The Aptima Herpes Simplex Viruses 1 & 2 assay (Aptima HSV 1 & 2 assay) is an in vitro diagnostic nucleic acid amplification test (NAAT), using real time transcription-mediated amplification (TMA), for the qualitative detection and differentiation of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) messenger RNA (mRNA) in clinician-collected swab specimens from anogenital skin lesions. The assay is intended for use with swab specimens placed in Aptima specimen transport medium (STM) or in viral transport media (VTM) that is immediately diluted into STM.

The Aptima HSV 1 & 2 assay is intended for use as an aid in the diagnosis of HSV-1 and/or HSV-2 infections in symptomatic male and female patients. The Aptima HSV 1 & 2 assay is indicated for use on the Panther® system.

Device Description

The Aptima Herpes Simplex Virus 1 & 2 assay (Aptima HSV assay) is a nucleic acid amplification test (NAAT) developed for use on the fully automated Panther system that utilizes target capture, transcription mediated amplification (TMA), and real-time detection of HSV-1, HSV-2, and an internal control (IC). The Aptima HSV assay amplifies and detects mRNAs for HSV-1 and HSV-2. These RNAs are expressed from the viral genome during the infection cycle, and are packaged inside HSV-1 and HSV-2 viral particles prior to virus release from infected cells. The Aptima HSV assay therefore detects virus-infected cells and the mature virus particles themselves.

The Aptima HSV assay involves three main steps, which all take place in a single tube on the Panther® system: target capture, target amplification by TMA, and detection of the amplification products (amplicon) by the fluorescent labeled probes (torches). The assay incorporates an IC in every test to monitor targeted nucleic acid capture, amplification and detection.

When the Aptima HSV assay is performed, the targeted viral mRNA and IC are isolated using magnetic microparticles and target-specific capture oligomers, in a process called target capture. The capture oligomers contain sequences complementary to specific regions of the targeted RNA (HSV mRNA or IC) as well as a string of deoxyadenosine residues. During the hybridization step, the sequence-specific regions of the capture oligomers bind to specific regions of the RNA target molecules. The microparticles, including the captured RNA target molecules bound to them, are pulled to the side of the reaction tube using magnets and the supernatant is aspirated. The particles are washed to remove residual specimen matrix that may contain amplification inhibitors. After target capture steps are completed, the specimens are ready for amplification.

Target amplification occurs via TMA, which is a transcription-based nucleic acid amplification method that utilizes two enzymes, MMLV (Moloney murine leukemia virus) reverse transcriptase and T7 RNA polymerase. The reverse transcriptase is used to generate a DNA copy (containing a promoter sequence for T7 RNA polymerase) of the target sequence. T7 RNA polymerase produces multiple copies of RNA amplicon from the DNA copy template. Detection is achieved using single-stranded nucleic acid torches that are present during the amplification of the target and hybridize specifically to the amplicon in real time. Each torch has a fluorophore and a quencher. The quencher suppresses the fluorescence of the fluorophore as it is designed to be in close proximity when not hybridized to the amplicon. When the torch binds to the amplicon, the quencher is moved farther away from the fluorophore and it will emit a signal at a specific wavelength when excited by a light source. More torch hybridizes when more amplicon is present. The increase in fluorescent signal from progressive amplification is detected by fluorometers within the Panther system. The Panther system can detect and discriminate between the three fluorescent signals corresponding to HSV-1, HSV-2 and IC amplification products. The fluorescence (measured in relative fluorescence units [RFU]) is monitored over time to produce a real-time fluorescence emergence curve for each reporter dye. The Panther system software compares the fluorescence emergence curves to fixed cut off times to report results (TTime) for HSV-1, HSV-2 and IC.

AI/ML Overview

Here's an analysis of the acceptance criteria and the study proving the device meets them, based on the provided text:

Acceptance Criteria and Device Performance for Aptima Herpes Simplex Viruses 1 & 2 Assay

1. Table of Acceptance Criteria and Reported Device Performance

The acceptance criteria are implied by the clinical performance targets presented in the study. While explicit pre-defined acceptance thresholds (e.g., "Sensitivity must be >90%") are not directly stated as pass/fail criteria, the reported performance metrics demonstrate the device's capability. For this analysis, I will use the clinical performance study results as the reported device performance against generally expected high standards for diagnostic accuracy.

Note: The document does not explicitly state the pre-defined "acceptance criteria" numerical targets. The reported performance below represents the observed results of the clinical study, which presumably met the internal performance requirements for the manufacturer and FDA review.

Table 1: Acceptance Criteria (Implied) and Reported Device Performance

Metric	Target (Implied Acceptance)	Reported Device Performance (Combined, VTM)	Reported Device Performance (Combined, STM)
HSV-1 Sensitivity	High sensitivity, typically >90% for diagnostic assays.	93.4% (95% CI: 85.5-97.2)	94.7% (95% CI: 87.1-97.9)
HSV-1 Specificity	High specificity, typically >95-98% for diagnostic assays.	99.8% (95% CI: 98.8 - >99.9)	99.6% (95% CI: 98.4-99.9)
HSV-2 Sensitivity	High sensitivity, typically >90% for diagnostic assays.	96.9% (95% CI: 94.0-98.4)	98.4% (95% CI: 96.1-99.4)
HSV-2 Specificity	High specificity, typically >95-98% for diagnostic assays.	97.5% (95% CI: 94.9-98.8)	92.8% (95% CI: 89.1-95.3)
Reproducibility	Consistent results across sites, operators, and reagent lots, especially at low concentrations.	Agreement with expected results generally high, with some variability at concentrations near or below LoD (e.g., 46.3% - 100%).	Agreement with expected results generally high, with some variability at concentrations near or below LoD (e.g., 46.3% - 100%).
Limit of Detection (LoD)	Low detection limit to ensure detection of low viral loads.	HSV-1: 60.6-186.9 TCID50/mL (depending on strain/media)	HSV-2: 18.2-128.8 TCID50/mL (depending on strain/media)
Interfering Substances	No significant impact on assay sensitivity or specificity.	No effect observed for tested substances at specified concentrations.	No effect observed for tested substances at specified concentrations.
Cross-Reactivity	No cross-reactivity with non-target microorganisms.	No evidence of cross-reactivity or microbial interference (except for Streptococcus pneumoniae at 1x10^6 CFU/mL where cross-reactivity was observed).	No evidence of cross-reactivity or microbial interference (except for Streptococcus pneumoniae at 1x10^6 CFU/mL where cross-reactivity was observed).
Co-Infection Detection	Ability to detect both HSV-1 and HSV-2 when present.	100% detection for both HSV-1 and HSV-2 in co-infected panels.	100% detection for both HSV-1 and HSV-2 in co-infected panels.

2. Sample size used for the test set and the data provenance

Clinical Test Set Sample Size:
- Total Subjects: 544 evaluable subjects (195 males and 349 females).
- Evaluated for HSV-1: 528 VTM specimens and 531 STM specimens.
- Evaluated for HSV-2: 533 VTM specimens and 535 STM specimens.
Data Provenance:
- Country of Origin: United States. The study was conducted at 17 US clinical sites.
- Retrospective or Prospective: Prospective. The study is described as a "prospective, multicenter clinical study."

3. Number of experts used to establish the ground truth for the test set and the qualifications of those experts

The document does not specify the number of experts used to establish the ground truth or their specific qualifications (e.g., "radiologist with 10 years of experience").

However, it describes the methods used for the composite reference method:

ELVIS HSV ID and D3 Typing Test system viral culture
A validated bidirectional PCR/sequencing procedure
A third FDA-cleared assay for HSV-1 and HSV-2 was used for final composite reference interpretation when the initial methods disagreed or when PCR/sequencing detected both types.

This suggests that the ground truth was established through a combination of highly reliable laboratory tests, implying a rigorous approach to defining true positive/negative cases, rather than relying solely on individual expert interpretation without further clarification.

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

The document describes an adjudication method for the ground truth:

"A third FDA-cleared assay for HSV-1 and HSV-2, was used to determine the final composite reference interpretation when the ELVIS D3 culture and PCR/sequencing results did not agree on the type of HSV detected or when PCR/sequencing detected both HSV-1 and HSV-2."

This indicates a hierarchical or tie-breaking system rather than a simple 2+1 or 3+1 consensus among human readers. It relies on a "composite reference method" combining results from multiple validated laboratory tests.

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

This is a diagnostic assay for detecting viral RNA, not an imaging device that involves human readers interpreting images with or without AI assistance. Therefore, no MRMC comparative effectiveness study involving human readers with AI assistance was performed or reported in this submission. The device (Aptima HSV 1 & 2 Assay) is designed to provide a direct qualitative result (positive/negative for HSV-1 and/or HSV-2) from a processed specimen.

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

Yes, a standalone performance study was done in the sense that the Aptima HSV 1 & 2 Assay (the "algorithm" in this context) directly processes specimens and generates results without requiring human interpretation for its output. The clinical performance study directly evaluated the accuracy of the assay's results against a composite reference standard. The "human-in-the-loop" would be the clinician collecting the sample and laboratory technicians running the test and reporting the results, but the analytical output itself is determined by the assay system.

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

The ground truth used was a composite reference method combining:

ELVIS HSV ID and D3 Typing Test system viral culture
A validated bidirectional PCR/sequencing procedure
A third FDA-cleared assay for HSV-1 and HSV-2 (used for tie-breaking/discrepancy resolution)

This is a robust form of ground truth based on multiple established laboratory diagnostic methods.

8. The sample size for the training set

The document does not report a sample size for a training set. This is expected for a diagnostic assay of this type, as it's not a machine learning or AI algorithm that requires a separate "training set" in the conventional sense. The assay's design and optimization (e.g., probe sequences, amplification conditions) would have been developed iteratively, but a distinct "training set" for performance evaluation is not applicable here. The analytical studies (LoD, cross-reactivity, etc.) and the clinical performance study represent the validation of the finalized assay.

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

As there is no explicit "training set" in the context of machine learning, this question is not directly applicable. The assay's components and parameters would have been optimized using internal development processes and validated through analytical studies. For these analytical studies (e.g., LoD, cross-reactivity), the "ground truth" (i.e., known-positive or known-negative samples, specific viral strains/concentrations) would have been established through well-characterized laboratory standards, spiked samples, and reference materials.

Summary

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Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002

June 15, 2017

Hologic, Inc. Ron Domingo Regulatory Affairs Manager 10210 Genetic Center Dr. San Diego, California 92121

Re: K162673

Trade/Device Name: Aptima Herpes Simplex Viruses 1 & 2 Assay Regulation Number: 21 CFR 866.3305 Regulation Name: Herpes simplex virus serological assays Regulatory Class: Class II Product Code: OOO Dated: May 16, 2017 Received: May 17, 2017

Dear Mr. Domingo:

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. Isting 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

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as set forth in the quality systems (OS) regulation (21 CFR Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

If vou desire specific advice for your device on our labeling regulation (21 CFR Part 801 and Part 809), please contact the Division of Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm. 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.

You may obtain other general information on your responsibilities under the Act from the Division of Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.

Sincerely,

Stephen J. Lovell –S for

Uwe Scherf, M.Sc., 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) K162673

Device Name Aptima Herpes Simplex Viruses 1 & 2 Assay

Indications for Use (Describe)

Warning

The Aptima HSV 1 & 2 assay is not FDA-cleared for use with cerebrospinal fluid (CSF) or for prenatal screening.

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

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

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510(k) SUMMARY Aptima Herpes Simplex Viruses 1 & 2 Assay

I. SUBMITTER

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

Contact Person:	Ron Domingo, MS, RACRegulatory Affairs Manager
	Ron.Domingo@Hologic.com
	Phone: (858) 410-8167
	Fax: (858) 410-5557

Date Prepared: June 14, 2017

II. DEVICE

	Proprietary Name of Device:	Aptima Herpes Simplex Viruses 1 & 2 Assay
	Common or Usual Name:	Nucleic acid amplification test for Herpes Simplex viruses
	Classification Name:	Herpes simplex virus serological assays
	Regulation Number:	21 CFR 866.3305
	Regulatory Class:	Class II
	Product Code:	OQO

III. PREDICATE DEVICE

The predicate device is the BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Q* Amplified DNA Assays (K103798; cleared March 18, 2011; BD Diagnostics Systems, Sparks, MD).

IV. DEVICE DESCRIPTION

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HSV-1 and HSV-2. These RNAs are expressed from the viral genome during the infection cycle, and are packaged inside HSV-1 and HSV-2 viral particles prior to virus release from infected cells. The Aptima HSV assay therefore detects virus-infected cells and the mature virus particles themselves.

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amplicon is present. The increase in fluorescent signal from progressive amplification is detected by fluorometers within the Panther system. The Panther system can detect and discriminate between the three fluorescent signals corresponding to HSV-1, HSV-2 and IC amplification products. The fluorescence (measured in relative fluorescence units [RFU]) is monitored over time to produce a real-time fluorescence emergence curve for each reporter dye. The Panther system software compares the fluorescence emergence curves to fixed cut off times to report results (TTime) for HSV-1, HSV-2 and IC.

Kit Components

The Aptima HSV assay is provided as a 100 test kit. The following 3 kits are required to perform the Aptima HSV assay on the Panther system:

Aptima Herpes Simplex Viruses 1 & 2 Assay Master Kit: Consists of 2 boxes containing eight reagents packaged by storage conditions (1 refrigerated and 1 room temperature box).
• Aptima Herpes Simplex Viruses 1 & 2 Controls Kit: Consists of 1 box containing a Positive and Negative Control.
Aptima Assay Fluids Kit: This ancillary kit is required to run the assay but is procured . separately. The Aptima Assay Fluids Kit consists of 1 box containing 3 components. The ancillary kit previously received market clearance for use in other commercialized Aptima assays (Ref: K003395).

The Aptima HSV assay utilizes two collection and handling kits, which are sold independently of the assay. The collection kits described below have previously received market clearance for use in other commercialized Aptima assays:

Aptima Multitest Swab Specimen Collection Kit (Ref: K032554) .
. Aptima Specimen Transfer Kit (Ref: K063664, K063451, K043224)

Instrumentation

The Aptima HSV assay has been designed for and validated on the Panther system. The Panther system is an integrated hardware and software system that fully automates all steps of nucleic

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acid testing necessary for diagnostic assays. The instrument provides automation of all assay steps including sample processing, amplification of nucleic acid, detection and quantitation, amplicon inactivation, and data reduction. The "Panther Instrument" refers to the analyzer and associated hardware combined with the software. The "Panther system" refers to the Panther Instrument operated in conjunction with the Aptima HSV Assay. The assay software in the computer workstation directs the analyzer modules to perform each sequential assay step.

The Panther system was previously FDA cleared for use with other Class II Aptima Assays (Ref: K122062 and K111409).

V. INDICATIONS FOR USE

Intended Use

Warning

The Aptima HSV 1 & 2 assay is not FDA-cleared for use with cerebrospinal fluid (CSF) or for prenatal screening.

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VI. COMPARISON OF TECHNOLOGICAL CHARACTERISTICS WITH THE PREDICATE DEVICE

A comparison of the Aptima Herpes HSV assay to the predicate BD ProbeTec Herpes Simplex Viruses (HSV 1 & 2) Q* Amplified DNA Assays (K103798) is summarized in Table 1 (similarities) and Table 2 (differences).

Item	Aptima HSV Assay(Subject Device)K162673	BD ProbeTec Assay(Predicate Device)K103798
Intended Use	The Aptima Herpes SimplexViruses 1 & 2 assay (Aptima HSV1 & 2 assay) is an in vitrodiagnostic nucleic acidamplification test (NAAT), usingreal time transcription-mediatedamplification (TMA), for thequalitative detection anddifferentiation of herpes simplexvirus type 1 (HSV-1) and type 2(HSV-2) messenger RNA (mRNA)in clinician-collected swab specimensfrom anogenital skin lesions. Theassay is intended for use with swabspecimens placed in Aptimaspecimen transport medium (STM)or in viral transport media (VTM)that is immediately diluted intoSTM.The Aptima HSV 1 & 2 assay isintended for use as an aid in thediagnosis of HSV-1 and/or HSV-2infections in symptomatic male andfemale patients. The Aptima HSV 1& 2 assay is indicated for use onthe Panther® system.WarningThe Aptima HSV 1 & 2 assay isnot FDA-cleared for use withcerebrospinal fluid (CSF) or for	The BD ProbeTec HerpesSimplex Viruses (HSV 1 & 2)QX Amplified DNA Assays,when tested with the BD ViperSystem in Extracted Mode, useStrand DisplacementAmplification technology forthe direct, qualitative detectionand differentiation of HerpesSimplex virus type 1 (HSV1)and Herpes Simplex virus type2 (HSV2) DNA in clinician-collected external anogenitallesion specimens. The assaysare indicated for use withsymptomatic individuals to aidin the diagnosis of anogenitalHSV1 and HSV2 infections.Warning: The BD ProbeTecHerpes Simplex Viruses (HSV1 & 2) QX Amplified DNAAssays (HSV QX Assays) arenot FDA cleared for use withcerebrospinal fluid (CSF). Theassays are not intended to beused for prenatal screening orfor individuals under the age of17 years.
Item	Aptima HSV Assay(Subject Device)K162673	BD ProbeTec Assay(Predicate Device)K103798
Qualitative/Quantitative Assay	Qualitative	Qualitative
Controls	Positive and negative controls.Provided as part of the Master Kit.	Per PI, positive and negativecontrols. Provided as part of aMaster Kit.
Patient Population	Symptomatic male and femalepatients	Symptomatic male and femalepatients
Specimen Types	Clinician-collected swab specimensfrom skin lesions in the anogenitalregion.	Clinician-collected externalanogenital lesion specimens.

Table 1: Comparison of Similarities Between Aptima HSV Assay and Predicate Device

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	Table 2: Comparison of Differences Between Aptima HSV Assav and Predicate Device

Item	Aptima HSV Assay(Subject Device)	BD ProbeTec Assay(Predicate Device)
Target	mRNA	DNA
Function	Qualitative detection anddifferentiation of mRNA fromherpes simplex viruses 1 & 2	Qualitative detection anddifferentiation of DNA fromherpes simplex viruses 1 & 2
Technology	The subject assay is comprised of anucleic acid amplification test(NAAT) for use on the automatedPanther system, that utilizes targetcapture to extract viral mRNA frompatient samples, and real-timetranscription mediatedamplification (TMA) and detectionof HSV-1, and HSV-2 mRNAs andan internal control RNA.	The predicate assay iscomprised of an amplificationtest for use on the automatedBD Viper System that utilizesnon-specific extraction of DNAfrom clinical specimens,followed by binding of DNA tomagnetic particles, washing ofthe bound nucleic acid andelution in an amplification-compatible buffer. Whenpresent, HSV1 and/or 2 DNA isdetected by StrandDisplacement Amplification(SDA) of type-specific targetsequences in the presence of afluorescently labeled detectorprobe.

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Item	Aptima HSV Assay(Subject Device)	BD ProbeTec Assay(Predicate Device)
Platform	The Panther system is an in vitrodiagnostic device intended for usein clinical laboratories by trainedlaboratory professionals for theautomation of NAATs. The Panthersystem provides automation of allassay steps, including sampleprocessing, amplification of targetnucleic acid, amplicon detection,data reduction, and ampliconinactivation.	BD Viper System is an in vitrodiagnostic device intended foruse in clinical laboratories bytrained laboratory professionalsfor the automation of NAATs.The BD Viper System providesautomated non-specificextraction and amplification ofDNA from clinical specimens.In Extracted Mode, it usesStrand DisplacementAmplification technology.When present, HSV 1 and/or 2amplicon is detected by StrandDisplacement Amplification.
Specimen Collection	• Aptima Multitest SwabSpecimen Collection Kit (STM)• Aptima Specimen Transfer Kit(for use with specimens collectedin VTM)	• BD ProbeTecQˣ CollectionKit for Endocervical orLesion Specimens• BD Universal ViralTransport Medium (VTM)w/Swab Collection KitCopan Universal TransportMedium w/Swab Collection Kit(same as BD VTM Kit)

VII. PERFORMANCE DATA

The following performance data were provided in support of the substantial equivalence determination.

Brief Description of Analytical (Non-Clinical) Studies

The following analytical studies (non-clinical) were conducted to support the clearance of the Aptima HSV Assay on the Panther system.

Analytical Sensitivity and Limit of Detection (LoD)

The LoD in clinical specimen matrix was determined by testing dilution panels for each of two strains of HSV-1 virus (Macintyre and HF strains) and two strains of HSV-2 viral (MS and G) in

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both Aptima Specimen Transport Media (STM) and Viral Transport Media (VTM) negative clinical pools in the Aptima HSV assay. LoD for the assay was based on results from the reagent lot with the highest concentration for the 95% predicted detection limit. Table 3 contains the probit-predicted 95% LoD values of the Aptima HSV assay for HSV-1 and HSV-2 in both specimen types.

HSV Type/Strain	Specimen Type	LoDTCID50/mL
HSV-1 MacIntyre	STM	60.6
HSV-1 MacIntyre	VTM	186.9
HSV-1 HF	STM	78.9
HSV-1 HF	VTM	159.3
HSV-2 MS	STM	18.2
HSV-2 MS	VTM	28.7
HSV-2 G	STM	18.8
HSV-2 G	VTM	128.8

Table 3: HSV 1 & 2 LoD in VTM and STM

Limit of Detection Verification

LoD of the Aptima HSV 1 & 2 assay was verified using two clinical isolates of HSV-1 and two clinical isolates of HSV-2 from HSV-positive clinical specimens that were cultured and quantitated in-house. Each isolate was tested with the Aptima HSV 1 & 2 assay using 60 replicates each at 1X LoD, 3X LoD, and 10X LoD, see Table 4 for LoD values. Testing was completed in both STM and VTM matrix for all four clinical isolates and was conducted using 3 lots of reagents. All replicates for all clinical isolates at all three concentrations tested were detected by the Aptima HSV 1 & 2 assay, demonstrating that the assay can accurately detect a range of both HSV-1 and HSV-2 isolates at the determined LoD.

HSV Type	Media Type	LoD (TCID50/mL)
HSV-1	STM	78.9
	VTM	186.9
HSV-2	STM	18.8
	VTM	128.8

Table 4: HSV 1 and HSV 2 LoD

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Limit of Detection in Neat STM.

The LoD in neat STM (Aptima Specimen Transport Media) was determined by testing panels that consisted of dilutions of HSV-1 and HSV-2 in vitro RNA transcripts (IVT) and dilutions of HSV-1 and HSV-2 viral particles in neat STM matrix. Through Probit analysis, results show the Aptima HSV Assay has a predicted 95% LoD of 68.3 copies/mL for HSV-1 IVT and 49.4 copies/mL for HSV-2 IVT. The assay has a predicted 95% LoD of 28.9 TCID%ML for HSV-1 MacIntyre virus and 0.54 TCID50/mL for HSV-2 MS virus.

Interfering Substances

Potentially interfering substances listed in Table 4 were tested in the Aptima HSV 1 & 2 assay. Panels were built in STM and evaluated for potential effects on both assay sensitivity and specificity. Sensitivity performance was evaluated separately for both HSV-1 and HSV-2 by spiking viral particles into substance-containing panels at 3X the LoD. HSV-negative panels containing each substance were also evaluated for specificity.

No effect on assay performance was observed in the presence of a representative brand of the following exogenous and endogenous substances tested at the concentrations indicated in Table 5.

Interfering Substance	Final Concentration
Vaginal lubricant	5% V/V
Spermicide/Contraceptive Jelly	4% W/V
Anti-fungal cream	5% W/V
Douche	5% V/V
Feminine Spray	5% W/V
Body Lotion	5% W/V
Powder	5% W/V
Glacial Acetic Acid Wash Solution	5% V/V
Hemorrhoidal Cream	5% W/V
Urine	5% V/V
Whole Blood	0.5% V/V
Leukocytes	$4x10^5$ Cells/mL
Mucus	0.3% W/V
Seminal Fluid	5% V/V
Feces	0.03% W/V
Protein	4% W/V
Antiviral drug (Acyclovir)	5% W/V

Table 5: Potential Interfering Substances

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Co-Infection

A co-infection study was evaluated to assess the ability of the Aptima HSV 1 & 2 assay to detect HSV 1 and HSV 2 analytes when both are present in one specimen. Low and high titer concentrations of HSV-1 viral particles in STM were tested in combination with low, moderate, and high concentrations of HSV-2 virus. Panel composition and concentrations are listed in Table 6. All testing resulted in 100% detection for both HSV-1 and HSV-2.

Panel Member	HSV-1 Concentration	HSV-2 Concentration
HSV-1 Low; HSV-2 Low	86.7 TCID50/mL1	1.62 TCID50/mL2
HSV-1 Low; HSV-2 Moderate	86.7 TCID50/mL1	540 TCID50/mL3
HSV-1 High; HSV-2 Low	28,900 TCID50/mL4	1.62 TCID50/mL2
HSV-1 High; HSV-2 Moderate	28,900 TCID50/mL4	540 TCID50/mL3
HSV-1 Low; HSV-2 High	86.7 TCID50/mL1	9,000 TCID50/mL5

Table 6: Co-infected Panel

¹ 3X LoD HSV-1.

2 3X LoD HSV-2.

3 1000X LoD HSV-2.

4 1000X LoD HSV-1.

5 16667X LoD HSV-2.

Cross-Reactivity

A study was performed to evaluate the cross-reactivity and interference with the Aptima HSV 1 & 2 assay in the presence of non-targeted microorganisms that could be present in clinical specimens. A panel consisting of 48 microorganisms (Table 7) was tested in STM at a test concentration of 1 x 105 units/mL for viruses and 1 x 106 units/mL for all other organisms. Organisms were tested in the absence of HSV or in the presence of either HSV-1 or HSV-2 at 3X LoD. Thirty replicates were tested per panel condition. No evidence of cross-reactivity or microbial interference was observed for any of the 48 test microorganisms tested with the Aptima HSV 1 & 2 assay, at the following concentrations.

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Microorganism	Concentration
Acinetobacter calcoaceticus	1x106 CFU/mL1,2
Acinetobacter lwoffii	1x106 CFU/mL1,2
Actinomyces israelii	1x106 RNA copies /mL2
Adenovirus type 1	1x105 TCID50/mL3
Alcaligenes faecalis	1x106 CFU/mL1
Atopobium vaginae	1x106 RNA copies /mL2
Bacteroides fragilis	1x106 CFU/mL1,2
Bifidobacterium adolescentis	1x106 CFU/mL1,2
BK virus	1x105 DNA copies/mL3
Bordetella bronchiseptica	1x106 CFU/mL1,2
Bordetella pertussis	1x106 CFU/mL1,2
Campylobacter jejuni	1x106 CFU/mL1,2
Candida glabrata	1x106 CFU/mL1,2
Clostridium difficile	1x106 CFU/mL1,2
Clostridium perfringens	1x106 CFU/mL1,2
Corynebacterium genitalium	1x106 CFU/mL1,2
Cryptococcus neoformans	1x106 CFU/mL1,2
Enterobacter aerogenes	1x106 CFU/mL1,2
Enterobacter cloacae	1x106 CFU/mL1,2
Enterococcus faecium	1x106 CFU/mL1,2
Enterococcus faecalis	1x106 CFU/mL1,2
Epstein-Barr virus	1x105 DNA copies/mL3
Escherichia coli	1x106 CFU/mL1,2
Fusobacterium nucleatum	1x106 CFU/mL1,2
Gardnerella vaginalis	1x106 CFU/mL1,2
Haemophilus ducreyi	1x106 CFU/mL1,2
Hepatitis B virus	1x105 IU/mL4,3
Klebsiella pneumoniae	1x106 CFU/mL1,2
Lactobacillus crispatus	1x106 CFU/mL1,2
Moraxella catarrhalis	1x106 CFU/mL1,2
Mycoplasma hominis	1x106 RNA copies /mL2
Mycoplasma orale	1x106 RNA copies /mL2
Neisseria gonorrhoeae	1x106 CFU/mL1,2
Neisseria meningitidis	1x106 CFU/mL1,2
Parvovirus B19	1x105 TCID50/mL3
Prevotella bivia	1x106 CFU/mL1,2
Microorganism	Concentration
Propionibacterium acnes	1x106 CFU/mL1,2
Proteus mirabilis	1x106 CFU/mL1,2
Proteus vulgaris	1x106 CFU/mL1,2
Pseudomonas aeruginosa	1x106 CFU/mL1,2
Staphylococcus aureus	1x106 CFU/mL1,2
Staphylococcus epidermidis	1x106 CFU/mL1,2
Staphylococcus saprophyticus	1x106 CFU/mL1,2
Streptococcus mitis	1x106 CFU/mL1,2
Streptococcus pneumoniae*	1x105 CFU/mL1,2
Streptococcus pyogenes	1x106 CFU/mL1,2
Varicella-zoster virus	1x105 DNA copies/mL3
West Nile virus	1x105 TCID50/mL3

Table 7: Cross-Reactivity and Microbial Interference Panel

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Table 7: Cross-Reactivity and Microbial Interference Panel

1 CFU = Colony Forming Units, 2Procured internally from Hologic, Inc., 3 Obtained from ZeptoMetrix Corporation (Buffalo NY), 4IU=International Units *Cross reactivity was observed in Streptococcus pneumoniae at 1x106 CFU/mL

Specimen Inhibition in Clinical Samples

An evaluation was conducted of the whole system failure rate leading to false negative or invalid results by testing low positive specimens. HSV negative VTM and STM samples were spiked with HSV-1 Macintyre or HSV-2 MS strain cultured virus at a test concentration at 3X LoD. Of 201 STM specimens tested, the false negative rate was 0% (0/201; 95% CI: 0.0-1.9) for HSV-1 and 0.5% (1/201; 95% CI: 0.1-2.8) for HSV-2. Of 202 VTM specimens tested, the false negative rate was 0% (0/202; 95% CI: 0.0-1.9) for HSV-1 and 0% (0/202; 95% CI: 0.0-1.9) for HSV-2.

Reproducibility and Within Laboratory Precision

To evaluate the reproducibility and within laboratory precision of the Aptima HSV assay, one HSV negative panel and six HSV positive panels were tested. The six HSV positive panels were spiked with HSV-1 cultured virus strain (MacIntyre) and/or HSV-2 cultured virus strain (MS). All panel members were made using STM. Three replicates of each panel was tested by three operators in two runs each in three reagent lots over 13 days.

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Results showed that the Aptima HSV assay has acceptable precision and is reproducible with various instruments, reagent lots, and operators.

Brief Description of Clinical Studies

Clinical testing of the Aptima HSV assay on the Panther system included performance and reproducibility testing. Substantial equivalence is based in part on the performance study.

Reproducibility

Aptima HSV 1 & 2 assay reproducibility was evaluated at three external US sites. Testing was performed using three lots of assay reagents and six operators (two at each site). At each site, testing was performed for at least six days. Panel members were created by spiking HSV-1 and/or HSV-2 viral particles into STM. Final HSV-1 concentrations ranged from 0 TCID50mL to 86.96 TCID50/mL and final HSV-2 concentrations ranged from 0 TCID50/mL to 1.63 TCID50/mL.

HSV-negative panel members and panel members containing low (<1 X LoD and 1-2 X LoD) and moderate levels (2-3 X LoD) of HSV-1 and HSV-2 were tested with the Aptima HSV 1 & 2 assay. Agreement with expected results was 100% for HSV-1 and HSV-2 in the negative and moderate positive panel members and < 100% in panel members with concentrations near or below the 95% LoD of the assay in STM spiked with viral particles (Tables 8). The signal variability of the Aptima HSV 1 & 2 assay in the low and moderate positive panel members and in the positive control is summarized in Tables 9 and 10.

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Conc		Target Conc(TCID50/mL)		Expected Result		N	Agreed (n)		Agreement (%)(95% CI)
HSV-1	HSV-2	HSV-1	HSV-2	HSV-1	HSV-2		HSV-1	HSV-2	HSV-1	HSV-2
Neg	Neg	0	0	Neg	Neg	108	108	108	100	100
									(96.6-100)	(96.6-100)
LPos	Neg	28.90	0	Pos	Neg	108	103	108	95.4	100
									(89.6-98.0)	(96.6-100)
Neg	LPos	0	0.54	Neg	Pos	108	108	105	100	97.2
									(96.6-100)	(92.1-99.1)
LPos	MPos	28.90	1.63	Pos	Pos	108	97	108	89.8	100
									(82.7-94.2)	(96.6-100)
MPos	LPos	86.96	0.54	Pos	Pos	108	108	108	100	100
									(96.6-100)	(96.6-100)
HNeg	Neg	3.00	0	Pos	Neg	108	50	108	46.3	100
									(37.2-55.7)	(96.6-100)
Neg	HNeg	0	0.20	Neg	Pos	108	108	86	100	79.6
									(96.6-100)	(71.1-86.1)

Table 8. Reproducibility - Agreement of Aptima HSV 1 & 2 Assay with Expected Results

Cl = confidence interval, Conc = concentration, HNeg = high negative (<1 X LoD), LPos = low positive (1-2 X LoD), MPos = moderate positive (2-3 X LoD), Neg = negative, Pos = positive

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			Between		BetweenOperators/Days¹	BetweenRuns	WithinRuns	Total
VirusConc	N	MeanTTime	SitesSD(%CV)	LotsSD(%CV)
HSV-1
LPos	103	24.68	0(0)	1.63(6.62)	0.75(3.04)	0.54(2.18)	0.88(3.55)	2.07(8.40)
LPos	97	23.91	0(0)	2.18(9.11)	0.86(3.58)	0(0)	1.60(6.71)	2.84(11.87)
MPos	108	22.96	0(0)	1.54(6.69)	0.38(1.65)	0.68(2.96)	0.94(4.11)	1.96(8.55)
HSV-2
LPos	105	25.49	0(0)	0.84(3.30)	0.70(2.74)	0(0)	2.52(9.87)	2.74(10.76)
LPos	108	25.34	0(0)	1.54(6.08)	0.86(3.41)	0.59(2.34)	2.67(10.53)	3.26(12.85)
MPos	108	22.91	0(0)	1.09(4.76)	0.35(1.53)	0.42(1.83)	1.06(4.64)	1.62(7.07)

Table 9. Reproducibility - Signal Variability of the Aptima HSV 1 & 2 Assay

Conc = concentration, CV = coefficient of variation, LPos = low positive (1-2 X LoD), MPos = moderate positive (2-3 X LoD), SD = standard deviation

Note: Variability from some factors may be numerically negative. This can occur if the variability due to those factors is very small. In these cases, SD and CV are shown as 0.

1Between Operators may be confounded with Between Days; therefore, Between Operators and Between Days estimates are combined in Between Operators/Days.

			BetweenSites	BetweenLots	BetweenOperators/Days1	Within Days	Total
		MeanTTime	SD(%CV)	SD(%CV)	SD(%CV)	SD(%CV)	SD(%CV)
Virus	N	20.11	0	1.30	0.40	1.09	1.75
HSV-1	36		(0)	(6.48)	(1.99)	(5.42)	(8.68)
HSV-2	36	22.16	0	1.61	0.71	1.38	2.24
			(0)	(7.27)	(3.21)	(6.22)	(10.09)

Table 10. Reproducibility - Signal Variability of the Aptima HSV 1 & 2 Assay in the Positive Control

CV = coefficient of variation, SD = standard deviation

Note: Variability from some factors may be numerically negative. This can occur if the variability due to those factors is very small. In these cases, SD and CV are shown as 0.

1Between Operators may be confounded with Between Days; therefore, Between Operators and Between Days estimates are combined in Between Operators/Days.

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Clinical Performance Study

A prospective, multicenter clinical study was conducted to establish the performance characteristics of the Aptima HSV 1 & 2 assay. A total of 544 evaluable subjects (195 males and 349 females) with active anogenital lesions were enrolled from 17 US clinical sites, including family planning, dermatology, pediatrics/adolescent, sexually transmitted infection, private practice, public health clinics, hospitals, universities, and clinical research sites. Two (2) swab specimens were collected from a single lesion from each subject: one was collected with a swab from a commercially available VTM collection kit and one was collected with a swab from the Aptima Multitest Swab Specimen Collection Kit (STM). Specimens were processed in accordance with the appropriate package insert instructions. Aptima HSV 1 & 2 assay testing was performed at 3 external sites. The performance of the Aptima HSV-1 & 2 assay was evaluated relative to a composite reference. The composite reference method used the results from ELVIS HSV ID and D3 Typing Test system viral culture and a validated bidirectional PCR/sequencing procedure. A third FDA-cleared assay for HSV-1 and HSV-2, was used to determine the final composite reference interpretation when the ELVIS D3 culture and PCR/sequencing results did not agree on the type of HSV detected or when PCR/sequencing detected both HSV-1 and HSV-2. The performance of the Aptima HSV 1 & 2 assay was estimated for detection of HSV-1 and HSV-2 separately using each specimen type. For HSV-1, of the 544 evaluable subjects, 528 VTM and 531 STM specimen types resulted in evaluable results for HSV-1 analysis (16 VTM and 13 STM were reported as unknown composite reference interpretation or withdrawn).

For HSV-2, of the 544 evaluable subjects, 533 VTM and 535 STM specimen types produced evaluable results for HSV-2 analysis (11 VTM and 9 STM were reported as unknown composite reference interpretation or withdrawn).

The performance of Aptima HSV 1 & 2 assay compared to the composite reference method is summarized for anogenital skin lesions in STM and VTM in Tables 11 and 12. Overall, for HSV-1, sensitivity was 93.4% for VTM and 94.7% for STM specimen types and specificity was 99.8% for VTM and 99.6% for STM specimen types (Table 11). For HSV-2, sensitivity was

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96.9% for VTM and 98.4% for STM specimen types and specificity was 97.5% for VTM and 92.8% for STM specimen types (Table 12).

SpecimenType	Gender	N	TP	FP	TN	FN	Prev(%)	Sensitivity %(95% CI)1	Specificity%(95% CI)1	PPV %(95% CI)2	NPV %(95% CI)2
VTM	Combined	528	71	13	451	54	14.4	93.4(85.5-97.2)	99.8(98.8->99.9)	98.6(93.0-100)	98.9(97.6-99.6)
	Male	192	19	1	170	2	10.9	90.5(71.1-97.3)	99.4(96.8-99.9)	95.0(78.6-99.8)	98.8(96.4-99.9)
	Female	336	52	0	281	3	16.4	94.5(85.1-98.1)	100(98.7-100)	100(93.7-100)	98.9(97.1-99.8)
STM	Combined	531	71	25	454	46	14.1	94.7(87.1-97.9)	99.6(98.4-99.9)	97.3(91.1-99.6)	99.1(97.9-99.8)
	Male	192	20	2	169	1	10.9	95.2(77.3-99.2)	98.8(95.8-99.7)	90.9(74.5-98.7)	99.4(97.2-100)
	Female	339	51	0	285	3	15.9	94.4(84.9-98.1)	100(98.7-100)	100(93.6-100)	99.0(97.2-99.8)

Table 11. Summary of HSV-1 Results by VTM and STM Specimen Type

STM = Aptima Multitest Swab specimen, Prev = prevalence, VTM = VTM sample

1Score CI

2PPV 95% CI computed from the exact 95% CI for the positive likelihood ratio, NPV 95% CI computed from the exact 95% CI for the negative likelihood ratio

3The sample had a negative culture result.

4Two samples had negative culture results, one had a non-typable HSV positive culture result, and two were HSV-1 positive by culture. 5Both specimens had negative culture results.

6 One specimen had a negative culture result, one had a non-typable HSV positive culture result, and two were HSV-1 positive by culture.

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SpecimenType	Gender	N	TP	FP	TN	FN	Prev(%)	Sensitivity %(95% CI)1	Specificity %(95% CI)1	PPV %(95% CI)2	NPV %(95% CI)2
VTM	Combined	533	248	73	270	84	48.0	96.9(94.0-98.4)	97.5(94.9-98.8)	97.3(94.7-98.8)	97.1(94.6-98.7)
	Male	194	79	2	110	3	42.3	96.3(89.8-98.7)	98.2(93.7-99.5)	97.5(92.0-99.7)	97.3(93.0-99.4)
	Female	339	169	5	160	5	51.3	97.1(93.5-98.8)	97.0(93.1-98.7)	97.1(93.8-99.0)	97.0(93.4-99.0)
STM	Combined	535	253	205	258	46	48.0	98.4(96.1-99.4)	92.8(89.1-95.3)	92.7(89.4-95.3)	98.5(96.3-99.6)
	Male	194	79	6	106	3	42.3	96.3(89.8-98.7)	94.6(88.8-97.5)	92.9(86.5-97.1)	97.2(92.8-99.4)
	Female	341	174	14	152	1	51.3	99.4(96.8-99.9)	91.6(86.3-94.9)	92.6(88.5-95.7)	99.3(96.6-100)

Table 12. Summary of HSV-2 Results by VTM and STM Specimen Type

STM = Aptima Multitest swab specimen, Prev = prevalence, VTM = VTM sample

1Score CI

2PV 95% CI computed from the exact 95% CI for the positive likelihood ratio, NPV 95% CI computed from the exact 95% CI for the negative likelihood ratio

3Six samples had negative culture results and one was HSV-1 positive by culture.

4All eight samples had negative culture results.

5Eighteen specimens had negative culture results and two were HSV-1 positive by culture.

6All four specimens had negative culture results.

Expected Values

Prevalence

The prevalence of HSV-1 and HSV-2 observed during a multi-center clinical study was calculated for the Aptima HSV 1 & 2 assay. The prevalence of HSV-1 and HSV-2 with the Aptima HSV 1 & 2 assay is summarized by age group, gender group, and specimen type in Table 13.

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Gender	Age Group	VTM		STM
		HSV-1	HSV-2	HSV-1	HSV-2
Male	All ages	10.4 (20/192)	41.8 (81/194)	11.5 (22/192)	43.8 (85/194)
	<2 years	0.0 (0/0)	0.0 (0/0)	0.0 (0/0)	0.0 (0/0)
	2 to 11 years	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)
	12 to 21 years	11.1 (2/18)	38.9 (7/18)	11.1 (2/18)	38.9 (7/18)
	22 to 30 years	14.0 (13/93)	34.0 (32/94)	14.9 (14/94)	35.8 (34/95)
	31 to 40 years	12.5 (5/40)	50.0 (20/40)	12.8 (5/39)	53.8 (21/39)
	41 to 50 years	0.0 (0/20)	52.4 (11/21)	0.0 (0/20)	52.4 (11/21)
	51 to 60 years	0.0 (0/14)	57.1 (8/14)	7.1 (1/14)	57.1 (8/14)
	> 60 years	0.0 (0/6)	50.0 (3/6)	0.0 (0/6)	66.7 (4/6)
Female	All ages	15.5 (52/336)	51.3 (174/339)	15.0 (51/339)	55.1 (188/341)
	<2 years	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)
	2 to 11 years	0.0 (0/0)	0.0 (0/0)	0.0 (0/0)	0.0 (0/0)
	12 to 21 years	23.8 (15/63)	56.3 (36/64)	23.1 (15/65)	59.1 (39/66)
	22 to 30 years	14.6 (25/171)	52.6 (90/171)	14.5 (25/172)	56.4 (97/172)
	31 to 40 years	11.1 (7/63)	46.0 (29/63)	12.5 (8/64)	47.6 (30/63)
	41 to 50 years	18.2 (4/22)	39.1 (9/23)	10.0 (2/20)	47.6 (10/21)
	51 to 60 years	7.7 (1/13)	46.2 (6/13)	7.1 (1/14)	57.1 (8/14)
	> 60 years	0.0 (0/3)	100 (4/4)	0.0 (0/3)	100 (4/4)
Combined	All ages	13.6 (72/528)	47.8 (255/533)	13.7 (73/531)	51.0 (273/535)
	<2 years	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)
	2 to 11 years	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)	0.0 (0/1)
	12 to 21 years	21.0 (17/81)	52.4 (43/82)	20.5 (17/83)	54.8 (46/84)
	22 to 30 years	14.4 (38/264)	46.0 (122/265)	14.7 (39/266)	49.1 (131/267)
	31 to 40 years	11.7 (12/103)	47.6 (49/103)	12.6 (13/103)	50.0 (51/102)
	41 to 50 years	9.5 (4/42)	45.5 (20/44)	5.0 (2/40)	50.0 (21/42)
	51 to 60 years	3.7 (1/27)	51.9 (14/27)	7.1 (2/28)	57.1 (16/28)
	> 60 years	0.0 (0/9)	70.0 (7/10)	0.0 (0/9)	80.0 (8/10)

Table 13. Gender and Age Distribution by VTM and STM Specimen Type¹

1 No subjects had positive Aptima HSV 1 & 2 assay results for both HSV-1 and HSV-2.

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Positive and Negative Predictive Values for Hypothetical Prevalence Rates

The estimated positive and negative predictive values (PPV and NPV) of the Aptima HSV 1 & 2 assay for detection of HSV-1 and HSV-2 across different hypothetical prevalence rates are shown for each specimen type in Table 14. These calculations are based on the overall estimated sensitivity and specificity for each specimen type as determined in the clinical performance study.

		HSV-1		HSV-2
	Prevalence (%)	PPV (%)	NPV (%)	PPV (%)	NPV (%)
Specimen Type
VTM	1	81.0	99.9	27.9	100
	2	89.6	99.9	43.9	99.9
	5	95.7	99.7	66.9	99.8
	10	97.9	99.3	81.0	99.6
	20	99.1	98.4	90.6	99.2
	30	99.5	97.3	94.3	98.6
	40	99.6	95.8	96.2	97.9
	50	99.8	93.8	97.5	96.9
STM	1	68.6	99.9	12.1	100
	2	81.5	99.9	21.8	100
	5	91.9	99.7	41.9	99.9
	10	96.0	99.4	60.3	99.8
	20	98.2	98.7	77.4	99.6
	30	98.9	97.8	85.4	99.3
	40	99.3	96.6	90.1	98.9
	50	99.5	94.9	93.2	98.4

Table 14. Prevalence vs Hypothetical PPV1 and NPV2 for Detection of HSV-1 and HSV-2 by Specimen Type

STM = Aptima Multitest swab specimen, VTM = VTM sample

1PPV was calculated using:

(Sensitivity x Prevalence) / (Sensitivity x Prevalence + [1 - Specificity] x [1 - Prevalence]).

2NPV was calculated using:

(Specificity x [1 - Prevalence]) / ([1 - Sensitivity] x Prevalence + Specificity x [1 - Prevalence]).

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TTime Distribution for Aptima HSV 1 & 2 Assay Positive Controls

The distribution of the TTime values for the Aptima HSV 1 & 2 assay positive control from all valid Aptima HSV 1 & 2 assay runs performed during the clinical performance study is presented in Table 15.

	TTime
Statistics	HSV-1	HSV-2
N	107	107
Mean	20.03	22.01
Median	19.8	21.7
SD	1.198	1.612
CV (%)	6.0	7.3
Minimum	18.1	19.5
Maximum	22.9	26.2

Table 15. Distribution of TTimes for Aptima HSV 1 & 2 Assay Positive Controls

CV = coefficient of variation, SD = standard deviation

VIII. CONCLUSIONS

The analytical and clinical study results demonstrate that the Aptima HSV assay on the Panther system is substantially equivalent to the predicate device that is currently marketed for the same intended use. Hardware and software verification and validation demonstrate that the Aptima HSV assay on the Panther system will perform as intended.

Regulation Number and Section

§ 866.3305 Herpes simplex virus serological assays.

(a)
Identification. Herpes simplex virus serological assays are devices that consist of antigens and antisera used in various serological tests to identify antibodies to herpes simplex virus in serum. Additionally, some of the assays consist of herpes simplex virus antisera conjugated with a fluorescent dye (immunofluorescent assays) used to identify herpes simplex virus directly from clinical specimens or tissue culture isolates derived from clinical specimens. The identification aids in the diagnosis of diseases caused by herpes simplex viruses and provides epidemiological information on these diseases. Herpes simplex viral infections range from common and mild lesions of the skin and mucous membranes to a severe form of encephalitis (inflammation of the brain). Neonatal herpes virus infections range from a mild infection to a severe generalized disease with a fatal outcome.(b)
Classification. Class II (special controls). The device is classified as class II (special controls). The special control for the device is FDA's revised guidance document entitled “Class II Special Controls Guidance Document: Herpes Simplex Virus Types 1 and 2 Serological Assays.” For availability of the guidance revised document, see § 866.1(e).