRP at the verified LoD are presented in Table 5-5.

Table 5-5. LoD Concentration, Positivity Rate and Mean Ct Values for SARS-CoV-2 for the First WHO International Standard for SARS-CoV-2 RNA in Clinical NS-UTM/VTM Matrix

Virus	LoDConcentration	SARS-CoV-2PositiveResults / TestReplicates	E Target		N2 Target		RdRP Target
1st WHOInternationalStandard forSARS-CoV-2RNA Virus	1000 IU/mL	20/20	20/20	34.3	19/20	39.5	20/20	36.5

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Analytical Inclusivity (Reactivity)

SARS-CoV-2 in silico Analyses

The inclusivity of Xpert Xpress CoV-2 plus primers was evaluated on June 30th, 2022 using in silico analysis of the assay amplicons in relation to 11,650,640 SARS-CoV-2 sequences available in the GISAID gene database for three targets, E, N2 and RdRP. The 11,650,640 SARS-CoV-2 sequences were separated into the lineages of interest based on the Pango Lineage assigned to each genome by GISAID, and those with ambiguous nucleotides were removed. Thus, the following inclusivity analyses focuses on the combined, non-ambiguous sequences from the variants of interest and variants of concern as of June 30th, 2022. These constituted 10.469.612 sequences for the E target. 10.587.381 sequences for the N2 target and 10,333,656 sequences for the RdRP target. Table 5-6 summarizes the effective predicted inclusivity for E, N2 and RdRP amplicons for the variants of interests and concern.

Table 5-6. Predicted Inclusivity for E, N2 and RdRP Amplicons for SARS-CoV-2 Variants of Interests and Concern

SARS-CoV-2Target Amplicon	Exact Match	1 Mismatch a	2 or MoreMismatches	PredictedInclusivity
E	10,420,248 of 10,469,612 total(99.5%)	48,562(0.5%)	802(0.01%)	100%
N2	10,386,068 of 10,587,381 total(98.1%)	196,336(1.9%)	4,977(0.05%)	99.95%
RdRP	10,247,146 of 10,333,656 total(99.2%)	85,373(0.8%)	1,137(0.01%)	100%

Single-nucleotide mismatches are predicted to not impact the performance of the test. a.

The in silico inclusivity of the Xpert Xpress CoV-2 plus probe oligonucleotides for E, N2 and RdRP were also assessed against the top 20 most frequent matches in the GISAID EpiCoV sequence database as of June 15th, 2022, which constituted 10,310, 839 for the E target, 10,428,014 for the N2 target and 10,178,602 for the RdRP target. For each of the probe oligonucleotides used in the Xpert Xpress CoV-2 plus test. Table 5-7 summarizes the number sequences as well as the corresponding percentage of sequences from this dataset with exact match, 1 mismatch/insertion, and 2 or more mismatches/insertions in the alignment.

Table 5-7. Predicted Inclusivity for E, N2 and RdRP Probes for SARS-CoV-2 Variants of Interests and Concern

SARS-CoV-2Target Probe	Exact Match	1 Mismatch/Insertion a	2 or MoreMismatches/Insertions	PredictedInclusivity
E	10,300,688 of 10,310,839 total(99.9%)	9,853(0.1%)	22(0.0002%)	100%
N2	10,351,581 of 10,428,014 total(99.3%)	72,957(0.7%)	0(0%)	100%
RdRP	0	10,140,254 of 10,178602 total(99.6%)	37,492(0.4%)	99.6%

Single-nucleotide mismatches/insertions are predicted to not impact the performance of the test. a.

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The in silico inclusivity of the Xpert Xpress CoV-2 plus primer/probe sequences for E, N2 and RdRP was further assessed against SAR-CoV-2 sequences submitted to the GISAID EpiCoV database up to 180 days prior to and including July 31, 2023 for SARS-CoV-2 isolates currently circulating in the United States. There were no instances where a specific primer/probe sequence was observed with greater than 0.5% frequency of mismatch during this 180-day period. The analysis predicted that the Xpert Xpress CoV-2 plus test will detect all currently circulating variants/lineages of SARS-CoV-2.

Moreover, based on the built-in redundancy of the Xpert Xpress CoV-2 plus test's SARS-CoV-2 amplification system (i.e., 3 independent targets, only 1 of 3 must be detected to assign a positive result), it is not anticipated that any of the evaluated sequences would be missed by the Xpert Xpress CoV-2 plus test.

SARS-CoV-2 Wet-Testing

In addition to the in silico analysis of the SARS-CoV-2 primers and probes for inclusivity, the inclusivity of the Xpert Xpress CoV-2 plus test was evaluated by bench testing against multiple strains of SARS-CoV-2 at levels near the analytical LoD (~3x LoD). A total of 61 strains comprised of 23 intact SARS-CoV-2 viral particles (18 inactivated viral cultures and 5 BSL-3 live viral cultures), and 38 SARS-CoV-2 in vitro RNA transcripts representing variant strains were tested in this study with the Xpert Xpress CoV-2 plus test. Three replicates were tested for each strain. All SARS-CoV-2 strains tested positive in all three replicates. Results are shown in Table 5-8.

CountNo.	SARS-CoV-2 Strain	TestedConcentration	SARS-CoV-2Test Results	Number of Positive ResultsObtained out of the Total Number ofValid Replicates
				E	N2	RdRP
1	2019-nCoV/Italy-INMI1 a	5 TCID50/mL	POS	3/3	3/3	3/3
2	England/204820464/2020 a	0.5 TCID50/mL	POS e	3/3	3/3	3/3
3	Hong Kong/VM20001061/2020 a	0.25 TCID50/mL	POS	3/3	3/3	3/3
4	South Africa/KRISP-K005325/2020 a	0.25 TCID50/mL	POS	3/3	3/3	3/3
5	USA/CA_CDC_5574/2020 a	0.25 TCID50/mL	POS	3/3	3/3	3/3
6	USA/MD-HP01542/2021 a	1.3e3 genomeequivalents/mL	POS	3/3	3/3	3/3
7	USA/GA-EHC-2811C/2021 a	1.3e3 genomeequivalents/mL	POS	3/3	3/3	3/3
8	NY-Wadsworth-103677-01/2020 a	0.15 TCID50/mL	POS	3/3	3/3	3/3
9	NY-Wadsworth-21006055-01/2021 a	0.04 TCID50/mL	POS	3/3	3/3	3/3
10	NY-Wadsworth-21025952-01/2021 a(Isolate 1)	0.625 TCID50/mL	POS	3/3	3/3	3/3
11	NY-Wadsworth-21018781-01/2021 a(Isolate 2)	0.625 TCID50/mL	POS	3/3	3/3	3/3
CountNo.	SARS-CoV-2 Strain	TestedConcentration	SARS-CoV-2Test Results		Number of Positive Results Obtained out of the Total Number of Valid Replicates
					E	N2	RdRP
12	NY-Wadsworth-21033899-01/2021 a	1.25 TCID50/mL	POS		3/3	3/3	3/3
13	NY-Wadsworth-33126-01/2020 a	0.15 TCID50/mL	POS		3/3	3/3	3/3
14	USA/CA-Stanford-15_S02/2021a	0.04 TCID50/mL	POS		3/3	3/3	3/3
15	USA/PHC658/2021a	25 TCID50/mL	POS		3/3	3/3	3/3
16	Japan/TY7-503/2021a	0.75 TCID50/mL	POS		3/3	3/3	3/3
17	hCoV-19/USA/MD-HP30386/2022 a	27.5 copies/mL	POS		3/3	3/3	3/3
18	hCoV-19/USA/COR-22-063113/2022 a	19 copies/mL	POS		3/3	3/3	3/3
19	Australia/VIC01/2020 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
20	Belgium/ULG/10004/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
21	England/205041766/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
22	England/MILK-9E05B3/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
23	England/SHEF-C05B2/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
24	France/HF2393/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
25	Hong Kong/HKU-211129-001/2021b	1.2e3 copies/mL	POS		3/3	3/3	3/3
26	Iceland/5/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
27	India/CT-ILSGS00361/2021 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
28	India/MH-NCCS-P1162000182735/2021b	1.2e3 copies/mL	POS		3/3	3/3	3/3
29	India/MH-SEQ-221_S66_R1_001/2021b	1.2e3 copies/mL	POS		3/3	3/3	3/3
30	Japan/Hu_DP_Kng_19-020/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
31	Japan/IC-0564/2021 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
32	Portugal/PT9543/2021b	1.2e3 copies/mL	POS		3/3	3/3	3/3
33	South Africa/KRISP-EC-K005299/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
34	Taiwan/NTU02/2020 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
35	USA/CA9/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
36	USA/CA-PC101P/2020 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
37	USA/CA-CZB-12943/2020 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
38	USA/MN2-MDH2/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
39	USA/NY-MSHSPSP-PV24650/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
40	USA/TX1/2020b	1.2e3 copies/mL	POS		3/3	3/3	3/3
41	USA/WA2/2020 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
42	USA/WA-CDC-UW21061750277/2021b	1.2e3 copies/mL	POS		3/3	3/3	3/3
43	Wuhan-Hu-1 b	1.2e3 copies/mL	POS		3/3	3/3	3/3
CountNo.	SARS-CoV-2 Strain	TestedConcentration	SARS-CoV-2Test Results	Number of Positive ResultsObtained out of the Total Number ofValid Replicates
				E	N2	RdRP
44	Germany/BavPat1/2020c	1.2e3 genomecopies/mL	POS	3/3	3/3	3/3
45	Singapore/2/2020c	300 genomeequivalents/mL	POS	3/3	3/3	3/3
46	USA/AZ1/2020c	300 genomeequivalents/mL	POS	3/3	3/3	3/3
47	USA/CA1/2020c	1.2e3 genomeequivalents/mL	POS	3/3	3/3	3/3
48	USA/CA2/2020c	1.2e3 genomeequivalents/mL	POS	3/3	3/3	3/3
49	USA/CA3/2020c	400 genomeequivalents/mL	POS	3/3	3/3	3/3
50	USA/CA4/2020c	1.2e3 genomeequivalents/mL	POS	3/3	3/3	3/3
51	USA/IL1/2020c	150 genomeequivalents/mL	POS	3/3	3/3	3/3
52	USA/New York-PV08001/2020c	1.2e3 genomecopies/mL	POS	3/3	3/3	3/3
53	USA/New York-PV08410/2020c	300 genomeequivalents/mL	POS	3/3	3/3	3/3
54	USA/New York-PV08449/2020c	300 genomeequivalents/mL	POS	3/3	3/3	3/3
55	USA/New York-PV09158/2020c	300 genomeequivalents/mL	POS	3/3	3/3	3/3
56	USA/WI1/2020c	1.2e3 genomeequivalents/mL	POS	3/3	3/3	3/3
57	hCoV-19/USA/MD-HP38861/2022 d	0.29 TCID50/mL	POS	3/3	3/3	3/3
58	hCoV-19/USA/MD-HP38960/2022 d	0.043 TCID50/mL	POS	3/3	3/3	3/3
59	hCoV-19/USA/CO-CDPHE-2102544747/2021 d	0.028 TCID50/mL	POS	3/3	3/3	3/3
60	hCoV-19/USA/MD-HP38288/2022 d	0.24 TCID50/mL	POS	3/3	3/3	3/3
61	hCoV-19/USA/MD-HP40900/2022 d	0.72 TCID50/mL	POS	3/3	3/3	3/3

Table 5-8. Analytical Reactivity (Inclusivity) of the Xpert Xpress CoV-2 plus Test

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Traditional 510(k) Submission

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Traditional 510(k) Submission

Inactivated viral culture fluid (intact viral particles) a.

Synthetic, in vitro RNA transcript, TWIST controls b.

Genomic RNA C.

BSL-3 live viral culture fluid (intact viral particles)
One of 3 replicates reported ERROR. The run was successfully repeated to obtain 3 valid replicates. e.

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Analytical Exclusivity (Specificity)

In Silico Analyses

The analytical specificity/cross-reactivity of the Xpert Xpress CoV-2 plus plan included evaluation of the SARS-CoV-2 test primer and probes with potentially cross-reactive microorganisms by in silico analysis was conducted by mapping the primers and probes of Xpert Xpress CoV-2 plus individually to the microorganism sequences downloaded from the NCBI database. E primers and probes are not specific for SARS-COV-2 and will detect Human and Bat SARS-coronavirus. No potential unintended cross reactivity with other organisms listed in Table 5-9 is expected based on the in silico analysis.

Microorganisms from the Same GeneticFamily	High Priority Organisms
Human coronavirus 229E	Adenovirus (e.g., C1 Ad. 71)
Human coronavirus OC43	Cytomegalovirus
Human coronavirus HKU1	Enterovirus (e.g., EV68)
Human coronavirus NL63	Epstein-Barr virus
SARS-coronavirus	Human Metapneumovirus (hMPV)
MERS-coronavirus	Influenza A & B
Bat coronavirus	Measles
	Mumps
	Parainfluenza virus 1-4
	Parechovirus
	Respiratory syncytial virus
	Rhinovirus
	Bacillus anthracis (Anthrax)
	Bordetella pertussis
	Bordetella parapertussis
	Chlamydia pneumoniae
	Chlamydia psittaci
	Corynebacterium diphtheriae
	Coxiella burnetii (Q-Fever)
	Escherichia coli
	Fusobacterium necrophorum
	Haemophilus influenzae
	Lactobacillus sp.
	Legionella non-pneumophila
	Legionella pneumophila
	Leptospira
	Moraxella catarrhalis
	Mycobacterium tuberculosis
	Mycoplasma genitalium
	Mycoplasma pneumoniae
	Neisseria elongata
	Neisseria meningitidis

Table 5-9. Microorganisms Analyzed in the in silico Analysis for the SARS-CoV-2 Target

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Microorganisms from the Same GeneticFamily	High Priority Organisms
	Pneumocystis jirovecii (PJP)
	Pseudomonas aeruginosa
	Staphylococcus aureus
	Staphylococcus epidermidis
	Staphylococcus salivarius
	Streptococcus pneumoniae
	Streptococcus pyogenes
	Aspergillus sp
	Candida albicans

Wet-Testing

In addition to the in silico analysis of the SARS-CoV-2 primers and probes for crossreactivity, the analytical specificity of the Xpert Xpress CoV-2 plus test was evaluated by bench-testing a panel of 62 microorganisms comprising (4) human coronaviruses, (1) MERScoronavirus, (1) SARS-coronavirus, (20) other respiratory viruses, (30) respiratory bacteria, (2) yeast strains, (3) fungal strain, and 1 human nasal wash fluid representing a diverse microbial flora in the human respiratory tract.

The intact viruses were tested at concentrations of ≥10° TCID50/mL, ≥10° CEID50/mL, and >105 copies/mL. Bacteria and yeast were tested at ≥10° CFU/mL. The bacteria Chlamydia pneumoniae and Mycoplasma pneumoniae were tested at concentrations of ≥10° IFU/mL and >106 CCU/mL. respectively. The live strains of Coxiella burnetii. Lactobacillus reuteri, and Neisseria meningitides were not available, therefore genomic DNA at ≥ 10° genome equivalent copies/mL was used. The in vitro transcribed (IVT) RNA sample was tested at > 10° genome equivalent/mL for human Coronavirus HKU1, and the genomic RNA sample was tested at > 10° genome equivalent/mL for SARS-coronavirus Urbani.

The microorganisms being evaluated for cross-reactivity were tested in groups or individually and spiked into negative simulated NPS/NS background matrix for testing. If a grouped microorganisms produced a SARS-CoV-2 positive result, then each member of the group was tested separately. If the individual non-target organism yielded a positive result, retesting was performed at lower concentrations until a concentration that no longer produce a false positive result was identified.

The results of the analytical specificity/exclusivity study demonstrate that the primer/probe sets included in the Xpert Xpress CoV-2 plus does not cross-react with the nucleic acids from non-intended respiratory microorganisms and phylogenetically related human coronavirus species. The one exception was the SARS-coronavirus, Urbani which yielded the expected test result of SARS-CoV-2 POSITIVE. We expected cross-reactivity for the E gene target with the SARS-coronavirus Urbani strain which has the highly conserved E gene target shared among coronaviruses in the B lineage Betacoronavirus. Results are shown in Table 5-10.

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Traditional 510(k) Submission

Table 5-10. Analytical Specificity (Exclusivity) of the Xpert Xpress CoV-2 plus Test
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Respiratory Microorganisms	TestGroup	Tested Concentration	FinalSARS-CoV-2Result Call- Number of Positive ResultsObtained from Total Numberof Replicates Tested
			E	N2	RdRP
Human coronavirus, 229EHuman coronavirus, OC43MERS-coronavirus	G1	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
Human coronavirus, NL63	G2	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
Human coronavirus, HKU1a	G3	1.1e6 genome cp/mL	NEG	0/3	0/3	0/3
SARS-coronavirus, Urbani a	G4	1.1e6 genome cp/mL	POS	3/3	0/3	0/3
Influenza A H1N1 (pdm2009),Michigan/272/2017		1.1e5 TCID50/mL
Influenza B (Victoria Lineage),Hawaii/01/2018 (NA D197N)	G5	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
RSV-A, Strain: 4/2015 Isolate #1		1.1e5 TCID50/mL
Adenovirus Type 1		1.1e5 TCID50/mL
Adenovirus Type 7ACytomegalovirusEchovirus	G6	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
Enterovirus, D68 strain US/KY/14-18953Epstein Barr Virus (Human Herpes Virus 4[Hhv-4])		1.1e5 TCID50/mL1.1e5 cp/mL
Herpes Simplex Virus (HSV) type 1Human metapneumovirus(hMPV-5, type B1)	G7	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
MeaslesMumps virusHuman parainfluenza Type 1Human parainfluenza Type 2		1.1e5 TCID50/mL
Human parainfluenza Type 3Human parainfluenza Type 4Rhinovirus, Type 1A b	G8	1.1e5 TCID50/mL4.5e4 TCID50/mL	NEG	0/3	0/3	0/3
Acinetobacter baumanniiBurkholderia cepaciaCandida albicansCandida parapsilosisBordetella pertussisChlamydia pneumoniaeCitrobacter freundiiCorynebacterium xerosis	G9	1.1e6 CFU/mL1.1e6 IFU/mL	NEG	0/3	0/3	0/3
Escherichia coliEnterococcus faecalisHemophilus influenzaeLegionella spp dMoraxella catarrhalisMycobacterium tuberculosis (avirulent)Mycoplasma pneumoniaeNeisseria mucosa	G10	1.1e6 CFU/mL1.1e6 CCU/mL	NEG	0/3	0/3	0/3
Propionibacterium acnes (= Cutibacteriumacnes) Z144	G11	1.1e6 CFU/mL	NEG	0/3	0/3	0/3
Pseudomonas aeruginosa, Z139Staphylococcus aureusStaphylococcus epidermidisStaphylococcus haemolyticus	G12	1.1e6 CFU/mL	NEG	0/3	0/3	0/3
Respiratory Microorganisms	TestGroup	Tested Concentration	FinalSARS-CoV-2Result Call-Out	Number of Positive ResultsObtained from Total Numberof Replicates Tested
				E	N2	RdRP
Streptococcus agalactiae		1.1e6 CFU/mL
Streptococcus pneumoniae		1.1e6 CFU/mL
Streptococcus pyogenes		1.1e6 CFU/mL
Streptococcus salivarius		1.1e6 CFU/mL
Streptococcus sanguinis		1.1e6 CFU/mL
Pneumocystis jirovecii (PJP)		1.1e6 CFU/mL
Lactobacillus reuteri, F275c	G13	1.1e6 genome cp/mL	NEG	0/3	0/3	0/3
Neisseria meningitides c	G13	1.1e6 genome cp/mL	NEG	0/3	0/3	0/3
Pooled human nasal wash	G14	N/A	NEG	0/3	0/3	0/3
Influenza C (Taylor/1233/1947)	G15	1.1e5 CEID50/mL	NEG	0/3	0/3	0/3
Rhinovirus, Type 1Ab	G16	1.1e5 TCID50/mL	NEG	0/3	0/3	0/3
Aspergillus flavus	G17	1.35e7 CFU/mL	NEG	0/3	0/3	0/3
Aspergillus fumigatus	G18	3.67e6 CFU/mL	NEG	0/3	0/3	0/3
Coxiella burnetii c	G19	2.5e6 genome cp/mL	NEG	0/3	0/3	0/3
Leptospira broomii	G20	2.9e6 CFU/mL	NEG	0/3	0/3	0/3
Parechovirus type 1	G21	3.39e5 TCID50/mL	NEG	0/3	0/3	0/3
Fusobacterium necrophorum	G22	6.52e6 CFU/mL	NEG	0/3	0/3	0/3
Mycoplasma genitalium c	G23	3.6e6 genome cp/mL	NEG	0/3	0/3	0/3

510(k) Summary

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Image /page/18/Picture/0 description: The image shows the logo for Cepheid, a molecular diagnostics company. Below the logo is the text "Xpert® Xpress CoV-2 plus". The logo consists of the company name in a stylized font, with a blue graphic above it.

Traditional 510(k) Submission

Microorganism in the form of genomic RNA were tested in Tris-EDTA+ ([NH]]>O2) buffer using an assay definition file (ADF) a. without sample preparation.

Rhinovirus, Type 1A was initially tested at 4.5 e4 TCIDsomL in test group G8 using virus lot 325725. It was re-tested b. individually (G16) at a higher concentration of 1.1e5 TCID50/mL using virus lot 326527.

c. Microorganisms in the form of genomic DNA were tested in simulated NPS/NS background matrix using the ADF with full sample preparation.

d. Legionella pneumophila was tested in this study.

Microbial Interference

A microbial interference study was performed to assess the inhibitory effects of commensal microorganisms potentially encountered in upper respiratory tract specimens on the performance of the Xpert Xpress CoV-2 plus test. A panel of 18 commensal microorganisms, consisting of 15 viral strains and 3 bacterial strains was tested. Contrived samples consisted of SARS-CoV-2 virus seeded at 1.15x-3x LoD into simulated nasopharyngeal swab (NPS)/ anterior nasal swab (NS) matrix in the presence of fifteen (15) commensal virus strains and three (3) commensal bacterial strains (Haemophilus influenzae, Staphylococcus aureus, Staphylococcus epidermidis) spiked at their respective concentrations listed in Table 5-11.

Replicates of 8 positive samples were tested with SARS-CoV-2 virus and each potential microbial interference strain combination. All 8 of 8 positive replicate samples were correctly identified as SARS-CoV-2 POSITIVE using the Xpert Xpress CoV-2 plus test. No interference by the listed above commensal viral or bacterial strains was reported at the concentrations tested.

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Commensal Strain	ConcentrationTested	Number of CorrectTest Results/Number Tested
Adenovirus Type 1C a	1x105 TCID50/mL	8/8
Adenovirus Type 2C b	1x105 TCID50/mL	8/8 c
Adenovirus Type 3Bb	1x105 TCID50/mL	8/8
Human Coronavirus-OC43 a	1x105 copies/mL	8/8
Human Coronavirus-229E b	1x105 TCID50/mL	8/8
Human Coronavirus-NL63 b	1x105 TCID50/mL	8/8
Human Coronavirus-HKU1b	1x105 copies/mL	8/8
Metapneumovirus 5, Type B1 a	1x105 TCID50/mL	8/8
Parainfluenza Type 1 a	1x105 TCID50/mL	8/8
Parainfluenza Type 2 a	1x105 TCID50/mL	8/8
Parainfluenza Type 3 a	1x105 TCID50/mL	8/8
Rhinovirus Type 1A a	1x105 PFU/mL	8/8
Influenza A H1N1 b	1x105 CEID50/mL	8/8
Influenza Bb	1x105 TCID50/mL	8/8 d
Respiratory Syncytial Virus A b	1x105 TCID50/mL	8/8
Haemophilus influenzae a	1x107 CFU/mL	8/8
Staphylococcus aureus a	1x107 CFU/mL	8/8
Staphylococcus epidermidis a	1x107 CFU/mL	8/8

Table 5-11: Microbial Interference Study Results

a. These commensal strains were tested with SARS-CoV-2 at a concentration of 3x LoD.

b. These commensal strains were tested with SARS-CoV-2 at a concentration of 1.15x LoD.

c. Two of 8 replicates reported as ERROR. The runs were successfully repeated to obtain 8 valid replicates.

d. One of 8 replicates reported as ERROR. The run was successfully repeated to obtain 8 valid replicates.

Potentially Interfering Substances

Substances that could be present in the nasopharynx (or introduced during specimen collection and handling) and potentially interfere with accurate detection of SARS-CoV-2 were evaluated with direct testing on the Xpert Xpress CoV-2 plus test. Potentially interfering substances in the nasal passage and nasopharynx may include, but are not limited to: blood, nasal secretions or mucus, and nasal and throat medications used to relieve congestion, nasal dryness, irritation, or asthma and allergy symptoms, as well as antibiotics and antivirals. Positive and negative samples were prepared in simulated nasopharyngeal swab (NPS)/ anterior nasal swab (NS) matrix. Negative samples (N = 8) were tested in the presence of each substance to determine the effect on the performance of the sample processing control (SPC). Positive samples (N = 8) were tested per substance with SARS-CoV-2 virus spiked at 3x LoD. The controls were samples with and without SARS-CoV-2 virus spiked at 3x LoD into simulated NPS/ NS matrix containing no potentially interfering substance.

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Image /page/20/Picture/1 description: The image contains the logo for Cepheid, a molecular diagnostics company. The logo features a stylized blue graphic above the company name, "Cepheid." Below the company name is the text "Xpert® Xpress CoV-2 plus", which likely refers to one of Cepheid's diagnostic tests. The text is in a smaller font size than the company name.

The 23 potentially substances, with active ingredients, that were evaluated are listed in Table 5-12. For substances that resulted in an INVALID test result, the concentration of the substance was reduced by half and re-tested. Interfering effects were observed for fluticasone propionate and mucin type I-S.

Substance ID	Substance/Class	Substance/Active Ingredient	Concentrations Tested
No substance	Control	Simulated NPS/NS Matrix	100% (v/v)
Afrin	Nasal Spray	Oxymetazoline (0.05%)	15% (v/v)
Albuterol Sulfate	Beta-adrenergicbronchodilator	Albuterol Sulfate (5mg/mL)	0.83 mg/mL (equivalentto 1 dose per day)
BD UniversalTransport Medium	Transport Media	BD Universal TransportMedium	100% (v/v)
Blood	Blood	Blood (Human)	2% (v/v)
Copan Swab M	Transport Media	Copan Swab M	100% (v/v)
FluMist	FluMist®	Live intranasal influenza virusvaccine	6.7% (v/v)
Fluticasone PropionateNasal Spray	Nasal corticosteroid	Fluticasone Propionate	5 µg/mL, 2.5 µg/mL
Ibuprofen	Analgesic (nonsteroidalanti-inflammatory drugs(NSAID))	Ibuprofen	21.9 mg/dL
Leukocytes	Leukocytes	Leukocytes (Human)	1 x 106 cells/mL
Menthol	Throat lozenges, oralanesthetic, and analgesic	Benzocaine, Menthol	1.7 mg/mL
Mucin type II	Mucin	Purified Mucin protein(Porcine submaxillary gland,type II)	0.1% (w/v)
Mucin type I-S	Mucin	Purified Mucin protein (Bovinesubmaxillary gland, type I-S)	2.5 mg/mL, 1.25 mg/mL
Mupirocin	Antibiotic, nasal ointment	Mupirocin (2%)	10 mg/mL
Human peripheralblood mononuclearcells (PBMC)	Human peripheral bloodmononuclear cells(PBMC)	Human peripheral bloodmononuclear cells (PBMC)	1x103 cells/µL
PHNY	Nasal Drops	Phenylephrine (1%)	15% (v/v)
Remel M4RT	Transport Media	Remel M4RT	100% (v/v)
Remel M5	Transport Media	Remel M5	100% (v/v)
Saline	Saline Nasal Spray	Sodium Chloride (0.65%)	15% (v/v)
Snuff	Tobacco	Nicotine	1% (w/v)
Tamiflu	Anti-viral drugs	Zanamivir	7.5 mg/mL
Tobramycin	Antibacterial, systemic	Tobramycin	4 µg/mL
Zicam	Nasal Gel	Luffa opperculata, Galphimiaglauca, Histaminumhydrochloricum Sulfur (0.05%)	15% (w/v)
Zinc	Zinc supplement	Zinc Gluconate	0.1 µg/mL

Table 5-12. Potentially Interfering Substances Tested

The results for the negative and positive samples are presented in Table 5-13.

The results from the SARS-CoV-2 negative samples showed that in the presence of fluticasone propionate nasal spray at 5 µg/mL and mucin type I-S at 2.5 mg/mL, 7 of 8 replicates correctly reported "SARS-CoV-2 NEGATIVE" test results, and 1 of 8 replicates reported "INVALID" test result for each substance. When the concentrations of fluticasone

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propionate nasal spray and mucin type I-S were reduced by half, 8/8 replicates provided valid test results and correctly reported "SARS-CoV-2 NEGATIVE" for each substance. No interference was observed at these lower concentrations.

The results from the SARS-CoV-2 positive samples showed that in the presence of fluticasone propionate nasal spray at 5 µg/mL, 7 of 8 replicates correctly reported "SARS-CoV-2 POSITIVE" test results and 1 of 8 replicate reported "INVALID" test result. When the concentration of the fluticasone propionate nasal spray was reduced to 2.5 µg/mL, 8/8 replicates provided valid test results and correctly reported "SARS-CoV-2 POSITIVE". No interference was observed at this lower concentration.

Table 5-13. SARS-CoV-2 Negative Samples Tested in the Presence of Potentially Interfering Substances

Substance	Concentration Tested	Number SARS-CoV-2Negative / Number of ValidReplicates Tested	Number SARS-CoV-2Positive / Number of ValidReplicates Tested
SARS-CoV-2 Negative(No Substance) Control	100% (v/v)	8/8	Not Applicable
SARS-CoV-2 Positive(No Substance) Control	100% (v/v)	Not Applicable	8/8
Afrin	15% (v/v)	8/8	8/8
Albuterol Sulfate	0.83 mg/mL	8/8	8/8
BD Universal TransportMedium	100 (v/v)	8/8	8/8
Blood	2% (v/v)	8/8	8/8
Copan Swab M	100 (v/v)	8/8	8/8
FluMist®	6.7% (v/v)	8/8	8/8
Fluticasone Propionate NasalSpray	5 µg/mL	7/8a	7/8a
Ibuprofen	21.9 mg/dL	8/8	8/8
Leukocytes	1 x 106 cells/mL	8/8	8/8
Menthol	1.7 mg/mL	8/8	8/8
Mucin (Type II)	0.1% (w/v)	8/8	8/8
Mucin (Type I-S)	1.25 mg/mL	7/8a	Not Applicable
Mupirocin	10 mg/mL	8/8	8/8
Human Peripheral BloodMononuclear Cells (PBMC)	1 x 103 cells/µL	8/8	8/8
PHNY	15% (v/v)	8/8	8/8
Remel M4RT	100% (v/v)	8/8	8/8
Remel M5	100% (v/v)	8/8	8/8
Saline	15% (v/v)	8/8	8/8
Snuff	1% (w/v)	8/8	8/8
Tamiflu	7.5 mg/mL	8/8	8/8
Tobramycin	4 µg/mL	8/8	8/8
Zicam	15% (w/v)	8/8	8/8
Zinc	0.1 µg/mL	8/8	8/8

a. One of 8 replicates reported INVALID test result, indicating interference from the substance was subsequently tested with 8 replicates at half the initial concentration.

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Image /page/22/Picture/0 description: The image shows the Cepheid logo, which includes a stylized blue graphic above the word "Cepheid." The text below the logo reads "Xpert® Xpress CoV-2 plus". The "Xpert" has a registered trademark symbol next to it.

Carryover Contamination

A study was conducted to assess whether the single-use, self-contained Xpert Xpress CoV-2 plus cartridge prevents specimen and amplicon carryover by testing a negative sample immediately after testing of a very high positive sample in the same GeneXpert module. The negative sample used in this study consisted of simulated NPS/NS matrix and the positive sample consisted of high SARS-CoV-2 virus concentration (inactivated SARS-CoV-2 USA-WA1/2020 at 5e4 copies/mL) spiked into negative NPS/NS matrix. The negative sample was tested in a GeneXpert module at the start of the study. Following the initial testing of the negative sample, the high positive sample was processed in the same GeneXpert module immediately followed by another negative sample. This was repeated 20 times in the same module, resulting in 20 positives and 21 negatives for the module. The study was repeated using a second GeneXpert module for a total of 40 positive and 42 negative samples. All 40 positive samples were correctly reported as SARS-CoV-2 POSITIVE and all 42 negative samples were correctly reported as SARS-CoV-2 NEGATIVE with the Xpert Xpress CoV-2 plus test. No specimen or amplicon carry-over contamination was observed in this study.

Reproducibility

The reproducibility of the Xpert Xpress CoV-2 plus test was established at 3 sites (2 external and 1 internal) using a 3-member panel including one negative sample, one low positive (~1.5x LoD) sample and one moderate positive (~3x LoD) sample. The negative sample consisted of simulated NS/NPS matrix without target microorganism or target RNA. The positive samples were contrived samples in a simulated matrix using inactivated NATtrol SARS-CoV-2 (ZeptoMetrix).

Testing was conducted over 6 days, using 3 lots of Xpert Xpress CoV-2 plus cartridges at 3 participating sites each with 2 operators to yield a total of 144 observations per panel member (3 Sites x 2 Operators x 3 Lots x 2 Days/Lot x 2 Runs x 2 Replicates = 144 observations/panel member).

The percent agreement of the qualitative results for SARS-CoV-2 detection for each panel member analyzed by each of the 6 operators and each site is shown in Table 5-14. In addition, the overall percent agreement for each sample (% total agreement) and the twosided Wilson Score confidence intervals (CI) are presented in the last column.

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Traditional 510(k) Submission

PanelMember	Site 1			Site 2			Site 3			% Total Agreementand 95% CI by PanelMember
	Op1	Op2	Site	Op1	Op2	Site	Op1	Op2	Site
Negative	100%(24/24)	95.8%(23/24)	97.9%(47/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(23/23)a	100%(47/47)	99.3% (142/143)[96.1% - 99.9%]
SARS-CoV-2Low Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100% (144/144)[97.4% - 100%]
SARS-CoV-2Mod Pos	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100%(24/24)	100%(24/24)	100%(48/48)	100% (144/144)[97.4% - 100%]

Table 5-14. Summary of the Reproducibility Results - % Agreement

The evaluation of reproducibility and within-laboratory precision of the underlying Ct values for E, N2 and RdRP obtained in the Xpert Xpress CoV-2 plus test was analyzed. The mean, standard deviation (SD), and coefficient of variation (CV) between-sites, between-operators, between-lots, between-days, between-runs and within-run for each panel member are presented in Table 5-15.

Sample	Analyte	N	Mean Ct	Between Sites		Between Operators		Between Lots		Between Days		Between Runs		Within Run		Total
				SD	CV (%)	SD	CV (%)	SD	CV (%)	SD	CV (%)	SD	CV (%)	SD	CV (%)	SD	CV (%)
Negative	SPC	143	31.1	0.29	0.9	0.16	0.5	0.42	1.3	0	0	0	0	1.47	4.7	1.56	5.0
SARS-CoV-2 Low Pos	E	144	34.0	0.06	0.2	0.19	0.6	0	0	0.14	0.4	0	0	0.55	1.6	0.60	1.8
	N2	144	37.5	0.12	0.3	0	0	0.2	0.5	0.11	0.3	0	0	0.68	1.8	0.73	1.9
	RdRP	144	36.3	0.07	0.2	0	0	0.25	0.7	0.06	0.2	0	0	0.61	1.7	0.66	1.8
SARS-CoV-2 Mod Pos	E	144	32.5	0.31	0.9	0.03	0.1	0	0	0.47	1.4	0	0	1.86	5.7	1.95	6.0
	N2	144	36.2	0.16	0.4	0	0	0.29	0.8	0.17	0.5	0	0	0.65	1.8	0.75	2.1
	RdRP	144	34.8	0.33	1.0	0	0	0	0	0.62	1.8	0	0	2.18	6.2	2.29	6.6

Table 5-15. Summary of Reproducibility Results – Nested ANOVA by Coefficient of Variation

510(k) Summary

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Single-Site Precision

The precision of the Xpert Xpress CoV-2 plus test was established at a single site using a 3member panel including one negative sample, one low positive (~1.5x LoD) sample and one moderate positive (~3x LoD) sample. The negative sample consisted of simulated NS/NPS matrix without target microorganism or target RNA. The positive samples were contrived samples in a simulated NS/NPS matrix using inactivated NATtrol SARS-CoV-2 (ZeptoMetrix).

Testing was conducted over 20 days, using one lot of Xpert Xpress CoV-2 plus cartridges at a single site and with 1 operator to yield a total of 80 observations per panel member (1 Site x 1 Operator x 1 Lot x 20 Days x 2 Runs x 2 Replicates = 80 observations/panel member). The results from the study are summarized in Table 5-16.

Panel Member	Agreement	% Total Agreement and 95% CI by Panel Member
Negative	80/80	100%(95.4%-100.0%)
SARS-CoV-2 Low Positive	80/80	100%(95.4%-100.0%)
SARS-CoV-2 Mod Positive	80/80	100%(95.4%-100.0%)

Table 5-16. Summary of the Single-Site Precision Results - % Agreement

5.4.2. Clinical Performance

The clinical performance of the Xpert Xpress CoV-2 plus test was evaluated in a multi-site, observational and method comparison study that included 32 geographically diverse sites in the United States (US). Of the 32 sites, 5 sites participated in specimen collection only, 26 sites performed Xpert testing and specimen collection, and 1 site performed Xpert testing as well as comparator and discrepant testing.

The performance of the Xpert Xpress CoV-2 plus test was evaluated using prospectively collected fresh (98.6%) and frozen (1.4%) clinical nasopharyngeal swab (NPS) and anterior nasal swab (NS) specimens in viral transport medium or universal transport medium. These specimens were collected in 2022 from individuals with signs and symptoms of respiratory tract infection and tested using Xpert Xpress CoV-2 plus side by side with a U.S. FDAcleared molecular respiratory panel that includes SARS-CoV-2, in a randomized and blinded fashion. A total of 4047 specimens (2029 NPS and 2018 NS) were evaluated from individuals with signs and symptoms of respiratory infection. Of these, 60 specimens vielded non-determinate results (INVALID, ERROR, and NO RESULT) with Xpert Xpress CoV-2 plus. Additionally, 237 specimens either yielded non-determinate result, or were not tested per comparator package insert. Therefore, 297 (60 + 237) specimens were excluded, and a total of 3750 (1879 NPS and 1871 NS) specimens that yielded valid results by both Xpert Xpress CoV-2 plus and the U.S. FDA-cleared molecular respiratory panel were included in

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the clinical performance evaluation. Available demographic data collected from study participants are presented in Table 5-17.

	NPS(N=1879)	NS(N=1871)	Overall(N=3750)
Gender
Female	1107 (58.9%)	1171 (62.6%)	2278 (60.7%)
Male	772 (41.1%)	700 (37.4%)	1472 (39.3%)
Age Group (Years)
<5	8 (0.4%)	64 (3.4%)	72 (1.9%)
6-21	389 (20.7%)	380 (20.3%)	769 (20.5%)
22-59	1228 (65.4%)	1167 (62.4%)	2395 (63.9%)
≥60	254 (13.5%)	260 (13.9%)	514 (13.7%)
Race
American Indian or Alaska Native	4 (0.2%)	4 (0.2%)	8 (0.2%)
Asian	44 (2.3%)	44 (2.4%)	88 (2.3%)
Black or African American	525 (27.9%)	524 (28.0%)	1049 (28.0%)
White	1236 (65.8%)	1222 (65.3%)	2458 (65.5%)
Native Hawaiian or Other PacificIslander	4 (0.2%)	0 (0%)	4 (0.1%)
Black or African American, White	7 (0.4%)	8 (0.4%)	15 (0.4%)
Other Mixed (each N≤5)	7 (0.4%)	2 (0.1%)	9 (0.2%)
Participant declined to answer, orunknown	52 (2.8%)	67 (3.6%)	119 (3.2%)
Ethnicity
Hispanic	170 (9.0%)	165 (8.8%)	335 (8.9%)
Non-Hispanic	1681 (89.5%)	1668 (89.2%)	3349 (89.3%)
Participant declined to answer, orunknown	28 (1.5%)	38 (2.0%)	66 (1.8%)
Specimen Testing
Fresh	1852 (98.6%)	1845 (98.6%)	3697 (98.6%)
Frozen	27 (1.4%)	26 (1.4%)	53 (1.4%)
Testing Environment
Laboratory/NPT	996 (53.0%)	971 (51.9%)	1967 (52.5%)
CW	883 (47.0%)	900 (48.1%)	1783 (47.5%)
Vaccine Status
Vaccinated	1336 (71.1%)	1344 (71.8%)	2680 (71.5%)
Not Vaccinated	523 (27.8%)	508 (27.2%)	1031 (27.5%)
Unknown	20 (1.1%)	19 (1.0%)	39 (1.0%)

Table 5-17. Demographic Data Summary of Participants Symptomatic of Respiratory Infection

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Image /page/26/Picture/0 description: The image shows the logo for Cepheid, a molecular diagnostics company. Below the logo is the text "Xpert® Xpress CoV-2 plus". This text likely refers to a rapid diagnostic test for COVID-19 developed by Cepheid. The logo and text are displayed in a clear and legible font.

Positive Percent Agreement (PPA) and Negative Percent (NPA) were determined by comparing the results of the Xpert Xpress CoV-2 plus test relative to the results of a U.S. FDA-cleared molecular respiratory panel for the SARS-CoV-2 target. Based on the specimens that yielded non-determinate results (INVALID, ERROR, and NO RESULT) with Xpert Xpress CoV-2 plus, the non-determinate rate was 1.6% (32/2029) with NPS and 1.4% (28/2018) with NS specimens.

A total of 3750 (1879 NPS and 1871 NS) specimens that yielded valid results by both Xpert Xpress CoV-2 plus and the U.S. FDA-cleared molecular respiratory panel were included in the clinical performance evaluation for SARS-CoV-2. The Xpert Xpress CoV-2 plus demonstrated an overall PPA and NPA of 98.1% and 98.3% for SARS-CoV-2, respectively (Table 5-18).

		Comparator Result
		Overall			NPS			NS
		Positive	Negative	Total	Positive	Negative	Total	Positive	Negative	Total
Xpert XpressCoV-2 plus	Positive	574	55	629	292	28b	320	282	27d	309
	Negative	11	3110	3121	9a	1550	1559	2c	1560	1562
	Total	585	3165	3750	301	1578	1879	284	1587	1871
PPA		98.1%(95% CI: 96.7% - 98.9%)			97.0%(95% CI: 94.4% - 98.4%)			99.3%(95% CI: 97.5% - 99.8%)
NPA		98.3%(95% CI: 97.7% - 98.7%)			98.2%(95% CI: 97.4% - 98.8%)			98.3%(95% CI: 97.5% - 98.8%)
NPS: nasopharyngeal swab specimen; NS: anterior nasal swab specimen; PPA: Positive Percent Agreement; NPA: Negative PercentAgreement; CI: 95% two-sided Confidence Interval
a Discrepant test results based on a U.S. FDA EUA SARS-CoV-2 molecular test: 7/9 SARS-CoV-2 negative; 1/7 SARS-CoV-2 positive; 1/9 invalid.
b Discrepant test results based on a U.S. FDA EUA SARS-CoV-2 molecular test: 8/28 SARS-CoV-2 positive; 19/28 SARS-CoV-2 negative; 1/28 invalid.
c Discrepant test results based on a U.S. FDA EUA SARS-CoV-2 molecular test: 2/2 SARS-CoV-2 negative.
d Discrepant test results based on a U.S. FDA EUA SARS-CoV-2 molecular test: 8/27 SARS-CoV-2 positive; 18/27 SARS-CoV-2

Table 5-18. Xpert Xpress CoV-2 plus Performance Results in Symptomatic Individuals

5.5. Conclusions

negative; 1/27 invalid.

The results of the non-clinical and clinical performance studies summarized above demonstrate that the Xpert Xpress CoV-2 plus test is substantially equivalent to the predicate device.

Regulation Number and Section

§ 866.3981 Device to detect and identify nucleic acid targets in respiratory specimens from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-target test.

(a)
Identification. A device to detect and identify nucleic acid targets in respiratory specimens from microbial agents that cause the SARS-CoV-2 respiratory infection and other microbial agents when in a multi-target test is an in vitro diagnostic device intended for the detection and identification of SARS-CoV-2 and other microbial agents when in a multi-target test in human clinical respiratory specimens from patients suspected of respiratory infection who are at risk for exposure or who may have been exposed to these agents. The device is intended to aid in the diagnosis of respiratory infection in conjunction with other clinical, epidemiologic, and laboratory data or other risk factors.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use in the labeling required under § 809.10 of this chapter must include a description of the following: Analytes and targets the device detects and identifies, the specimen types tested, the results provided to the user, the clinical indications for which the test is to be used, the specific intended population(s), the intended use locations including testing location(s) where the device is to be used (if applicable), and other conditions of use as appropriate.
(2) Any sample collection device used must be FDA-cleared, -approved, or -classified as 510(k) exempt (standalone or as part of a test system) for the collection of specimen types claimed by this device; alternatively, the sample collection device must be cleared in a premarket submission as a part of this device.
(3) The labeling required under § 809.10(b) of this chapter must include:
(i) A detailed device description, including reagents, instruments, ancillary materials, all control elements, and a detailed explanation of the methodology, including all pre-analytical methods for processing of specimens;
(ii) Detailed descriptions of the performance characteristics of the device for each specimen type claimed in the intended use based on analytical studies including the following, as applicable: Limit of Detection, inclusivity, cross-reactivity, interfering substances, competitive inhibition, carryover/cross contamination, specimen stability, precision, reproducibility, and clinical studies;
(iii) Detailed descriptions of the test procedure(s), the interpretation of test results for clinical specimens, and acceptance criteria for any quality control testing;
(iv) A warning statement that viral culture should not be attempted in cases of positive results for SARS-CoV-2 and/or any similar microbial agents unless a facility with an appropriate level of laboratory biosafety (
e.g., BSL 3 and BSL 3+, etc.) is available to receive and culture specimens; and(v) A prominent statement that device performance has not been established for specimens collected from individuals not identified in the intended use population (
e.g., when applicable, that device performance has not been established in individuals without signs or symptoms of respiratory infection).(vi) Limiting statements that indicate that:
(A) A negative test result does not preclude the possibility of infection;
(B) The test results should be interpreted in conjunction with other clinical and laboratory data available to the clinician;
(C) There is a risk of incorrect results due to the presence of nucleic acid sequence variants in the targeted pathogens;
(D) That positive and negative predictive values are highly dependent on prevalence;
(E) Accurate results are dependent on adequate specimen collection, transport, storage, and processing. Failure to observe proper procedures in any one of these steps can lead to incorrect results; and
(F) When applicable (
e.g., recommended by the Centers for Disease Control and Prevention, by current well-accepted clinical guidelines, or by published peer-reviewed literature), that the clinical performance may be affected by testing a specific clinical subpopulation or for a specific claimed specimen type.(4) Design verification and validation must include:
(i) Detailed documentation, including performance results, from a clinical study that includes prospective (sequential) samples for each claimed specimen type and, as appropriate, additional characterized clinical samples. The clinical study must be performed on a study population consistent with the intended use population and compare the device performance to results obtained using a comparator that FDA has determined is appropriate. Detailed documentation must include the clinical study protocol (including a predefined statistical analysis plan), study report, testing results, and results of all statistical analyses.
(ii) Risk analysis and documentation demonstrating how risk control measures are implemented to address device system hazards, such as Failure Modes Effects Analysis and/or Hazard Analysis. This documentation must include a detailed description of a protocol (including all procedures and methods) for the continuous monitoring, identification, and handling of genetic mutations and/or novel respiratory pathogen isolates or strains (
e.g., regular review of published literature and periodic in silico analysis of target sequences to detect possible mismatches). All results of this protocol, including any findings, must be documented and must include any additional data analysis that is requested by FDA in response to any performance concerns identified under this section or identified by FDA during routine evaluation. Additionally, if requested by FDA, these evaluations must be submitted to FDA for FDA review within 48 hours of the request. Results that are reasonably interpreted to support the conclusion that novel respiratory pathogen strains or isolates impact the stated expected performance of the device must be sent to FDA immediately.(iii) A detailed description of the identity, phylogenetic relationship, and other recognized characterization of the respiratory pathogen(s) that the device is designed to detect. In addition, detailed documentation describing how to interpret the device results and other measures that might be needed for a laboratory diagnosis of respiratory infection.
(iv) A detailed device description, including device components, ancillary reagents required but not provided, and a detailed explanation of the methodology, including molecular target(s) for each analyte, design of target detection reagents, rationale for target selection, limiting factors of the device (
e.g., saturation level of hybridization and maximum amplification and detection cycle number, etc.), internal and external controls, and computational path from collected raw data to reported result (e.g., how collected raw signals are converted into a reported signal and result), as applicable.(v) A detailed description of device software, including software applications and hardware-based devices that incorporate software. The detailed description must include documentation of verification, validation, and hazard analysis and risk assessment activities, including an assessment of the impact of threats and vulnerabilities on device functionality and end users/patients as part of cybersecurity review.
(vi) For devices intended for the detection and identification of microbial agents for which an FDA recommended reference panel is available, design verification and validation must include the performance results of an analytical study testing the FDA recommended reference panel of characterized samples. Detailed documentation must be kept of that study and its results, including the study protocol, study report for the proposed intended use, testing results, and results of all statistical analyses.
(vii) For devices with an intended use that includes detection of Influenza A and Influenza B viruses and/or detection and differentiation between the Influenza A virus subtypes in human clinical specimens, the design verification and validation must include a detailed description of the identity, phylogenetic relationship, or other recognized characterization of the Influenza A and B viruses that the device is designed to detect, a description of how the device results might be used in a diagnostic algorithm and other measures that might be needed for a laboratory identification of Influenza A or B virus and of specific Influenza A virus subtypes, and a description of the clinical and epidemiological parameters that are relevant to a patient case diagnosis of Influenza A or B and of specific Influenza A virus subtypes. An evaluation of the device compared to a currently appropriate and FDA accepted comparator method. Detailed documentation must be kept of that study and its results, including the study protocol, study report for the proposed intended use, testing results, and results of all statistical analyses.
(5) When applicable, performance results of the analytical study testing the FDA recommended reference panel described in paragraph (b)(4)(vi) of this section must be included in the device's labeling under § 809.10(b) of this chapter.
(6) For devices with an intended use that includes detection of Influenza A and Influenza B viruses and/or detection and differentiation between the Influenza A virus subtypes in human clinical specimens in addition to detection of SARS-CoV-2 and similar microbial agents, the required labeling under § 809.10(b) of this chapter must include the following:
(i) Where applicable, a limiting statement that performance characteristics for Influenza A were established when Influenza A/H3 and A/H1-2009 (or other pertinent Influenza A subtypes) were the predominant Influenza A viruses in circulation.
(ii) Where applicable, a warning statement that reads if infection with a novel Influenza A virus is suspected based on current clinical and epidemiological screening criteria recommended by public health authorities, specimens should be collected with appropriate infection control precautions for novel virulent influenza viruses and sent to State or local health departments for testing. Viral culture should not be attempted in these cases unless a BSL 3+ facility is available to receive and culture specimens.
(iii) Where the device results interpretation involves combining the outputs of several targets to get the final results, such as a device that both detects Influenza A and differentiates all known Influenza A subtypes that are currently circulating, the device's labeling must include a clear interpretation instruction for all valid and invalid output combinations, and recommendations for any required followup actions or retesting in the case of an unusual or unexpected device result.
(iv) A limiting statement that if a specimen yields a positive result for Influenza A, but produces negative test results for all specific influenza A subtypes intended to be differentiated (
i.e., H1-2009 and H3), this result requires notification of appropriate local, State, or Federal public health authorities to determine necessary measures for verification and to further determine whether the specimen represents a novel strain of Influenza A.(7) If one of the actions listed at section 564(b)(1)(A) through (D) of the Federal Food, Drug, and Cosmetic Act occurs with respect to an influenza viral strain, or if the Secretary of Health and Human Services determines, under section 319(a) of the Public Health Service Act, that a disease or disorder presents a public health emergency, or that a public health emergency otherwise exists, with respect to an influenza viral strain:
(i) Within 30 days from the date that FDA notifies manufacturers that characterized viral samples are available for test evaluation, the manufacturer must have testing performed on the device with those influenza viral samples in accordance with a standardized protocol considered and determined by FDA to be acceptable and appropriate.
(ii) Within 60 days from the date that FDA notifies manufacturers that characterized influenza viral samples are available for test evaluation and continuing until 3 years from that date, the results of the influenza emergency analytical reactivity testing, including the detailed information for the virus tested as described in the certificate of authentication, must be included as part of the device's labeling in a tabular format, either by:
(A) Placing the results directly in the device's labeling required under § 809.10(b) of this chapter that accompanies the device in a separate section of the labeling where analytical reactivity testing data can be found, but separate from the annual analytical reactivity testing results; or
(B) In a section of the device's label or in other labeling that accompanies the device, prominently providing a hyperlink to the manufacturer's public website where the analytical reactivity testing data can be found. The manufacturer's website, as well as the primary part of the manufacturer's website that discusses the device, must provide a prominently placed hyperlink to the website containing this information and must allow unrestricted viewing access.