The Aptima Combo 2 Assay is a target amplification nucleic acid probe test that utilizes target capture for the in vitro qualitative detection and differentiation of ribosomal RNA (rRNA) from Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (GC) to aid in the diagnosis of chlamydial and/or gonococcal disease using the Panther System as specified. On the Panther System, the assay may be used to test the following specimens from symptomatic and asymptomatic individuals: clinician-collected endocervical, PreservCyt® Solution liquid Pap specimens, vaginal, throat, rectal, and male urethral swab specimens; patient-collected vaginal swab specimens1, and female and male urine specimens.

The Aptima Combo 20 Assay is a target amplification nucleic acid probe test that utilizes target capture for the in vitro qualitative detection and differentiation of ribosomal RNA (rRNA) from Chlamydia trachomatis (CT) and/or Neisseria gonorrhoeae (GC) to aid in the diagnosis of chlamydial and/or gonococcal urogenital disease using the Tigris® DTS® Automated Analyzer or semi-automated instrumentation as specified. The assay may be used to test the following specimens from symptomatic individuals: clinician-collected endocervical, vaginal and male urethral swab specimens; and female and male urine specimens. The assay may be used to test the following specimens from asymptomatic individuals: clinician-collected endocervical, vaginal and male urethral swab specimens; patient-collected vaginal swab specimens'; and female and male urine specimens. The assay is also intended for use with the testing of gynecological specimens, from both symptomatic and asymptomatic patients, collected in the PreservCyt® Solution.

Device Description

The Aptima Combo 2 Assay combines the technologies of target capture, TMA, and DKA. Specimens are collected and transferred into their respective specimen transport tubes. The transport solutions in these tubes release the rRNA targets and protect them from degradation during storage. When the Aptima Combo 2 Assay is performed in the laboratory, the target rRNA molecules are isolated from specimens by use of capture oligomers via target capture that utilizes magnetic microparticles. The capture oligomers contain sequences complementary to specific regions of the target molecules as well as a string of deoxyadenosine residues. A separate capture oligomer is used for each target. During the hybridization step, the sequence specific regions of the capture oligomers bind to specific regions of the target molecules. The capture oligomer:target complex is then captured out of solution by decreasing the temperature of the reaction to room temperature. This temperature reduction allows hybridization to occur between the deoxyadenosine region on the capture oligomer and the poly-deoxythymidine molecules that are covalently attached to the magnetic particles. The microparticles, including the captured target molecules bound to them, are pulled to the side of the reaction vessel using magnets and the supernatant is aspirated. The particles are washed to remove residual specimen matrix that may contain amplification reaction inhibitors. After the target capture steps are completed, the specimens are ready for amplification.

Target amplification assays are based on the ability of complementary oligonucleotide primers to specifically anneal and allow enzymatic amplification of the target nucleic acid strands. The Aptima Combo 2 Assay replicates a specific region of the 23S rRNA from CT and a specific region of the 16S rRNA from GC via DNA intermediates. A unique set of primers is used for each target molecule. Detection of the rRNA amplification product sequences (amplicon) is achieved using nucleic acid hybridization. Single-stranded nucleic acid chemiluminescent probes, which are complementary to a region of each target amplicon, are labeled with different acridinium ester molecules. The updated version of the Aptima Combo 2 assay incorporates a second CT probe, complementary to a unique region of the existing CT amplicon. This tandem probe provides detection coverage for the variant strains of C. trachomatis that emerged in 2019. The labeled probes combine with amplicon to form stable hybrids. The Selection Reagent differentiates hybridized from unhybridized probe, eliminating the generation of signal from unhybridized probe. During the detection step, light emitted from the labeled hybrids is measured as photon signals in a luminometer, and are reported as Relative Light Units (RLU). In DKA, differences in the kinetic profiles of the CT and GC labeled probes allow for the differentiation of signal; kinetic profiles are derived from measurements of photon output during the detection read time. The chemiluminescent detection for CT signal has very rapid kinetics and has the "flasher" kinetic type. The chemiluminescent detection reaction for GC signal is relatively slower and has the "glower" kinetic type. Assay results are determined by a cut-off based on the total RLU and the kinetic curve type.

AI/ML Overview

The provided text describes a 510(k) premarket notification for a modified diagnostic device, the Aptima Combo 2 Assay, used for detecting Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC). The modification involves a change in the Probe reagent to improve detection of emerging CT variants.

Here's an analysis of the acceptance criteria and the study that proves the device meets them, based on the provided document:

1. Table of Acceptance Criteria and Reported Device Performance

The FDA 510(k) submission process for in vitro diagnostic devices focuses on demonstrating substantial equivalence to a legally marketed predicate device. For modifications to an existing cleared device, the key acceptance criteria revolve around showing that the changes do not negatively impact the assay's performance, safety, and effectiveness, and specifically address the reason for the modification (improved CT variant detection).

While explicit enumerated "acceptance criteria" with numerical thresholds are not presented in a table format as might be seen for a new device, the document implies the following performance criteria were met:

Performance Characteristic	Acceptance Criteria (Implied)	Reported Device Performance (Updated AC2 Assay)
Limit of Detection (LoD) for FI-nvCT	LoD for the Finnish variant of Chlamydia trachomatis (FI-nvCT) should be within acceptable limits (typically very low concentration).	Determined to be less than one IFU per assay in urine, ThinPrep, and simulated swab matrix specimens on both Panther and Tigris systems. Detection capabilities confirmed across multiple CT variants. This indicates high sensitivity for the targeted variants.
Clinical Comparability - CT	High positive and negative percent agreement with the current (predicate) AC2 assay for CT detection. Data should support that the updated assay performs similarly to the predicate on clinical samples.	Positive Percent Agreement (PPA): 100% (95% C.I.: 92.7% - 100%) Negative Percent Agreement (NPA): 98.9% (95% C.I.: 96.9% - 99.6%) Overall agreement was >99.0% for CT. This demonstrates strong concordance with the predicate device.
Clinical Comparability - GC	High positive and negative percent agreement with the current (predicate) AC2 assay for GC detection. Data should support that the updated assay performs similarly to the predicate on clinical samples.	Positive Percent Agreement (PPA): 100% (95% C.I.: 92.4% - 100%) Negative Percent Agreement (NPA): 99.6% (95% C.I.: 98.0% - 99.9%) Overall agreement was >99.0% for GC. This demonstrates strong concordance with the predicate device, also showing no negative impact on GC detection.
Clinical Panel Agreement (CT/GC)	High agreement (100% or very close) to expected panel results for both wild type CT, FI-nvCT, and GC across various concentrations. Consistency across instruments, lots, operators, days, and runs.	100% (97.6-100%) total CT and GC agreement to the expected panel result for the updated AC2 assay. For the current AC2 assay, 100% agreement except for the moderate (0.2 IFU/mL) FI-nvCT only panel (98.2% CT agreement, 99.1% GC agreement). This indicates the updated assay improves detection of the FI-nvCT variant while maintaining performance for other targets. Variability was comparable.
Microorganism Cross-Reactivity and Microbial Interference	No significant impact on detection capabilities or analytical specificity from a panel of common microorganisms. No false positives or interference should occur.	None of the 86 microorganisms tested were found to have an impact on the detection capabilities or analytical specificity of the updated version of the AC2 assay. This demonstrates high specificity and robustness against common interfering substances/microorganisms.
Overall Risk Profile	No new hazards introduced, and the overall residual risk does not increase compared to currently marketed products. Risks should be reduced as far as possible and meet pre-defined acceptability criteria.	Based on risk analysis and verification activities, all risks are reduced as far as possible and meet pre-defined acceptability criteria. No hazards fell within "Undesirable" or "Unacceptable" residual risk regions. Device modifications do not introduce any new hazards or increase the overall residual risk.

2. Sample Sizes Used for the Test Set and Data Provenance

Limit of Detection (LoD) Study:
- Sample Size: 30 replicates of each dilution were tested for each specimen type (urine, ThinPrep, simulated swab matrix). This was done with 3 reagent lots on both Panther and Tigris systems.
- Total replicates: 30 (replicates/dilution) * 3 (specimen types) * 3 (lots) * 2 (systems) = 540 replicates per target organism (CT/GC).
- Data Provenance: In vitro transcripts diluted in negative urine specimens, negative ThinPrep specimens, and simulated swab matrix specimens. This is laboratory-derived analytical data, not directly stated to be from a particular country. It is essentially prospective in nature for validating the new formulation.
Clinical Comparability Study:
- Sample Size: Not explicitly stated as a total count, but implied by the comparison tables below Table 4 and Table 5.
  - CT: 49 CT positive, 273 CT negative, 3 discordant (updated positive, current negative). Total: 325 remnant samples.
  - GC: 47 GC positive, 275 GC negative, 1 discordant (updated positive, current negative). Total: 323 remnant samples.
- Data Provenance: Remnant swab specimens collected from patients undergoing CT and/or GC screening. The origin country is not specified, but it's retrospective use of existing clinical samples.
CT/GC Clinical Panel Agreement Study:
- Sample Size: 20 prepared CT/GC clinical panels. Each panel was tested in triplicate, in two runs per day, on three Panther systems, by two operators, using three lots of reagents over seven days.
- Total tests: 20 (panels) * 3 (replicates) * 2 (runs/day) * 3 (systems) * 2 (operators) * 3 (lots) * 7 (days) = This calculation seems off and yields a very large number. More simply, if each panel was tested in triplicate across the stated conditions, the total number of data points specifically for these panels would be substantial. The document states "Each of the 20 panels were tested in triplicate...over seven days", implying multiple runs under varying conditions, leading to hundreds of data points (e.g., 20 panels * 3 replicates * 2 runs * 3 systems * 3 lots * 2 operators * 7 days if all combinations were run, which is impractical; more likely "across" these variables). Let's interpret "in triplicate...over seven days" as at least 3 runs per panel across variable conditions for a total of 60 tests (20 panels * 3 replicates) per condition (e.g., per system/lot/operator combination). The key is "The results show 100%...total CT and GC agreement to the expected panel result for the updated AC2 assay."
- Data Provenance: Prepared clinical panels containing known concentrations of wild type CT, FI-nvCT, and GC in urine specimens. This is an expertly constructed analytical study, effectively prospective in its execution for validation.

3. Number of Experts Used to Establish Ground Truth for the Test Set and Qualifications

The document does not mention the use of human experts (e.g., radiologists) for establishing ground truth for a test set. This is a molecular diagnostic assay, not an imaging device. Ground truth for these studies is typically established by:

Analytical studies: Known concentrations of purified nucleic acids (e.g., in vitro transcripts) or spiked microorganisms.
Clinical comparability: Agreement with a legally marketed predicate device (the "current" AC2 assay) on remnant clinical samples. The predicate device itself was cleared based on its own performance studies.
Clinical panels: Known concentrations of target organisms in simulated or real matrices.

Therefore, the concept of "experts" in the context of ground truth establishment for this specific device would relate to the scientific team involved in designing the analytical panels and interpreting the molecular results, rather than clinical experts adjudicating cases.

4. Adjudication Method for the Test Set

Not applicable in the typical sense for this type of device. There's no "adjudication" of images or clinical cases by multiple readers. The output of the device is a qualitative "positive" or "negative" result based on Relative Light Units (RLU) and kinetic curve type. Equivalence is determined by statistical agreement (percent agreement) between the test device and the predicate or expected panel results.

5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done

No. An MRMC study is relevant for imaging devices where human readers interpret medical images, often with and without AI assistance, to assess diagnostic performance. This document concerns a molecular diagnostic assay where a laboratory instrument provides a qualitative result. There is no human "reading" of the assay outcome in the same way an image is read.

6. If a Standalone (Algorithm Only Without Human-in-the-Loop Performance) Was Done

Yes, implicitly. The performance data presented (LoD, clinical comparability, clinical panel agreement, cross-reactivity) are all "standalone" in the sense that they demonstrate the analytical and clinical performance of the assay system itself (reagent + instrument) without any direct human interpretation of raw data (beyond standard laboratory procedures for operating the instrument and interpreting its final qualitative output, which is not "in-the-loop" AI assistance). The device is intended to provide a diagnostic result directly.

7. The Type of Ground Truth Used

Analytical Studies (LoD, Cross-Reactivity, Microbial Interference): "Spiked" or "known concentration" ground truth. For example, FI-nvCT in vitro transcripts at varying known concentrations, or panels of known microorganisms.
Clinical Comparability: The results from the predicate device (current AC2 assay) on remnant clinical samples served as the de-facto ground truth for evaluating equivalence. While not "absolute" ground truth (like pathology), it's the standard for substantial equivalence in device modifications.
Clinical Panel Agreement: Expected panel result based on known concentrations of spiked organisms. This is a form of engineered, highly controlled ground truth.

8. The Sample Size for the Training Set

The document does not directly disclose the sample size for the training set. This is a common characteristic of medical device submissions, as the focus is on validation (test set performance) rather than the proprietary details of model training (if applicable to this type of assay, though molecular assays often rely on established biochemical principles rather than "training" in the machine learning sense). The development of the reformulated probe reagent would have involved laboratory optimization and characterization experiments, which are analogous to a "training" phase.

9. How the Ground Truth for the Training Set Was Established

Since this is a molecular diagnostic assay and not an AI/ML product requiring extensive "training" data, the concept of "ground truth for the training set" isn't directly applicable in the same way. The development process for the reformulated probe would have involved:

Understanding the genetic variants: Identifying the specific mutations in the C. trachomatis variants (e.g., FI-nvCT) that caused detection issues with the previous probe.
Rational design: Designing new probe sequences that would bind effectively to both the original target and the new variants.
Iterative testing and optimization: Extensive laboratory testing of various probe formulations using cultured organisms, synthetic nucleic acids, and potentially some clinical samples to ensure desired performance (sensitivity, specificity, variant detection) during the development phase. This iterative process, guided by known target sequences and laboratory results, serves as the "ground truth" for optimizing the assay components.

In summary, the submission demonstrates that the modified Aptima Combo 2 Assay maintains or improves its performance compared to the predicate device, particularly in its ability to detect emerging CT variants, without compromising its overall accuracy or introducing new risks. The studies are analytical and comparative, appropriate for a molecular diagnostic device modification rather than an imaging AI solution.

Summary

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Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). The logo consists of two parts: the symbol of the Department of Health & Human Services on the left and the FDA acronym along with the full name of the agency on the right. The FDA part of the logo is in blue, with the acronym in a square and the full name written out next to it.

May 17, 2020

Hologic, Inc. Anila Tarte Regulatory Affairs Specialist 10210 Genetic Center Drive San Diego, California 92121

Re: K200866

Trade/Device Name: Aptima Combo 2 Assay (Panther System) and Aptima Combo 2 Assay (Tigris System) Regulation Number: 21 CFR 866.3393 Regulation Name: Nucleic Acid Detection System for Non-Viral Microorganism(s) Causing Sexually Transmitted Infections. Regulatory Class: Class II Product Code: QEP, LSL, MKZ Dated: March 30, 2020 Received: April 1, 2020

Dear Anila Tarte:

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. 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 located 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.

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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) for devices or post-marketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-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 4, Subpart A) for combination products; 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 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.

Steven Gitterman, M.D., Ph.D. Deputy Director Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

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HOLOGIC®

510(k) SUMMARY

Aptima Combo 2® Assay (Panther® and Tigris® DTS® System)

I. SUBMITTER

Hologic, Inc. 10210 Genetic Center Drive San Diego, CA 92121

Contact Information:

	Anila Tarte
	Regulatory Affairs Specialist
Phone:	858-410-8055
Email:	anila.tarte@hologic.com

Date Prepared: May 14, 2020

II. DEVICES

Proprietary Name:	Aptima Combo 2® Assay (Panther® System)
Classification Name:	Nucleic Acid Detection System for Non-Viral Microorganism(s)Causing Sexually Transmitted Infections
Regulation Number:	866.3393
Regulatory Class:	Class II
Product Code:	QEP
Subsequent Product Code:	MKZ, LSL
Proprietary Name:	Aptima Combo 2® Assay (Tigris® DTS® System)
Classification Name:	Nucleic Acid Detection System for Non-Viral Microorganism(s)Causing Sexually Transmitted Infections
Regulation Number:	866.3393
Regulatory Class:	Class II
Product Code:	OEP

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III. PREDICATE DEVICE

The predicate device is the Aptima Combo 2 Assay on Panther and Tigris Systems (K200436; cleared 03/24/2020). The predicate device has not been subject to a design-related recall.

IV. DEVICE DESCRIPTIONS

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second CT probe, complementary to a unique region of the existing CT amplicon. This tandem probe provides detection coverage for the variant strains of C. trachomatis that emerged in 2019. The labeled probes combine with amplicon to form stable hybrids. The Selection Reagent differentiates hybridized from unhybridized probe, eliminating the generation of signal from unhybridized probe. During the detection step, light emitted from the labeled hybrids is measured as photon signals in a luminometer, and are reported as Relative Light Units (RLU). In DKA, differences in the kinetic profiles of the CT and GC labeled probes allow for the differentiation of signal; kinetic profiles are derived from measurements of photon output during the detection read time. The chemiluminescent detection for CT signal has very rapid kinetics and has the "flasher" kinetic type. The chemiluminescent detection reaction for GC signal is relatively slower and has the "glower" kinetic type. Assay results are determined by a cut-off based on the total RLU and the kinetic curve type.

V. DESCRIPTION OF DEVICE MODIFICATION

The clearance of this Special 510(k) application supports a change in formulation to the Probe reagent contained in the Aptima Combo 2 assay. Reformulation of the Probe reagent was necessary to detect recently emerged variants of Chlamydia trachomatis (CT) that were discovered outside of the U.S. using the Panther System or Tigris System. The updated version of the Aptima Combo 2 Assay (termed "updated AC2 assay") includes dual (redundant) CT detection probe, which not only identifies all recent variants of CT, but is also intended to provide diagnostic protection against future genetic variants within the AC2 probe region.

Table 1: Aptima Combo 2 Assay - Catalog Numbers
-------------------------------------------------	--

Kit Description	Current KitCat. No.	Updated KitCat. No.
Aptima Combo 2 Assay, 100-Test Kit (Panther System)	302923	PRD-05576
Aptima Combo 2 Assay, 250-Test Kit (Panther System)	303094	PRD-05571
Aptima Combo 2 Assay, 250-Test Kit (Tigris System)	301130	PRD-05572
Aptima Combo 2 Assay, 1000-Test Kit (Tigris System) *	301130B	PRD-05572B

AC2 for Tigris 1000-Test Kit is packaged to contain four 250-Test Kits

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VI. INDICATIONS FOR USE

Intended Use - Aptima Combo 2 Assay (Panther)

'Patient-collected vaginal swab specimens are an option for screening women when a pelvic exam is not otherwise indicated. The Aptima Multitest Swab Specimen Collection Kit has not been evaluated for home use.

Intended Use - Aptima Combo 2 Assay (Tigris)

IPatient-collected vaginal swab specimens are an option for screening women when a pelvic exam is not otherwise indicated. The Aptima Multitest Swab Specimen Collection Kit is not for home use.

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VII. COMPARISON OF TECHNOLOGICAL CHARACTERISTICS WITH THE PREDICATE DEVICES

A comparison of the subject device to the predicate devices is summarized in Table 2 (AC2 assay on Panther) and Table 3 (AC2 assay on Tigris). Use of the updated AC2 assay does not change the principles of procedure, intended use, or primary technological characteristics. The similarities and differences between the subject and predicate devices are further discussed following the substantial equivalence tables. This discussion is the same for each assay.

Table 2: Comparison Between Predicate Device and Subject Device - AC2 Assay on the Panther System

Item	Predicate DeviceAC2 Assay (Panther)K200436	Subject Device AC2Assay (Panther)K200866
TechnologyPrinciple ofOperation	Target Capture (TC), Transcription-MediatedAmplification (TMA), HybridizationProtection Assay (HPA)	Same
Platform	Automated Panther System	Same
Function	Detection and differentiation of rRNA fromChlamydia trachomatis and Neisseriagonorrhoeae	Same
OrganismsDetected	Chlamydia trachomatis (CT) and/orNeisseria gonorrhoeae (GC)	Same
PatientPopulation	Symptomatic and asymptomatic individuals	Same
Intended Use	The Aptima Combo 2 Assay is a targetamplification nucleic acid probe test thatutilizes target capture for the in vitroqualitative detection and differentiation ofribosomal RNA (rRNA) from Chlamydiatrachomatis (CT) and/or Neisseriagonorrhoeae (GC) to aid in the diagnosis ofchlamydial and/or gonococcal urogenitaldisease using the Panther® System asspecified On the Panther System, the assaymay be used to test the following specimensfromsymptomatic and asymptomatic individuals:clinician-collected endocervical, vaginal,throat, rectal, and male urethral swabspecimens clinician-collected gynecological	The Aptima Combo 2® Assay is a targetamplification nucleic acid probe test thatutilizes target capture for the in vitroqualitative detection and differentiationof ribosomal RNA (rRNA) fromChlamydia trachomatis (CT) and/orNeisseria gonorrhoeae (GC) to aid inthe diagnosis of chlamydial and/orgonococcal disease using the Panther®System as specified. On the PantherSystem, the assay may be used to testthe following specimens fromsymptomatic and asymptomaticindividuals: clinician-collectedendocervical, PreservCyt® Solutionliquid pan specimens vaginal throat
Same*

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Item	Predicate DeviceAC2 Assay (Panther)K200436	Subject Device AC2Assay (Panther)K200866
	specimens collected in the PreservCyt®Solution, patient-collected vaginal swabspecimens,1 and female and male urinespecimens.	rectal, and male urethral swabspecimens, patient-collected vaginalswab specimens,1 and female and maleurine specimens.
	1Patient-collected vaginal swab specimens are anoption for screening women when a pelvic examis not otherwise indicated. The Aptima MultitestSwab Specimen Collection Kits have not beenevaluated for home use.	1Patient-collected vaginal swab specimensare an option for screening women when apelvic exam is not otherwise indicated. TheAptima Multitest Swab Specimen CollectionKit has not been evaluated for home use.

Edits to the intended use were made for clarification purposes only. Specimen types were aligned and/or grouped into respective clinician-collected and patient-collected specimen types

	Table 3: Comparison Between Predicate Device and Subject Device - AC2 Assay
on the Tigris System

Item	Predicate DeviceAC2 Assay (Tigris)K200436	Subject DeviceAC2 Assay (Tigris)K200866
TechnologyPrinciple ofOperation	Target Capture (TC), Transcription-MediatedAmplification (TMA), Hybridization Protection Assay(HPA)	Same
Platform	Automated Tigris System	Same
Function	Detection and differentiation of rRNA from Chlamydiatrachomatis and Neisseria gonorrhoeae	Same
OrganismsDetected	Chlamydia trachomatis (CT) and/or Neisseriagonorrhoeae (GC)	Same
PatientPopulation	Symptomatic and asymptomatic individuals	Same
Intended Use	The Aptima Combo 2 Assay is a target amplificationnucleic acid probe test that utilizes target capture for the invitro qualitative detection and differentiation of ribosomalRNA (rRNA) from Chlamydia trachomatis (CT) and/orNeisseria gonorrhoeae (GC) to aid in the diagnosis ofchlamydial and/or gonococcal urogenital disease using theTigris® DTS® Automated Analyzer. On the Tigris DTSsystem, the assay may be used to test the followingspecimens from symptomatic individuals: clinician-collected endocervical, vaginal and male urethral swabspecimens; and female and male urine specimens. Theassay may be used to test the following specimens fromasymptomatic individuals: clinician-collectedendocervical, vaginal and male urethral swab specimens;patient-collected vaginal swab specimens1; and female and	Same

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Item	Predicate DeviceAC2 Assay (Tigris)K200436	Subject DeviceAC2 Assay (Tigris)K200866
	male urine specimens. The assay is also intended for usewith the testing of gynecological specimens, from bothsymptomatic and asymptomatic patients, collected in thePreservCyt® Solution.
	1Patient-collected vaginal swab specimens are an option for screeningwomen when a pelvic exam is not otherwise indicated. The vaginal andmultitest swab specimen collection kits are not for home use.

Similarities

Both the predicate and subject devices utilize the same technology and principles of operation, mechanisms of action, and run on the same automated instrument systems. There are no changes to the assay kit configuration, intended use, results interpretation, or existing performance of the assay. Additionally, the proposed changes do not affect the existing Aptima Controls Kit, Aptima ancillary or collection kits, or the software and hardware associated with the use of the Panther or Tigris systems.

Differences

Changes to the user interface are minimal and include updated packaging and labeling to differentiate between the current AC2 assay kit and the updated AC2 assay kit. Package insert changes include new kit catalog numbers and updates to the Analytical Performance section demonstrating detection of all recent CT variants using the updated AC2 assay.

VIII. Design Control Activities

Hologic's overall product development activities are conducted per procedure, 'Product Development Procedure' which is in conformance with the design control requirements as specified in 21 CFR 820.30. Verification testing was performed to confirm clinical comparability between the current Probe and reformulated Probe reagents. The completed verification studies demonstrate that the reformulated Probe reagent does not impact assay performance, assay safety and effectiveness, and confirms that the modified assay meets the design input requirements.

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Hologic risk analysis activities are conducted per Product Safety Risk Management Procedure which is in conformance with ISO 14971:2007. Based on the results of the risk analysis and verification activities, and in accordance with ISO 14971:2007, all risks are reduced as far as possible and meet the pre-defined acceptability criteria. There were no hazards that fell within the "Undesirable" or "Unacceptable" residual risk regions. The device modifications do not introduce any new hazards or increase the overall residual risk as compared to the currently marketed products.

IX. ASSAY PERFORMANCE

Performance of the updated AC2 Assay was evaluated. The result of this evaluation demonstrated that existing performance and claims of the assay were not impacted due to the reformulated Probe reagent.

Brief Description of Non-Clinical Data

The following analytical (non-clinical) studies were conducted to support the clearance of the updated AC2 assay on the Panther System and Tigris System.

Limit of Detection – Panther and Tigris

The analytical sensitivity for the Finnish variant of Chlamydia trachomatis (FI-nvCT) was determined by testing dilutions of in vitro transcripts in negative urine specimens, negative ThinPrep specimens, and simulated swab matrix specimens. Thirty replicates of each dilution were tested on both the Panther system and Tigris system with each of three reagent lots of the updated AC2 assay for a total of 90 replicates per specimen type. The analytical sensitivity was determined to be less than one IFU per assay in urine, ThinPrep, and simulated swab matrix specimens. The detection capabilities of the updated version of the AC2 assay were confirmed across multiple CT variants.

Clinical Comparability

The clinical specimen agreement between the current version and updated version of the AC2 assay was evaluated using remnant swab specimens collected from patients undergoing CT

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and/or GC screening. A single replicate of each specimen was tested with both the current version and the updated version of the AC2 assay on the Panther System.

Table 4 and Table 5 show clinical comparison results for the CT and GC positive and negative percent agreement, respectively. The overall agreement was >99.0% for both CT and GC.

		Current AC2 Assay
		CT Positive	CT Negative
Updated AC2Assay	CT Positive	49	3
	CT Negative	0	273
Positive Percent Agreement (95% C.I.): 100% (92.7% - 100%)
Negative Percent Agreement (95% C.I.): 98.9% (96.9% - 99.6%)

Table 4: Clinical Specimen Comparison Results - CT

Table 5: Clinical Specimen Comparison Results - GC

		Current AC2 Assay
		GC Positive	GC Negative
Updated AC2Assay	GC Positive	47	1
	GC Negative	0	275
Positive Percent Agreement (95% C.I.): 100% (92.4% - 100%)
Negative Percent Agreement (95% C.I.): 99.6% (98.0% - 99.9%)

CT/GC Clinical Sample Agreement

The clinical panel agreement study evaluated the equivalence between the current and updated versions of the AC2 assay using 20 prepared CT/GC clinical panels containing 0 to 2,500 IFU/mL of wild type CT, 0 to 500 IFU/mL of FI-nvCT, and 0 to 125,000 CFU/mL of GC in urine specimens. Each of the 20 panels were tested in triplicate in two runs per day, on three Panther systems, by two operators, using three lots of reagents over seven days.

The results show 100% (97.6-100%) total CT and GC agreement to the expected panel result for the updated AC2 assay. Results also show 100% (97.6-100%) total CT and GC agreement to the expected panel result for the current AC2 assay, with the exception of the moderate (0.2 IFU/mL) FI-nvCT only panel, which had 98.2% (93.5-99.5%) CT agreement and 99.1% (94.9-99.8%) GC agreement. The percent agreement to the expected result for the detection of wild

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type CT and GC is comparable between the current AC2 assay and the updated AC2 assay. In addition, the observed variability of the current AC2 and the updated AC2 assays was comparable between instruments, lots, operators, days, and runs.

Microorganism Cross-Reactivity and Microbial Interference

The analytical specificity and microbial interference of the updated version of the AC2 assay was evaluated using 86 microorganisms consisting primarily of viral, bacterial, and yeast strains. Each pool of microorganisms was tested with and without the presence of FI-nvCT in vitro transcripts at a concentration of 3x LoD. None of the microorganisms tested were found to have an impact on the detection capabilities or analytical specificity of the updated version of the AC2 assay.

Pool ID	Microorganism	Pool ID	Microorganism
0	SVSM (Control)
1	Acinetobacter lwoffiiActinomyces israeliiAlcaligenes faecalisAnaerococcus vaginalis	12	Proteus vulgarisShigella dysenteriaeShigella flexneriShigella sonneri
2	Arcanobacterium haemolyticumAtopobium vaginaeBacteroides fragilisBacteroides oralis	13	Stenotrophomonas maltophiliaStreptococcus agalactiaeStreptococcus anginosusStreptococcus pyogenes
3	Bifidobacterium adolescentisBordatella parapertussisCampylobacter jejuniCampylobacter rectus	14	Ureaplasma parvumUreaplasma urealyticumVeillonella parvulaBurkholderia cepacia
4	Citrobacter koseriCorynebacterium diptheriaCorynebacterium genitaliumCorynebacterium pseudodiptheriticum	15	Clostridium difficilePrevotella biviaCandida albicansCryptococcus neoformans
5	Eggerthella lentaEnterobacter cloacaeEnterococcus faecalisEscherichia coli	16	Entamoeba histolyticaGiardia lambliaPentatrichomonas hominisTrichomonas vaginalis

Table 6: Cross-Reactivity Microorganisms

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Pool ID	Microorganism	Pool ID	Microorganism
6	Fusobacterium necrophorumFusobacterium nucleatumGardnerella vaginalisHelicobacter pylori	17	Adenovirus Type 07ACoronavirus 229ECoxsackievirus B3Echovirus Type 11
7	Haemophilus ducreyiHaemophilus parahaemolyticusHaemophilus parainfluenzaeKlebsiella pneumoniae	18	Enterovirus Type 68Epstein-Barr virusHepatitis B VirusHepatitis C Virus
8	Lactobacillus acidophilusLegionella (Tatlockia) micdadeiLegionella jordanisLeptotrichia buccalis	19	HIVHPV 16 (SiHa cells)HPV 18 (HeLa cells)Human Metapneumovirus Type 20
9	Listeria monocytogenesMegasphaera Type 1Mobiluncus curtisiiMoraxella catarrhalis	20	HSV IHSV IIInfluenza A H3N2Influenza B Massachusetts/2/12
10a	Mycoplasma genitaliumMycoplasma hominisMycoplasma pneumoniae	21	Norovirus Group IIRespiratory Syncytial virus Type BRhinovirus A16
10b	Neisseria gonorrhoeae
11	Peptostreptococcus microsPropionibacterium acnesStaphylococcus aureusStaphylococcus epidermidis	22	Chlamydia pneumoniaeChlamydia psittaciChlamydia psittaci

X. CONCLUSIONS

A comparison of the intended use, technological characteristics, and results from the analytical performance studies demonstrate that the updated AC2 assay on the Panther and Tigris systems performs comparably to the predicate device.

Regulation Number and Section

§ 866.3393 Device to detect nucleic acids from non-viral microorganism(s) causing sexually transmitted infections and associated resistance marker(s).

(a)
Identification. A device to detect nucleic acids from non-viral microorganism(s) causing sexually transmitted infections and associated resistance marker(s) is an in vitro diagnostic device intended for the detection and identification of nucleic acids from non-viral microorganism(s) and their associated resistance markers in clinical specimens collected from patients suspected of sexually transmitted infections. The device is intended to aid in the diagnosis of non-viral sexually transmitted infections in conjunction with other clinical and laboratory data. These devices do not provide confirmation of antibiotic susceptibility since mechanisms of resistance may exist that are not detected by the device.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use for the labeling required under § 809.10 of this chapter must include a detailed description of targets the device detects, the results provided to the user, the clinical indications appropriate for test use, and the specific population(s) for which the device is intended.
(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 discussion of the performance characteristics of the device for all claimed specimen types based on analytical studies, including Limit of Detection, inclusivity, cross-reactivity, interfering substances, competitive inhibition, carryover/cross contamination, specimen stability, within lab precision, and reproducibility, as appropriate;
(iii) Detailed descriptions of the test procedure, the interpretation of test results for clinical specimens, and acceptance criteria for any quality control testing;
(iv) Limiting statements indicating 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) Reliable 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
(D) If appropriate (
e.g., recommended by the Centers for Disease Control and Prevention, by current well-accepted clinical guidelines, or by published peer reviewed research), that the clinical performance is inferior in a specific clinical subpopulation or for a specific claimed specimen type; and(v) If the device is intended to detect antimicrobial resistance markers, limiting statements, as appropriate, indicating that:
(A) Negative results for claimed resistance markers do not indicate susceptibility of detected microorganisms, as resistance markers not measured by the assay or other potential mechanisms of antibiotic resistance may be present;
(B) Detection of resistance markers cannot be definitively linked to specific microorganisms and the source of a detected resistance marker may be an organism not detected by the assay, including colonizing flora;
(C) Detection of antibiotic resistance markers may not correlate with phenotypic gene expression; and
(D) Therapeutic failure or success cannot be determined based on the assay results, since nucleic acid may persist following appropriate antimicrobial therapy.
(4) Design verification and validation must include:
(i) Detailed device description documentation, including methodology from obtaining sample to result, design of primer/probe sequences, rationale for target sequence selection, and computational path from collected raw data to reported result (
e.g., how collected raw signals are converted into a reported result).(ii) Detailed documentation of analytical studies, including, Limit of Detection, inclusivity, cross-reactivity, microbial interference, interfering substances, competitive inhibition, carryover/cross contamination, specimen stability, within lab precision, and reproducibility, as appropriate.
(iii) Detailed documentation and performance results from a clinical study that includes prospective (sequential) samples for each claimed specimen type and, when determined to be appropriate by FDA, additional characterized clinical samples. The study must be performed on a study population consistent with the intended use population and compare the device performance to results obtained from FDA accepted comparator methods. Documentation from the clinical studies must include the clinical study protocol (including a predefined statistical analysis plan) study report, testing results, and results of all statistical analyses.
(iv) A detailed description of the impact of any software, including software applications and hardware-based devices that incorporate software, on the device's functions.