Predicate Devices

Reference Devices

AI/ML (Artificial Intelligence / Machine Learning)

No
The summary describes a standard multiplexed nucleic acid-based in vitro diagnostic test and an automated system for sample processing and detection. There is no mention of AI or ML being used for data analysis, interpretation, or any other function within the device or its associated system.

Therapeutic

No.
The device is an in vitro diagnostic test intended as an aid in the diagnosis of gastrointestinal illness by detecting and identifying nucleic acids from various pathogens. It is not designed for treatment or therapy.

Diagnostic

Yes

Explanation: The "Intended Use / Indications for Use" section explicitly states, "The BioCode Gastrointestinal Pathogen Panel (GPP) is a qualitative multiplexed nucleic acid-based in vitro diagnostic test intended for use with the BioCode MDx 3000 Instrument" and "The BioCode GPP is indicated as an aid in the diagnosis of gastrointestinal illness".

SaMD (Software as a Medical Device)

No

The device is an in vitro diagnostic test that requires a specific hardware instrument (BioCode MDx 3000 Instrument) for operation, including PCR amplification, target capture, signal generation, and optical detection. It is not solely software.

IVD (In Vitro Diagnostic)

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

Explicitly Stated in Intended Use: The very first sentence of the "Intended Use / Indications for Use" section clearly states: "The BioCode Gastrointestinal Pathogen Panel (GPP) is a qualitative multiplexed nucleic acid-based in vitro diagnostic test intended for use with the BioCode MDx 3000 Instrument."
Purpose of the Test: The test is designed to detect and identify nucleic acids from pathogens in stool samples to aid in the diagnosis of gastrointestinal illness. This is a classic function of an in vitro diagnostic device, which is used to examine specimens from the human body to provide information for diagnosis, treatment, or prevention of disease.
Use with an Instrument: The device is intended for use with the BioCode MDx 3000 Instrument, which is a system designed for laboratory testing.
Analysis of Biological Samples: The test analyzes nucleic acids extracted from stool samples, which are biological specimens from the human body.

All of these points align with the definition and purpose of an in vitro diagnostic device.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The BioCode Gastrointestinal Pathogen Panel (GPP) is a qualitative multiplexed nucleic acid-based in vitro diagnostic test intended for use with the BioCode MDx 3000 Instrument. The BioCode GPP is capable of the simultaneous detection and identification of nucleic acids from multiple bacteria, viruses, and parasites extracted directly from unpreserved stool samples or stool preserved in Cary-Blair transport medium obtained from individuals with signs and/or symptoms of gastrointestinal infection. The following bacteria, parasites, and viruses are identified using the BioCode Gastrointestinal Pathogen Panel:

· Campylobacter (C. jejuni/C. coli)
· Clostridium difficile (C. difficile) toxin A/B (Fresh samples only)
· Salmonella spp
· Vibrio (V. parahaemolyticus/V. vulnificus/ V. cholerae), including specific identification of Vibrio parahaemolyticus
· Yersinia enterocolitica
· Enteroaggregative Escherichia coli (EAEC)
· Enterotoxigenic Escherichia coli (ETEC) lt/st
· E. coli 0157 serogroup
Shiga-like toxin-producing Escherichia coli (STEC) stx1/stx2
Shigella/ Enteroinvasive Escherichia coli (EIEC)
· Cryptosporidium spp (C. parvum/C. hominis)
Entamoeba histolytica
· Giardia lamblia (also known as G. intestinalis and G. duodenalis)
Adenovirus F 40/41
Norovirus GI/GII
Rotavirus A

The BioCode GPP is indicated as an aid in the diagnosis of gastrointestinal illness and results are meant to be used in conjunction with other clinical, laboratory, and epidemiological data.

Positive results do not rule out co-infection with organisms not included in the BioCode GPP. The agent detected may not be the definite cause of the disease. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease.

Concomitant culture is necessary for organism recovery and further typing of bacterial agents.

This device is not intended to monitor or guide treatment for C. difficile infection.

Due to the small number of positive specimens collected for certain organisms during the prospective clinical study, performance characteristics for Adenovirus 40/41, Campylobacter, E. coli 0157, Shigella/EIEC, Yersinia enterocolitica, and Giardia lamblia were established additionally with retrospective clinical specimens. Performance characteristics for Entamoeba histolytica, Giardia lamblia, Yersinia enterocolitica and Vibrio (V. parahaemolyticus, V. vulnificus, and V. cholerae) were established primarily using contrived clinical specimens.

Product codes (comma separated list FDA assigned to the subject device)

PCH, OOI

Device Description

The BioCode Gastrointestinal Pathogen Panel (GPP) is a multiplexed nucleic acid-based test designed to be used with the BioCode MDx 3000 system. The BioCode MDx 3000 is an automated system that integrates PCR amplification, target capture, signal generation and optical detection for multiple gastrointestinal pathogens from a single stool specimen, either unpreserved or in Cary Blair. Stool specimens are processed and nucleic acids extracted with the easyMAG and MagNa Pure. Once the PCR plate is set up and sealed, all other operations are automated on MDx 3000. The BioCode MDx 3000 Gastrointestinal Infection Panel simultaneously tests for 17 pathogens (see table below) from unpreserved stool specimens or stool collected in Cary-Blair transport medium. Results from the BioCode Gastrointestinal Pathogen Panel (GPP) test are available within less than 4 hours.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Not Found

Indicated Patient Age Range

Not Found

Intended User / Care Setting

Prescription Use (Part 21 CFR 801 Subpart D)

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

Clinical Investigational Study:
A clinical investigational study was performed in which a total of 466 leftover, de-identified samples (275 frozen unpreserved and 191 inoculated Cary-Blair) that were prospectively collected for the clinical study that resulted in the K180041 BioCode GPP clearance were extracted using the MagNA Pure 96 and the easyMAG, and tested on the MDx 3000 system. Fifty-three (53) freshly collected leftover samples were used for the C. difficile testing. In addition, a total of 120 samples were contrived and tested to determine the performance characteristics for Entamoeba histolytica, Yersinia enterocolitica and Vibrio (V. parahaemolyticus, V. vulnificus, and V. cholerae).

Reference Methods from BioCode GPP original submission:

Adenovirus 40/41: Composite result of PCR/sequencing
Campylobacter (C. jejuni, C. coli): Culture
Clostridium difficile (C. difficile) toxin A/B: FDA cleared NAT
Cryptosporidium (C. parvum, C. hominis): PCR/sequencing
Entamoeba histolytica: PCR/sequencing
Escherichia coli (E. coli) 0157: Enrichment culture
Enteropathogenic E. coli (EPEC): Composite result of PCR/sequencing
Enterotoxigenic E. coli (ETEC) LT/ST: Composite result of PCR/sequencing
Enteroaggregative E. coli (EAEC): Composite result of PCR/sequencing
Giardia lamblia /intestinalis: Composite result of PCR/sequencing
Norovirus GI/GII: Composite result of PCR/sequencing
Rotavirus A: Composite result of PCR/sequencing
Salmonella: Enrichment culture
Shiga-like Toxin producing E. coli (STEC) stx1/stx2: Enrichment culture/cleared antigen test
Shigella (S. boydii, S. sonnei, S. flexneri, S. dysenteriae)/EIEC: Enrichment culture
Vibrio spp. (V. cholerae, V. parahaemolyticus, V. vulnificus): Culture
Yersinia enterocolitica: Culture

Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

Clinical Performance Study:
A clinical investigational study was performed with a total of 466 leftover, de-identified archived samples (275 frozen unpreserved and 191 inoculated Cary-Blair) and 53 freshly collected leftover samples (for C. difficile) tested with the BioCode MDx 3000 system after extraction using MagNA Pure 96 and easyMAG. Additionally, 120 contrived samples were tested for Entamoeba histolytica, Yersinia enterocolitica, and Vibrio.

Sample Size: Archived Samples: 466 (275 frozen unpreserved, 191 inoculated Cary-Blair). Fresh Samples (for C. difficile): 53. Contrived Samples: 120.
Key Results:
- Archived Samples Performance (compared to historical Reference results), detailed in a table for each target and extraction method (EasyMAG and MP96), showing Positive Percent Agreement (PPA) and Negative Percent Agreement (NPA). Sample sizes varied per target.
- Fresh Samples Performance (C. difficile and other targets), detailed in a table for Unpreserved (Fresh) samples, showing PPA and NPA.
- Contrived Specimen Results for Vibrio parahaemolyticus, Vibrio spp. (not parahaemolyticus), Yersinia enterocolitica, Entamoeba histolytica, and all other targets (N/A for PPA).
- Clinical Co-infection Summary: 246/466 (52.8%) had +1 Target, 78/466 (16.7%) had +2 Targets, and 13/466 (2.8%) had +3 Targets.

Analytical Performance Studies:

Reproducibility Study:
- Sample Size: Not explicitly stated as a number of samples, but involved 7 contrived samples, extracted in triplicate and each assayed in singlet, with 30 runs (2 runs/day for 5 days by 3 operators on 3 instruments).
- Key Results: Reproducibility was > 99%. All results were as expected with the exception of two false negative results for Cryptosporidium parvum. Detailed tables show % Agreement with Expected Result for various organisms at 1.5x LoD (Low) and 3x LoD (Medium) concentrations, and qualitative/quantitative MFI reproducibility across instruments/operators.
Single vs. Multi-spike Study:
- Key Results: Demonstrated equivalent LoD when spiking with single versus multiple organisms (Clostridium difficile, Salmonella enterica ssp. enterica, Giardia intestinalis, Rotavirus A). Both single and multi-spiked samples achieved ≥95% detection of 20 replicates (≥19 out of 20) at the same concentrations. LoD for the remaining organisms were performed in pairs, except for norovirus.
Limit of Detection (LoD) Study:
- Study Type: Analytical.
- Sample Size: 20 replicates for confirmation of LoD.
- Key Results: LoD for each stock was defined as the lowest concentration with ≥95% detection of 20 replicates (19 out of 20), determined separately for unpreserved stool and Cary-Blair preserved stool. LoDs were the same or within 2-fold for each extraction system (easyMAG and MagNA Pure 96) for various bacteria, parasites, and viruses. For Norovirus GI and GII, positive clinical specimens were used for serial dilutions, and LoDs were compared between easyMAG and MagNA Pure 96 extractions for unpreserved stool and Cary-Blair stool.

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

Clinical Sensitivity (Positive Agreement): TP/(TP + FN). TP = true positive by both the EasyMag and the MagNA Pure 96; FN = false negative by the MagNA Pure 96 only.
Clinical Specificity (Negative Agreement): TN/(TN + FP). TN = true negative or negative by the EasyMag and the MagNA Pure 96; FP = false positive by the MagNA Pure 96 only.
Exact binomial two-sided 95% confidence interval was calculated for PPA and NPA in clinical studies.
Reproducibility: Expressed as % Agreement with Expected Result and MFI Reproducibility (Mean, StDev, %CV, Min, Max).
Limit of Detection (LoD) Detection: Expressed as the number of positive detections out of 20 replicates (e.g., 20/20, 19/20).

Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.

K180041 – BioCode Gastrointestinal Pathogen Panel (GPP)

Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).

Not Found

Regulation Number and Section

§ 866.3990 Gastrointestinal microorganism multiplex nucleic acid-based assay.

(a)
Identification. A gastrointestinal microorganism multiplex nucleic acid-based assay is a qualitativein vitro diagnostic device intended to simultaneously detect and identify multiple gastrointestinal microbial nucleic acids extracted from human stool specimens. The device detects specific nucleic acid sequences for organism identification as well as for determining the presence of toxin genes. The detection and identification of a specific gastrointestinal microbial nucleic acid from individuals exhibiting signs and symptoms of gastrointestinal infection aids in the diagnosis of gastrointestinal infection when used in conjunction with clinical evaluation and other laboratory findings. A gastrointestinal microorganism multiplex nucleic acid-based assay also aids in the detection and identification of acute gastroenteritis in the context of outbreaks.(b)
Classification. Class II (special controls). The special controls are set forth in FDA's guideline document entitled: “Class II Special Controls Guideline: Gastrointestinal Microorganism Multiplex Nucleic Acid-Based Assays for Detection and Identification of Microorganisms and Toxin Genes from Human Stool Specimens.” For availability of the guideline document, see § 866.1(e).

Summary

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

June 5, 2019

Applied Biocode. Inc. Robert Tullio Regulatory Consultant 10020 Pioneer Blvd. Suite 102 Santa Fe Springs, California 90067-0

Re: K190585

Trade/Device Name: Biocode Gastrointestinal Pathogen Panel (GPP) Regulation Number: 21 CFR 866.3990 Regulation Name: Gastrointestinal microorganism multiplex nucleic acid-based assay Regulatory Class: Class II Product Code: PCH, OOI Dated: March 4, 2019 Received: March 6, 2019

Dear Robert Tullio:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database located at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

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

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

1

requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

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

Enclosure

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Indications for Use

510(k) Number (if known) K190585

Device Name

BioCode Gastrointestinal Pathogen Panel (GPP)

Indications for Use (Describe)

The BioCode Gastrointestinal Pathogen Panel (GPP) is a qualitative multiplexed nucleic acid-based in vitro diagnostic test intended for use with the BioCode MDx 3000 Instrument. The BioCode GPP is capable of the simultaneous detection and identification of nucleic acids from multiple bacteria, viruses, and parasites extracted directly from unpreserved stool samples or stool preserved in Cary-Blair transport medium obtained from individuals with signs and/or symptoms of gastrointestinal infection. The following bacteria, parasites, and viruses are identified using the BioCode Gastrointestinal Pathogen Panel:

· Campylobacter (C. jejuni/C. coli)
· Clostridium difficile (C. difficile) toxin A/B (Fresh samples only)
· Salmonella spp
· Vibrio (V. parahaemolyticus/V. vulnificus/ V. cholerae), including specific identification of Vibrio parahaemolyticus
· Yersinia enterocolitica
· Enteroaggregative Escherichia coli (EAEC)
· Enterotoxigenic Escherichia coli (ETEC) lt/st
· E. coli 0157 serogroup
Shiga-like toxin-producing Escherichia coli (STEC) stx1/stx2
Shigella/ Enteroinvasive Escherichia coli (EIEC)
· Cryptosporidium spp (C. parvum/C. hominis)
Entamoeba histolytica
· Giardia lamblia (also known as G. intestinalis and G. duodenalis)
Adenovirus F 40/41
Norovirus GI/GII
Rotavirus A

The BioCode GPP is indicated as an aid in the diagnosis of gastrointestinal illness and results are meant to be used in conjunction with other clinical, laboratory, and epidemiological data.

Positive results do not rule out co-infection with organisms not included in the BioCode GPP. The agent detected may not be the definite cause of the disease. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease.

Concomitant culture is necessary for organism recovery and further typing of bacterial agents.

This device is not intended to monitor or guide treatment for C. difficile infection.

Due to the small number of positive specimens collected for certain organisms during the prospective clinical study, performance characteristics for Adenovirus 40/41, Campylobacter, E. coli 0157, Shigella/EIEC, Yersinia enterocolitica, and Giardia lamblia were established additionally with retrospective clinical specimens. Performance characteristics for Entamoeba histolytica, Giardia lamblia, Yersinia enterocolitica and Vibrio (V. parahaemolyticus, V. vulnificus, and V. cholerae) were established primarily using contrived clinical specimens.

3

X Prescription Use (Part 21 CFR 801 Subpart D)

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

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510(k) SUMMARY

Introduction: According to the requirements of 21 CFR 807.92, the following provides sufficient information to understand the basis for a determination of substantial equivalence.

Submitted by:

Applied BioCode, Inc. 10020 Pioneer Blvd. Suite 102 Santa Fe Springs, CA 90670

Contact:

Robert Di Tullio Regulatory Consultant rditullio@apbiocode.com Telephone: 310 801 1235 Fax: 323 372 3816

Date Submitted:

March 5, 2019

Trade Name:

BioCode Gastrointestinal Pathogen Panel (GPP)

Classification Name and Regulation Number:

Gastrointestinal microorganism multiplex nucleic acid-based assay (21 CFR 866.3990)

Predicate Device:

K180041 – BioCode Gastrointestinal Pathogen Panel (GPP)

Intended Use:

The BioCode Gastrointestinal Pathogen Panel (GPP) is a qualitative multiplexed nucleic acid-based in vitro diagnostic test intended for use with the BioCode MDx 3000 Instrument. The BioCode GPP is capable of the simultaneous detection and identification of nucleic acids from multiple bacteria, viruses, and parasites extracted directly from unpreserved stool samples or stool preserved in Cary-Blair transport medium obtained from individuals with signs and/or symptoms of gastrointestinal infection. The following bacteria, parasites, and viruses are identified using the BioCode Gastrointestinal Pathogen Panel:

" Campylobacter (C. jejuni/C. coli)
Clostridium difficile (C. difficile) toxin A/B (Fresh samples only)
l Salmonella spp
" Vibrio (V. parahaemolyticus/V. vulnificus/ V. cholerae), including specific identification of Vibrio parahaemolyticus
Yersinia enterocolitica
" Enteroaggregative Escherichia coli (EAEC)

5

" Enterotoxigenic Escherichia coli (ETEC) It/st
י E. coli 0157 serogroup
. Shiga-like toxin-producing Escherichia coli (STEC) stx1/stx2
. Shigella/ Enteroinvasive Escherichia coli (EIEC)
" Cryptosporidium spp (C. parvum/C. hominis)
. Entamoeba histolytica
" Giardia lamblia (also known as G. intestinalis and G. duodenalis)
" Adenovirus F 40/41
" Norovirus GI/GII
. Rotavirus A

The BioCode GPP is indicated as an aid in the diagnosis of gastrointestinal illness and results are meant to be used in conjunction with other clinical, laboratory, and epidemiological data.

Positive results do not rule out co-infection with organisms not included in the BioCode GPP. The agent detected may not be the definite cause of the disease. Negative results in the setting of clinical illness compatible with gastroenteritis may be due to infection by pathogens that are not detected by this test or non-infectious causes such as ulcerative colitis, irritable bowel syndrome, or Crohn's disease. Concomitant culture is necessary for organism recovery and further typing of bacterial agents. This device is not intended to monitor or guide treatment for C. difficile infection.

Due to the small number of positive specimens collected for certain organisms during the prospective clinical study, performance characteristics for Adenovirus 40/41, Campylobacter, E. coli 0157, Shigella/EIEC, Yersinia enterocolitica, and Giardia lamblia were established additionally with retrospective clinical specimens. Performance characteristics for Entamoeba histolytica, Giardia lamblia, Yersinia enterocolitica and Vibrio (V. parahaemolyticus, V. vulnificus, and V. cholerae) were established primarily using contrived clinical specimens.

Device Description:

The BioCode Gastrointestinal Pathogen Panel (GPP) is a multiplexed nucleic acid-based test designed to be used with the BioCode MDx 3000 system. The BioCode MDx 3000 is an automated system that integrates PCR amplification, target capture, signal generation and optical detection for multiple gastrointestinal pathogens from a single stool specimen, either unpreserved or in Cary Blair. Stool specimens are processed and nucleic acids extracted with the easyMAG and MagNa Pure. Once the PCR plate is set up and sealed, all other operations are automated on MDx 3000. The BioCode MDx 3000 Gastrointestinal Infection Panel simultaneously tests for 17 pathogens (see table below) from unpreserved stool specimens or stool collected in Cary-Blair transport medium. Results from the BioCode Gastrointestinal Pathogen Panel (GPP) test are available within less than 4 hours.

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Bacteria, Viruses, Diarrheagenic E. coli/Shigella, and Parasites Detected by the BioCode MDx Gastrointestinal Pathogen Panel (GPP)

Bacteria	Parasites
■ Campylobacter spp. (C. jejuni, C. coli)	■ Cryptosporidium spp.
■ Clostridium difficile toxin A/B (Fresh samples only)	■ Entamoeba histolytica
■ Enteroaggregative E. coli (EAEC)	■ Giardia lamblia
■ Enterotoxigenic E. coli (ETEC): LT/ST	Viruses
■ Shiga-toxin producing E. coli (STEC): stx1/stx2	■ Adenovirus 40/41
■ E.coli O157	■ Norovirus GI/GII
■ Shigella spp. /Enteroinvasive E.coli (EIEC)	■ Rotavirus A
■ Salmonella spp.	RNA Internal Control
■ Vibrio parahaemolyticus
■ Vibrio spp (not parahaemolyticus)
■ Yersinia enterocolitica

Device Comparison:

Comparison of the Applied BioCode GPP with the Predicate Device

Characteristic	Proposed Device	Predicate
Name	BioCode Gastrointestinal Pathogen Panel (GPP)	BioCode
Gastrointestinal
Pathogen Panel
(GPP)
Common
Name	Gastrointestinal Microorganism Multiplex Nucleic acid-based
assay	Gastrointestinal
Microorganism
Multiplex Nucleic
acid-based assay
510(k) No.	N/A	K180041
Regulation	21CFR 866.3990	21CFR 866.3990
Product Code	PCH, OOI	PCH, OOI
Device Class	II	II
Similarities

Intended Use	The BioCode Gastrointestinal Pathogen Panel (GPP) is a
qualitative, multiplexed nucleic acid-based in vitro diagnostic test
intended for use with the BioCode MDx 3000 Instrument. The
BioCode GPP is capable of the simultaneous detection and
identification of nucleic acids from multiple bacteria, viruses, and
parasites extracted directly from unpreserved stool samples or
stool preserved in Cary-Blair transport medium obtained from
individuals with signs and/or symptoms of gastrointestinal
infection. The following bacteria, parasites, and viruses are
identified using the BioCode Gastrointestinal Pathogen Panel:
Campylobacter ( C. jejuni/C. coli ) Clostridium difficile ( C. difficile ) toxin A/B (Fresh samples only) Salmonella spp Vibrio ( V. parahaemolyticus/V. vulnificus/ V. cholerae ),
including specific identification of Vibrio parahaemolyticus Yersinia enterocolitica Enteroaggregative Escherichia coli (EAEC) Enterotoxigenic Escherichia coli (ETEC) lt/st E. coli O157 serogroup Shiga-like toxin-producing Escherichia coli (STEC) stx1/stx2 Shigella/ Enteroinvasive Escherichia coli (EIEC) Cryptosporidium spp ( C. parvum/C. hominis ) Entamoeba histolytica Giardia lamblia (also known as G. intestinalis and G. duodenalis ) Adenovirus F 40/41 Norovirus GI/GII Rotavirus A The BioCode GPP is indicated as an aid in the diagnosis of
specific agents of gastrointestinal illness and results are meant to
be used in conjunction with other clinical, laboratory, and
epidemiological data. For In Vitro Diagnostic Use Only. For
Prescription Use Only.

Positive results do not rule out co-infection with organisms
not included in the BioCode GPP. The agent detected may not be
the definite cause of the disease. Negative results in the setting
of clinical illness compatible with gastroenteritis may be due to
infection by pathogens that are not detected by this test or non-
infectious causes such as ulcerative colitis, irritable bowel
syndrome, or Crohn's disease. Concomitant culture is necessary
for organism recovery and further typing of bacterial agents. This
device is not intended to monitor or guide treatment for C. difficile infection. | Same |

7

Premarket Notification 510(k)

8

	Premarket Notification 510(k)
--	-------------------------------	--

Characteristic	Proposed Device	Predicate
Instrument	Nucleic Acid Purification System
BioCode MDx 3000	Same
Sample Type	Unpreserved stool and stool in Cary Blair Medium	Same
Controls	Externally Sourced - Zeptometrix	Same
Methodology	Multiplex RT-PCR and probe hybridization to biotinylated PCR
product(s) followed by fluorescence detection and decoding of
barcoded magnetic beads (BMB) that are coupled to target-
specific probes	Same
Calibrators	Internal Calibration	Same
	Difference
Sample
Extraction	Roche MagNA Pure 96	easyMAG

Summary of Performance Characteristics of the Biocode GPP.

Clinical Performance

A clinical investigational study was performed in which a total of 466 leftover, de-identified samples (275 frozen unpreserved and 191 inoculated Cary-Blair) that were prospectively collected for the clinical study that resulted in the K180041 BioCode GPP clearance were extracted using the MagNA Pure 96 and the easyMAG, and tested on the MDx 3000 system. Fifty-three (53) freshly collected leftover samples were used for the C. difficile testing. In addition, a total of 120 samples were contrived and tested to determine the performance characteristics for Entamoeba histolytica, Yersinia enterocolitica and Vibrio (V. parahaemolyticus, V. vulnificus, and V. cholerae). The results of the clinical investigational study follow.

Archived Samples
Total Specimen Count	466
Gender
Male	248/466 (53.22%)
Female	211/466 (45.28%)
Unknown	7/466 (1.50%)
Age Category
≤ 5 yrs	85/466 (18.24%)
6-21 yrs	91/466 (19.53%)
22-59 yrs	194/466 (41.63%)
60+yrs	89/466 (19.10%)
Unknown	7/466 (1.50%)

Table. Demographic data for archived specimens (frozen unpreserved and inoculated Cary-Blair)

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Fresh Samples
Total Specimen Count	53
Gender
Male	9/53 (16.98%)
Female	44/53 (83.02%)
Age Category
≤ 5 yrs	1/53 (1.89%)
6-21 yrs	5/53 (9.43%)
22-59 yrs	23/53 (43.40%)
60+yrs	24/53 (45.28%)

Table. Demographic data for fresh specimens (unpreserved)

Clinical sensitivity (positive agreement) was calculated as TP/(TP + FN). TP = true positive by both the EasyMag and the MagNA Pure 96; FN = false negative by the MagNA Pure 96 only. Clinical specificity (negative agreement) was calculated as TN/(TN + FP). TN = true negative or negative by the EasyMag and the MagNA Pure 96; FP = false positive by the MagNA Pure 96 only. The exact binomial two-sided 95% confidence interval was calculated. The results stratified by sample type and storage method are presented in the table below.

Table. Performance compared to historical Reference results. Stratified by sample type and extraction method. N=290 samples. 1 Invalid for EasyMAG (Cary-Blair sample); 3 Invalid for MP96 (1 Cary-Blair, 2 Unpreserved).

		EasyMAG		MP96
Target	Specimen Type	Positive
Agreement
PPA (%)	Negative
Agreement
NPA (%)	Positive
Agreement
PPA (%)	Negative
Agreement
NPA (%)
Campylobacter spp.	Inoculated
Cary-Blair	3/3 (100%)	135/140 (96%)	3/3 (100%)	134/140 (96%)
	Unpreserved	1/1 (100%)	139/145 (96%)	1/1 (100%)	135/143 (94%)
	All Archived	4/4 (100%)	274/285 (96%)	4/4 (100%)	269/283 (95%)
Clostridium difficile	Inoculated
Cary-Blair	9/12 (75%)	131/131 (100%)	9/12 (75%)	131/131 (100%)
	Unpreserved	12/13
(92%)	131/133 (98%)	12/13 (92%)	127/131 (97%)
	All Archived	21/25
(84%)	262/264 (99%)	21/25 (84%)	258/262 (98%)
E. coli 0157	Inoculated
Cary-Blair	1/1 (100%)	141/142 (99%)	0/1 (N/A)	141/142 (99%)
	Unpreserved	0/0 (N/A)	144/146 (99%)	0/0 (N/A)	142/144 (99%)
	All Archived	1/1 (100%)	285/288 (99%)	0/1 (N/A)	283/286 (99%)
Enteroaggregative E.
coli (EAEC)	Inoculated
Cary-Blair	14/14
(100%)	128/129 (99%)	13/14 (93%)	127/129 (98%)	Unpreserved	13/14
(93%)	131/132 (99%)	13/14 (93%)	130/131 (99%)
	Enteroaggregative E.
coli (EAEC)	Inoculated
Cary-Blair	14/14
(100%)	128/129 (99%)	13/14 (93%)	127/129 (98%)
Unpreserved		13/14
(93%)	131/132 (99%)	13/14 (93%)	130/131 (99%)

10

		EasyMAG		MP96
Target	Specimen Type	Positive
Agreement
PPA (%)	Negative
Agreement
NPA (%)	Positive
Agreement
PPA (%)	Negative
Agreement
NPA (%)
	All Archived	27/28
(96%)	259/261 (99%)	26/28 (93%)	257/260 (99%)
Enterotoxigenic E. coli
(ETEC)	Inoculated
Cary-Blair	3/6 (50%)	137/137 (100%)	3/6 (50%)	137/137 (100%)
	Unpreserved	6/7 (86%)	139/139 (100%)	6/7 (86%)	138/138 (100%)
	All Archived	9/13 (69%)	276/276 (100%)	9/13 (69%)	275/275 (100%)
Shiga toxin-producing
E. coli (STEC)	Inoculated
Cary-Blair	1/3 (33%)	139/140 (99%)	1/3 (33%)	139/140 (99%)
	Unpreserved	0/0 (N/A)	144/146 (99%)	0/0 (N/A)	142/144 (99%)
	All Archived	1/3 (33%)	283/286 (99%)	1/3 (33%)	281/284 (99%)
Salmonella spp.	Inoculated
Cary-Blair	8/10 (80%)	129/133 (97%)	9/10 (90%)	126/133 (95%)
	Unpreserved	10/10
(100%)	132/136 (97%)	10/10 (100%)	131/134 (98%)
	All Archived	18/20
(90%)	261/269 (97%)	19/20 (95%)	257/267 (96%)
Shigella/ EIEC	Inoculated
Cary-Blair	3/3 (100%)	134/140 (96%)	3/3 (100%)	134/140 (96%)
	Unpreserved	2/2 (100%)	136/144 (94%)	2/2 (100%)	135/142 (95%)
	All Archived	5/5 (100%)	270/284 (95%)	5/5 (100%)	269/282 (95%)
Vibrio
parahaemolyticus	Inoculated
Cary-Blair	0/0 (N/A)	142/143 (99%)	0/0 (N/A)	142/143 (99%)
	Unpreserved	0/0 (N/A)	145/146 (99%)	0/0 (N/A)	143/144 (99%)
	All Archived	0/0 (N/A)	287/289 (99%)	0/0 (N/A)	285/287 (99%)
Vibrio spp. (not
parahaemolyticus)	Inoculated
Cary-Blair	0/0 (N/A)	143/143 (100%)	0/0 (N/A)	143/143 (100%)
	Unpreserved	0/0 (N/A)	146/146 (100%)	0/0 (N/A)	144/144 (100%)
	All Archived	0/0 (N/A)	289/289 (100%)	0/0 (N/A)	287/287 (100%)
Yersinia enterocolitica	Inoculated
Cary-Blair	0/0 (N/A)	141/143 (99%)	0/0 (N/A)	141/143 (99%)
	Unpreserved	0/0 (N/A)	146/146 (100%)	0/0 (N/A)	144/144 (100%)
	All Archived	0/0 (N/A)	287/289 (99%)	0/0 (N/A)	285/287 (99%)
Cryptosporidium spp.	Inoculated
Cary-Blair	3/4 (75%)	139/139 (100%)	2/4 (50%)	139/139 (100%)
	Unpreserved	3/4 (75%)	142/142 (100%)	3/4 (75%)	140/140 (100%)
	All Archived	6/8 (75%)	281/281 (100%)	5/8 (63%)	279/279 (100%)
Entamoeba histolytica	Inoculated
Cary-Blair	0/0 (N/A)	143/143 (100%)	0/0 (N/A)	143/143 (100%)
Target	Specimen Type	EasyMAG
Positive
Agreement
PPA (%)	EasyMAG
Negative
Agreement
NPA (%)	MP96
Positive
Agreement
PPA (%)	MP96
Negative
Agreement
NPA (%)
Giardia lamblia	Unpreserved	0/0 (N/A)	146/146 (100%)	0/0 (N/A)	144/144 (100%)
	All Archived	0/0 (N/A)	289/289 (100%)	0/0 (N/A)	287/287 (100%)
	Inoculated
Cary-Blair	1/1 (100%)	141/142 (99%)	1/1 (100%)	141/142 (99%)
Giardia lamblia	Unpreserved	1/1 (100%)	142/145 (98%)	1/1 (100%)	140/143 (98%)
Giardia lamblia	All Archived	2/2 (100%)	283/287 (99%)	2/2 (100%)	281/285 (99%)
Adenovirus 40/41	Inoculated
Cary-Blair	1/2 (50%)	142/142 (100%)	1/2 (50%)	138/141 (98%)
	Unpreserved	5/6 (83%)	137/140 (98%)	5/6 (83%)	134/138 (97%)
	All Archived	6/8 (75%)	279/282 (99%)	6/8 (75%)	272/279 (97%)
Norovirus (GI/GII)	Inoculated
Cary-Blair	19/19
(100%)	124/124 (100%)	19/19 (100%)	122/124 (98%)
	Unpreserved	19/19
(100%)	125/127 (98%)	19/19 (100%)	124/125 (99%)
	All Archived	38/38
(100%)	249/251 (99%)	38/38 (100%)	246/249 (99%)
Rotavirus A	Inoculated
Cary-Blair	11/11
(100%)	130/132 (98%)	11/11 (100%)	132/133 (99%)
	Unpreserved	9/9 (100%)	135/137 (99%)	9/9 (100%)	130/135 (96%)
	All Archived	20/20
(100%)	264/269 (99%)	20/20 (100%)	262/268 (98%)

11

Footnotes (D = Detected, N = Not Detected)

| | | | Historical
Results
(K180041) | MP96
Extraction
(K190585) | | |
|---------------------------------------------|--------------------|---------------|------------------------------------------------|------------------------------------------|---------------|------------------------------------------------------------------------------------------------------------------------------------|
| Target | Speci
men
ID | Refere
nce | BioC
ode
GPP | easy
MAG | MP
96 | Comment |
| | 01-
0027 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq C. jejuni, but contig too short;
concordant with historical GPP result |
| Campylobacter
spp | 01-
0048 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 01-
0137 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 01-
0237 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 01-
0352 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 02-
0159 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 02-
0183 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 02-
0367 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq not detected, concordant with
historical GPP result |
| | 03-
0153 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq not detected, concordant with
historical GPP result |
| | | | Historical
Results
(K180041) | MP96
Extraction
(K190585) | | |
| Target | Specimen
ID | Reference | BioCode
GPP | easy
MAG | MP
96 | Comment |
| Clostridium
difficile | 03-
0308 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 03-
0334 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; concordant with
historical GPP result |
| | 02-
0069 | N | D | N | D | Reference assay (culture) not as sensitive, PCR/Seq C. jejuni; MP96 concordant
with historical GPP result |
| | 02-
0193 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 03-
0364 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 01-
0031 | N | D | D | D | Reference Ct beyond cut-off (38.7); concordant with historical GPP Result |
| | 01-
0063 | N | D | D | D | Reference Ct beyond cut-off with ref (38.3); concordant with historical GPP Result |
| | 02-
0201 | D | N | D | N | Late Ct with ref (36.6) tested fresh, low positive; see repeat results for additional
informations |
| | 02-
0350 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 03-
0053 | D | D | N | D | Late Ct with ref (36.2) tested fresh; low positive; see repeat results for additional
informations |
| | 03-
0351 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 01-
0002 | D | N | N | N | Late Ct with ref (36.6) tested fresh; concordant with historical GPP Result (frozen) |
| | 02-
0378 | D | N | N | N | Late Ct with ref (36.6) tested fresh; concordant with historical GPP Result (fresh) |
| | 03-
0353 | D | D | N | N | Late Ct with ref (35.5) tested frozen; possible sample degradation |
| E. coli 0157 | 02-
0080 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq O157; concordant with
historical GPP result |
| | 02-
0082 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq O157; concordant with
historical GPP result |
| | 02-
0206 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq 0157; concordant with
historical GPP result |
| | 02-
0279 | D | D | N | N | Possible sample degradation; current testing concordant |
| | 03-
0140 | N | D | D | D | Late Ct with ref (37.1) on repeat testing; concordant with historical GPP Result |
| | 02-
0183 | N | N | D | N | Possible low positive; see repeat results for additional information |
| Enteroaggregative E. coli (EAEC) | 03-
0004 | D | D | D | N | Possible low positive; see repeat results for additional information |
| | 02-
0367 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 03-
0046 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| Enterotoxigenic
E. coli (ETEC) | 02-
0203 | D | N | N | N | Reference result could not be confirmed by 2 additional rounds of PCR/Seq;
concordant with historical GPP Result |
| | 02-
0203 | D | N | N | N | Reference result could not be confirmed by 2 additional rounds of PCR/Seq;
concordant with historical GPP Result |
| | 02-
0207 | D | N | N | N | Late Ct (35.2) Reference result could not be confirmed by 2 additional rounds of
PCR/Seq; concordant with historical GPP Result |
| | 03-
0004 | D | N | N | N | Reference result could not be confirmed by 2 additional rounds of PCR/Seq;
concordant with historical GPP Result |
| | 02-
0391 | D | D | N | N | Late Ct (37.8) with ref tested fresh; possible sample degradation |
| Salmonella spp. | 01-
0091 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.enterica; concordant with
historical GPP result |
| | | | Historical
Results
(K180041) | MP96
Extraction
(K190585) | | |
| Target | Speci
men
ID | Refere
nce | BioC
ode
GPP | easy
MAG | MP
96 | Comment |
| | 01-
0137 | N | N | D | N | Possible Contamination; see repeat results |
| | 01-
0278 | D | D | N | D | See repeat results for additional information |
| | 01-
0346 | D | D | N | N | Possible sample degradation; current testing concordant |
| | 02-
0195 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0213 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.enterica; concordant with
historical GPP result |
| | 02-
0260 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0378 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 03-
0116 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.enterica; concordant with
historical GPP result |
| | 03-
0147 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.enterica; concordant with
historical GPP result |
| | 03-
0173 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.typhi; concordant with
historical GPP result |
| | 03-
0189 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.enterica; concordant with
historical GPP result |
| | 03-
0206 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq S.typhi; concordant with
historical GPP result |
| Shiga toxin-
producing E. coli
(STEC) | 01-
0098 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq STEC; concordant with historical
GPP result |
| | 02-
0367 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq STEC; concordant with historical
GPP result |
| | 02-
0393 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq STEC; concordant with historical
GPP result |
| | 02-
0279 | D | D | D | N | See repeat results for additional information |
| | 02-
0081 | D | D | N | N | Possible sample degradation; current testing concordant |
| | 01-
0019 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0020 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0027 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0099 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0199 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| Shigella/EIEC | 01-
0201 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0202 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0259 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 02-
0176 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 03-
0056 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 03-
0090 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 03-
0238 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | | | Historical
Results
(K180041) | MP96
Extraction
(K190585) | | |
| Target | Specimen
ID | Reference | Biocode
GPP | easy
MAG | MP
96 | Comment |
| | 03-
0380 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq Shigella spp; concordant with
historical GPP result |
| | 01-
0319 | N | N | D | N | Possible low positive; see repeat results for additional information |
| Vibrio
parahaemolyticus | 03-
0194 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq V.parahaemolyticus; concordant
with historical GPP result |
| | 03-
0351 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq V.parahaemolyticus; concordant
with historical GPP result |
| Yersinia
enterocolitica | 01-
0024 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq V.parahaemolyticus; concordant
with historical GPP result |
| | 03-
0307 | N | D | D | D | Reference assay (culture) not as sensitive, PCR/seq not detected; concordant with
historical GPP result |
| | 01-
0212 | D | D | D | N | See repeat results for additional information |
| Cryptosporidium
parvum | 01-
0265 | D | D | N | N | Possible sample degradation; current testing concordant |
| | 01-
0285 | D | D | N | N | Possible sample degradation; current testing concordant |
| | 01-
0002 | N | D | D | D | Reference assay (PCR/Seq) not as sensitive; concordant with historical GPP result |
| Giardia
intestinalis | 01-
0300 | N | D | D | D | Reference assay (PCR/Seq) not as sensitive; concordant with historical GPP result |
| | 02-
0258 | N | D | D | D | Reference assay (PCR/Seq) not as sensitive; concordant with historical GPP result |
| | 03-
0140 | N | D | D | D | Reference assay (PCR/Seq) not as sensitive; concordant with historical GPP result |
| Human
adenovirus
40/41 | 01-
0128 | N | N | D | D | concordant with historical GPP result |
| | 01-
0021 | N | N | D | N | Possible Contamination; see repeat results for additional information |
| | 02-
0288 | N | N | D | N | Possible Contamination; see repeat results for additional information |
| | 01-
0137 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0161 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0183 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0379 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0391 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 03-
0400 | N | N | N | D | Possible Contamination; see repeat results for additional information |
| | 02-
0137 | D | N | N | N | Reference result could not be confirmed by 2 additional rounds of PCR/Seq;
concordant with historical GPP Result |
| | 02-
0147 | D | D | N | N | Possible sample degradation; current testing concordant |
| Norovirus GI/GII | 01-
0004 | N | D | D | D | concordant with historical GPP result |
| | 01-
0036 | N | N | D | N | Low positive based on MFI (3150) |
| | 02-
0376 | N | N | N | D | Low positive based on MFI (1265) |
| | 02-
0394 | N | N | N | D | Low positive based on MFI (2249) |
| Human rotavirus
A | 01-
0128 | N | N | D | D | Current testing concordant |
| Target | Specimen ID | Reference | Historical Results
(K180041)
BioCode GPP | MP96 Extraction
(K190585)
easy MAG | MP96
MP 96 | Comment |
| | 01-0337 | N | D | D | D | concordant with historical GPP result |
| | 02-0175 | N | D | D | D | concordant with historical GPP result |
| | 02-0246 | N | D | D | N | Possible low positive; see repeat results for additional information |
| | 01-0098 | N | D | N | D | Possible low positive; see repeat results for additional information |
| | 01-0202 | N | N | N | D | Possible Contamination/low positive; see repeat results for additional information |
| | 03-0400 | N | N | N | D | Possible Contamination; see repeat results for additional information |

12

13

14

15

Comparator (Reference) methods from BioCode GPP original submission

Target Pathogen/Toxin

Target Pathogen/Toxin	Reference Method
Adenovirus 40/41	Composite result of PCR/sequencing
Campylobacter (C. jejuni, C. coli)	Culture
Clostridium difficile (C. difficile) toxin A/B	FDA cleared NAT
Cryptosporidium (C. parvum, C. hominis)	PCR/sequencing
Entamoeba histolytica	PCR/sequencing
Escherichia coli (E. coli) 0157	Enrichment culture
Enteropathogenic E. coli (EPEC)	Composite result of PCR/sequencing
Enterotoxigenic E. coli (ETEC) LT/ST	Composite result of PCR/sequencing
Enteroaggregative E. coli (EAEC)	Composite result of PCR/sequencing
Giardia lamblia /intestinalis	Composite result of PCR/sequencing
Norovirus GI/GII	Composite result of PCR/sequencing
Rotavirus A	Composite result of PCR/sequencing
Salmonella	Enrichment culture
Shiga-like Toxin producing E. coli (STEC) stx1/stx2	Enrichment culture/cleared antigen test
Shigella (S. boydii, S. sonnei, S. flexneri, S. dysenteriae)/EIEC	Enrichment culture
Vibrio spp. (V. cholerae, V. parahaemolyticus, V. vulnificus)	Culture
Yersinia enterocolitica	Culture

| Target | Specimen
Type | (n) | Positive Agreement | | Negative Agreement | |
|-----------------------------------------------|--------------------------|-----|--------------------|-------------|--------------------|----------------|
| | | | PPA (%) | 95% CI | NPA (%) | 95% CI |
| Campylobacter
spp.a | Inoculated
Cary-Blair | 190 | 22/23 (95.65) | 79.0 – 99.2 | 164/167 (98.20) | 94.9 – 99.4 |
| | Unpreserved
(Frozen) | 274 | 27/27 (100) | 87.5 - 100 | 244/247 (98.80) | 96.5 - 99.6 |
| | All Archived | 464 | 49/50 (98.0) | 89.5 - 99.6 | 408/414 (98.60) | 96.9 – 99.3 |
| Clostridium
difficile b | Inoculated
Cary-Blair | 190 | 10/11 (90.91) | 62.3 – 98.4 | 178/179 (99.44) | 96.9 – 99.9 |
| | Unpreserved
(Frozen) | 274 | 21/22 (95.45) | 78.2 – 99.2 | 250/252 (99.20) | 97.2 – 99.8 |
| Target | Specimen Type | (n) | Positive Agreement | | Negative Agreement | |
| | All Archived | 464 | 31/33 (93.94) | 80.4 - 98.3 | 428/431 (99.30) | 98.0 - 99.8 |
| | Inoculated
Cary-Blair | 190 | 3/3 (100) | 43.9 - 100 | 187/187 (100) | 98.0 - 100 |
| E. coli O157 | Unpreserved
(Frozen) | 274 | 14/14 (100) | 78.5 - 100 | 260/260 (100) | 98.5 - 100 |
| | All Archived | 464 | 17/17 (100) | 81.6 - 100 | 447/447 (100) | 99.1 - 100 |
| | Inoculated
Cary-Blair | 190 | 15/17 (88.24) | 65.7 - 96.7 | 171/173 (98.8) | 95.9 - 99.7 |
| Enteroaggregative
E. coli (EAEC) c | Unpreserved
(Frozen) | 274 | 29/29 (100) | 88.3 - 100 | 244/245 (99.59) | 97.7 - 99.9 |
| | All Archived | 464 | 44/46 (95.65) | 85.5 - 98.8 | 416/418 (99.50) | 98.3 - 99.9 |
| | Inoculated
Cary-Blair | 190 | 3/5 (60.00) | 23.1 - 88.2 | 185/185 (100) | 98.0 - 100 |
| Enterotoxigenic
E. coli (ETEC) d | Unpreserved
(Frozen) | 274 | 13/13 (100) | 77.2 - 100 | 261/261 (100) | 98.5 - 100 |
| | All Archived | 464 | 16/18 (88.89) | 67.2 - 96.9 | 446/446 (100) | 99.1 - 100 |
| Shiga toxin-
producing E. coli
(STEC) e | Inoculated
Cary-Blair | 190 | 12/13 (92.31) | 66.7 - 98.6 | 177/177 (100) | 97.9 - 100 |
| | Unpreserved
(Frozen) | 274 | 29/30 (96.67) | 83.3 - 99.4 | 243/244 (99.60) | 97.7 - 99.9 |
| | All Archived | 464 | 41/43 (95.35) | 84.5 - 98.7 | 420/421 (99.80) | 98.7 - 100 |
| | Inoculated
Cary-Blair | 190 | 17/17 (100) | 81.6 - 100 | 169/173 (97.70) | 94.2 - 99.1 |
| Salmonella spp.f | Unpreserved
(Fresh) | 274 | 25/27 (92.59) | 76.6 - 97.9 | 246/247 (99.60) | 97.7 - 99.9 |
| | All Archived | 464 | 42/44 (95.45) | 84.9 - 98.7 | 415/420 (98.80) | 97.2 - 99.5 |
| Shigella/ EIEC g | Inoculated
Cary-Blair | 190 | 9/9 (100) | 70.1 - 100 | 181/181 (100) | 97.9 - 100 |
| | Unpreserved
(Frozen) | 274 | 20/22 (90.91) | 72.2 - 97.5 | 252/252 (100) | 98.5 - 100 |
| | All Archived | 464 | 29/31 (93.55) | 79.3 - 98.2 | 433/433 (100) | 99.1 - 100 |
| Vibrio
parahaemolyticus h | Inoculated
Cary-Blair | 190 | 1/1 (100) | 20.7 - 100 | 189/189 (100) | 98.0 - 100 |
| | Unpreserved
(Frozen) | 274 | 1/1 (100) | 20.7 - 100 | 272/273 (99.6) | 98.0 - 99.9 |
| | All Archived | 464 | 2/2 (100) | 34.2 - 100 | 461/462 (99.8) | 99.8 - 100 |
| Vibrio spp. (not
parahaemolyticus)i | Inoculated
Cary-Blair | 190 | N/A | N/A | 190/190 (100) | 98.0 - 100 |
| | Unpreserved
(Frozen) | 274 | 0/1 (0%) | N/A | N/A | 273/274 (99.6) |
| | All Archived | 464 | 0/1 (0%) | N/A | 463/464 (99.8) | 98.8 - 100 |
| Yersinia
enterocolitica j | Inoculated
Cary-Blair | 190 | 3/3 (100) | 43.9 - 100 | 187/187 (100) | 98.0 - 100 |
| | Unpreserved
(Frozen) | 274 | 3/3 (100) | 43.9 - 100 | 269/271 (99.26) | 97.3 - 99.8 |
| | All Archived | 464 | 6/6 (100) | 61.0 - 100 | 456/458 (99.6) | 98.4 - 99.9 |
| | Specimen
Type | (n) | Positive Agreement | | Negative Agreement | |
| Target | | | PPA (%) | 95% CI | NPA (%) | 95% CI |
| Cryptosporidium
spp.k | Inoculated
Cary-Blair | 190 | 11/12 (91.67) | 64.6 – 98.5 | 178/178 (100) | 97.9 - 100 |
| | Unpreserved
(Frozen) | 274 | 22/24 (91.67) | 74.2 - 97.7 | 248/250 (99.20) | 97.1 - 99.8 |
| | All Archived | 464 | 33/36 (91.67) | 78.2 – 97.1 | 426/428 (99.5) | 98.3 - 99.9 |
| Entamoeba
histolytica | Inoculated
Cary-Blair | 190 | N/A | N/A | 190/190 (100) | 98.0 - 100 |
| | Unpreserved
(Frozen) | 274 | N/A | N/A | 274/274 (100) | 98.6 - 100 |
| | All Archived | 464 | N/A | N/A | 464/464 (100) | 99.2 - 100 |
| Giardia lamblial | Inoculated
Cary-Blair | 190 | 3/3 (100) | 43.90 - 100 | 187/187 (100) | 98.0 - 100 |
| | Unpreserved
(Frozen) | 274 | 14/14 (100) | 78.5 - 100 | 255/260 (98.1) | 95.6 - 99.2 |
| | All Archived | 464 | 17/17 (100) | 81.6 - 100 | 442/447 (98.9) | 97.4 - 99.5 |
| Adenovirus 40/41m | Inoculated
Cary-Blair | 190 | 7/10 (70.00) | 39.7 – 89.2 | 177/180 (98.32) | 95.2 - 99.4 |
| | Unpreserved
(Frozen) | 274 | 11/14 (78.60) | 52.4 - 92.4 | 252/260 (96.90) | 94.0 - 98.4 |
| | All Archived | 464 | 21/24 (87.50) | 69.0 - 95.7 | 429/440 (97.5) | 95.6 - 98.6 |
| Norovirus
(GI/GII)n | Inoculated
Cary-Blair | 190 | 19/19 (100) | 83.2 - 100 | 168/171 (98.20) | 95.0 - 99.4 |
| | Unpreserved
(Frozen) | 274 | 21/22 (95.45) | 78.2 – 99.2 | 248/252 (98.40) | 96.0 - 99.4 |
| | All Archived | 464 | 37/41 (90.24) | 77.5 - 96.1 | 416/423 (98.3) | 96.6 - 99.2 |
| Rotavirus Ao | Inoculated
Cary-Blair | 190 | 12/13 (92.31) | 66.7 – 98.6 | 176/177 (99.44) | 96.9 - 99.9 |
| | Unpreserved
(Frozen) | 274 | 15/15 (100) | 79.6 - 100 | 255/259 (98.5) | 96.1 - 99.4 |
| | All Archived | 464 | 27/28 (96.43) | 82.3 - 99.4 | 431/436 (98.9) | 97.3 - 99.5 |

Table. Summary of Clinical Investigational Study Results (Archived Specimens) stratified by sample type and storage

16

17

Sixty-four (64) archived samples with discordant results were retested twice with both easyMag and/or MagNA Pure 96 systems.

a - Campylobacter spp. One (1) false negative retested became true positives retested, 2 were true negative, one became true positive, two remained false positive not retested due to insufficient volume to retest.

b - Clostridium difficile: Of the 2 false negatives retested, 1 became true positive, the other one was true negative. Of the 3 false positives retested, 2 were true negative and 1 was true positive.

c - EAEC: Of the 2 false negatives retested, 1 was false positive. Of the 3 false positive. Of the 3 false positives retested, 2 were true negative and 1 was true positive.

d - ETEC: Two (2) false negatives tested were true negative.

e - STEC: Of 3 false positives retested, 2 were true positive and 1 was true negative.

f - Salmonella spp. Of the 2 false negatives retested, one was false positive and the other one was true negative. Of the 5 false positives retested, 4 were true negative and 1 was true positive.

g - Shigella/EIEC: Of the 2 false negatives retested, 1 was true positive and the other remained false negative.

h - Vibrio parahaemolyticus: The 1 false positive retested was true negative.

i - Vibrio spp. The 1 false negative retested remained false negative. Detected as Vibrio parahaemolyticus by MP96.

j - Yersinia enterocolitica: Two (2) false positives retested were true negative.

k - Cryptosporidium spp: Of the 3 false negative and 2 were false negative and 2 were false negative. The 2 false positives retested remained false positive.

18

l - Giardia lamblia: Of the 5 false positives retested, 1 became true negative. The remaining 2 were not retested due insufficient volume to retest.

m- Adenovirus 40/41: All 6 false negatives. Of the 11 false positives, 9 were retested and became true negative, and the remaining 2 were not retested due insufficient volume to retest.

n - Norovirus G1/G2: The 1 false negative retested was true negatives retested, 6 were true negative and one became true positive.

o - Rotavirus: The 1 false negative retested was true negatives retested, 1 was true positive, 3 were true negative, and 1 remained false positive.

| Target | Sample Name | Sample
Type | Extraction | Original Run | Reflex Testing
Result | Final Result |
|---------------------------|-------------|--------------------------|---------------|--------------|--------------------------|--------------|
| | 01-0079 | Inoculated
Cary-Blair | EasyMag | IC Invalid | IC Valid | Valid |
| Internal Control
(MS2) | 01-0073 | Inoculated
Cary-Blair | MagNA
Pure | IC Invalid | IC Invalid | Invalid |
| | 02-0211 | Unpreserved | MagNa
Pure | IC Invalid | IC Invalid | Invalid |
| | 02-0228 | Unpreserved | EasyMag | IC Invalid | IC Valid | Valid |
| | Mayo-G1_124 | Unpreserved | MagNA
Pure | IC Invalid | IC Valid | Valid |

Table. Results of Reflex Testing of Invalid Samples (Archived Samples)

Note. Five (5) archived samples were invalid. After relflex testing, two samples (1 inoculated Cary-Blair and 1 unpreserved) were still invalid.

Target	Specimen	(n)	Positive Agreement		Negative Agreement
	Type		PPA (%)	95% CI	NPA (%)	95% CI
Campylobacter spp.	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Clostridium difficile	Unpreserved
(Fresh)	52	30/30 (100)	88.6-100	22/22 (100)	85.1-100
E. coli 0157	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Enteroaggregative E.
coli (EAEC)	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Enterotoxigenic
E. coli (ETEC)	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Shiga toxin-producing
E. coli (STEC)	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Salmonella spp.	Unpreserved
(Fresh)	47	1/1 (100)	20.7-100	46/46 (100)	92.3-100
Shigella/ EIEC	Unpreserved
(Fresh)	47	1/1 (100)	20.7-100	46/46 (100)	92.3-100
Vibrio
parahaemolyticus	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Vibrio spp. (not
parahaemolyticus)	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100
Yersinia enterocolitica	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4-100

Table. Summary of Clinical Investigational Study Results of Fresh Specimens

19

Target	Specimen	(n)	Positive Agreement		Negative Agreement
	Type		PPA (%)	95% CI	NPA (%)	95% CI
Cryptosporidium spp.	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4- 100
Entamoeba histolytica	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4- 100
Giardia lamblia¹	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4- 100
Adenovirus 40/41	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4- 100
Norovirus
(GI/GII)	Unpreserved
(Fresh)	47	1/1 (100)	20.7 -100	46/46 (100)	92.3- 100
Rotavirus A	Unpreserved
(Fresh)	47	N/A	N/A	47/47(100)	92.4- 100

Note. Fifty-three (53) fresh unpreserved samples were tested. The Internal Control (MS2) result of the 5 C. difficile positives and 1 C. difficile negative were invalid. Therefore, one sample for C. difficile target was invalid. For the remaining targets, 6 samples were invalid.

Table. Summary of Contrived Specimen Results

Target	Specimen Type	(n)	Positive Agreement		Negative Agreement
			PPA (%)	95% CI	NPA (%)	95% CI
Vibrio parahaemolyticus a	Unpreserved (Frozen)	119	28/30 (93.3)	78.7 – 98.2	89/89 (100)	95.9 – 100
Vibrio spp. (not parahaemolyticus) b	Unpreserved (Frozen)	118	29/29 (100)	88.3 - 100	89/89 (100)	95.9 - 100
Yersinia enterocolitica	Unpreserved (Frozen)	118	30/30 (100)	88.6 – 100	88/88 (100)	95.8 – 100
Entamoeba histolytica	Unpreserved (Frozen)	118	30/30 (100)	88.6 – 100	88/88 (100)	96.8 – 100
All other targets	Unpreserved (Frozen)	118	N/A	N/A	118/118 (100)	96.8 – 100

Note. One hundred twenty (120) contrived samples were tested. ª The Internal control (MS2) result of one of the Vibrio parahaemolyticus positives was invalid. * One (1) of the Vibrio spp. positives was negative and invalid (IC negative) with both the easyMag and the MagNA Pure 96.

Table. Clinical Co-infection Summary

Co-infection summary
+1 Target	246/466 (52.8%)
+2 Targets	78/466 (16.7%)
+3 Targets	13/466 (2.8%)

Analytical Performance

20

The results of the analytical studies summarized in the following paragraphs met the acceptance criteria and successfully demonstrated the analytical performance characteristics of the proposed BioCode GPP using the MagNA Pure 96 extraction system.

REPRODUCIBILITY STUDY

A study was performed to assess the Reproducibility of the BioCode GPP using samples extracted with the MagNA Pure 96. This study was designed to assess intra-assay (within run), Inter-assay (run-to-run), day-to-day and instrument-to-instrument (operator) reproducibility. One lot of reagents was assayed at Applied BioCode on 3 instruments by 3 operators, 2 runs per day per operator for 5 days (total of 30 runs). The reproducibility panel consisted of 7 contrived samples (sample 7 a negative control) extracted in triplicate and each assayed in singlet. The samples consisted of combinations of 12 representative targets at 1.5x LoD (Low) and 3x LoD (Medium). Reproducibility was > 99%.

All results are as expected with the exception of two false negative results for Cryptosporidium parvum (one low positive and one medium positive).


Organism Tested	Target
Probe	Concentration Tested	Expected
Results	Instrument1

Operator 1 | Instrument2
Operator 2 | Instrument3
Operator 3 | All
Instruments/
Operators |
| Campylobacter
jejuni spp. jejuni
ATCC 33292 | Campy | Medium Positive 3xLoD
1.05 x 103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
5.25 x 102 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Clostridium
difficile
(toxinotype III;
Nap1)
Zeptometrix
0801619cf | tcdB | Medium Positive 3xLoD
1.25 x 103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
6.23 x 102 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Enteroaggregative
E. coli 092:H33
(EAEC)
STEC TW04440 | EAEC | Medium Positive 3xLoD
2.10 x 103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
1.05 x 103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Organism Tested | Target
Probe | Concentration Tested | Expected
Results | Instrument1
Operator 1 | Instrument2
Operator 2 | Instrument3
Operator 3 | All
Instruments/
Operators |
| Enterotoxigenic E.
coli 078:H11
H10407 (ETEC)
ATCC 35401 | ST-1a | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| | | Medium Positive 3xLoD
8.40 x 10² CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
4.20 x 10² CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Salmonella
enterica ssp.
enterica
ATCC 14028 | Salm | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| | | Medium Positive 3xLoD
3.30 x 10³ CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
1.65 x 10³ CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Shiga-like toxin
producing E. coli
(STEC)
ATCC BAA-2217 | stx2 | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| | | Medium Positive 3xLoD
3.75 x 10³ CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
1.88 x10³ CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Shigella sonnei
ATCC 29930 | Shig | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| | | Medium Positive 3xLoD
6.60 x 10² CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | Low Positive 1.5xLoD
3.30 x 10² CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Vibrio
parahaemolyticus
ATCC 17802 | V.para | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| | | Medium Positive 3xLoD
1.95 x10¹ CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | | | % Agreement with Expected Result | | | | |
| Organism Tested | Target
Probe | Concentration Tested | Expected
Results | Instrument1
Operator 1 | Instrument2
Operator 2 | Instrument3
Operator 3 | All
Instruments/
Operators |
| Yersinia
enterocolitica
ATCC 23715 | | Low Positive 1.5xLoD
9.75x10° CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Yersinia
enterocolitica
ATCC 23715 | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Yersinia
enterocolitica
ATCC 23715 | | Medium Positive 3xLoD
2.25 x103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Yersinia
enterocolitica
ATCC 23715 | Y.ent | Low Positive 1.5xLoD
1.13 x103 CFU/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Yersinia
enterocolitica
ATCC 23715 | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Cryptosporidium
parvum
waterborne P102 | | Medium Positive 3xLoD
9.30 x 103 oocysts/mL | Detected | 30/30
(100%) | 30/30
(100%) | 29/30
(97%) | 89/90 (99%) |
| | Crypto | Low Positive 1.5xLoD
4.65 x 103 oocysts/mL | Detected | 30/30
(100%) | 30/30
(100%) | 29/30
(97%) | 89/90 (99%) |
| | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Giardia intestinalis
(aka G. lamblia)
waterborne P101 | | Medium Positive 3xLoD
2.70 x 103 cysts/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | G.lam | Low Positive 1.5xLoD
1.35 x 103 cysts/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Giardia intestinalis
(aka G. lamblia)
waterborne P101 | | None | Not
Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |
| Rotavirus
ATCC VR-2018 | | Medium Positive 3xLoD
3.75 x 103 TCID50/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| | Rota | Low Positive 1.5xLoD
1.88 x 102 TCID50/mL | Detected | 30/30
(100%) | 30/30
(100%) | 30/30
(100%) | 90/90 (100%) |
| Organism Tested | Target Probe | Concentration Tested | Expected Results | % Agreement with Expected Result | | | |
| | | | | Instrument1 - Operator 1 | Instrument2 - Operator 2 | Instrument3 - Operator 3 | All Instruments/ Operators |
| | | None | Not Detected | 150/150
(100%) | 150/150
(100%) | 150/150
(100%) | 450/450
(100%) |

Table. Reproducibility of BioCode GPP with MP96 extractions- Qualitative results.

21

22

23

Table. Reproducibility of BioCode GPP with MP96 extractions- Quantitative results

| Organism | Target
Probe | Test Level | Instrument/Operator | MFI Reproducibility | | | | |
|---------------------------------------------------------------------------------------|-----------------|--------------------------------------------------------------|---------------------------|---------------------|-------|-------|-------|-------|
| | | | | Mean | StDev | %CV | Min | Max |
| Campylobacter jejuni spp. jejuni
ATCC 33292 | Campy | Moderate
Positive
3xLoD
$1.05 x 10^3$
CFU/mL | C2- Operator 1 | 26128 | 4013 | 15 | 8265 | 29801 |
| | | | C3- Operator 2 | 28562 | 2652 | 9 | 22187 | 33021 |
| | | | C4- Operator 3 | 24154 | 2714 | 11 | 18890 | 28822 |
| | | | All Instruments/Operators | 26281 | 3637 | 14 | 8265 | 33021 |
| | | Low Positive
1.5xLoD
$5.25 x 10^2$
CFU/mL | C2- Operator 1 | 24831 | 3563 | 14 | 14326 | 30367 |
| | | | C3- Operator 2 | 25831 | 4285 | 17 | 14406 | 31649 |
| | | | C4- Operator 3 | 21655 | 4326 | 20 | 10566 | 30156 |
| | | | All Instruments/Operators | 24106 | 4407 | 14 | 10566 | 31649 |
| Clostridium difficile
(toxinotype III;
Nap1)
Zeptometrix
0801619cf | tcdB | Moderate
Positive
3xLoD
$1.25 x 10^3$
CFU/mL | C2- Operator 1 | 27589 | 2276 | 8 | 22487 | 31505 |
| | | | C3- Operator 2 | 22672 | 3726 | 16 | 17112 | 32047 |
| | | | C4- Operator 3 | 22780 | 2838 | 11 | 16926 | 27598 |
| | | | All Instruments/Operators | 24347 | 3762 | 15 | 16926 | 32047 |
| | | Low Positive
1.5xLoD
$6.23 x 10^2$
CFU/mL | C2- Operator 1 | 25007 | 1761 | 7 | 20436 | 27552 |
| | | | C3- Operator 2 | 20116 | 3513 | 17 | 13752 | 27461 |
| | | | C4- Operator 3 | 20902 | 2561 | 11 | 15608 | 27662 |
| | | | All Instruments/Operators | 22008 | 3438 | 16 | 13752 | 27662 |
| Enteroaggregati ve E. coli
092:H33 (EAEC)
STEC TW04440 | EAEC | Moderate
Positive
3xLoD
$2.10 x 10^3$
CFU/mL | C2- Operator 1 | 30474 | 1672 | 5 | 25616 | 33694 |
| | | | C3- Operator 2 | 28424 | 3220 | 14 | 20108 | 34967 |
| | | | C4- Operator 3 | 25129 | 2198 | 9 | 18787 | 28489 |
| | | | All Instruments/Operators | 28009 | 3281 | 12 | 18787 | 34967 |
| | | Low Positive
1.5xLoD
$1.05 x 10^3$
CFU/mL | C2- Operator 1 | 29161 | 1492 | 5 | 26343 | 31716 |
| | | | C3- Operator 2 | 25402 | 3630 | 14 | 18723 | 32890 |
| | | | C4- Operator 3 | 22836 | 2538 | 11 | 15300 | 27002 |
| | | | All Instruments/Operators | 25800 | 3734 | 15 | 15300 | 32890 |
| Enterotoxigenic E. coli 078:H11
H10407 (ETEC)
ATCC 35401 | ST-1a | Moderate
Positive
3xLoD
$8.40 x 10^2$
CFU/mL | C2- Operator 1 | 35769 | 1009 | 3 | 33677 | 38306 |
| | | | C3- Operator 2 | 34358 | 4831 | 14 | 25398 | 42656 |
| | | | C4- Operator 3 | 28730 | 3638 | 13 | 22254 | 35493 |
| | | | All Instruments/Operators | 32952 | 4648 | 14 | 22254 | 42656 |
| | | Low Positive
1.5xLoD
$4.20 x 10^2$
CFU/mL | C2- Operator 1 | 32795 | 2240 | 7 | 28536 | 36716 |
| | | | C3- Operator 2 | 30726 | 3040 | 10 | 24219 | 35644 |
| | | | C4- Operator 3 | 24907 | 2765 | 11 | 20236 | 30170 |
| | | | All Instruments/Operators | 29476 | 4291 | 15 | 20236 | 36716 |
| | Salm | | C2- Operator 1 | 15291 | 1496 | 10 | 12588 | 18918 |
| Organism | Target
Probe | Test Level | Instrument/Operator | Mean | StDev | %CV | Min | Max |
| Salmonella
enterica ssp.
enterica
ATCC 14028 | | Moderate
Positive
3xLoD
3.30 x 103
CFU/mL | C3- Operator 2 | 13931 | 2645 | 19 | 6284 | 18489 |
| | | C4- Operator 3 | 15701 | 1456 | 9 | 13306 | 19184 | |
| | | All Instruments/Operators | 14974 | 2068 | 14 | 6284 | 19184 | |
| | | Low Positive
1.5xLoD
1.65 x 103
CFU/mL | C2- Operator 1 | 17247 | 1108 | 6 | 15153 | 19682 |
| | | C3- Operator 2 | 14880 | 1682 | 11 | 10847 | 19127 | |
| | | C4- Operator 3 | 16234 | 1455 | 7 | 13301 | 19674 | |
| | | All Instruments/Operators | 16120 | 1721 | 10 | 10847 | 19682 | |
| Shiga-like toxin
producing E. coli
(STEC)
ATCC BAA-2217 | stx2 | Moderate
Positive
3xLoD
3.75 x 103
CFU/mL | C2- Operator 1 | 27217 | 1112 | 4 | 23606 | 30023 |
| | | C3- Operator 2 | 31888 | 1316 | 4 | 29183 | 34448 | |
| | | C4- Operator 3 | 26310 | 1515 | 6 | 22970 | 30028 | |
| | | All Instruments/Operators | 28472 | 2785 | 10 | 22970 | 34448 | |
| | | Low Positive
1.5xLoD
1.88 x103
CFU/mL | C2- Operator 1 | 27467 | 1547 | 6 | 23914 | 30339 |
| | | C3- Operator 2 | 31460 | 1501 | 5 | 28002 | 34121 | |
| | | C4- Operator 3 | 25293 | 2097 | 8 | 20340 | 29403 | |
| | | All Instruments/Operators | 28073 | 3089 | 11 | 20340 | 34121 | |
| Shigella sonnei
ATCC 29930 | Shig | Moderate
Positive
3xLoD
6.60 x 102
CFU/mL | C2- Operator 1 | 17531 | 1173 | 7 | 15327 | 19469 |
| | | C3- Operator 2 | 15369 | 3781 | 25 | 7573 | 21561 | |
| | | C4- Operator 3 | 16454 | 2056 | 13 | 11910 | 21114 | |
| | | All Instruments/Operators | 16451 | 2697 | 16 | 7573 | 21561 | |
| | | Low Positive
1.5xLoD
3.30 x 102
CFU/mL | C2- Operator 1 | 13919 | 1862 | 13 | 9233 | 17022 |
| | | C3- Operator 2 | 12242 | 2503 | 20 | 6435 | 16434 | |
| | | C4- Operator 3 | 12832 | 2184 | 17 | 8352 | 17489 | |
| | | All Instruments/Operators | 12998 | 2283 | 18 | 6435 | 17489 | |
| Vibrio
parahaemolytic
us ATCC 17802 | V.para | Moderate
Positive
3xLoD
1.95 x101
CFU/mL | C2- Operator 1 | 23605 | 2305 | 10 | 17750 | 29798 |
| | | C3- Operator 2 | 21458 | 3112 | 15 | 13138 | 27526 | |
| | | C4- Operator 3 | 17486 | 2821 | 16 | 8543 | 22789 | |
| | | All Instruments/Operators | 20850 | 3739 | 18 | 8543 | 29798 | |
| | | Low Positive
1.5xLoD
9.75x100
CFU/mL | C2- Operator 1 | 22628 | 2402 | 11 | 17145 | 26762 |
| | | C3- Operator 2 | 18000 | 6067 | 34 | 6043 | 27526 | |
| | | C4- Operator 3 | 16263 | 3060 | 19 | 6989 | 23597 | |
| | | All Instruments/Operators | 18964 | 4922 | 26 | 6043 | 27526 | |
| Yersinia
enterocolitica
ATCC 23715 | Y.ent | Moderate
Positive
3xLoD
2.25 x103
CFU/mL | C2- Operator 1 | 14365 | 1847 | 13 | 10209 | 17892 |
| | | C3- Operator 2 | 15303 | 1786 | 12 | 10813 | 17893 | |
| | | C4- Operator 3 | 14256 | 2325 | 16 | 9523 | 19982 | |
| | | All Instruments/Operators | 14641 | 2033 | 14 | 9523 | 19982 | |
| | | Low Positive
1.5xLoD
1.13 x103
CFU/mL | C2- Operator 1 | 14741 | 1694 | 11 | 11853 | 19628 |
| | | C3- Operator 2 | 15592 | 2014 | 13 | 12978 | 20765 | |
| | | C4- Operator 3 | 14732 | 2253 | 16 | 10750 | 19994 | |
| | | All Instruments/Operators | 15022 | 2019 | 13 | 10750 | 20765 | |
| Organism | Target
Probe | Test Level | Instrument/Operator | Mean | StDev | %CV | Min | Max |
| Cryptosporidium parvum
waterborne
P102 | Crypto | Moderate
Positive
3xLoD
$9.30 x 10^3$
oocysts/mL | C2- Operator 1 | 18731 | 4323 | 23 | 9904 | 28443 |
| | | | C3- Operator 2 | 20843 | 3978 | 19 | 11526 | 28678 |
| | | | C4- Operator 3* | 14867 | 4457 | 31 | 5669 | 26611 |
| | | | All Instruments/Operators | 18222 | 4873 | 26 | 5669 | 28678 |
| | | Low Positive
1.5xLoD
$4.65 x 10^3$
oocysts/mL | C2- Operator 1 | 13816 | 5170 | 37 | 2572 | 24148 |
| | | | C3- Operator 2 | 15402 | 3996 | 26 | 8166 | 22933 |
| | | | C4- Operator 3* | 10785 | 3409 | 33 | 4413 | 18053 |
| | | | All Instruments/Operators | 13363 | 4631 | 35 | 2572 | 24148 |
| Giardia
intestinalis (aka
G. lamblia)
waterborne
P101 | G.lam | Moderate
Positive
3xLoD
$2.70 x 10^3$
cysts/mL | C2- Operator 1 | 14327 | 4487 | 31 | 7696 | 30050 |
| | | | C3- Operator 2 | 21743 | 5006 | 23 | 8932 | 31538 |
| | | | C4- Operator 3 | 16963 | 3780 | 22 | 11527 | 25036 |
| | | | All Instrument/Operators | 17677 | 5377 | 30 | 7696 | 31538 |
| | | Low Positive
1.5xLoD
$1.35 x 10^3$
cysts/mL | C2- Operator 1 | 17340 | 3668 | 21 | 11338 | 25626 |
| | | | C3- Operator 2 | 24751 | 4701 | 19 | 12478 | 31282 |
| | | | C4- Operator 3 | 19821 | 3772 | 20 | 12677 | 25949 |
| | | | All Instruments/Operators | 20637 | 5081 | 25 | 11338 | 31282 |
| Rotavirus
ATCC VR-2018 | Rota | Moderate
Positive
3xLoD
$3.75 x 10^3$
TCID50/mL | C2- Operator 1 | 39523 | 2086 | 5 | 35835 | 44023 |
| | | | C3- Operator 2 | 39162 | 3400 | 9 | 28863 | 44274 |
| | | | C4- Operator 3 | 32899 | 2956 | 8 | 25107 | 38361 |
| | | | All Instruments/Operators | 37195 | 4170 | 13 | 25107 | 44274 |
| | | Low Positive
1.5xLoD
$1.88 x 10^2$
TCID50/mL | C2- Operator 1 | 36515 | 5114 | 14 | 16127 | 40941 |
| | | | C3- Operator 2 | 36312 | 3863 | 11 | 26491 | 41547 |
| | | | C4- Operator 3 | 31925 | 3280 | 10 | 21114 | 36158 |
| | | | All Instruments/Operators | 34917 | 4628 | 11 | 16127 | 41547 |

24

25

*The one replicate with 'Not detected' result was not included in the calculations.

SINGLE VS. MULTI-SPIKE

To demonstrate that the LoD is equivalent when spiking with single versus multiple organisms, LoD for representative organisms (S. enterica, C. difficile, G. lamblia, and Rotavirus) were performed as single targets and with all representative organisms combined, using both extraction systems. The results of spiking single versus multiple organisms showed equivalent LoD (see Table below). LoD for the remaining organisms were performed in pairs, except for norovirus which was tested from positive clinical specimens.

Single and multi-spiked samples both achieved ≥95% detection of 20 replicates (≥19 out of 20) at same concentrations for the challenge organisms (concentrations indicated in the tables below).

Table. Comparison of results for limit of detection testing with single or multiple spiked samples for easyMAG extractions assayed with BioCode GPP.

26

| Strain | Single
Spike/
Multiple
Spike | Source | Target
Probe | Unpreserved
Stool LoD | Unpreserve
d Stool
Detection | Cary-Blair Stool
LoD | Cary-Blair
Stool
Detection |
|---------------------------------------------|---------------------------------------|--------------------|-----------------|----------------------------|------------------------------------|----------------------------|----------------------------------|
| Clostridium
difficile
(toxinotype 0) | Single
Spike | ATCC 9689 | tcdB | $9.50 x 10^1$
CFU/mL | 20/20 | $9.50 x 10^1$
CFU/mL | 20/20 |
| | Multi
Spike | | | | 20/20 | | 19/20 |
| Salmonella
enterica ssp.
enterica | Single
Spike | ATCC 14028 | Salm | $1.10 x 10^3$
CFU/mL | 20/20 | $1.10 x 10^3$
CFU/mL | 20/20 |
| | Multi
Spike | | | | 20/20 | | 20/20 |
| Giardia
intestinalis
(aka G. lamblia) | Single
Spike | waterborne
P101 | G.lam | $9.00 x 10^2$
cysts/mL | 20/20 | $9.00 x 10^2$
cysts/mL | 20/20 |
| | Multi
Spike | | | | 20/20 | | 20/20 |
| Rotavirus A | Single
Spike | ATCC VR-
2018 | Rota | $2.50 x 10^3$
TCID50/mL | 20/20 | $1.25 x 10^3$
TCID50/mL | 20/20 |
| | Multi
Spike | | | | 20/20 | | 20/20 |

Table. Comparison of results for limit of detection testing with single or multiple spiked samples for MP96 extractions assayed with BioCode GPP.

| Strain | Single
Spike/
Multiple
Spike | Source | Target
Probe | Unpreserved
Stool LoD | Unpreserve
d Stool
Detection | Cary-Blair Stool
LoD | Cary-Blair
Stool
Detection |
|---------------------------------------------|---------------------------------------|--------------------|-----------------|----------------------------|------------------------------------|----------------------------|----------------------------------|
| Clostridium
difficile
(toxinotype 0 ) | Single
Spike
Multi
Spike | ATCC 9689 | tcdB | $9.50 x 10^1$
CFU/mL | 20/20
20/20 | $9.50 x 10^1$
CFU/mL | 20/20
20/20 |
| Salmonella
enterica ssp.
enterica | Single
Spike
Multi
Spike | ATCC 14028 | Salm | $1.10 x 10^3$
CFU/mL | 20/20
20/20 | $1.10 x 10^3$
CFU/mL | 20/20
20/20 |
| Giardia
intestinalis
(aka G. lamblia) | Single
Spike
Multi
Spike | waterborne
P101 | G.lam | $9.00 x 10^2$
cysts/mL | 20/20
20/20 | $9.00 x 10^2$
cysts/mL | 20/20
20/20 |
| Rotavirus A | Single
Spike
Multi
Spike | ATCC VR-
2018 | Rota | $1.25 x 10^3$
TCID50/mL | 20/20
20/20 | $1.25 x 10^3$
TCID50/mL | 20/20
20/20 |

LIMIT OF DETECTION (LoD)

27

A study was performed to assess the performance of the BioCode GPP on the BioCode MDx 3000 at the Limit of Detection (LoD) for both unpreserved Stool and Cary-Blair specimens. In this study the Gl Panel was tested with quantified bacteria, virus or parasite stocks (except norovirus which used clinical samples). For initial screening, four replicates of each concentration (near LoD for the predicate) in negative stool and Cary-Blair were extracted on the easyMAG and MagNA Pure 96 Systems and tested in singlet with the BioCode GPP on the BioCode MDx 3000 system. The LoD was confirmed by extracting 20 replicates of each sample type/extraction method and testing each in singlet for a total of 20 replicates at or near presumptive LoD. LoD for each stock was defined as the lowest concentration with ≥95% detection of 20 replicates (19 out of 20), and was determined separately unpreserved stool and Cary-Blair preserved stool. The LoDs determined from the quantitated stocks or clinical samples are presented below and in the labeling. The LoDs were the same or within 2-fold for each extraction system.

		EasyMag		MagNA Pure 96
Strain	Source	Unpreserved Stool
LoD	Detection	Unpreserved Stool
LoD	Detection
Campylobacter coli	ATCC 33559	2.81 x 101 CFU/mL	19/20	2.81 x 101 CFU/mL	20/20
Campylobacter jejuni
spp. jejuni	ATCC 33292	3.50 x 102 CFU/mL	19/20	3.50 x 102 CFU/mL	19/20
Clostridium difficile
(toxinotype 0 )	ATCC 9689	9.50 x 101 CFU/mL	20/20	9.50 x 101 CFU/mL	20/20
Clostridium difficile
(toxinotype III; Nap1)	Zeptometrix
0801619cf	4.15 x 102 CFU/mL	20/20	4.15 x 102 CFU/mL	20/20
Enteroaggregative E.
coli 092:H33 (EAEC)	STEC TW04440	7.00 x 102 CFU/mL	20/20	7.00 x 102 CFU/mL	20/20
Enteroinvasive E. coli
029:NM (EIEC)	ATCC 43892	3.60 x 102 CFU/mL	20/20	1.80 x 102 CFU/mL	20/20
Enterotoxigenic E. coli
078:H11 H10407
(ETEC)	ATCC 35401	2.80 x 102 CFU/mL	20/20	2.80 x 102 CFU/mL	20/20
Salmonella bongori	SGSC 4900	1.40 x 103 CFU/mL	20/20	1.40 x 103 CFU/mL	20/20
Salmonella enterica
ssp. enterica	ATCC 14028	1.10 x 103 CFU/mL	20/20	1.10 x 103 CFU/mL	20/20
Shiga-like toxin
producing E. coli (STEC)	ATCC BAA-2217	1.25 x 103 CFU/mL	20/20	1.25 x 103 CFU/mL	20/20
E. coli 0157	ATCC 700376	1.65 x 103 CFU/mL	20/20	1.65 x 103 CFU/mL	20/20
Shigella sonnei	ATCC 29930	2.20 x 102 CFU/mL	20/20	2.20 x 102 CFU/mL	20/20
Vibrio cholerae	ATCC 25870	2.45 x 102 CFU/mL	20/20	2.45 x 102 CFU/mL	20/20
Vibrio
parahaemolyticus	ATCC 17802	6.50 x 100 CFU/mL	20/20	6.50 x 100 CFU/mL	20/20

Table. Comparison of results for limit of detection testing for Unpreserved Stool extracted with the easyMAG or MP96 and assayed with BioCode GPP.

28

		EasyMag		MagNA Pure 96
Strain	Source	Unpreserved Stool
LoD	Detection	Unpreserved Stool
LoD	Detection
Yersinia enterocolitica	ATCC 23715	$7.50 x 10^2$ CFU/mL	20/20	$7.50 x10^2$ CFU/mL	20/20
Cryptosporidium
parvum	waterborne
P102C	$3.10 x 10^3$ oocysts/mL	20/20	$3.10 x10^3$ oocysts/mL	20/20
Entamoeba histolytica
HB-301:NIH	BEI NR-178	$1.55 x 10^{-1}$ cysts/mL	20/20	$1.55 x10^{-1}$ cysts/mL	20/20
Giardia intestinalis (aka
G. lamblia)	waterborne
P101	$9.00 x 10^2$ cysts/mL	20/20	$9.00 x10^2$ cysts/mL	20/20
Adenovirus 40 (dugan)	Zeptometrix
0810084	$2.00 x 10^{-1}$ TCID50/mL	20/20	$1.00 x10^{-1}$ TCID50/mL	20/20
Adenovirus 41 (TAK)	Zeptometrix
0810085	$9.4 x 10^{-2}$ TCID50/mL	20/20	$4.70 x10^{-2}$ TCID50/mL	20/20
Rotavirus A	ATCC VR-2018	$2.5 x 10^3$ TCID50/mL	20/20	$1.25 x10^3$ TCID50/mL	20/20

Table. Comparison of results for limit of detection testing for Cary-Blair Stool extracted with the easyMAG or MP96 and assayed with BioCode GPP.

Strain	Source	Cary-Blair Stool LoD	Detection	Cary-Blair Stool LoD	Detection
Campylobacter coli	ATCC 33559	$5.61 x 10^1$ CFU/mL	20/20	$2.81 x 10^1$ CFU/mL	20/20
Campylobacter jejuni
spp. jejuni	ATCC 33292	$3.50 x10^2$ CFU/mL	20/20	$7.00 x10^2$ CFU/mL	20/20
Clostridium difficile
(toxinotype 0 )	ATCC 9689	$9.50 x 10^1$ CFU/mL	20/20	$9.50 x 10^1$ CFU/mL	20/20
Clostridium difficile
(toxinotype III; Nap1)	Zeptometrix
0801619cf	$4.15 x10^2$ CFU/mL	20/20	$4.15 x10^2$ CFU/mL	20/20
Enteroaggregative E.
coli 092:H33 (EAEC)	STEC TW04440	$7.00 x10^2$ CFU/mL	20/20	$7.00 x10^2$ CFU/mL	20/20
Enteroinvasive E. coli
029:NM (EIEC)	ATCC 43892	$3.60 x10^2$ CFU/mL	20/20	$1.80 x10^2$ CFU/mL	20/20
Enterotoxigenic E. coli
078:H11 H10407
(ETEC)	ATCC 35401	$2.80 x10^2$ CFU/mL	20/20	$2.80 x10^2$ CFU/mL	19/20
Salmonella bongori	SGSC 4900	$1.40 x10^3$ CFU/mL	19/20	$1.40 x10^3$ CFU/mL	20/20
Salmonella enterica
ssp. enterica	ATCC 14028	$1.10 x10^3$ CFU/mL	20/20	$1.10 x10^3$ CFU/mL	20/20
Shiga-like toxin
producing E. coli (STEC)	ATCC BAA-2217	$1.25 x10^3$ CFU/mL	20/20	$1.25 x10^3$ CFU/mL	20/20
E. coli 0157	ATCC 700376	$3.30 x10^3$ CFU/mL	20/20	$1.65 x10^3$ CFU/mL	20/20

29

	Premarket Notification 510(k)
--	-------------------------------	--

		EasyMag		MagNA Pure 96
Strain	Source	Cary-Blair Stool LoD	Detection	Cary-Blair Stool LoD	Detection
Shigella sonnei	ATCC 29930	2.20 x102 CFU/mL	20/20	2.20 x102 CFU/mL	20/20
Vibrio cholerae	ATCC 25870	2.45 x102 CFU/mL	20/20	2.45 x102 CFU/mL	20/20
Vibrio
parahaemolyticus	ATCC 17802	6.50 x100 CFU/mL	20/20	6.50 x100 CFU/mL	20/20
Yersinia enterocolitica	ATCC 23715	7.50 x102 CFU/mL	20/20	7.50 x102 CFU/mL	20/20
Cryptosporidium
parvum	waterborne
P102C	3.10 x103 oocysts/mL	20/20	3.10 x103 oocysts/mL	20/20
Entamoeba histolytica
HB-301:NIH	BEI NR-178	1.55 x10-1 cysts/mL	20/20	1.55 x10-1 cysts/mL	20/20
Giardia intestinalis (aka
G. lamblia)	waterborne
P101	9.00 x102 cysts/mL	20/20	9.00 x102 cysts/mL	20/20
Adenovirus 40 (dugan)	Zeptometrix
0810084	2.00 x10-1 TCID50/mL	20/20	1.00 x10-1 TCID50/mL	20/20
Adenovirus 41 (TAK)	Zeptometrix
0810085	4.70 x10-2 TCID50/mL	20/20	4.70 x10-2 TCID50/mL	20/20
Rotavirus A	ATCC VR-2018	1.25 x103 TCID50/mL	20/20	1.25 x103 TCID50/mL	20/20

For Norovirus GI and GII targets, positive clinical specimens were used, and serial dilutions (initial 10-fold dilution series followed by finer dilutions) were performed. Four replicates of each dilution in negative unpreserved stool and Cary-Blair stool were extracted with the easyMAG and MagNA Pure 96 Systems and tested with the BioCode GPP on the BioCode MDx 3000 system. The LoD was confirmed by extracting 20 replicates of each sample type with each extraction method and testing at or near presumptive LoD. For unpreserved stool, LoD with the easyMag extraction was 2-fold and 8.3-fold lower than the MagNA Pure 96 extraction for Norovirus GI and Norovirus GII, respectively. For Cary-Blair stool, LoD with the easyMag extraction was less than 2-fold lower than the MagNA Pure 96 extraction for both Norovirus Gl and GII.

Table. Norovirus - Comparison of results for limit of detection testing for Unpreserved Stool extracted with the easyMAG or MP96 and assayed with BioCode GPP.

| Target | Source | Target
Probe | EasyMag | | MagNA Pure 96 | |
|---------------|-----------------------|-----------------|-------------------------------|-----------|-------------------------------|-----------|
| | | | Unpreserved
Stool Dilution | Detection | Unpreserved
Stool Dilution | Detection |
| Norovirus GI | Clinical Sample ID#60 | NoVG1 | 1:10,000 | 20/20 | 1:5,000 | 19/20 |
| Norovirus GII | Clinical Sample ID#54 | NoVG2 | 1:250,000 | 20/20 | 1:30,000 | 20/20 |

Table. Norovirus - Comparison of results for limit of detection testing for Cary-Blair Stool extracted with the easyMAG or MP96 and assayed with BioCode GPP.

30

| Target | Source | Target
Probe | EasyMag | | MagNA Pure 96 | |
|---------------|-----------------------|-----------------|------------------------------|-----------|------------------------------|-----------|
| | | | Cary-Blair
Stool Dilution | Detection | Cary-Blair
Stool Dilution | Detection |
| Norovirus GI | Clinical Sample ID#60 | NoVG1 | 1:50,000 | 20/20 | 1:80,000 | 19/20 |
| Norovirus GII | Clinical Sample ID#54 | NoVG2 | 1:100,000 | 20/20 | 1:80,000 | 20/20 |

Conclusion:

The intended use and fundamental scientific technology of the BioCode GPP using the MagNA Pure 96 is substantially equivalent to the predicate device. Clinical and non-clinical studies have established that the BioCode GPP with the MagNA Pure 96 is substantially equivalent to the predicate device.