cobas CT/NG for use on cobas 6800/8800 systems

K173887 · Roche Molecular Systems, Inc. · LSL · Mar 21, 2018 · Microbiology

Device Facts

• Record ID: K173887
• Device Name: cobas CT/NG for use on cobas 6800/8800 systems
• Applicant: Roche Molecular Systems, Inc.
• Product Code: LSL · Microbiology
• Decision Date: Mar 21, 2018
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3390
• Device Class: Class 2

Intended Use

The cobas® CT/NG on the cobas® 6800/8800 system is an automated, qualitative in vitro nucleic acid diagnostic test, that utilizes real-time polymerase chain reaction (PCR), for the direct detection of Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (NG) DNA in male and female urine, clinician-instructed self-collected vaginal swab specimens (collected in a clinical setting), clinician-collected vaginal swab specimens, and endocervical swab specimens, all collected in cobas® PCR Media (Roche Molecular Systems, Inc.), and cervical specimens collected in PreservCyt® solution. This test is intended as an aid in the diagnosis of chlamydial and gonococcal disease in both symptomatic and asymptomatic individuals.

Device Story

Automated, qualitative in vitro nucleic acid diagnostic test; utilizes real-time PCR for direct detection of CT and NG DNA. Input: male/female urine, vaginal swabs, endocervical swabs, cervical specimens in PreservCyt. Process: fully automated nucleic acid extraction/purification; PCR amplification using target-specific primers/probes for CT cryptic plasmid/ompA gene and NG DR-9 region; DNA internal control monitors process. Output: qualitative results (positive/negative/invalid) displayed on system screen, exported, or printed. Used in clinical laboratory settings; operated by trained laboratory personnel. Healthcare providers use results to aid diagnosis of chlamydial and gonococcal disease; enables timely clinical decision-making and patient management.

Clinical Evidence

Multi-site, prospective clinical study (n=5,053 evaluable subjects). Performance compared to Patient Infected Status (PIS) based on combined results of FDA-cleared NAATs. Sensitivity for CT ranged from 92.5% to 100% across specimen types; specificity ranged from 98.8% to 99.8%. Sensitivity for NG ranged from 89.5% to 100%; specificity ranged from 99.7% to 100%. Reproducibility study confirmed high precision across sites, lots, and operators.

Technological Characteristics

Automated nucleic acid extraction/purification; real-time PCR amplification; TaqMan probe-based detection (fluorescence resonance energy transfer). Targets: CT cryptic plasmid/ompA gene; NG DR-9 region. System: cobas 6800/8800. Connectivity: integrated system software for data management. Sterilization: not applicable (reagents). Software: automated result assignment.

Indications for Use

Indicated for the qualitative detection of Chlamydia trachomatis and/or Neisseria gonorrhoeae DNA in male and female urine, clinician-instructed self-collected vaginal swabs, clinician-collected vaginal swabs, endocervical swabs, and cervical specimens in PreservCyt solution. Intended for symptomatic and asymptomatic individuals as an aid in diagnosis.

Regulatory Classification

Identification

Neisseria spp. direct serological test reagents are devices that consist of antigens and antisera used in serological tests to identify Neisseria spp. from cultured isolates. Additionally, some of these reagents consist of Neisseria spp. antisera conjugated with a fluorescent dye (immunofluorescent reagents) which may be used to detect the presence of Neisseria spp. directly from clinical specimens. The identification aids in the diagnosis of disease caused by bacteria belonging to the genus Neisseria, such as epidemic cerebrospinal meningitis, meningococcal disease, and gonorrhea, and also provides epidemiological information on diseases caused by these microorganisms. The device does not include products for the detection of gonorrhea in humans by indirect methods, such as detection of antibodies or of oxidase produced by gonococcal organisms.

Predicate Devices

• cobas® CT/NG v2.0 Test (K163184)

Submission Summary (Full Text)

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Image /page/0/Picture/0 description: The image shows the logos of the Department of Health & Human Services and the U.S. Food & Drug Administration (FDA). The Department of Health & Human Services logo is on the left, and the FDA logo is on the right. The FDA logo is a blue square with the letters "FDA" in white, followed by the words "U.S. Food & Drug Administration" in blue.

March 21, 2018

Roche Molecular Systems, Inc. Nobuko Nakajima Manager, Regulatory Affairs 4300 Hacienda Drive Pleasanton, California 94588-2722

Re: K173887

Trade/Device Name: cobas CT/NG for use on cobas 6800/8800 systems Regulation Number: 21 CFR 866.3390 Regulation Name: Neisseria spp. direct serological test reagents Regulatory Class: Class II Product Code: LSL, MKZ, OOI Dated: December 20, 2017 Received: December 21, 2017

Dear Nobuko Nakajima:

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 (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. 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.

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 803); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR

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Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/MedicalDevices/DeviceRegulationandGuidance/) and CDRH Learn (http://www.fda.gov/Training/CDRHLearn). 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 (http://www.fda.gov/DICE) for more information or contact DICE by email ([e-mail]) or phone [phone] or 301-796-7100).

Sincerely,

# Tamara V. Feldblyum -S for

Uwe Scherf, Ph.D. Director Division of Microbiology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health

Enclosure

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### Indications for Use

510(k) Number (if known) K173887

### Device Name

cobas® CT/NG for use on the cobas® 6800/8800 Systems

### Indications for Use (Describe)

The cobas® CT/NG on the cobas® 6800/8800 system is an automated, qualitative in vitro nucleic acid diagnostic test, that utilizes real-time polymerase chain reaction (PCR), for the direct detection of Chlamydia trachomatis (CT) and/or Neisseria gonorthoeae (NG) DNA in male and female urine, clinician-instructed self-collected vaginal swab specimens (collected in a clinical setting), clinician-collected vaginal swab specimens, and endocervical swab specimens, all collected in cobas® PCR Media (Roche Molecular Systems, Inc.), and cervical specimens collected in PreservCyt® solution. This test is intended as an aid in the diagnosis of chlamydial and gonococcal disease in both symptomatic and asymptomatic individuals.

Ancillary Collection Kits:

The cobas® PCR Media Dual Swab Sample Kit is used to collect and transport endocervical and vaginal swab specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for gynecological specimens.

Note: This kit has been validated for use with the following tests:

· cobas® CT/NG v2.0 Test

- · cobas® CT/NG for use on cobas® 6800/8800 Systems
The cobas® PCR Media Uni Swab Sample Kit is used to collect and transport human specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for human specimens.

Note: This kit has been validated for use with the following tests:

- · cobas® CT/NG v2.0 Test
• cobas® CT/NG for use on cobas® 6800/8800 Systems

· cobas® Cdiff Test for use on the cobas® 4800 System

The cobas® PCR Urine Sample Kit is used to collect and transport urine specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for urine specimens. Use this cobas® CT/NG on cobas® 6800/8800 Systems or the cobas® CT/NG v2.0 Test.

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

X Prescription Use (Part 21 CFR 801 Subpart D)

Over-The-Counter Use (21 CFR 801 Subpart C)

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# cobas® CT/NG for use on the cobas® 6800/8800 Systems 510(k) Summary

This summary of 510(k) safety and effectiveness information is being submitted in accordance with the requirements of 21 CFR 807.92.

| Submitter Name | Roche Molecular Systems, Inc. |
|----------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Address | 4300 Hacienda Drive<br>Pleasanton, CA, 94588-2722 |
| Contact | Nobuko Nakajima<br>Phone: [phone]<br>FAX: [phone]<br>Email: [e-mail] |
| Date Prepared | December 4, 2017 |
| Proprietary Name | cobas® CT/NG<br>for use on cobas® 6800/8800 systems |
| Common Name | Real-time PCR assay, in vitro nucleic acid amplification test for the quantitative<br>detection of Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (NG) |
| Classification Name | Sec. 866.3390 Neisseria spp. direct serological test reagents<br>Sec. 866.3120 Chlamydia serological reagents,<br>Sec. 862.2570 Real Time Nucleic Acid Amplification System |
| Product Codes | LSL<br>MKZ<br>OOI |
| Predicate Devices | cobas® CT/NG v2.0 Test (K163184) |
| Establishment Registration | Roche Molecular Systems, Inc. (2243471) |

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#### 1. DEVICE DESCRIPTION

cobas® CT/NG is a new qualitative test performed on the cobas® 6800 System and cobas® 8800 System. cobas® CT/NG enables the detection of CT/NG DNA in endocervical, vaginal, urine and cervical specimens of infected female patients and urine specimens in infected male patients. Target-specific primers and two probes are used to detect but not discriminate between the CT cryptic plasmid and the ompA gene. Additionally, target-specific primers and two probes are used to detect but not discriminate between two conserved sequences in the NG DR-9 region. The DNA Internal Control, used to monitor the entire sample preparation and PCR amplification process, is introduced into each specimen during sample processing. In addition, the test utilizes a low titer positive and a negative control.

#### Principles of the procedure 1.1.

cobas® CT/NG is based on fully automated sample preparation (nucleic acid extraction and purification) followed by PCR amplification and detection. The cobas® 6800/8800 Systems consist of the sample supply module, the transfer module, the processing module, and the analytic module. Automated data management is performed by the cobas® 6800/8800 software which assigns test results for all tests as positive, negative or invalid. Results can be reviewed directly on the system screen, exported, or printed as a report.

Nucleic acid from patient samples, external controls and added internal control DNA (DNA-IC) molecules is simultaneously extracted. In summary, bacterial nucleic acid is released by addition of proteinase and lysis reagent to the sample. The released nucleic acid binds to the silica surface of the added magnetic glass particles. Unbound substances and impurities, such as denatured protein, cellular debris and potential PCR inhibitors are removed with subsequent wash steps and purified nucleic acid is eluted from the magnetic glass particles with elution buffer at elevated temperature.

Selective amplification of target nucleic acid from the sample is achieved by the use of targetspecific forward and reverse primers which are selected from highly conserved plasmid and genomic regions of CT and NG. A region on the CT cryptic plasmid and the ompA gene (dual target) and two conserved sequences of the NG DR-9 region are amplified by cobas® CT/NG.

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Selective amplification of DNA-IC is achieved by the use of sequence-specific forward and reverse primers which are selected to have no homology with either the CT or NG target regions. A thermostable DNA polymerase enzyme is used for PCR amplification. The target and DNA-IC sequences are amplified simultaneously utilizing a universal PCR amplification profile with predefined temperature steps and number of cycles. The master mix includes deoxyuridine triphosphate (dUTP), instead of deoxythimidine triphosphate (dTTP), which is incorporated into the newly synthesized DNA (amplicon). Any contaminating amplicon from previous PCR runs are eliminated by the AmpErase enzyme, which is included in the PCR master mix, during the first thermal cycling step. However, newly formed amplicons are not eliminated since the AmpErase enzyme is inactivated once exposed to temperatures above 55℃.

The cobas® CT/NG master mix contains two detection probes specific for the CT target sequences, two detection probes specific for the NG target sequences and one for the DNA-IC. The probes are labeled with target specific fluorescent reporter dyes allowing simultaneous detection of CT targets, NG targets and DNA-IC in three different target channels. When not bound to the target sequence, the fluorescent signal of the intact probes is suppressed by a quencher dye. During the PCR amplification step, hybridization of the probes to the specific single-stranded DNA template results in cleavage of the 5' to 3' exonuclease activity of the DNA polymerase resulting in separation of the reporter and quencher dyes and the generation of a fluorescent signal. With each PCR cycle, increasing amounts of cleaved probes are generated and the cumulative signal of the reporter dye increases concomitantly. Real-time detection and discrimination of PCR products is accomplished by measuring the fluorescence of the released reporter dyes for the CT and NG targets and DNA-IC, respectively.

### Figure 1: cobas® CT/NG on the cobas® 6800/8800 system

| [IVD] | | |
|-------------------------------------------------------------|----------------------------------------------------------------------|--------------------------------------------------------------------|
| cobas CT/NG Positive Control Kit | cobas CT/NG | cobas Buffer Negative Control Kit |
| Positive control kit for use on the cobas 6800/8800 Systems | Qualitative nucleic acid test for use on the cobas 6800/8800 Systems | Buffer negative control kit for use on the cobas 6800/8800 Systems |
| | | |
| cobas | cobas | cobas |

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#### 2. INTENDED USE

The cobas® CT/NG on the cobas® 6800/8800 system is an automated, qualitative in vitro nucleic acid diagnostic test, that utilizes real-time polymerase chain reaction (PCR), for the direct detection of Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (NG) DNA in male and female urine, clinician-instructed self-collected vaginal swab specimens (collected in a clinical setting), clinician-collected vaginal swab specimens, and endocervical swab specimens, all collected in cobas® PCR Media (Roche Molecular Systems, Inc.), and cervical specimens collected in PreservCyt® solution. This test is intended as an aid in the diagnosis of chlamydial and gonococcal disease in both symptomatic and asymptomatic individuals.

#### Ancillary Collection Kits 2.1.

The cobas® PCR Media Dual Swab Sample Kit is used to collect and transport endocervical and vaginal swab specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for gynecological specimens.

Note: This kit has been validated for use with the following tests:

- cobas® CT/NG v2.0 Test .
- cobas® CT/NG for use on cobas® 6800/8800 Systems .

The cobas® PCR Media Uni Swab Sample Kit is used to collect and transport human specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for human specimens.

Note: This kit has been validated for use with the following tests:

- cobas® CT/NG v2.0 Test .
- cobas® CT/NG for use on cobas® 6800/8800 Systems .
- cobas® Cdiff Test for use on the cobas® 4800 System .

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The cobas® PCR Urine Sample Kit is used to collect and transport urine specimens. The cobas® PCR Media serves as a nucleic acid stabilizing transport and storage medium for urine specimens. Use this collection kit only with either cobas® CT/NG on cobas® 6800/8800 Systems or the cobas® CT/NG v2.0 Test.

#### TECHNOLOGICAL CHARACTERISTICS 3.

The primary technological characteristics and intended use of the RMS cobas® CT/NG for use on the cobas® 6800/8800 systems are substantially equivalent to other legally marketed nucleic acid amplification tests intended for the qualitative detection of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG).

As indicated in Table 1, the cobas® CT/NG for use on the cobas® 6800/8800 systems is substantially equivalent to significant characteristics of the identified predicate device, the currently cleared cobas® CT/NG v2.0 Test (K163184).

| | Predicate Device:<br>cobas® CT/NG v2.0 Test<br>(K163184) | Submitted Device<br>cobas® CT/NG for use on the<br>cobas® 6800/8800 systems |
|------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Intended Use | The cobas® CT/NG v2.0 Test is an<br>automated, in vitro nucleic acid amplification<br>test for the qualitative detection of<br>Chlamydia trachomatis (CT) and/or<br>Neisseria gonorrhoeae (NG) DNA in<br>urogenital specimens. The Test utilizes the<br>Polymerase Chain Reaction (PCR) for the<br>detection of Chlamydia trachomatis and<br>Neisseria gonorrhoeae DNA in male and<br>female urine, self-collected vaginal swab<br>specimens (collected in a clinical setting),<br>clinician-collected vaginal swab specimens,<br>and endocervical swab specimens, all<br>collected in cobas® PCR Media (Roche<br>Molecular Systems, Inc.), and cervical<br>specimens collected in PreservCyt® solution.<br>This test is intended as an aid in the<br>diagnosis of chlamydial and gonococcal<br>disease in both symptomatic and<br>asymptomatic individuals. | The cobas® CT/NG on the cobas® 6800/8800<br>system is an automated, qualitative in vitro<br>nucleic acid diagnostic test, that utilizes real-time<br>polymerase chain reaction (PCR), for the direct<br>detection of Chlamydia trachomatis (CT) and/or<br>Neisseria gonorrhoeae (NG) DNA in male and<br>female urine, clinician instructed self-collected<br>vaginal swab specimens (collected in a clinical<br>setting), clinician-collected vaginal swab<br>specimens, and endocervical swab specimens,<br>all collected in cobas® PCR Media (Roche<br>Molecular Systems, Inc.), and cervical<br>specimens collected in PreservCyt® solution.<br>This test is intended as an aid in the diagnosis of<br>chlamydial and gonococcal disease in both<br>symptomatic and asymptomatic individuals. |
| | Predicate Device:<br>cobas® CT/NG v2.0 Test<br>(K163184) | Submitted Device<br>cobas® CT/NG for use on the<br>cobas® 6800/8800 systems |
| | Ancillary Collection Kits: | Ancillary Collection Kits: |
| | The cobas® PCR Media Dual Swab Sample<br>Kit is used to collect and transport<br>endocervical and vaginal swab specimens.<br>The cobas® PCR Media serves as a nucleic<br>acid stabilizing transport and storage<br>medium for gynecological specimens. Use<br>this collection kit only with the cobas®<br>CT/NG v2.0 Test. | The cobas® PCR Media Dual Swab Sample Kit<br>is used to collect and transport endocervical and<br>vaginal swab specimens. The cobas® PCR<br>Media serves as a nucleic acid stabilizing<br>transport and storage medium for gynecological<br>specimens. Note: This kit has been validated for<br>use with the following tests: cobas® CT/NG v2.0<br>Test cobas® CT/NG for use on<br>cobas® 6800/8800 Systems. |
| Intended Use,cont. | The cobas® PCR Media Uni Swab Sample<br>Kit is used to collect and transport human<br>specimens. The cobas® PCR Media serves<br>as a nucleic acid stabilizing transport and<br>storage medium for human specimens. Use<br>this collection kit with cobas® CT/NG v2.0<br>Test and cobas® Cdiff Tests. | The cobas® PCR Media Uni Swab Sample Kit is<br>used to collect and transport human specimens.<br>The cobas® PCR Media serves as a nucleic acid<br>stabilizing transport and storage medium for<br>human specimens Note: This kit has been<br>validated for use with the following tests:<br>cobas® CT/NG v2.0 Test cobas® CT/NG for use<br>on cobas® 6800/8800 Systems cobas® Cdiff<br>Test for use on the cobas® 4800 System |
| | The cobas® PCR Urine Sample Kit is used<br>to collect and transport urine specimens.<br>The cobas® PCR Media serves as a nucleic<br>acid stabilizing transport and storage<br>medium for urine specimens. Use this<br>collection kit only with the cobas® CT/NG<br>v2.0 Test. | The cobas® PCR Urine Sample Kit is used to<br>collect and transport urine specimens. The<br>cobas® PCR Media serves as a nucleic acid<br>stabilizing transport and storage medium for<br>urine specimens Use this collection kit only with<br>either cobas® CT/NG on cobas® 6800/8800<br>Systems or the cobas® CT/NG v2.0 Test. |
| Sample Types | • Male and female urine,<br>• Self-collected/clinician-collected<br>vaginal swab specimens in<br>cobas® PCR Media,<br>• Endocervical swab specimens in<br>cobas® PCR Media,<br>• Cervical specimens in PreservCyt®<br>solution | Same |
| Subject Status | Asymptomatic and symptomatic | Same |
| Sample Collection<br>Devices | cobas® PCR Media Dual Swab Sample Kit<br>cobas® PCR Media Uni Swab Sample Kit<br>cobas® PCR Urine Sample Kit | Same |
| CT Analyte targets | CT cryptic plasmid DNA<br>CT ompA gene | Same |
| NG Analyte targets | NG genomic DNA | Same |
| | Predicate Device:<br>cobas® CT/NG v2.0 Test<br>(K163184) | Submitted Device<br>cobas® CT/NG for use on the<br>cobas® 6800/8800 systems |
| Sample<br>Preparation<br>Procedure | Semi-automated | Automated |
| Amplification<br>Technology | Real-time PCR | Same |
| Detection<br>Chemistry | Paired reporter and quencher fluorescence<br>labeled probes (TaqMan Technology) using<br>fluorescence resonance energy transfer<br>(FRET) | Same |
| Result Analysis | Based on PCR cycle threshold analysis | Same |
| Analyzer | cobas® 4800 System | cobas® 6800/8800 systems |

Comparison of the cobas® CT/NG for use on the cobas® 6800/8800 systems with Table 1: the Predicate Device

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#### 4. NON-CLINICAL PERFORMANCE EVALUATION

#### 4.1. Limit of Detection (LoD)

Analytical sensitivity (Limit of Detection or LoD) was determined by analyzing a dilution series of quantified cultures of Chlamydia trachomatis (serovars D and I) and Neisseria gonorrhoeae isolates 2948 (ATCC 19424) and 891. CT and NG cultures were diluted into a matrix of pooled negative specimens of each sample type and 70-78 replicates were tested for each level in each specimen type. All levels were analyzed across 3 unique lots of reagents. LoD for each specimen type is shown in Table 2 as the target concentration which can be detected in ≥ 95% of the replicates for all lots.

| Specimen Types | C. trachomatis | | N. gonorrhoeae | | | | | |
|------------------------------------|-----------------|---------------------|-----------------|---------------------|-----------------|---------------------|-----------------|---------------------|
| | Serovar D | | Serovar I | | Strain 2948 | | Strain 891 | |
| | LOD<br>(IFU/mL) | Mean<br>Ct<br>Value | LOD<br>(IFU/mL) | Mean<br>Ct<br>Value | LOD<br>(CFU/mL) | Mean<br>Ct<br>Value | LOD<br>(CFU/mL) | Mean<br>Ct<br>Value |
| Endocervical Swab<br>in cobas® PCR | 0.3 | 36.6 | 1.4 | 37.1 | 0.4 | 36.3 | 0.08 | 37.5 |
| Vaginal Swab in | 0.3 | 37.3 | 1.4 | 37.0 | 0.4 | 36.3 | 0.08 | 37.0 |
| cobas® PCR | 0.2 | 37.8 | 1.3 | 37.1 | 0.2 | 36.3 | 0.04 | 38.3 |

| Table 2: Analytical sensitivity (Limit of Detection) |
|------------------------------------------------------|
| |

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| Cervical Samples<br>collected into<br>PreservCyt®<br>Solution | 0.6 | 37.4 | 2.9 | 37.4 | 0.2 | 36.7 | 0.08 | 37 |
|---------------------------------------------------------------|-----|------|-----|------|-----|------|------|----|
|---------------------------------------------------------------|-----|------|-----|------|-----|------|------|----|

IFU = Inclusion Forming Unit; quantification of the same C. trachomatis culture using DFA method equates 1 IFU to 6.6 signal generating units (SGU) for Serovar D, and 13.9 SGU for serovar I, where SGU includes Elementary Bodies as well as Reticulate Bodies of C. trachomatis

CFU = Colony Forming Units

#### 4.2. Inclusivity

Inclusivity testing was performed for 13 additional CT serovars, the Swedish new variant strain (nvCT) and an additional 43 independently isolated strains of NG using one lot of reagents. Testing was performed using CT and NG cultures diluted into pools of negative specimens. Results are shown in Table 3 and Table 4 for CT serovars and NG strains, respectively. Twenty replicates per dilution level were tested for each strain in each specimen type.

| | | Swab* Specimens | | Urine Specimens | | PreservCyt Specimens | |
|-------------------------|--|-----------------|-------|-----------------|-------|----------------------|-------|
| Serovar Type or Variant | | IFU/mL | % Pos | IFU/mL | % Pos | IFU/mL | % Pos |
| A | | 1.4 | 100 | 0.7 | 100 | 1.4 | 100 |
| B | | 5.9 | 100 | 2.9 | 100 | 5.9 | 100 |
| Ba | | 18.3 | 100 | 9.1 | 100 | 18.3 | 100 |
| C | | 0.6 | 100 | 0.3 | 100 | 0.6 | 100 |
| E | | 6.4 | 100 | 3.2 | 100 | 6.4 | 100 |
| F | | 3.2 | 100 | 1.6 | 100 | 3.2 | 100 |
| G | | 2.9 | 100 | 1.5 | 100 | 2.9 | 100 |
| H | | 9.7 | 100 | 4.8 | 100 | 9.7 | 100 |
| J | | 1.4 | 100 | 0.7 | 100 | 1.4 | 100 |
| K | | 2.0 | 100 | 1.0 | 100 | 2.0 | 100 |
| LGV Type 1 | | 5.9 | 100 | 3.0 | 100 | 5.9 | 100 |
| LGV Type 2 | | 12.8 | 100 | 6.4 | 100 | 12.8 | 100 |
| LGV Type 3 | | 0.7 | 100 | 0.4 | 100 | 0.7 | 100 |
| nyCT | | 0.7 | 100 | 0.3 | 100 | 0.7 | 100 |

Inclusivity Testing for CT Serovars Table 3:

* Vaginal swab samples were used as a representative swab sample type for vaginal and endocervical swab specimens.

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| Numbers of NG Strains | Swab* Specimens | |
|-----------------------|----------------------|-------|
| | CFU/mL | % Pos |
| 39 | 0.4 | ≥ 95 |
| 4 | 1.0 | ≥ 95 |
| Total = 43 | | |
| Numbers of NG Strains | Urine Specimens | |
| | CFU/mL | % Pos |
| 41 | 0.2 | ≥ 95 |
| 2 | 0.5 | 100 |
| Total = 43 | | |
| Numbers of NG Strains | PreservCyt Specimens | |
| | CFU/mL | % Pos |
| 42 | 0.4 | ≥ 95 |
| 1 | 1.0 | 100 |
| Total = 43 | | |

Table 4: Inclusivity Testing for NG strains

* Vaginal swab samples were used as a representative swab sample type for vaginal and endocervical swab specimens.

#### 4.3. Precision

In-house precision was examined using a panel composed of CT and NG cultures diluted into a pool of negative endocervical swab specimen matrix collected in cobas® PCR Media, a pool of negative urine matrix plus cobas® PCR Media and a pool of negative cervical specimen matrix collected in PreservCyt® Solution. Endocervical swabs were intended to represent all swab samples collected in cobas® PCR Media (endocervical and vaginal). Four levels were tested using CT serovar D and NG strain 2948 (ATCC 19424) as the target organisms.

The precision panel was designed to include members with high negative, low and moderate concentrations of CT and NG for each panel matrix, corresponding to ~0.3x, ~1x and ~3x LoD. Testing was performed with three lots of cobas® CT/NG reagents and two instruments for a total of 24 runs. A description of the precision panels and the study performance hit rate is shown in Table 5. All negative panel members tested negative throughout the study. Analysis of standard deviation and percent coefficient of variation of the Ct values from valid tests performed on

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positive panel members (see Table 6 and Table 7) yielded overall CV (%) ranges from 1.62% to 4.05% for CT and from 1.17% to 3.55% for NG. Testing occurred over 12 days, using 2 instruments, with 2 runs per day. Each run consisted of 3 replicates of each sample.

| Level | N Tested | N positive CT | N positive NG | Hit Rate | | 95% CI CT | | 95% CI NG | |
|------------------------------------------------------|----------|---------------|---------------|----------|------|-----------|-----|-----------|-----|
| | | | | CT | NG | LL | UL | LL | UL |
| Endocervical Swab in cobas® PCR Media | | | | | | | | | |
| Negative | 72 | 0 | 0 | 0% | 0% | 0.0 | 5.0 | 0.0 | 5.0 |
| High Negative | 72 | 51 | 32 | 71% | 44% | 59 | 81 | 33 | 57 |
| Low | 72 | 69 | 68 | 96% | 94% | 88 | 99 | 86 | 98 |
| Moderate | 72 | 72 | 72 | 100% | 100% | 95 | 100 | 95 | 100 |
| Cervical samples collected into PreservCyt® Solution | | | | | | | | | |
| Negative | 72 | 0 | 0 | 0% | 0% | 0.0 | 5.0 | 0.0 | 5.0 |
| High Negative | 72 | 38 | 47 | 53% | 65% | 41 | 65 | 53 | 76 |
| Low | 72 | 72 | 69 | 100% | 96% | 95 | 100 | 88 | 99 |
| Moderate | 72 | 72 | 72 | 100% | 100% | 95 | 100 | 95 | 100 |
| cobas® PCR Media with Urine | | | | | | | | | |
| Negative | 72 | 0 | 0 | 0% | 0% | 0.0 | 5.0 | 0.0 | 5.0 |
| High Negative | 72 | 56 | 56 | 78% | 78% | 66 | 87 | 66 | 87 |
| Low | 72 | 71 | 72 | 99% | 100% | 92 | 100 | 95 | 100 |
| Moderate | 72 | 72 | 72 | 100% | 100% | 95 | 100 | 95 | 100 |

Table 5: Summary of within-laboratory precision

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| Level<br>(Hit Rate) | Mean Ct | Between<br>instrument | | Between<br>lot | | Within run | | Between<br>run | | Between<br>day | | Total | |
|------------------------------------------------------|---------|-----------------------|---------|----------------|---------|------------|---------|----------------|---------|----------------|---------|-------|---------|
| | | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% |
| Endocervical Swab in cobas® PCR Media | | | | | | | | | | | | | |
| High<br>Negative<br>(71%) | 39.7 | 0.0 | 0.0 | 0.0 | 0.0 | 1.2 | 3.21 | 0.0 | 0.00 | 0.3 | 0.8 | 1.3 | 3.3 |
| Low<br>(96%) | 38.5 | 0.0 | 0.0 | 0.0 | 0.1 | 1.1 | 2.96 | 0.0 | 0.00 | 0.4 | 1.2 | 1.2 | 3.2 |
| Moderate<br>(100%) | 36.9 | 0.0 | 0.0 | 0.2 | 0.6 | 0.5 | 1.45 | 0.0 | 0.18 | 0.0 | 0.0 | 0.6 | 1.6 |
| Cervical Samples collected into PreservCyt® Solution | | | | | | | | | | | | | |
| High<br>Negative<br>(53%) | 38.3 | 0.6 | 1.5 | 0.5 | 1.3 | 1.1 | 2.92 | 0.0 | 0.00 | 0.0 | 0.0 | 1.3 | 3.5 |
| Low<br>(100%) | 36.9 | 0.2 | 0.5 | 0.2 | 0.7 | 0.6 | 1.85 | 0.0 | 0.00 | 0.0 | 0.0 | 0.7 | 2.0 |
| Moderate<br>(100%) | 35.6 | 0.0 | 0.0 | 0.2 | 0.5 | 0.5 | 1.46 | 0.0 | 0.24 | 0.0 | 0.0 | 0.5 | 1.5 |
| cobas® PCR Media with Urine | | | | | | | | | | | | | |
| High<br>Negative<br>(78%) | 38.9 | 0.0 | 0.0 | 0.1 | 0.3 | 1.2 | 3.22 | 0.3 | 1.01 | 0.0 | 0.0 | 1.3 | 3.3 |
| Low<br>(99%) | 38.3 | 0.1 | 0.2 | 0.0 | 0.0 | 1.5 | 3.97 | 0.0 | 0.00 | 0.2 | 0.7 | 1.5 | 4.0 |
| Moderate<br>(100%) | 37.1 | 0.0 | 0.0 | 0.0 | 0.0 | 1.0 | 2.84 | 0.0 | 0.00 | 0.2 | 0.7 | 1.0 | 2.9 |

### Table 6: Overall mean, standard deviations and coefficients of variation (%) for cycle threshold, CT positive panel members

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| Level<br>(Hit Rate) | Mea<br>n Ct | Between<br>instrumen<br>t | | Between<br>lot | | Within run | | Between<br>run | | Between<br>day | | Total | |
|------------------------------------------------------|-------------|---------------------------|---------|----------------|---------|------------|---------|----------------|---------|----------------|---------|----------|---------|
| | | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% | SD | CV<br>% |
| Endocervical Swab in cobas® PCR Media | | | | | | | | | | | | | |
| High<br>Negative<br>(44% ) | 39.1 | 0.0<br>0 | 0.00 | 0.3<br>1 | 0.79 | 0.8<br>4 | 2.14 | 0.7<br>2 | 1.85 | 0.5<br>7 | 1.46 | 1.2<br>8 | 3.28 |
| Low (94%) | 38.1 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 1.2<br>7 | 3.34 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 1.2<br>7 | 3.34 |
| Moderate<br>(100%) | 36.5 | 0.0<br>0 | 0.00 | 0.2<br>4 | 0.67 | 0.6<br>9 | 1.89 | 0.0<br>0 | 0.00 | 0.1<br>5 | 0.40 | 0.7<br>4 | 2.04 |
| Cervical Samples collected into PreservCyt® Solution | | | | | | | | | | | | | |
| High<br>Negative<br>(65%) | 39.0 | 0.3<br>4 | 0.87 | 0.0<br>0 | 0.00 | 1.1<br>1 | 2.85 | 0.0<br>8 | 0.20 | 0.4<br>5 | 1.16 | 1.2<br>5 | 3.21 |
| Low (96%) | 38.0 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 1.2<br>5 | 3.28 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 1.2<br>5 | 3.28 |
| Moderate<br>(100%) | 35.8 | 0.0<br>0 | 0.00 | 0.2<br>8 | 0.78 | 0.7<br>6 | 2.13 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 0.8<br>1 | 2.27 |
| cobas® PCR Media with Urine | | | | | | | | | | | | | |
| High<br>Negative<br>(78%) | 39.1 | 0.0<br>0 | 0.00 | 0.2<br>6 | 0.66 | 1.3<br>5 | 3.46 | 0.0<br>0 | 0.00 | 0.1<br>8 | 0.45 | 1.3<br>9 | 3.55 |
| Low<br>(100%) | 36.7 | 0.1<br>4 | 0.38 | 0.1<br>6 | 0.42 | 0.7<br>1 | 1.92 | 0.0<br>0 | 0.00 | 0.0<br>0 | 0.00 | 0.7<br>4 | 2.00 |
| Moderate<br>(100%) | 34.9 | 0.0<br>0 | 0.00 | 0.1<br>6 | 0.47 | 0.3<br>7 | 1.06 | 0.0<br>6 | 0.18 | 0.0<br>0 | 0.00 | 0.4<br>1 | 1.17 |

### Table 7: Overall mean, standard deviations and coefficients of variation (%) for cycle threshold, NG positive panel members

#### Analytical specificity/Cross-reactivity 4.4.

A panel of 149 bacteria, fungi and viruses, including those commonly found in the male and female urogenital tract, 20 representatives of non-gonorrhoeae Neisseria strains and other phylogenetically unrelated organisms, were tested with cobas®CT/NG to assess analytical specificity. The organisms listed in Table 8 were spiked at concentrations of approximately 1 x 106 units*/mL for bacteria and approximately 1 x 105 units*/mL for viruses into pools of negative vaginal swab specimens in cobas® PCR Media, urine stabilized in cobas® PCR Media

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and cervical specimens in PreservCyt® Solution. Testing was performed with each potential interfering organism alone as well as with each organism mixed with CT and NG cultures at ~3x LoD. Results indicated that none of these organisms interfered with the detection of CT and NG or produced false positive results in the CT/NG negative matrices. (N=3 across the tested specimen types).

*All bacteria were quantified as Colony Forming Units (CFU) except Chlamydophila pneumonia and Chlamydophila psittaci which were quantified as Elementary Bodies (EB). All viruses were quantified as units/mL as determined by TCID50 Endpoint Dilution Assay. Trichomonas vaginalis and HPV16 were quantified as cells/mL.

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| Achromobacter xerosis | Gemella haemolysans | Neisseria subflava |
|--------------------------------|-----------------------------------------|----------------------------------------|
| Acinetobacter calcoaceticus | Haemophilus ducreyi | Neisseria weaverii |
| Acinetobacter Iwoffi | Haemophilus influenzae | Pantoea agglomerans |
| Actinomyces israelii | Helicobacter pylori | Paracoccus denitrificans |
| Aerococcus viridans | Herpes simplex virus I | Peptostreptococcus anaerobius |
| Aeromonas hydrophila | Herpes simplex virus II** | Peptostreptococcus<br>asaccharolyticus |
| Alcaligenes faecalis | HPV16* | Peptostreptococcus magnus |
| Atopobium vaginae | Kingella dentrificans | Plesiomonas shigelloides |
| Bacillus subtilis | Kingella kingae | Propionibacterium acnes |
| Bacteriodes fragilis | Klebsiella oxytoca | Proteus mirabilis |
| Bacteroides caccae | Klebsiella pneumoniae | Proteus penneri |
| Bacteroides ureolyticus | Lactobacillus acidophillus | Proteus vulgaris |
| Bergeriella denitrificans | Lactobacillus brevis | Providencia rettgeri |
| Bifidobacterium adolescentis | Lactobacillus crispatus | Providencia stuartii |
| Bifidobacterium breve | Lactobacillus jensenii | Pseudomonas aeruginosa |
| Bifidobacterium longum | Lactobacillus lactis | Pseudomonas fluorescens |
| Blautia product | Lactobacillus leichmannii | Pseudomonas putida |
| Branhamella catarrhalis | Lactobacillus oris | Rahnella aquatilis |
| Brevibacterium linens | Lactobacillus parabuchnerri | Rhizobium radiobacter |
| Campylobacter coli | Lactobacillus reuteri | Rhodospirillum rubrum |
| Campylobacter jejuni | Lactobacillus vaginalis | Saccharomyces cerevisiae |
| Candida albicans | Lactococcus lactis cremoris | Salmonella choleraesuis |
| Candida glabrata | Legionella pneumophila | Salmonella minnesota |
| Candida parapsilosis | Leuconostoc paramesenteroides | Salmonella typhimurium |
| Candida tropicalis | Listeria monocytogenes | Serratia denitrificans |
| Chlamydophila pneumoniae | Micrococcus luteus | Serratia marcescens |
| Chlamydophila psittaci | Moraxella lacunata | Shigella dysenteriae |
| Chromobacter violaceum | Moraxella osloensis | Staphylococcus aureus |
| Citrobacter freundii | Morganella morganii | Staphylococcus epidermidis |
| Clostridium difficile | Mycobacterium smegmatis | Staphylococcus saprophyticus |
| Clostridium perfringens | Mycoplasma genitalium*** | Streptococcus agalactiae |
| Corynebacterium genitalium | Mycoplasma hominis | Streptococcus anginosus |
| Corynebacterium xerosis | Neisseria cinerea | Streptococcus bovis |
| Cryptococcus neoformans | Neisseria elongata subsp. elongata | Streptococcus dysgalactiae |
| Cytomegalovirus** | Neisseria elongata subsp. nitroreducens | Streptococcus equinis |
| Deinococcus radiodurans | Neisseria flava | Streptococcus mitis |
| | | |
| Derxia gummosa | Neisseria flavescens | Streptococcus mutans |
| Eikenella corrodens | Neisseria kochi | Streptococcus pneumoniae |
| Enterobacter aerogenes | Neisseria lactamica | Streptococcus pyogenes |
| Enterobacter cloacae | Neisseria macacae | Streptococcus salivarius |
| Enterococcus avium | Neisseria meningitidis Serogroup A | Streptococcus sanguis |
| Enterococcus casseliflavus | Neisseria meningitidis Serogroup B | Streptomyces griseinus |
| Enterococcus faecalis | Neisseria meningitidis Serogroup C | Trichomonas vaginalis |
| Enterococcus faecium | Neisseria meningitidis Serogroup D | Trueperella pyogenes |
| Erysipelothrix rhusiopathiae | Neisseria meningitidis Serogroup W135 | Ureaplasma urealyticum |
| Escherichia coli | Neisseria meningitidis Serogroup Y | Veillonela parvula |
| Escherichia fergusonii | Neisseria mucosa | Vibrio cholerae |
| Flavobacterium meningosepticum | Neisseria perflava | Vibrio parahaemolyticus |
| Fusobacterium nucleatum | Neisseria polysaccharea | Yersinia enterocolitica |
| Gardnerella vaginalis | Neisseria sicca | - |

Table 8: Microorganisms tested for analytical specificity/cross reactivity

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HPV16 was tested as CaSki cells.

** Organism was tested at a concentration of 1 x 104 Units/mL.

***Organism was tested at a concentration of 1 x 105 CFU/mL.

#### 4.5. Interference

The effects of over-the-counter or prescription products that may be present in urogenital specimens (Table 9), were evaluated. Testing was done using pooled clinical specimens (vaginal swab, urine and PreservCyt® specimens) with spiking of potential interferents at levels expected from normal patient usage. Interferents were tested in CT/NG negative specimen pools as well as in specimen pools with CT/NG at ~3x LoD. CT serovars D and I and NG strains 2948 (ATCC 19424) and 891 were used in this study. Five replicates each of CT/NG negative and CT/NG positive samples were tested with each product in each specimen type, except for RepHresh™ Odor Eliminating Vaginal Gel and RepHresh™ Clean Balance Gel, which were tested with 2 replicates each to verify interference that had been observed with Replens™ Long-Lasting Vaginal Moisturizer, a product with a similar formulation.

Of the over-the-counter (OTC) and prescription products tested, Metronidazole Vaginal Gel, Replens™ Long-Lasting Vaginal Moisturizer, RepHresh™ Odor Eliminating Vaginal Gel and RepHresh™ Clean Balance produced false negative or invalid results in at least one replicate of the samples tested.

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| Product Name | Vaginal<br>Swabs | Urine | PreservCyt<br>Solution |
|------------------------------------------------------------------------------|------------------|-------|------------------------|
| | mg/mL | mg/mL | mg/mL |
| Clindamycin Phosphate Vaginal Cream | 7.1 | 3.4 | 1.6 |
| Equate tioconazole 1 | 3.7 | 1.7 | 0.8 |
| Equate Vagicaine Anti-Itch Cream | 4.1 | 2.0 | 0.9 |
| Estrace | 3.8 | 2.0 | 1.0 |
| K-YTM Ultra Gel | 5.7 | 2.7 | 1.2 |
| Metronidazole Vaginal Gel | 0.1* | 0.1* | 0.2* |
| Monistat 3 Vaginal Antifungal Combination Pack | 3.7 | 1.7 | 0.7 |
| Monistat® Complete Care Itch Relief Cream | 3.7 | 1.8 | 0.9 |
| 7 Day Vaginal Cream | 3.9 | 1.8 | 0.8 |
| Norforms Suppositories | 3.4 | 1.7 | 0.7 |
| Premarin | 6.1 | 3.1 | 1.4 |
| ReplensTM Long-Lasting Vaginal Moisturizer | 0.05* | 0.05* | 0.2* |
| Summer's Eve Feminine Deodorant Spray | 6.4 | 3.1 | 2.0 |
| VCF - Vaginal Contraceptive Foam | 2.1 | 1.0 | 0.4 |
| Yeast Gard Advanced | 3.7 | 1.7 | 1.0 |
| Azo Standard (urine only) | N/A | 0.1 | N/A |
| RepHreshTM Odor Eliminating Vaginal Gel | ‡ | ‡ | ‡ |
| RepHreshTM Clean Balance Gel | ‡ | ‡ | ‡ |
| * Concentrations above this level may cause interference in clinical samples | | | |

Table 9: List of substances with concentrations tested that do not interference with test performance in urogenital specimens

# RepHresh™ products were tested using simulated swab specimen. Concentrations of product that did not interfe…