The Aptima Neisseria gonorrhoeae (GC) Assav is an in vitro qualitative nucleic acid amplification (NAAT) the detection of ribosomal RNA (rRNA) from Neisseria gonorrhoeae (GC) to aid in the diagnosis of gonocccal urogenital disease using the Panther System. The assay may be used to test male urine specimens from symptomatic and asymptomatic individuals.

Device Description

The Aptima GC assay is a target amplification nucleic acid probe test for in vitro qualitative detection of ribosomal RNA (rRNA) from Neisseria gonorrhoeae (GC). The Aptima Neisseria gonorrhoeae Assay combines the technologies of target capture, transcription-mediated amplification (TMA), and hybridization protection assay (HPA). Specimens are collected and transferred into their respective specimen transport tubes. The transport solution in these tubes releases the rRNA target and protects it from degradation during storage. When the Aptima Neisseria gonorrhoeae Assay is performed in the laboratory, the target rRNA molecule is isolated from the specimens by use of a capture oligomer via target capture that utilizes magnetic microparticles. The capture oligomer contains a sequence complementary to a specific region of the target molecule as well as a string of deoxyadenosine residues. During the hybridization step, the sequence specific region of the capture oligomer binds to a specific region of the target molecule. 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 micro particles, including the captured target molecule 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 Hologic TMA reaction replicates a specific region of the 16S rRNA from GC via DNA intermediates. A unique set of primers is used for the target molecule. Detection of the rRNA amplification product sequences (amplicon) is achieved using nucleic acid hybridization. A single-stranded chemiluminescent DNA probe, which is complementary to a region of the target amplicon, is labeled with an acridinium ester molecule. The labeled DNA probe combines with amplicon to form stable RNA:DNA 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 RNA:DNA hybrids is measured as photon signals in a luminometer, and are reported as Relative Light Units (RLU). The device reagents are identical to the Aptima Neisseria gonorrhoeae Assay reagents for use on the Tigris DTS system but are intended for use on the Panther system with different specimen type indications. The Panther and Tigris DTS systems use the same principles of operation.

AI/ML Overview

Here's a breakdown of the acceptance criteria and study details for the Aptima Neisseria gonorrhoeae Assay, based on the provided FDA 510(k) summary:

This device is an in vitro qualitative nucleic acid amplification test (NAAT) for the detection of Neisseria gonorrhoeae (GC) rRNA to aid in the diagnosis of gonococcal urogenital disease in male urine specimens using the Panther System. It does not appear to involve AI assistance or human reader studies.

1. Acceptance Criteria and Reported Device Performance

The provided document does not explicitly state pre-defined acceptance criteria for clinical performance (sensitivity and specificity) as a set of numerical thresholds. Instead, it states that the study data demonstrate that performance of the Aptima Neisseria gonorrhoeae Assay on the Panther system is substantially equivalent to that of currently FDA-cleared assays for male urine specimens.

However, analytical studies do present performance metrics that implicitly act as acceptance criteria, for example, for Limit of Detection (LoD), precision, and specificity (no interference).

Here's a summary of reported device performance based on the provided text:

Table of Reported Device Performance

Metric	Acceptance Criteria (Implicit from Study Design/Results)	Reported Device Performance
Clinical Performance	Substantially equivalent to currently FDA-cleared assays for male urine specimens in terms of sensitivity and specificity.	Overall Sensitivity (Prosp. Clinical Study): 98.4% (95% CI: 94.4%-99.6%) for male urine. Overall Specificity (Prosp. Clinical Study): 99.9% (95% CI: 99.7%-100%) for male urine.
Analytical Sensitivity (LoD)	Target concentration detectable in 95% of replicates for urine specimens.	LoD for ATCC 49226: 0.04933 CFU/mL LoD for WHO X/NCTC 13820: 0.03986 CFU/mL (Note: Initial general statement indicated LoD below 125 CFU/mL, but specific strain data gives more precise values).
Within-Lab Precision	100% agreement to expected results for positive and negative panel members. CV and SD values for RLU.	Agreement to Expected Result: 100% for all four panel members (Low, Moderate, High Positive, Negative). Total CV for RLU: Low positive (19.43%), Moderate positive (16.99%), High positive (15.29%), Negative (92.04%). (Individual SD and CV components for Lot, Instrument, Operator, Day, Run, Within-Run also provided).
Analytical Specificity	No interference from 155 culture isolates (87 urogenital, 68 phylogenetic) or various external/internal substances.	No interference observed with any of the tested substances or organisms (including 155 isolates, various bodily fluids, and common medications/substances).
Carryover	Low overall carryover rate.	Overall Carryover Rate: 0.07% (95% CI: 0.02–0.25%).
Run-Size Validity	100% agreement with expected results; no front-to-back positional effects.	Negative and Positive panel member results produced 100% agreement with expected results, with no difference in performance between the front and back of the runs.
Control Validity	Run controls meet performance criteria and properly control run validity over the 24-hour timeframe.	Acceptance criteria for this study were met. Control RLUs were within the expected range for 0 and 30 hours. Run controls met performance criteria.
Control Effectiveness	GC Controls correctly predict sample results under fault conditions not detected by instrument process controls.	GC Controls correctly predicted sample results in 8 out of 8 tested conditions. Results met acceptance criteria.
Environmental Conditions	Meets performance requirements at specified temperature (15-30°C) and humidity (20-85%) limits.	Negative and Positive panel member results produced 100% agreement with expected results under environmental limits. Device meets performance requirements.
Reproducibility	100% agreement with expected results for all panel members across sites, operators, and reagent lots. CV and SD values for RLU.	Agreement to Expected Result: 100% for all panel members. Total CV for RLU: Negative (12.0%), Low Positive (42.4%), Positive (7.5%). (Individual SD and CV components for Sites, Operators, Lots, Runs, Within-Runs also provided).

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

Clinical Test Set:
- Total Enrolled Subjects: 2085 male subjects.
- Evaluable Subjects: 1959 male subjects (126 subjects not evaluable).
- Specimens Included in Performance Analysis: 1958 male urine specimens (one specimen with final GC equivocal result was excluded).
- Data Provenance: Prospective, multi-center clinical study conducted at 11 geographically and ethnically diverse US clinical sites.
  - Sites included obstetrics and gynecology, family planning, and STI clinics.
  - Specimens were collected from symptomatic and asymptomatic men.

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

The document does not explicitly state the number of experts used to establish ground truth for the clinical test set.
The ground truth (Patient Infected Status - PIS) was established using up to 3 FDA-cleared NAATs (Nucleic Acid Amplification Tests). This implies that a consensus or reference standard approach using multiple existing, cleared diagnostic methods was used, rather than individual expert review or pathological examination for individual cases.

4. Adjudication Method for the Test Set

The document states that the Patient Infected Status (PIS) was established using "up to 3 FDA-cleared NAATs". This indicates a composite reference standard approach.
The specific adjudication rule (e.g., 2/3 positive, 3/3 positive) is not detailed, but the use of multiple FDA-cleared methods implies a robust, multi-test consensus for the ground truth.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

No, an MRMC comparative effectiveness study was not done.
This device is an in vitro diagnostic (IVD) assay (a qualitative nucleic acid amplification test), not an imaging AI device that assists human readers. Therefore, there is no "human-in-the-loop" component or an effect size for human readers improving with AI assistance.

6. Standalone Performance

Yes, standalone performance was done.
The described clinical study evaluates the performance of the "Aptima Neisseria gonorrhoeae Assay on the Panther System" directly against a composite reference standard (PIS derived from FDA-cleared NAATs). This is a standalone performance assessment of the algorithm/device itself, without human intervention in the result interpretation.

7. Type of Ground Truth Used

The ground truth used for the clinical test set was a composite reference standard, referred to as "Patient Infected Status (PIS)".
PIS was established by testing male urine specimens with "up to 3 FDA-cleared NAATs". This is a highly robust method, relying on multiple well-validated diagnostic tests to determine the true infection status.

8. Sample Size for the Training Set

The document does not specify the sample size for the training set.
This document is a 510(k) summary for premarket notification, focusing on the validation of the device for its intended use. Information regarding the development and training of the assay (e.g., specific molecular sequences, probe design) is generally proprietary and not included in this type of submission. The focus is on the performance of the final, locked version of the device.

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

The document does not specify how the ground truth for any potential training set was established, nor explicitly mention a distinct training set.
For an IVD such as this, the "training" (or development and optimization) typically involves extensive analytical studies (e.g., primer design, probe specificity, optimization of reaction conditions, LoD determination, cross-reactivity testing) rather than machine learning-style "training data" with a "ground truth" in the same sense as an AI imaging algorithm. The core of the assay relies on well-established molecular biology principles and target identification.

Summary

{0}------------------------------------------------

Image /page/0/Picture/0 description: The image shows the logo of the U.S. Food & Drug Administration (FDA). On the left is the Department of Health & Human Services logo. To the right of that is the FDA logo, which is a blue square with the letters "FDA" in white. To the right of the blue square is the text "U.S. FOOD & DRUG ADMINISTRATION" in blue.

January 26, 2024

Hologic, Inc. Jon Kukowski Regulatory Affairs Specialist 10210 Genetic Center Drive San Diego, California 92121

Re: K231329

Trade/Device Name: Aptima Neisseria gonorrhoeae Assay Regulation Number: 21 CFR 866.3390 Regulation Name: Neisseria Spp. Direct Serological Test Reagents Regulatory Class: Class II Product Code: LSL, QEP Dated: December 28, 2023 Received: December 28, 2023

Dear Jon Kukowski:

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 (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.

{1}------------------------------------------------

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" (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 QS 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).

{2}------------------------------------------------

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 Center for Devices and Radiological Health

Enclosure

{3}------------------------------------------------

Indications for Use

510(k) Number (if known) K231329

Device Name Aptima Neisseria gonorrhoeae Assay

Indications for Use (Describe)

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)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.

DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

Department of Health and Human Services Food and Drug Administration Office of Chief Information Officer Paperwork Reduction Act (PRA) Staff PRAStaff(@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

{4}------------------------------------------------

510(k) Summary

Contact Details

Applicant Name: Hologic, Inc. Applicant Address: 10210 Genetic Center Drive San Diego CA 92121 United States Applicant Contact Telephone: 858-410-8245 Applicant Contact: Jon Kukowski Applicant Contact Email: jonathan.kukowski@hologic.com

Device Name

Device Trade Name: Aptima Neisseria gonorrhoeae Assay Common Name: Neisseria spp. direct serological test reagents Classification Name: DNA-Reagents, Neisseria Regulation Number: 866.3390 Regulatory Class: Class II Product Code: LSL

Legally Marketed Predicate Devices

Predicate #: K063664 Predicate Trade Name: Aptima Neisseria gonorrhoeae Assay Product Code: LSL

Device Description Summary

Clearance of this pre-market application will allow the Aptima Neisseria gonorrhoeae Assay to be performed on the Panther system using male urine specimens.

Specimens are collected and transferred into their respective specimen transport tubes. The transport solution in these tubes releases the rRNA target and protects it from degradation during storage. When the Aptima Neisseria gonorrhoeae Assay is performed in the laboratory, the target rRNA molecule is isolated from the specimens by use of a capture oligomer via target capture

{5}------------------------------------------------

that utilizes magnetic microparticles. The capture oligomer contains a sequence complementary to a specific region of the target molecule as well as a string of deoxyadenosine residues. During the hybridization step, the sequence specific region of the capture oligomer binds to a specific region of the target molecule. 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 micro particles, including the captured target molecule 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 Hologic TMA reaction replicates a specific region of the 16S rRNA from GC via DNA intermediates. A unique set of primers is used for the target molecule. Detection of the rRNA amplification product sequences (amplicon) is achieved using nucleic acid hybridization. A single-stranded chemiluminescent DNA probe, which is complementary to a region of the target amplicon, is labeled with an acridinium ester molecule. The labeled DNA probe combines with amplicon to form stable RNA:DNA 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 RNA:DNA hybrids is measured as photon signals in a luminometer, and are reported as Relative Light Units (RLU).

The device reagents are identical to the Aptima Neisseria gonorrhoeae Assay reagents for use on the Tigris DTS system but are intended for use on the Panther system with different specimen type indications. The Panther and Tigris DTS systems use the same principles of operation.

{6}------------------------------------------------

Intended Use

The Aptima Neisseria gonorrhoeae (GC) Assay is an in vitro qualitative nucleic acid amplification (NAAT) test for the detection of ribosomal RNA (rRNA) from Neisseria gonorrhoeae (GC) to aid in the diagnosis of gonococcal urogenital disease using the Panther System. The assay may be used to test urine from symptomatic and asymptomatic male individuals.

Indications for Use Comparison

This pre-market application is to clear the Aptima Neisseria gonorrhoeae Assay on an additional platform, the Panther system, for use with male urine specimen type indications. The analytical and clinical study results demonstrate that the Aptima Neisseria gonorrhoeae Assay on the Panther system performs comparably to the predicate device in detecting rRNA of GC from the designated specimen type and support a substantial equivalence decision.

Technological Comparison

The Aptima Neisseria gonorrhoeae Assay incorporates the technologies of target capture, Transcription-Mediated Amplification (TMA), and Hybridization Protection Assay (HPA), which are described in previous 510(k) submissions for the Aptima Neisseria gonorrhoeae Assay including K043144, cleared in March 2005. The Aptima Neisseria gonorrhoeae Assay and system technologies remain unchanged in this submission, including biochemical (e.g., in vitro nucleic acid probe test) and physical detection (e.g., chemiluminescence) technologies.

Non-Clinical and/or Clinical Tests Summary & Conclusions

The following studies were conducted to support performance of the Aptima Neisseria gonorrhoeae Assay on the Panther system using male urine specimens.

Analytical Studies

Analytical Sensitivity

Studies were conducted to verify the analytical sensitivity (concentration of analyte giving ≥95% positivity) of the Aptima Neisseria gonorrhoeae Assay on the Panther system by testing panels made by spiking GC organisms into male urine pools, Results showed that the LoD on the Panther was below 125 CFU/mL (250 fg GC rRNA/assay).

{7}------------------------------------------------

Limit of Detection Study

The limit of detection (LoD) was tested and confirmed with sensitivity panels prepared using two strains of GC organisms spiked into pooled negative urine. Testing evaluated one antibiotic susceptible strain; Neisseria gonorrhoeae ATCC 49226 (GP1803), and one antibiotic resistant stain; Neisseria gonorrhoeae WHO X/NCTC 13820 (GP2730). Sensitivity panels were tested on three Panther instruments with two reagent lots. At least 20 replicates were run for each concentration for each reagent lot for each strain. LoD, defined as the target concentration that can be detected in 95% of the replicates tested for urine specimens is 0.04933 CFU/mL for ATCC 49226 and 0.03986 CFU/mL for stain X/NCTC 13820.

Within-lab Precision Study

Precision was measured using positive panel members consisting of Neisseria gonorrhea cells at three different concentrations approximately: 3x LoD (Low Positive), >3x and <5x LoD (Moderate Positive), and >10x LoD (High positive). Cells were spiked into pooled negative urine matrix (Urine mixed 1:1 with Urine Transport Media). The negative panel member consisted of unspiked negative urine matrix. Testing was conducted over the course of at least 20 nonconsecutive days using two lots of reagents on three Panther systems by three operators performing at least two daily runs. Agreement to expected results was 100% for all four panel members. The precision of the signal for each panel is shown in table below.

PanelMember	N.gonorrhoeaeconcentration(CFU/mL)	N	MeankRLU	Agreement toExpected Result	Between-LotSD	Between-LotCV(%)	Between-InstrumentSD	Between-InstrumentCV(%)	Between-OperatorSD	Between-OperatorCV(%)	Between-DaySD	Between-DayCV(%)	Between-RunSD	Between-RunCV(%)	Within-RunSD	Within-RunCV(%)	TotalSD	TotalCV(%)
Lowpositive	0.1431	162	520.1	100%	65.05	12.51	38.39	7.38	0.00	0.00	14.84	2.85	20.11	3.87	62.34	11.99	101.07	19.43
Moderatepositive	0.2417	162	809.4	100%	90.32	11.16	52.74	6.52	7.71	0.95	15.00	1.85	24.09	2.98	84.36	10.42	137.55	16.99
Highpositive	0.5426	162	1529.4	100%	182.18	11.91	78.41	5.13	19.09	1.25	51.45	3.36	0.00	0.00	111.09	7.26	233.86	15.29
Negative	0.000	162	4.2	100%	1.19	28.36	0.87	20.80	0.51	12.28	0.00	0.00	0.00	0.00	3.52	84.17	3.85	92.04
Note: The analysis was performed using SAS PROC MIXED, which applies a lower boundary of 0 to all variance components in the model by default. If a variancecomponent is 0, then SD=0 and CV=0%.

{8}------------------------------------------------

Analytical Specificity

A total of 155 culture isolates were evaluated using the Aptima Neisseria gonorrhoeae Assay on the DTS system or Panther system. These isolates included 87 organisms that may be isolated from the urogenital tract and 68 additional organisms that represent a phylogenetic cross-section of organisms. The tested organisms included bacteria, fungi, yeast, parasites and viruses. All organisms except C. psittaci, C. pneumoniae, U. urealyticum, C. trachomatis and the viruses were tested at 1.0 x 106 cells/ assay. C. psittaci VR601 was tested at 8.0 x 104 cells/assay and C. psittaci VR125 was tested at 1.0 x 105 cells/assay. C. pneumoniae was tested at 4 x 103 cells/assay and U. urealyticum was tested at 6.7 x 106 cells/assay. C. trachomatis was tested at 1.0 x 104 IFU/mL. The viruses were tested as follows: (a) herpes simplex virus I: 2.5 x 104 TCID50/assay, (b) herpes simplex virus II: 6.0 x 104 TCID50/assay, (c) human papillomavirus 16: 2.9 x 106 DNA copies/assay and (d) cytomegalovirus: 4.8 x 105 cells/assay.

Interfering Substances

For a nucleic acid amplification assay, analytical specificity with respect to potentially interfering substances is largely determined by the chemistry of the assay (e.g. oligonucleotide sequences) rather than by the platform. Because the reagents for the Aptima Neisseria gonorrhoeae Assay are identical between DTS and Panther systems, data generated on the DTS system supports the performance of the assay on Panther system.

The following interfering substances were individually spiked into urine specimens: 10% blood, contraceptive jelly, spermicide, moisturizer, hemorrhoidal anesthetic, body oil, powder, antifungal cream, vaginal lubricants, feminine spray and leukocytes (1.0 x 10° cells/mL). The following interfering substances were individually spiked into urine specimens: 30% blood, urine analytes, protein, glucose, ketones, bilirubin, nitrate, urobilinogen, pH 4 (acidic), pH 9 (alkaline), leukocytes (1.0 x 106 cells/mL), cellular debris, vitamins, minerals, acetaminophen, aspirin and ibuprofen. All were tested for potential assay interference in the absence and presence of GC at the estimated rRNA equivalent of 50 GC cells/assay (250 fg/assay). The rRNA equivalents were calculated based on the genome size and estimated DNA:RNA ratio/cell of each organism. No interference was observed with any of the tested substances. No inhibitors of amplification were observed in the Aptima Neisseria gonorrhoeae Assay.

{9}------------------------------------------------

Mucin and seminal fluid were tested on the Panther system by spiking into pooled negative urine negative in the presence of Neisseria gonorrhoeae ATCC 49226. Mucin at (0.1% Vol/vol) and seminal fluid (at 1% vol/vol) did not interfere with the assay.

Carryover Study

A multi-run analytical study was conducted using spiked panels on three Panther Systems. Carryover was assessed using approximately 20% high titer GC samples (>2 x 10 cells/mL, or rRNA equivalent) dispersed between negative samples. Testing was carried out using 5 runs on each of three Panther Systems with a total of 2941 negative samples. The overall carryover rate was 0.07% with a 95% confidence interval of 0.02–0.25%.

Run-Size Validity

This study was to confirm that there are no front-to-back positional effects within a run for the Aptima Neisseria gonorrhoeae Assay on the Panther system. Panels and controls with known concentrations were tested and results were examined for front-to-back effects when compared to expected results. Negative and Positive panel member results produced 100% agreement with the expected results, with no difference in performance between the front and back of the runs.

Control Validity

This study was to demonstrate that Aptima Neisseria gonorrhoeae Assay run controls meet performance criteria and properly control for run validity over the timeframe allowed by the Panther software. The acceptance criteria for this study were met. Results at time-point 0 hour and time-point 30 hour (24 hours + 25%) yielded the expected results for the study and the control RLUs were within the expected range. Run controls met performance criteria and properly control for run validity over the 24-hours control validity timeframe allowed by the Panther software.

Control Effectiveness

This study was to demonstrate that the Aptima Neisseria gonorrhoeae Assay run controls are properly invalidated under fault conditions that are not detected by instrument process controls and that may exist during assay processing. The performance of the GC Controls correctly

{10}------------------------------------------------

predicted the sample results in 8 out of 8 of the conditions tested for the assay when run on the Panther system. Results met the acceptance criteria of the study and demonstrated that performance of the Aptima Neisseria gonorrhoeae Assay controls properly determine run and test validity under fault conditions that are not detected by Panther system process controls.

Environmental Conditions

This study was to demonstrate that the Aptima Neisseria gonorrhoeae Assay on the Panther system meets performance requirements at the limits of Panther system environmental conditions for external ambient temperature (15-30°C) and relative humidity (20-85%) specifications. Target panels at or below the sensitivity claims for the assays were tested. Negative and Positive panel member results produced 100% agreement with the expected results. The Aptima Neisseria gonorrhoeae Assay on the Panther system meets performance requirements at the limits of Panther system environmental conditions for external ambient temperature (15-30°C) and relative humidity (20-85%) specifications.

Reproducibility Study

Aptima Neisseria gonorrhoeae Assay reproducibility was evaluated on the Panther system at two external US laboratories and at Hologic. Testing was performed using two lots of assay reagents and a total of six operators (two at each site, testing was performed over at least six days.

Reproducibility panel members were created using clinical urine specimens in urine transport medium (UTM). The GC positive panel members were created by diluting GC positive clinical urine specimens with volume from pooled GC negative clinical urine specimens to achieve the appropriate targeted RLU results (low RLU positive or positive). The negative panel member was created using pooled GC negative clinical urine specimens.

The agreement with expected results was 100% for all panel members.

The table below shows the signal variability of assay RLU results for each panel member between sites, between operators, between lots, between runs, within runs, and overall (Total). Only samples with valid results were included in the analyses.

{11}------------------------------------------------

PanelMember	TargetRLU(x1000)	N	MeanRLU(x1000)	BetweenSites		BetweenOperators		BetweenLots		BetweenRuns		WithinRuns		Total
				SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)	SD	CV(%)
Negative	<50	108	2.1	0.0	0.0	0.0	1.4	0.0	0.0	0.0	0.0	0.2	11.9	0.2	12.0
LowPositive	100 to<2000	105	926.1	81.2	8.8	0.0	0.0	98.4	10.6	86.2	9.3	361.5	39.0	392.9	42.4
Positive	2000 to<12000	107	6196.9	247.5	4.0	23.7	0.4	315.1	5.1	136.5	2.2	187.4	3.0	463.5	7.5

CV = coefficient of variation. RLU = relative light unit. SD = standard deviation.

Notes: The RLU value reported by the software is the total measured RLU divided by 1000 with the digits after the decimal point

Variability from some factors may be numerically negative. In these cases, SD and CV are shown as 0.0. 'Invalid results were excluded from the analyses for low positive (n=1) panel members.

Clinical Study

A prospective, multi-center clinical study was conducted to establish the clinical performance characteristics of the Aptima Neisseria gonorrhoeae Assay on the Panther system. Specimens were collected from 2085 symptomatic and asymptomatic men enrolled at 11 geographically and ethnically diverse US clinical sites, including obstetrics and gynecology, family planning, and STI clinics. Subjects were classified as symptomatic if symptoms were reported by the subject. Subjects were classified as asymptomatic if the subject did not report symptoms.

One hundred twenty six (126) enrolled subjects were not evaluable (12 were withdrawn and 114 did not have at least one specimen with a valid non-excluded Aptima Neisseria gonorrhoeae Assay result and a conclusive infected status). The average age among evaluable study subjects was 35.6 years (range = 14 to 84 years).

Symptoms were reported in 42.1% (825/1959) of the evaluable subjects.

One first-catch urine specimen was collected from each male subject. All specimens were collected by the subject at the clinical sites.

Specimens were tested with the Aptima Neisseria gonorrhoeae Assay on the Panther system. Specimens with initial equivocal or invalid Aptima GC assay results or instrument processing errors were retested, volume permitting; valid retest results were included in the performance analyses. Male urine specimens were tested with up to 3 FDA-cleared NAATs to establish the patient infected status (PIS).

Performance of the Aptima Neisseria gonorrhoeae Assay was estimated relative to the PIS.

{12}------------------------------------------------

Of the specimens collected, 2042 were processed in valid Aptima Neisseria gonorrhoeae Assay runs, including 85 (4.2%) that had to be retested due to invalid results. Overall, 2030 (99.4%) had final valid results, and 12 (0.6%) had final invalid results and were excluded from the analyses. A total of 1958 male urine specimens from evaluable subjects were included in the analyses comparing Aptima Neisseria gonorrhoeae Assay results to the PIS: one specimen with final GC equivocal result was excluded from the performance analyses. Overall, the calculated sensitivity is 98.4% (95% CI: 94.4%-99.6%) for male urine. Overall specificity estimate is 99.9% (95% CI: 99.7%-100%) for male urine.

The study data demonstrate that performance of the Aptima Neisseria gonorrhoeae Assay on the Panther system is substantially equivalent to that of currently FDA-cleared assays for male urine specimens.

Conclusion

Analytical and clinical studies performed to evaluate male urine specimen type for use with the Aptima Neisseria gonorrhoeae Assay on the Panther system demonstrated that the device performs as intended and performs as safe and as effective compared to the predicate device. All study results demonstrated that the performance of male urine specimens for use with the Aptima Neisseria gonorrhoeae Assay on the Panther system is consistent with current expectations for GC testing, , and that the assay is safe and effective for its intended use.

Regulation Number and Section

§ 866.3390

Neisseria spp. direct serological test reagents.(a)
Identification. Neisseria spp. direct serological test reagents are devices that consist of antigens and antisera used in serological tests to identifyNeisseria spp. from cultured isolates. Additionally, some of these reagents consist ofNeisseria spp. antisera conjugated with a fluorescent dye (immunofluorescent reagents) which may be used to detect the presence ofNeisseria spp. directly from clinical specimens. The identification aids in the diagnosis of disease caused by bacteria belonging to the genusNeisseria, 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.(b)
Classification. Class II (performance standards).