The GenePOC CDiff assay performed on the revogene instrument is a qualitative in vitro diagnostic test that utilizes automated sample processing and real-time polymerase chain reaction (PCR) to detect the toxin B (tcdB) gene of toxigenic Clostridium difficile( C. difficile) in unformed (liquid or soft) stool specimens obtained from patients suspected of having C. difficile infection (CDI). The GenePOC CDiff assay is intended to aid in the diagnosis of CDI.

Device Description

The GenePOC™ CDiff assay is a single-use test for the qualitative detection of the toxin B (tcdB) gene of toxigenic Clostridium difficile (C. difficile) in unformed (liquid or soft) stool specimens. The GenePOC™ CDiff assay kit is comprised of the disposable CDiff microfluidic cartridges (PIE), Disposable Transfer Loops (DTL), Sample Buffer Tubes (SBT), and Disposable Transfer Tools (DTT; pipette). These components are used to suspend the sample, extract, amplify, and detect C. difficile nucleic acid. A Process Control (PrC) is also incorporated into each PIE to verify sample processing and amplification steps. The PrC allows for the verification of potential inhibitor substances as well as microfluidic, instrument or reagent failure. The GenePOC™ CDiff assay is designed to be used on the revogene™. The revogene™ is an instrument that automates sample homogenization, sample dilution, cells lysis, DNA amplification and detection of the amplified PCR products.

Each GenePOC™ CDiff assay kit provides components for 24 tests. User intervention is required for sample preparation, transferring the stool specimen with the DTL into the SBT, using the DTT to transfer the sample into the PIE, and loading/unloading the PIE into the revogene™ carousel. Each PIE is a completely integrated closed device in which a sample is dispensed and processed through different microfluidic chambers and channels that allow for the sample processing and subsequent real-time PCR steps.

Upon completion of a run, the results are computed by the revogene™ from measured fluorescent signals and embedded calculation algorithms. The output results include positive, negative, indeterminate, and unresolved. Upon completion of a run, the user removes the used cartridges and disposes of them in normal biological waste. Results may be viewed, printed, transferred, and/or stored by the user.

AI/ML Overview

The provided document describes the regulatory submission for the GenePOC CDiff assay, a diagnostic test for Clostridium difficile infection (CDI). The document focuses on the analytical and clinical performance of the device to demonstrate its substantial equivalence to a predicate device.

Here's a breakdown of the acceptance criteria and study details based on the provided text:

1. Table of Acceptance Criteria and Reported Device Performance

The document does not explicitly state pre-defined acceptance criteria in a formal table. However, the performance characteristics sections imply the criteria for various analytical and clinical aspects. The reported performance is presented in the tables and text within the document.

Let's infer the acceptance criteria from the reported results, especially concerning agreement percentages and confidence intervals. Typically, for in vitro diagnostics, high sensitivity and specificity are desired, often above certain thresholds (e.g., 85% or 90% for sensitivity, and 95% or more for specificity). For reproducibility, high agreement is also expected (e.g., >95%).

Metric	Acceptance Criterion (Implied)	Reported Device Performance (GenePOC CDiff)
Analytical Performance
Reproducibility
Between-Laboratory Reproducibility:
Low Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	97.2% (95% CI: 93.6-99.1%)
Moderate Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	98.3% (95% CI: 95.2-99.7%)
True Negative Samples (Overall Agreement)	Very high agreement (e.g., >95%)	100% (95% CI: 97.5-100%)
CVs for Ct Values	Low variability (e.g., <10%)	Consistently below 7.54% (Overall)
Between-Lot Reproducibility:
Low Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	95.0% (95% CI: 90.7-97.7%)
Moderate Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	98.3% (95% CI: 95.2-99.7%)
True Negative Samples (Overall Agreement)	Very high agreement (e.g., >95%)	100% (95% CI: 97.5-100%)
Within-Laboratory Precision:
Low Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	94.4% (95% CI: 89.3-97.6%)
Moderate Positive Samples (Overall Agreement)	High agreement (e.g., >90%)	96.5% (95% CI: 92.1-98.9%)
True Negative Samples (Overall Agreement)	Very high agreement (e.g., >95%)	100% (95% CI: 96.9-100%)
Detection Limit (LoD)	Lowest concentration for 95%+ detection	1,500 CFU/mL of SB for C. difficile strains (95% or greater detection)
Analytical Inclusivity	All strains detected at 2-3xLoD	All 20 toxigenic C. difficile strains detected at 3,750 CFU/mL SB (2-3xLoD)
Analytical Specificity	Minimal to no false positives	Limited specific false positives at high concentrations for some Clostridium species. No reactivity at lower concentrations. One case of false positive for Enterococcus faecalis at high concentration.
Interference	No interference	No interference from 30 tested organisms; interference from Tums and Stomaax at high concentrations; no interference from 5 endogenous agents.
Carry-Over & Cross-Contamination	Absence of carry-over/cross-contamination	Demonstrated absence.
Clinical Performance
Sensitivity (Fresh Specimens)	High sensitivity (e.g., >80%)	80.5% (95% CI: 72.0-87.4%)
Specificity (Fresh Specimens)	High specificity (e.g., >95%)	97.1% (95% CI: 95.5-98.2%)
Sensitivity (Frozen Specimens)	High sensitivity (e.g., >80%)	87.3% (95% CI: 82.1-91.4%)
Specificity (Frozen Specimens)	High specificity (e.g., >95%)	97.3% (95% CI: 96.3-98.1%)
Unresolved Rate (After Repeat Testing)	Low rate (e.g., <1%)	Fresh: 0.1% (1/798); Frozen: 0.0% (0/1665)
Indeterminate Rate (After Repeat Testing)	Low rate (e.g., <1%)	Fresh: 0.0% (0/798); Frozen: 0.1% (1/1665)

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

Clinical Study Test Set (Patients):
- Total Samples: 2,461 fully compliant samples (797 fresh + 1,664 frozen).
- Data Provenance: Prospective multicenter trial at 7 geographically diverse clinical trial sites (US and Canada).
- Retrospective/Prospective: The study design included testing fresh samples (prospectively collected and tested immediately) and retrospectively collected samples that were stored frozen (prospectively collected but tested after freezing).
Analytical Performance Test Sets:
- Precision/Reproducibility: 1,022 samples (383 low positive, 384 moderate positive, and 255 negative samples) were tested across different studies (Between-Laboratory, Between-Lot, Within-Laboratory). These were spiked stool samples. The provenance of the negative stool pool for preparing these panels is not specified by country, but the panel strains are ATCC (American Type Culture Collection), indicating standardized reference strains.
- Detection Limit (LoD): Not specified in terms of sample size, but involved testing replicates (multiple runs, instruments, operators) of specific C. difficile strains.
- Analytical Inclusivity: 20 strains of toxigenic C. difficile from various geographic origins.
- Analytical Specificity: 58 various analytes (1 yeast, 6 viruses, 50 bacteria, and human DNA).
- Interference (Non-Target Organisms): Tests involved 30 organisms.
- Interference (Exogenous/Endogenous Substances): 21 potentially interfering substances.

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

This information is not explicitly stated in the provided text.
For the clinical study, the reference method (ground truth) was "Combined Direct and Enriched Culture" followed by cytotoxicity testing (CCNA) on isolated C. difficile colonies. While this is a laboratory-based gold standard, the document does not specify the number of laboratory experts/technicians involved in performing and interpreting these reference methods or their specific qualifications. It also doesn't mention expert clinical adjudication for the patient diagnosis itself, as the device is for in vitro diagnostic testing based on lab results.

4. Adjudication Method for the Test Set

For the clinical study, the "ground truth" (reference method) was established as follows:
- A specimen was considered positive for toxigenic C. difficile if C. difficile was recovered by either direct or enriched culture AND if bacterial isolates tested positive by cytotoxicity testing (CCNA).
- A specimen was considered negative only if it tested negative by both direct and combined culture (i.e., direct and enriched culture).
This is a laboratory-based algorithm for establishing ground truth, not a multi-reader clinical adjudication process. The document does not describe an adjudication method involving multiple human readers for the final clinical diagnosis or the interpretation of the reference method.

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

No, an MRMC comparative effectiveness study was not done.
This device is an in vitro diagnostic (IVD) assay that automates sample processing, DNA amplification, and detection of a specific gene. It provides a "positive," "negative," "indeterminate," or "unresolved" result. It is not an AI-based image analysis tool or a system that assists human readers in interpreting complex data where reader improvement could be measured. Therefore, the concept of human readers improving with AI assistance is not applicable to this type of device. The study evaluates the performance of the automated assay itself against a laboratory reference method.

6. If a Standalone (i.e., algorithm only without human-in-the-loop performance) Was Done

Yes, this was a standalone performance study. The GenePOC CDiff assay operates on the revogene instrument, which automates the entire process from sample loading to result computation. The results are "computed by the revogene™ from measured fluorescent signals and embedded calculation algorithms." The performance metrics (sensitivity, specificity, reproducibility) are reported for the device as a whole, essentially as an algorithm-only (automated instrument) performance, without human "in-the-loop" interpretation of the primary signals. Human intervention is limited to sample preparation and loading/unloading.

7. The Type of Ground Truth Used (Expert Consensus, Pathology, Outcomes Data, etc.)

The ground truth used for the clinical study was a laboratory-based reference method: "Combined Direct and Enriched Culture" for C. difficile recovery, followed by cytotoxicity testing (CCNA) on the isolated bacterial strain to confirm toxin production. This is a robust and widely accepted method for confirming toxigenic C. difficile in stool samples.

8. The Sample Size for the Training Set

The document does not mention a separate "training set" for an algorithm. This suggests that the device's algorithms and parameters were likely developed and validated internally by the manufacturer during product development, possibly using a series of development and internal validation studies. The clinical and analytical studies described are primarily for performance demonstration and likely served as a "test set" for regulatory submission, rather than a training set for a machine learning model.
For an IVD like this, the "training" (calibration, optimization of thresholds) would typically be done during the engineering and design phases and locked down before clinical validation.

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

As a dedicated "training set" is not explicitly mentioned or described, the method for establishing its ground truth is also not provided. If an internal training or development set was used, it would have likely relied on similar laboratory reference methods to establish the true positive/negative status of samples.

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). 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.

November 22, 2017

GenePOC Inc. Guy Sevigny Senior Regulatory Affairs Specialist 360 rue Franquet Quebec, G1P 4N3 Ca

Re: K172569

Trade/Device Name: GenePOC CDiff Regulation Number: 21 CFR 866.3130 Regulation Name: Clostridium difficile toxin gene amplification assay Regulatory Class: Class II Product Code: OZN Dated: August 25, 2017 Received: August 25, 2017

Dear Guy Sevigny:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food. Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

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

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR

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

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

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/MedicalDevices/DeviceRegulationandGuidance/) and CDRH Learn (http://www.fda.gov/Training/CDRHLearn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (http://www.fda.gov/DICE) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

Steven R. Gitterman -S for

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

Enclosure

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Indications for Use

510(k) Number (if known)

K172569

Device Name GenePOC CDiff

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)

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510(K) SUMMARY

A. GENERAL INFORMATION

Submission Date:	November 17, 2017
Submitter Information:
Submitted By:	GenePOC Inc.360 rue FranquetQuébec (Québec) G1P 4N3 Canada
Contact Person:	Guy Sevigny

Contact Person:	Guy Sevigny
	Sr. Regulatory Affairs Specialist
	GenePOC Inc.
	Telephone: +1 418 650-3535 ext. 261
	Email: guy.sevigny@genepoc.ca

B. PURPOSE FOR SUBMISSION:

To obtain a substantial equivalence determination for the GenePOC™ CDiff assay

C. MEASURAND:

tcdB gene of toxigenic Clostridium difficile (C. difficile)

D. TYPE OF TEST:

Real-time Polymerase chain reaction (rtPCR)

E. DEVICE INFORMATION:

1. Trade Name: GenePOC™ CDiff
1. Regulation: 21 CFR 866.3130 - Clostridium difficile toxin gene amplification assay
1. Classification: Class II
1. Product Code: OZN
1. Panel: 83, Microbiology

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F. INTENDED USE:

1. Intended Use and Indications for Use:
  The GenePOC™ CDiff assay performed on the revogene™ instrument is a qualitative in vitro diagnostic test that utilizes automated sample processing and real-time polymerase chain reaction (PCR) to detect the toxin B (tcdB) gene of toxigenic Clostridium difficile (C. difficile) in unformed (liquid or soft) stool specimens obtained from patients suspected of having C. difficile infection (CDI). The GenePOC™ CDiff assay is intended to aid in the diagnosis of CDI.
1. Special conditions for use statement(s): Prescription Use Only
1. Special instrument requirements: revogene™M

G. DEVICE DESCRIPTION:

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H. SUBSTANTIAL EQUIVALENCE INFORMATION:

1. Predicate Device Name: BD MAX CDIFF ASSAY, BD MAX INSTRUMENT
1. Predicate 510(k) Number: K130470

3. Comparison with Predicate:

Item	GenePOCTM CDiff	BD MAX Cdiff Assay (Predicate Device)
K Number	Subject of submission	K130470
Intended Useand Indicationsfor Use	The GenePOC CDiff assay performedon the revogeneTM instrument is aqualitative in vitro diagnostic test thatutilizes automated sample processingand real-time polymerase chainreaction (PCR) to detect the toxin B( tcdB ) gene of toxigenic Clostridiumdifficile ( C. difficile ) in unformed(liquid or soft) stool specimensobtained from patients suspected ofhaving C. difficile infection (CDI). TheGenePOC CDiff assay is intended toaid in the diagnosis of CDI.	The BD MAX Cdiff Assay performed onthe BD MAXTM System is an automated invitro diagnostic test for the direct,qualitative detection of the Clostridiumdifficile toxin B gene ( tcdB ) in humanliquid or soft stool specimens from patientssuspected of having C. difficile infection(CDI). The test, performed directly on thespecimen, utilizes real-time polymerasechain reaction (PCR) for the amplificationof C. difficile toxin B gene DNA andfluorogenic target-specific hybridizationprobes for the detection of the amplifiedDNA. The BD MAX Cdiff Assay isintended to aid in the diagnosis of CDI.
SIMILARITIES
Classification	Class II	Same
Product Code	OZN	Same
DNA Target	Presence of the toxin B ( tcdB ) gene	Same
Specimen type	Unformed (liquid or soft) stool	Same
Assay format	• Amplification: Real Time PCR• Detection: Fluorogenic target-specific hybridization	Same
Detectionprobes	TaqMan® Probe	Same
Samplepreparation	Automated by revogeneTM	Automated by BD MAXTM system
Interpretation oftest results	Automated (Diagnostic software of therevogeneTM)	Automated (Diagnostic software of BDMAXTM system)
Internal ProcessControl	To help monitor presence of potentialinhibitory substances as well as anysystem or reagent failures	Specimen Processing Control (SPC)
DIFFERENCES
Instrument	revogeneTM	BD MAXTM

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I. STANDARDS/GUIDANCE DOCUMENTS REFERENCED:

CLSI Guideline EP25-A, Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline.

J. TEST PRINCIPLE:

The GenePOC revogene™ automates and integrates nucleic acid extraction and amplification, and detection of the target sequence in complex samples using real-time PCR. A liquid or soft stool specimen is collected using a standard stool collection device. Using the DTL, a disposable 5uL inoculating loop dipped into the stool specimen, stool material is transferred into SBT. After vortexing, approximately 150 uL of the inoculated sample buffer is transferred into the GenePOC microfluidic cartridge using the DTT. The loaded CDiff cartridge is placed into the revogene™ for further sample processing. No operator intervention is necessary once the clinical sample is loaded onto the revogene™.

Each CDiff microfluidic cartridge (PIE) is a completely integrated and self-contained device. Each sample is sequentially transferred by centrifugation from one microfluidic chamber to the next and all reagents specific for the PCR reaction are incorporated and dried within the PCR wells. The stepwise process includes sample homogenization, specimen dilution and lysis of cells followed by the subsequent real-time PCR steps within 1 PCR well in the cartridge. An internal Process Control (PrC) is contained in the homogenization chamber and is therefore present in every test to verify critical steps of the analytical process (including sample homogenization, dilution, sample lysis, and nucleic acid amplification and detection) for the presence of potential inhibitory substances as well as system or reagent failures. The amplified products are detected in real time using target-specific TaqMan® chemistrybased probes. The CDiff specific designed primers and probe detect a target region of 262 base pairs (bp) of the toxin B gene (tcdB) of Clostridium difficile. The results are computed by the system from measured fluorescent signals and embedded calculation algorithms. Results may be viewed, printed, transferred, and/or stored by the user.

K. PERFORMANCE CHARACTERISTICS:

1. Analytical Performance

a. Precision/Reproducibility

For the Reproducibility and Precision study, a total of 1,022 samples (383 low positive, 384 moderate positive, and 255 negative samples) were tested with the GenePOCTM CDiff assay. Single site and multi-site precision studies were conducted to determine Between-Laboratory Reproducibility, Between-Lot Reproducibility, and Within-Laboratory Precision. A five members panel comprised of 1 negative panel member and 4 negative stool samples spiked with either a low positive (2,438 CFU/mL and 2,925 CFU/mL of SB) or a moderate positive (3,750 CFU/mL and 4,500 CFU/mL of SB) final concentration of C. difficile Toxinotype 0 (ATCC 432551M; ribotype 087) strain or with C. difficile Toxinotype IIIb (ATCC BAA-1805™; NAP1/ribotype 027) strain respectively. The negative panel member was prepared by using a pool of toxigenic C. difficile negative stools.

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For all factors evaluated and the different panel members tested, the variation of results is consistently below 7.54% (%CV values).

Between-Laboratory Reproducibility:

The Between-Laboratory Reproducibility study was performed with one reagent lot on five days (consecutive or not) by two operators at three selected sites. Two runs were performed by each operator on each day. The overall agreement of assay results was 97.2% (95% CI = 93.6-99.1%) for Low Positive (LP) samples (both toxigenic C. difficile strains), 98.3% (95% CI = 95.2-99.7%) for Moderate Positive (MP) samples (both toxigenic C. difficile strains) and 100% (95% CI = 97.5-100%) for True Negative (TN) samples.

GenePOC™ CDiff Assay Results and Percent Agreement for Between-Laboratory Reproducibility Qualitative Analysis

PanelType	Strain	Site 01	Site 02	Site 03	AssayResults/Total	Agreement(%)[95% CI]
LowPositive	ATCC 43255TM	27/30	28/30	30/30	85/90	94.4[87.5%-98.2%]
	ATCC BAA-1805TM	30/30	30/30	30/30	90/90	100[96.7%-100%]
	Overall AssayResults/Total(% Agreement)[95% CI]	57/60(95%)[86.1-99.0%]	58/60(96.7%)[88.5-99.6]	60/60(100%)[95.1-100]	175/180(97.2%)	97.2[93.6%-99.1%]
ModeratePositive	ATCC 43255TM	29/30	29/30	29/30	87/90	96.7[90.6%-99.3%]
	ATCC BAA-1805TM	30/30	30/30	30/30	90/90	100[96.7%-100%]
	Overall AssayResults/Total(% Agreement)[95% CI]	59/60(98.3%)[91.1-100%]	59/60(98.3%)[91.1-100%]	59/60(98.3%)[91.1-100%]	177/180(98.3%)	98.3[95.2%-99.7%]
Negative	Overall AssayResults/Total(% Agreement)[95% CI]	40/40(100%)[92.8-100%]	40/40(100%)[92.8%-100%]	40/40(100%)[92.8%-100%]	120/120(100%)	100[97.5%-100%]

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C. difficile
PanelType	Strain	MeanCt(CDiff)	N	Between-Laboratory		Between-Operators		Between-Days		ResidualError		Overall
				SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
LowPositive	ATCC43255TM	38.48	85	0.23	0.61	0.00	0.00	0.00	0.00	1.54	4.00	1.56	4.05
LowPositive	ATCCBAA-1805TM	36.37	90	1.12	3.07	0.00	0.00	0.00	0.00	2.50	6.89	2.74	7.54
ModeratePositive	ATCC43255TM	37.68	87	0.46	1.23	0.00	0.00	0.00	0.00	1.56	4.14	1.63	4.32
ModeratePositive	ATCCBAA-1805TM	36.57	90	0.54	1.48	0.19	0.52	0.16	0.43	1.54	4.21	1.65	4.52
PrC
PanelType	Strain	MeanCt(PrC)	N	Between-Laboratory		Between-Operators		Between-Days		ResidualError		Overall
				SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
Negative	N/A	33.09	120	0.45	1.36	0.19	0.58	0.07	0.20	0.89	2.70	1.02	3.09

Results for the Ct Values Analysis of Between-Laboratory Reproducibility Study

Between-Lot Reproducibility:

The Between-Lot Reproducibility study was performed at one site with three lots for a total of 15 days of testing (5 days per reagent lot) and two runs performed by each operator on each day. The overall agreement of assay results was 95.0% (95% CI = 90.7- 97.7%) for Low Positive (LP) samples (both toxigenic C. difficile strains), 98.3% (95% CI = 95.2-99.7%) for Moderate Positive (MP) samples (both toxigenic C. difficile strains) and 100% (95% CI = 97.5-100%) for True Negative (TN) samples.

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GenePOCTM CDiff Assay Results and Percent Agreement for Between-Lot Reproducibility Qualitative Analysis

Panel Type	Strain	Kit lot #1(L1906151)	Kit lot #2(L1906091)	Kit lot #3(L1906141)	AssayResults/Total	Agreement(%)[95% CI]
LowPositive	ATCC 43255 TM	27/30	27/30	28/30	82/90	91.1[83.2%-96.1%]
	ATCC BAA-1805TM	30/30	29/30	30/30	89/90	98.9[94.0%-100%]
	Overall AssayResults/Total(% Agreement)[95% CI]	57/60(95%)[86.1-99.0%]	56/60(93.3%)[83.8-98.2%]	58/60(96.7%)[88.5-99.6%]	171/180(95%)	95.0[90.7%-97.7%]
ModeratePositive	ATCC 43255TM	29/30	30/30	28/30	87/90	96.7[90.6%-99.3%]
	ATCC BAA-1805TM	30/30	30/30	30/30	90/90	100[96.7%-100%]
	Overall AssayResults/Total(% Agreement)[95% CI]	59/60(98.3%)[91.1-100%]	60/60(100%)[95.1-100%]	58/60(96.7%)[88.5-99.6%]	177/180(98.3%)	98.3[95.2%-99.7%]
Negative	Overall AssayResults/Total(% Agreement)[95% CI]	40/40(100%)[92.8-100%]	40/40(100%)[92.8-100%]	40/40(100%)[92.8-100%]	120/120(100%)	100[97.5%-100%]

Results for the Ct Values Analysis of Between-Lot Reproducibility Studv

C. difficile
PanelType	Strain	MeanCt(CDiff)	N	Between-Lot		Between-Operators		Between-Days		ResidualError		Overall
				SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
LowPositive	ATCC43255TM	38.57	82	0.08	0.20	0.10	0.26	0.15	0.40	1.71	4.43	1.72	4.46
	ATCCBAA-1805TM	37.02	89	0.59	1.60	0.46	1.24	0.00	0.00	1.83	4.96	1.98	5.35
ModeratePositive	ATCC43255TM	37.55	87	0.70	1.86	1.02	2.71	0.00	0.00	1.77	4.72	2.16	5.75
	ATCCBAA-1805TM	36.86	90	0.16	0.42	0.72	1.96	0.18	0.48	1.07	2.90	1.31	3.56
PrC
PanelType	Strain	MeanCt(PrC)	N	Between-Lot		Between-Operators		Between-Days		ResidualError		Overall
				SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
Negative	N/A	33.21	120	0.34	1.02	0.23	0.68	0.18	0.55	0.78	2.35	0.90	2.71

Within-Laboratory Precision:

The Within-Laboratory Precision of the assay was performed at one site and tested specimens with one reagent lot for a total of 12 days of testing with two runs performed by each operator on each day. The overall agreement of assay results was 94.4% (95% CI = 89.3-97.6%) for Low Positive (LP) samples (both toxigenic C. difficile strains), 96.5% (95% CI = 92.1-98.9%) for Moderate Positive (MP) samples (both toxigenic C. difficile strains) and 100% (95% CI = 96.9-100%) for True Negative

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(TN) samples.

Panel Type	Strain	Assay Results/Total	Agreement (%)[95% CI]
Low Positive	ATCC 43255TM	64/72	88.9[79.3%-95.1%]
	ATCC BAA-1805TM	71/71	100[95.9%-100%]
	Overall Assay Results/Total(% Agreement)[95% CI]	135/143(94.4%)	94.4[89.3%-97.6%]
Moderate Positive	ATCC 43255TM	69/72	95.8[88.3%-99.1%]
	ATCC BAA-1805TM	70/72	97.2[90.3%-99.7%]
	Overall Assay Results/Total(% Agreement)[95% CI]	139/144(96.5%)	96.5[92.1%-98.9%]
Negative	Overall Assay Results/Total(% Agreement)[95% CI]	95/95(100%)	100[96.9%-100%]

GenePOC™ CDiff Assay Results and Percent Agreement for Within-Laboratory Precision Qualitative Analysis

b. Linearity/Assay Reportable Range

Not applicable.

Traceability, Stability, Expected Values (controls, calibrators, or methods) ن

There are three types of controls for the GenePOC™ CDiff assay including an internal process control (PrC), positive external control (PEC) and negative external control (NEC). The PrC is provided in each GenePOC™ CDiff assay. The PrC is extracted, amplified, and detected along with each specimen tested and verifies the efficacy of the dilution, cell lysis, PCR amplification, and detection processes. For the PEC and NEC, GenePOC recommends using commercially available control materials (e.g., ATCC 43255TM, a C. difficile strain bearing the tcdB gene for PEC, and ATCC 43593, a nontoxigenic C. difficile strain for NEC). It is recommended that the positive bacterial strain be freshly prepared in saline to a turbidity of 0.5 McFarland from isolated colonies and subsequently diluted ½ in saline before its addition into the SBT with the DTL. The negative strain could be prepared the same way while added directly to the SBT without the dilution step.

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d. Detection Limit

The LoD was established and confirmed in two separate studies. The confirmation of the LoD included three GenePOC™ CDiff reagent lots, six revogene™ instruments, and two operators. There were 4 runs per instrument per day, and the study was conducted over four days. Strain C. difficile ATCC 432551M and strain C. difficile ATCC BAA-1805TM were used in the study. The LoD was defined as the lowest concentration at which 95% or greater of all replicates tested positive. The LoD of the GenePOC™ CDiff assay was 1,500 CFU/mL of SB for each strain.

e. Analytical Inclusivity

The analytical reactivity (inclusivity) of the GenePOC™ CDiff assay was evaluated by testing 20 strains of toxigenic C. difficile from various geographic origins representing eight different toxinotypes. Three lots of GenePOC™ CDiff assay kits were used for sample testing and n=1 sample/strain/reagent lot. All toxigenic C. difficile strains tested were detected at 3,750 CFU/mL SB (2 - 3xLOD).

C. difficile Strain	Toxinotype, Toxin
ATCC 9689	Toxinotype 0, A+, B+
ATCC 700792	Toxinotype 0, A+, B+
ATCC 17858	Toxinotype 0, A+, B+
ATCC BAA-1382	Toxinotype 0, A+, B+
ATCC 51695	Toxinotype 0, A+, B+
ATCC 43600	Toxinotype 0, A+, B+
ATCC 43599	Toxinotype 0, A+, B+
ATCC 43596	Toxinotype 0, A+, B+
ATCC 43594	Toxinotype 0, A+, B+
ATCC 17857	Toxinotype 0, A+, B+
ATCC 43598	Toxinotype VIII, A-, B+
CCUG 8864	Toxinotype X, A-, B+
ATCC BAA-1870	Toxinotype IIIb, NAP1, A+, B+
ATCC BAA-1812	Toxinotype XII, A+, B+
ATCC BAA-1803	Toxinotype IIIc, NAP1, A+, B+
ATCC BAA-1814	Toxinotype XXII, A+, B+
ATCC BAA-1804	Toxinotype 0, A+, B+
ATCC BAA-1875	Toxinotype V, A+, B+
ATCC BAA-2155	Toxinotype XXII, A+, B+
ATCC BAA-1873	Toxinotype 0, A+, B+

f. Analytical Specificity

A total of 58 various analytes (1 yeast, 6 viruses, 50 bacteria, and human DNA) found in clinical unformed stool specimens (different from toxigenic C. difficile) were selected. These included:

Commensal and pathogenic microorganisms (bacteria, yeasts and viruses) from the . intestinal tract:
Species phylogenetically related to C. difficile;
C. difficile non-pathogenic strains; and
Human DNA.

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Under the condition of the study, Clostridium sordellii was detected by the CDiff assay at approximately 10° CFU/mL of SB for one replicate out of 3, but was found non-reactive at a load of 10° CFU/mL of SB. Clostridium novyi and Clostridium scindens produced false positive reactions in 1 replicate out of 6 tested at approximately 106 CFU/mL of SB. No reactivity was observed for three replicates tested at 105 CFU/mL of SB. Enterococcus faecalis produced false positive reaction in one replicate out of three tested at approximately 107 CFU/mL of SB. No reactivity was observed for three replicates tested at 106 CFU/mL of SB.

Name	Identification	Name	Identification
Bacteria (non-toxigenic C. difficile and other Clostridium species)
Clostridium difficile(non-toxigenic)	ATCC 43593	Clostridium perfringens	ATCC 13124
Clostridium difficile(non-toxigenic)	ATCC 43601	Clostridium scindens	ATCC 35704
Clostridium bifermentans	ATCC 638	Clostridium septicum	ATCC 12464
Clostridium butyricum	ATCC 860	Clostridium sordellii	ATCC 9714
Clostridium haemolyticum	ATCC 9650	Clostridium sporogenes	ATCC 15579
Clostridium novyi	ATCC 19402	Flavonifractor plautii(anc. design. Clostridiumorbiscindens)	ATCC 49531
Bacteria (potentially present in gastrointestinal tract)
Abiotrophia defectiva	ATCC 49176	Peptostreptococcusanaerobius	ATCC 27337
Acinetobacter baumannii	ATCC 19606	Plesiomonas shigelloides	ATCC 14029
Aeromonas hydrophila	ATCC 7966	Porphyromonasasaccharolytica	ATCC 25260
Alcaligenes faecalis subsp.faecalis	ATCC 15554	Prevotella melaninogenica	ATCC 25845
Bacillus cereus	ATCC 14579	Proteus mirabilis	ATCC 33583
Bacteroides fragilis	ATCC 25285	Providencia alcalifaciens	ATCC 9886
Campylobacter jejuni subsp. jejuni	ATCC 33560	Pseudomonas aeruginosa	ATCC 35554
Campylobacter jejuni(anc. design. Campylobactercoli)	ATCC 43479	Salmonella enterica subsp.arizonae	ATCC 13314
Citrobacter freundii	ATCC 8090	Salmonella enterica subsp.enterica serovarCholeraesuis	ATCC 7001
Edwardsiella tarda	ATCC 15947	Salmonella enterica subsp.enterica serovarTyphimurium	ATCC 14028
Enterobacter aerogenes	ATCC 13048	Serratia liquefaciens	ATCC 27592
Enterobacter cloacae subsp.cloacae	ATCC 13047	Serratia marcescens subsp.marcescens	ATCC 13880
Enterococcus faecalis	ATCC 19433	Shigella boydii	ATCC 9207
Escherichia coli	ATCC 11775	Shigella dysenteriae	CCRI-7792
Escherichia coli O157:H7	CCRI-22391	Shigella sonnei	ATCC 29930
Name	Identification	Name	Identification
Helicobacter pylori	ATCC 43504	Staphylococcus aureussubsp. aureus	ATCC 33592
Klebsiella oxytoca	ATCC 8724	Staphylococcus epidermidis	ATCC 14990
Lactobacillus acidophilus	ATCC 4356	Streptococcussp.(S.agalactiae)	ATCC 12973
Listeria monocytogenes	ATCC 7644	Vibrio parahaemolyticus	ATCC 17802
Viruses
Human Adenovirus 1 (DNA)	ATCC VR-1D	Enterovirus (RNA)	ATCC VR-1823D
Rotavirus (RNA)	ATCC VR-2018DQ	Echovirus (RNA)	ATCC VR-1734D
Norovirus (RNA)	ATCC VR-3235SD (GII)	Human Herpesvirus 5(Cytomegalovirus) (DNA)	ATCC VR-538D
Yeast
Candida albicans	ATCC 10231
Other Organisms
Human DNA	Kit, TQMN control genomic DNA

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g. Interference with Non-Target Organisms

The potentially inhibitory effect of 30 organisms, that may be present in the normal intestinal flora and which are not targeted by the test, was assessed using organisms selected from the cross-reactivity study. Each organism category (i.e., 27 bacteria, 1 yeast, 2 viruses) was represented with a special attention to include the most frequent causative agents of intestinal tract infections. Groups of 2 to 6 organisms were prepared in toxigenic C. difficile-negative liquid stool matrix, and tested in duplicate in presence of either 3,750 CFU/mL of SB of the toxigenic C. difficile ATCC® 43255™ strain or 4,500 CFU/mL of SB of the toxigenic C. difficile ATCC® BAA-1805™ strain, to assess their potential interference on detection of toxigenic C. difficile or PrC. Each organism within group was diluted to reach a load of ≥10° CFU/mL of SB for bacterium and yeast, and ≥10° copies/mL of SB for viruses.

None of the 30 organisms present at ≥10° CFU/mL of SB for bacteria and yeast and ≥10° copies/mL of SB for viruses interfered with detection of PrC and with the toxigenic C. difficile ATCC® BAA-1805™ strain. Two groups showed a potentially inhibitory effect on detection of the toxigenic C. difficile strain ATCC® 43255™. Nevertheless, when each bacterium from these groups was tested individually at a load of ≥10° CFU/ mL of SB in presence of C. difficile strain ATCC® 43255™, none interfered.

h. Interference with Exogenous and Endogenous Substances

Interference on the GenePOC™ CDiff assay was evaluated with 21 potentially interfering exogenous and endogenous substances in absence and in presence of 2 C. difficile strains ATCC® 43255™ and ATCC® BAA- 1805™ tested at 2-3xLoD (3,750 CFU/mL of SB or 4,500 CFU/mL of SB respectively) across 3 reagent lots.

N=16 exogenous substances occasionally used or found in the intestinal tract were tested including: Nystatin, Personnelle Hydrocortisone cream, Preparation H®, Tums®,

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Stomaax®, Life BRAND™ Heavy Mineral Oil USP, Mesalazine, Trojan® with spermicidal Lubricant Condoms, Pepto Bismol TM, Imodium®, Senokot®, Vancomycin, Metronidazole, Aleve®, Equate™ Flushable Moist Wipes, and Wet Ones®. Interference was observed with Tums and Stomaax® at concentrations of > 0.5 mg/mL of SB and > 0.5 uL/mL of SB respectively.

N=5 endogenous agents were tested including: Triglyceride Mix (C2-C10), Palmitic acid, Stearic acid, Whole blood, and Mucus. No interference was observed.

Carry-Over and Cross Contamination Studies i.

The within-run and between-run carry-over and cross- contamination were assessed using positive samples prepared in a toxigenic C. difficile-negative liquid stool matrix to reach a final concentration of >107 CFU/mL of SB of the toxigenic C. difficile ATCC® 43255TM strain. True negative samples, prepared with the toxigenic C. difficile-negative liquid stool matrix only, were also tested.

For the within-run study, a total of 10 runs were performed by two operators with the CDiff assay on one revogene. Four (4) high positive samples and 4 negative samples were tested by alternating positive and negative samples in each run. For the between-run study, a run of 8 replicates of high positive samples followed by a run of 8 replicates of negative samples were performed by two operators, for a total of 10 runs on one revogene.

Absence of carry-over and cross-contamination was demonstrated.

Sample Storage j.

Collected specimens should be stored between 2°C and 25°C during transport.

Stool specimens can be stored at 25℃ for up to 2 days, or at 2-8℃ for up to 4 days. Inoculated SBT can be stored at 25℃ for up to 2 days, or at 2-8℃ for up to 3 days.

Store the CDiff kit at 2-25°C. The expiration date is indicated on the box kit's label.

Do not open a pouch until ready to perform testing. Use the PIE within 1 hour after opening the pouch.

2. Comparison Studies

a. Method Comparison with predicate device Not applicable

b. Matrix Comparison

Not applicable.

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3. Clinical Studies

GenePOC conducted a prospective multicenter trial at 7 geographically diverse clinical trial sites (US and Canada). Unformed (liquid or soft) stool specimens were collected from patients suspected of having C. difficile infection (CDI). Samples were tested with both the reference method (Combined Direct and Enriched Culture) and the GenePOC™ CDiff assay. The direct culture method consisted in the transfer of a swab of the stool specimen from Anaerobic Transport Medium to pre-reduced selective anaerobic media, a standard cycloserine cefoxitin and fructose agar plate (CCFA), followed by cytotoxicity testing on characterized C. difficile colonies isolated from stool. For the enriched culture method, the same swab that was utilized to inoculate the CCFA plate was used to inoculate a cycloserine cefoxitin mannitol broth with taurocholate and lysozyme (CCMB-TAL) tube. The enrichment broth was sub-cultured on another CCFA plate and followed the same procedure used for the direct method. A specimen was considered positive for toxigenic C. difficile if C. difficile was recovered from stool either by direct or enriched culture and if bacterial isolates tested positive by CCNA. If C. difficile was isolated from the direct culture and the isolate tested positive by cytotoxicity testing, the enrichment culture was not further analyzed. Specimens were classified as negative for toxigenic C. difficile only if they tested negative by both direct, and combined culture i.e. direct and enriched culture.

The study design consisted of testing fresh samples and prospectively testing retrospectively collected samples that were stored frozen.

Subjects	All SubjectsN=2461	FreshN=797	FrozenN=1664
Source of specimens
In-patient	1804 (73,3%)	617 (77,4%)	1187 (71,3%)
Out-patient	420 (17,1%)	123 (15,4%)	297 (17,8%)
Emergency	234 (9,5%)	57 (7,2%)	177 (10,6%)
Missing	3 (0,1%)	0 (0,0%)	3 (0,2%)
Age Class
< 2	9 (0,4%)	4 (0,5%)	5 (0,3%)
3-18	105 (4,3%)	30 (3,8%)	75 (4,5%)
19-60	1199 (48,7%)	399 (50,1%)	800 (48,1%)
> 60	1148 (46,6%)	364 (45,7%)	784 (47,1%)

The study population demographics are presented below.

Two thousand four hundred and sixty-three (2,463) specimens were used to establish the performance of the GenePOC CDiff test by comparison with Combined Direct and Enriched Culture method. All 2,463 freshly collected specimens were tested in culture, 798 were tested with GenePOC™ CDiff assay as fresh specimens and a subset of 1,665 specimens were frozen before testing with the GenePOC™M CDiff assay.

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Of the 798 fresh and 1.665 frozen eligible specimens that were compliant at the specimen and PCR level, 9 and 13 were respectively reported unresolved at initial testing (1,1% for the fresh specimens and 0,8% for the frozen specimens) and only one fresh specimen remained unresolved following repeat testing. The unresolved rate after repeat testing was 0.1% (1/798) for the fresh specimens and 0.0% (0/1665) for the frozen specimens.

Of the 798 fresh and 1,665 frozen eligible specimens that were compliant at the specimen and PCR level, 12 and 28 were respectively reported indeterminate at initial testing (1,5% for the fresh specimens and 1,7% for the frozen specimens) and only one frozen specimen remained indeterminate following repeat testing. The indeterminate rate after repeat testing was 0,0% (0/798) for the fresh specimen and 0,1% (1/1665) for the frozen specimens.

A total of 2,461 fully compliant fresh (n=797) and frozen (n=1,664) samples were tested with both the reference method (Combined Direct and Enriched Culture) and the GenePOC™ CDiff assay. The GenePOC™ CDiff assay sensitivity and specificity obtained with the fresh stool specimens were 80.5% (91/113) and 97.1% (664/684), respectively. The GenePOC™ CDiff assay sensitivity and specificity obtained with the frozen stool specimens were 87.3% (192/220) and 97.3% (1,405/1,444) respectively. In addition, the GenePOC™ CDiff assay agreement with the Direct Culture method was 95.5% (63/66) for the PPA and 93.4% (683/731) for the NPA for the fresh stool specimens, whereas for the frozen stool specimens, 95.2% (160/168) for the PPA and 95.3% (1,425/1,496) for the NPA were obtained

Overall performance (all sites combined) with the GenePOC™ CDiff assay in comparison with the Direct Culture and Reference Method (Combined Direct and Enriched Culture) obtained with fresh specimens

Overall performance		Direct Culture		Total	Reference Method		Total
		Positive	Negative		Positive	Negative
GenePOCTMCDiff test	Positive	63	48	111	91	20B	111
	Negative	3	683	686	22C	664	686
Total		66	731	797	113	684	797
PPAA		95.5% [87.3 - 99.1%]		n/a
NPA		93.4% [91.4 - 95.1%]		n/a
Sensitivity		n/a		80.5% [72.0 - 87.4%]
Specificity		n/a		97.1% [95.5 - 98.2%]

A Numbers between parentheses indicate exact binomial 95% CI.

B Of the 20 specimens with false-positive GenePOC CDiff test results relative to the combined direct and enrichment culture 8 were positive and 4 were negative by a second NAAT method (Sites' Routine PCR Assay).

C Of the 22 specimens with false-negative GenePOC CDiff test results relative to the combined direct and enrichment culture 13 were negative and 4 were positive by a second NAAT method (Sites' Routine PCR Assay).

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Overall performance (all sites combined) with the GenePOC™ CDiff assay in comparison with the Direct Culture and Reference Method (combined Direct and Enriched Culture) obtained with frozen specimens

Overall performance		Direct Culture		Total	Reference Method		Total
		Positive	Negative		Positive	Negative
GenePOCTM	Positive	160	71	231	192	39B	231
CDiff test	Negative	8	1,425	1,433	28C	1,405	1,433
	Total	168	1,496	1,664	220	1,444	1,664
PPAA		95.2% [90.8 - 97.9%]			n/a
NPA		95.3% [94.1 - 96.3%]			n/a
Sensitivity		n/a			87.3%	[82.1 - 91.4%]
Specificity		n/a			97.3%	[96.3 - 98.1%]

A Numbers between parentheses indicate exact binomial 95% CI.

B Of the 39 specimens with false-positive GenePOC CDiff test results relative to the combined direct and enrichment culture 17 were positive and 15 were negative by a second NAAT method (Sites' Routine PCR Assay).

C Of the 28 specimens with false-negative GenePOC CDiff test results relative to the combined direct and enrichment culture 14 were negative and 12 were positive by a second NAAT method (Sites' Routine PCR Assay).

1. Clinical Cut-off
  Not applicable.

L. CONCLUSION:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

Regulation Number and Section

§ 866.3130 Clostridium difficile toxin gene amplification assay.

(a)
Identification. AClostridium difficile toxin gene amplification assay is a device that consists of reagents for the amplification and detection of target sequences inClostridium difficile toxin genes in fecal specimens from patients suspected of havingClostridium difficile infection (CDI). The detection of clostridial toxin genes, in conjunction with other laboratory tests, aids in the clinical laboratory diagnosis of CDI caused byClostridium difficile. (b)
Classification. Class II (special controls). The special controls are set forth in FDA's guideline document entitled: “Class II Special Controls Guideline: Toxin Gene Amplification Assays for the Detection ofClostridium difficile; Guideline for Industry and Food and Drug Administration Staff.” See § 866.1(e) for information on obtaining this document.