(90 days)
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No
The device is an enzyme immunoassay, which is a biochemical test, not a software or imaging-based device that would typically incorporate AI/ML. The summary explicitly states "Not Applicable (Immunoassay, not a machine learning device)" for the description of training and test sets.
No
The device is an in vitro diagnostic test that aids in the diagnosis of gastrointestinal infections by detecting specific antigens in fecal specimens. It does not provide any therapy or treatment.
Yes
The "Intended Use / Indications for Use" section explicitly states that "The test is indicated as an aid in the diagnosis of gastrointestinal infection when giardiasis, cryptosporidiosis and amebiasis is suspected." This directly aligns with the definition of a diagnostic device.
No
The device is an enzyme immunoassay kit, which is a laboratory test involving physical reagents and a microassay plate, not a software-only device.
Yes, this device is an IVD (In Vitro Diagnostic).
Here's why:
- Intended Use: The intended use explicitly states that the test is for the "simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens." This is a classic description of an in vitro diagnostic test, as it analyzes a human specimen (fecal) outside of the body to provide information for diagnosis.
- Device Description: The description details an "enzyme immunoassay" that uses antibodies to detect antigens in a human fecal specimen. This process is performed in vitro (in a lab setting, using reagents and a microassay plate).
- Anatomical Site: The test is performed on "Human fecal specimens," which are collected from the human body but analyzed in vitro.
- Performance Studies: The document describes various performance studies (Prospective, Retrospective, Reproducibility, Analytical Sensitivity, Analytical Specificity, etc.) which are standard for evaluating the performance of IVD devices. These studies assess the device's ability to accurately detect the target analytes in human specimens.
- Predicate Device: The mention of a "Predicate Device(s)" with a K number (K051929; GIARDIA/CRYPTOSPORIDIUM CHEK®) is a strong indicator that this device is being submitted for regulatory review as an IVD, as predicate devices are used for comparison in the regulatory process for new IVDs.
All of these elements align with the definition and characteristics of an In Vitro Diagnostic device.
N/A
Intended Use / Indications for Use
The TECHLAB® TRI-COMBO PARASITE SCREEN test is an enzyme immunoassay for the simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens. The test is indicated as an aid in the diagnosis of gastrointestinal infection when giardiasis, cryptosporidiosis and amebiasis is suspected. The test does not differentiate between the three parasites and follow-up testing is required for all positive results to confirm the specific diagnosis.
Product codes (comma separated list FDA assigned to the subject device)
MHJ
Device Description
The TRI-COMBO PARASITE SCREEN test is an enzyme immunoassay for the simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens. The test uses monoclonal and polyclonal antibodies to cell-surface antigens of Giardia, Cryptosporidium and E. histolytica. The microassay plate in the kit contains immobilized monoclonal antibodies against the antigens, and the Conjugate consists of polyclonal antibodies against the antigens. In the assay, an aliquot of a diluted fecal specimen is transferred to a microassay well. The immobilized monoclonal antibodies bind the Giardia, Cryptosporidium and/or E. histolytica antigens if they are present. Upon addition, Conjugate then binds to the antigen/ antibody complex. Any unbound materials are removed during the washing steps. Following the addition of Substrate, a color is detected due to the enzyme-antibody-antigen complexes that formed in the presence of antigens and conjugate.
Mentions image processing
Not Found
Mentions AI, DNN, or ML
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Input Imaging Modality
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Anatomical Site
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Indicated Patient Age Range
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Intended User / Care Setting
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Description of the training set, sample size, data source, and annotation protocol
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Description of the test set, sample size, data source, and annotation protocol
Prospective Study:
- Sample Size: 754 (total 14 microscopy positive samples (13 Giardia and one E. histolytica), 740 negative samples).
- Data Source: 3 independent sites.
- Annotation Protocol: Microscopy for Giardia, Cryptosporidium and E. histolytica; molecular comparison, FDA cleared antigen test for E. histolytica, PCR with sequencing.
Retrospective Study:
- Sample Size: 96 archived specimens.
- Data Source: One clinical site, samples collected from an E. histolytica endemic area.
- Annotation Protocol: Microscopy and PCR.
Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
Prospective Study
- Study Type: Clinical performance study comparing the TRI-COMBO PARASITE SCREEN test to microscopy for Giardia, Cryptosporidium and E. histolytica.
- Sample Size: 754
- Key Results:
- Sensitivity: 92.9% (95% Confidence Limits: 68.5% - 98.7%)
- Specificity: 98.1% (95% Confidence Limits: 96.9% - 98.9%)
- Specimens Positive for Giardia by microscopy/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 12/13
- Specimens Positive for E. histolytica by microscopy/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 1/1
- The fourteen TRI-COMBO PARASITE SCREEN positives that were microscopy negative were confirmed to be positive for Giardia with an alternate FDA cleared antigen test or by PCR with sequencing.
Retrospective Study
- Study Type: Clinical performance study comparing the TRI-COMBO PARASITE SCREEN test to Microscopy and PCR.
- Sample Size: 96
- Key Results:
- Sensitivity: 94.4% (95% Confidence Limits: 87.7% - 97.9%)
- Specificity: 100% (95% Confidence Limits: 61.0% - 100%)
- Specimens Positive for Giardia/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 41/41
- Specimens Positive for Cryptosporidium/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 27/30
- Specimens Positive for E. histolytica/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 28/30
- Eight specimens were dual positive for Giardia and E. histolytica by the Microscopy and PCR and tested positive in the TRI-COMBO PARASITE SCREEN test. Three specimens were dual positive for Giardia and Cryptosporidium by Microscopy and PCR and tested positive in the TRI-COMBO PARASITE SCREEN test.
Prospective Study (compared to microscopy and molecular testing)
- Study Type: Clinical performance study comparing the TRI-COMBO PARASITE SCREEN test to microscopy and molecular testing.
- Sample Size: 754
- Key Results:
- Positive Percent Agreement: 62.1% (44.0% - 77.3%)
- Negative Percent Agreement: 98.8% (97.7% - 99.4%)
- Specimens Positive for Giardia/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 17/24
- Specimens Positive for E. histolytica/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 1/5
- Specimens Positive for Cryptosporidium/Specimens Detected by the TRI-COMBO PARASITE SCREEN: 0/0
- These nine specimens were tested with an alternate FDA cleared antigen test resulting in 9/9 Giardia determined to be antigen positive.
- These eleven specimens were tested with an alternate FDA cleared antigen test resulting in 6/7 Giardia and 4/4 E. histolytica were determined to be antigen negative.
Reproducibility
- Study Type: Reproducibility study.
- Sample Size: 20 human fecal specimens.
- Key Results: Results were consistent among different locations and exhibited a correlation of 99.9%. The samples produced the expected results 99.9% of the time.
Analytical Sensitivity (Limit of Detection)
- Study Type: LoD determination study.
- Key Results:
- LoD for Giardia: 8450 cysts/mL of feces (169 cysts detected per test).
- LoD for Cryptosporidium: 47,962 oocysts/mL of feces (959 oocysts detected per test).
- LoD for E. histolytica: 1.676 PZs/mL (34 PZs detected per test).
- For fecal matrix/Cary Blair: Giardia 34,155 cysts/mL (427 cysts detected); Cryptosporidium 99,456 oocysts/mL (1243 oocysts detected); E. histolytica 4655 PZs/mL (58 PZs detected per test).
- For fecal matrix/C&S: Giardia 37,095 cysts/mL (464 cysts detected); Cryptosporidium 122,299 oocysts/mL (1529 oocysts detected); E. histolytica 3948 PZs/mL (49 PZs detected per test).
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
Prospective Study (compared to microscopy):
- Sensitivity: 92.9%
- Specificity: 98.1%
Retrospective Study (compared to microscopy and PCR):
- Sensitivity: 94.4%
- Specificity: 100%
Prospective Study (compared to microscopy and molecular testing):
- Positive Percent Agreement: 62.1%
- Negative Percent Agreement: 98.8%
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.
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
§ 866.3220
Entamoeba histolytica serological reagents.(a)
Identification. Entamoeba histolytica serological reagents are devices that consist of antigens and antisera used in serological tests to identify antibodies toEntamoeba histolytica in serum. Additionally, some of these reagents consist of antisera conjugated with a fluorescent dye (immunofluorescent reagents) used to identifyEntamoeba histolytica directly from clinical specimens. The identification aids in the diagnosis of amebiasis caused by the microscopic protozoan parasiteEntamoeba histolytica and provides epidemiological information on diseases caused by this parasite. The parasite may invade the skin, liver, intestines, lungs, and diaphragm, causing disease conditions such as indolent ulcers, an amebic hepatitis, amebic dysentery, and pulmonary lesions.(b)
Classification. Class II (special controls). The device is exempt from the premarket notification procedures in subpart E of part 807 of this chapter subject to § 866.9.
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Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002
July 10, 2017
TECHLAB, INC. DONNA LINK DIRECTOR REGULATORY AND COMPLIANCE 2001 KRAFT DRIVE BLACKSBURG VA 24060-6358
Re: K171078
Trade/Device Name: Tri-combo Parasite Screen Regulation Number: 21 CFR 866.3220 Regulation Name: Entamoeba histolyticaserological reagents Regulatory Class: II Product Code: MHJ, MHI and KHW Dated: April 10, 2017 Received: April 11, 2017
Dear Ms. Link:
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 Parts 801 and 809); medical device reporting (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 Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.
1
If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Industry and Consumer Education at its toll-free number (800) 638 2041 or (301) 796-7100 or at its Internet address
http://www.fda.gov/MedicalDevices/Resourcesfor You/Industry/default.htm. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to
http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.
You may obtain other general information on your responsibilities under the Act from the Division of Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address
http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.
Sincerely yours,
Ribhi Shawar -S For
Uwe Scherf, M.Sc., Ph.D. Director Division of Microbiology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health
Enclosure
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Indications for Use
510(k) Number (if known) K171078
Device Name
TRI-COMBO PARASITE SCREEN
Indications for Use (Describe)
The TECHLAB® TRI-COMBO PARASITE SCREEN test is an enzyme immunoassay for the simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens. The test is indicated as an aid in the diagnosis of gastrointestion when giardiasis, cryptosporidiosis is suspected. The test does not differentiate between the three parasites and follow-up testing is required for all positive results to confirm the specific diagnosis.
| Type of Use (Select one or both, as applicable) | |
|---|---|
| X Prescription Use (Part 21 CFR 801 Subpart D) | Over-The-Counter Use (21 CFR 801 Subpart C) |
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TRI-COMBO PARASITE SCREEN 510(k) SUMMARY
This summary of 510(k) safety and effectiveness is being submitted in accordance with the requirements of 21 CFR 807.92.
Applicant/Contact Information:
| Date Prepared: | July 10, 2017 |
|---|---|
| Name: | TECHLAB, Inc. |
| Address: | 2001 Kraft Drive |
| Corporate Research Center | |
| Blacksburg, VA 24060 USA | |
| Contact Person: | Donna T. Link |
| Phone Number: | 540-953-1664 |
| Email: | dlink@techlab.com |
1.1 Manufacturing Facility Address
TECHLAB, Inc. 20 Corporate Drive Radford, VA 24141 USA
1.2 Product and Trade Name of the Device
TRI-COMBO PARASITE SCREEN
1.3 Common Name or Classification Name
Giardia spp., Cryptosporidium spp., and E. histolytica detection test
1.4 Classification and Regulation
Class II
21 CFR 866.3220; Entamoeba histolytica serological reagents
1.5 Product Code
MHJ - Cryptosporidium spp.
1.6 Panel
83 Microbiology
4
Intended Use
The TECHLAB® TRI-COMBO PARASITE SCREEN test is an enzyme immunoassay for the simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens. The test is indicated as an aid in the diagnosis of gastrointestinal infection when qiardiasis, cryptosporidiosis and amebiasis is suspected. The test does not differentiate between the three parasites and follow-up testing is required for all positive results to confirm the specific diagnosis.
Explanation
The most common method used to detect giardiasis, cryptosporidiosis and amebiasis has been diagnosis by ova and parasite (O&P) light microscopy. However, accuracy of O&P results depends upon the skill of the technician and also relies on the presence of intact cysts in the feces, which may not be present in all samples. In addition, it is only rarely possible to distinguish between the pathogenic and non-pathogenic species of Entamoeba using microscopy. The success rate of fecal examination by microscopy varies between 50 and 70%, and multiple specimens are usually necessary to establish a diagnosis. When infection is present but parasites are not detected by microscopy, sampling and testing of duodenal fluid may detect trophozoites, but this method is invasive and expensive. Detection of the organism and antigens by ELISA provides an alternative method of diagnosis and is sensitive and specific. The TRI-COMBO PARASITE SCREEN ELISA procedure is straightforward to perform and exhibits increased sensitivity compared to microscopic examination. Large numbers of specimens can be tested rapidly and objectively, and the procedure is less labor-intensive than most microscopy methods.
Device Description
The TRI-COMBO PARASITE SCREEN test is an enzyme immunoassay for the simultaneous qualitative detection of Giardia spp., Cryptosporidium spp. and/or E. histolytica antigen in human fecal specimens. The test uses monoclonal and polyclonal antibodies to cell-surface antigens of Giardia, Cryptosporidium and E. histolytica. The microassay plate in the kit contains immobilized monoclonal antibodies against the antigens, and the Conjugate consists of polyclonal antibodies against the antigens. In the assay, an aliquot of a diluted fecal specimen is transferred to a microassay well. The immobilized monoclonal antibodies bind the Giardia, Cryptosporidium and/or E. histolytica antigens if they are present. Upon addition, Conjugate then binds to the antigen/ antibody complex. Any unbound materials are removed during the washing steps. Following the addition of Substrate, a color is detected due to the enzyme-antibody-antigen complexes that formed in the presence of antigens and conjugate.
Materials Provided
- . Conjugate (7 mL) – Antibodies against antigens of Giardia spp., Cryptosporidium spp., and E. histolytica in a buffered protein solution containing 0.05% ProClin® 300
- . Diluent (50 mL) – Buffered protein solution containing 0.02% thimerosal. The Diluent is also to be used as the negative control solution
- . Stop Solution (7 mL) – 0.6 N sulfuric acid. Caution: Avoid contact with skin: flush with water immediately if contact occurs
- . Giardia Positive Control (3.5 mL) – Giardia antigen in a buffered protein solution with 0.02% thimerosal
- Cryptosporidium Positive Control (3.5 mL) Cryptosporidium antigen in a buffered protein . solution with 0.02% thimerosal
- . E. histolytica Positive Control (3.5 mL) – E. histolytica antigen in a buffered protein solution with 0.02% thimerosal
5
- Substrate (14 mL) – solution containing tetramethylbenzidine and peroxide
- Wash Buffer Concentrate (50 mL) 20X concentrate containing phosphate buffered saline, . detergent, and 0.2% thimerosal
- . Microassay Plate - 12 strips, each consisting of 8 wells coated with antibodies to Giardia spp., Cryptosporidium spp., and E. histolytica antigen (stored with desiccant)
- Disposable plastic transfer pipettes - Quantity 100
- . Plastic adhesive sheets - Quantity 2 Sheets
- Wash Solution Label - Quantity 1
Predicate Device Information
The predicate device (GIARDIA/CRYPTOSPORIDIUM CHEK®) and the TRI-COMBO PARASITE SCREEN test use the same EIA (enzyme immunoassay) technology and are substantially equivalent in principle. The following tables show a comparison of both devices' similarities and differences.
| Similarities | ||||
|---|---|---|---|---|
| ltem | Device (K171028) | Predicate (K051929) | ||
| Intended Use | The TECHLAB® TRI-COMBO PARASITE | |||
| SCREEN test is an enzyme immunoassay for | ||||
| the simultaneous qualitative detection of | ||||
| Giardia spp., Cryptosporidium spp. and/or E. | ||||
| histolytica antigen in human fecal specimens. | ||||
| The test is indicated as an aid in the diagnosis | ||||
| of gastrointestinal infection when giardiasis, | ||||
| cryptosporidiosis and amebiasis is suspected. | ||||
| The test does not differentiate between the | ||||
| three parasites and follow-up testing is | ||||
| required for all positive results to confirm the | ||||
| specific diagnosis. | The GIARDIA/ | |||
| CRYPTOSPORIDIUM CHEK test is | ||||
| an enzyme immunoassay for the | ||||
| qualitative detection of Giardia cyst | ||||
| and Cryptosporidium oocyst antigen | ||||
| in human fecal specimens. It is | ||||
| indicated for use as an aid in the | ||||
| diagnosis of patients with diarrhea | ||||
| suspected of Giardia and/or | ||||
| Cryptosporidium gastrointestinal | ||||
| infections | ||||
| Technology | Enzyme Linked Immunoassay (ELISA) | Same | ||
| Antibody Format | Monoclonal capture Ab Polyclonal secondary | |||
| Ab | Same | |||
| Type of Test | Qualitative | Same | ||
| Format/Tests | Microassay Well Plate (96 tests) | Same | ||
| Controls | Positive and negative control are included in | |||
| the kit | Same | |||
| Interpretation | Spectrophotometrically and visually | Same |
| Differences | ||
|---|---|---|
| Item | Device | Predicate |
| Analyte Detected | Giardia spp., Cryptosporidium | |
| spp., and E. histolytica specific | ||
| antigens | Giardia spp., and Cryptosporidium spp., | |
| specific antigens | ||
| Acceptable Specimen Type | Fecal specimens in Cary-Blair | |
| and C&S Transport Media | Specimens in preservation media of 10% | |
| buffered formalin or Sodium Acetate | ||
| Formalin (SAF) |
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Summary of Performance Data
Prospective Study
The performance of the TRI-COMBO PARASITE SCREEN test was evaluated at 3 independent sites. The three sites yield a total of 14 microscopy positive samples (13 Giardia and one E. histolytica). The remaining 740 samples were negative. Of the 740 negative samples, four were E. histolytica positive by molecular comparison only but were negative by the TRI-COMBO PARASITE SCREEN test, negative by microscopy, and negative by FDA cleared antigen test for E. histolytica. Table 1 summarizes the performance observed, which is primarily a study to evaluate specificity due to the low number of positive specimens (see retrospective study for evaluation of sensitivity). Prospective testing was also read visually and performance was not significantly different from spectrophotometric readings.
Table 1. Summary of prospective clinical performance comparing the TRI-COMBO PARASITE SCREEN test to microscopy for Giardia. Cryptosporidium and E. histolytica
| TRI-COMBO PARASITE SCREEN
| (N = 754) | Microscopy | |
|---|---|---|
| Positive | Negative | |
| Positive | 13 | 14* |
| Negative | 1 | 726 |
| 95% Confidence Limits | ||
| Sensitivity | 92.9% | 68.5% - 98.7% |
| Specificity | 98.1% | 96.9% - 98.9% |
| Specimens Positive for Giardia by microscopy/
| Specimens Detected by the TRI-COMBO PARASITE SCREEN | 12/13 |
|---|---|
| Specimens Positive for E. histolytica by microscopy/ | |
| Specimens Detected by the TRI-COMBO PARASITE SCREEN | 1/1 |
*The fourteen TRI-COMBO PARASITE SCREEN positives that were microscopy negative were confirmed to be positive for Giardia with an alternate FDA cleared antigen test or by PCR with sequencing.
Retrospective Study
Testing consisted of 96 archived specimens previously collected and frozen from one clinical site. The Frozen samples are included in the bank based on being characterized as microscopy and PCR positive. The specimens were collected from an E. histolytica endemic area and contained specimens also positives for Giardia and Cryptosporidium. Table 2 summarizes the performance observed. Retrospective testing was also read visually and performance was not different from spectrophotometric readings.
Table 2. Summary of retrospective clinical performance comparing the TRI-COMBO PARASITE SCREEN test to Microscopy and PCR
| TRI-COMBO PARASITE SCREEN | Microscopy and PCR | |
|---|---|---|
| (N = 96) | Positive | Negative |
| Positive | 85 | 0 |
| Negative | 5 | 6 |
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| 95% Confidence Limits | ||
|---|---|---|
| Sensitivity | 94.4% | 87.7% - 97.9% |
| Specificity | 100% | 61.0% - 100% |
| Specimens Positive for Giardia/ | Specimens Detected by the TRI-COMBO PARASITE SCREEN | 41/41 |
| Specimens Positive for Cryptosporidium/ | Specimens Detected by the TRI-COMBO PARASITE SCREEN | 27/30 |
| Specimens Positive for E. histolytica/ | Specimens Detected by the TRI-COMBO PARASITE SCREEN | 28/30 |
| Note: Eight specimens were dual positive for Giardia and E. histolytica by the Microscopy and PCR and | ||
| tested positive in the TRI-COMBO PARASITE SCREEN test. Three specimens were dual positive for | ||
| Giardia and Cryptosporidium by Microscopy and PCR and tested positive in the TRI-COMBO PARASITE | ||
| SCREEN test. |
The prospective study results were analyzed by considering composite results from multiple tests that consisted of light microscopy, molecular testing of a commercial FDA cleared device and PCR with sequencing for the identification of Giardia spp., Cryptosporidium spp., in addition to identification and subspeciation of E. histolytica. This testing was mainly done because identification of E. histolytica organisms cannot be determined solely by microscopy because it is morphologically indistinguishable from the non-pathogenic E. dispar. Use of an alternate molecular testing is needed to confirm Entamoeba speciation. The molecular testing algorithm used provides a comparator method that is highly sensitive at the detection of Giardia spp., Cryptosporidium spp. and E. histolytica. The performance is summarized in table 3 and is presented as positive percent agreement and negative percent agreement.
Table 3. Summary of prospective clinical performance comparing the TRI-COMBO PARASITE SCREEN test to microscopy and molecular testing
| TRI-COMBO PARASITE SCREEN
| (N = 754) | Microscopy and Molecular testing | |
|---|---|---|
| Positive | Negative | |
| Positive | 18 | 9* |
| Negative | 11** | 716 |
| Positive Percent Agreement | 62.1% | 44.0% - 77.3% |
| Negative Percent Agreement | 98.8% | 97.7% - 99.4% |
| Specimens Positive for Giardia/
| Specimens Detected by the TRI-COMBO PARASITE SCREEN | 17/24 |
|---|---|
| Specimens Positive for E. histolytica/ | |
| Specimens Detected by the TRI-COMBO PARASITE SCREEN | 1/5 |
| Specimens Positive for Cryptosporidium/ | |
| Specimens Detected by the TRI-COMBO PARASITE SCREEN | 0/0 |
' These nine specimens were tested with an alternate FDA cleared antigen test resulting in 99 Giardia determined to be antigen positive.
**These eleven specimens were tested with an alternate FDA cleared antigen test resulting in 6/7 Giardia and 4/4 E. histolytica were determined to be antigen negative.
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Reproducibility
The reproducibility of the TRI-COMBO PARASITE SCREEN test was determined using 20 human fecal specimens coded to prevent their identification during testing. Testing was performed at 2 independent laboratories and on-site at TECHLAB, Inc. The samples were tested twice a day over a 5-day period by multiple technicians at each site using 2 different kit lots. Positive and negative controls were run with each panel of the masked samples. The results from each laboratory were submitted to TECHLAB, Inc. and compared with in-house results. The results were consistent among the different locations and exhibited a correlation of 99.9%. The samples produced the expected results 99.9% of the time.
Analytical Sensitivity
Limit of Detection (LoD) – cutoff points for Giardia cysts, Cryptosporidium oocysts and E. histolytica pathogenic zymodemes in fecal specimens for the TRI-COMBO PARASITE SCREEN test. Determined following CLSI document EP17-A2.
CLSI. Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures: Aproved Guideline - Second Edition. CLSI document EP17-A2. Wayne, PA: Clinical and Laboratory Standards Institute; 2012.
The analytical sensitivity of the test was determined by using purified Giardia cysts, Cryptosporidium oocysts, and E. histolytica pathogenic zymodemes in a sample matrix. The concentration of Giardia cysts, Cryptosporidium oocysts, and E. histolytica pathogenic zymodemes in fecal matrix of which specimens were positive by the TRI-COMBO PARASITE SCREEN test 95% of the time was the assay limit-of-detection (LoD). Test results determined the LoD for the assay to be 8450 cysts/mL of feces for Giardia (equivalent to 169 cysts detected per test), 47,962 oocysts/mL of feces for Cryptosporidium (equivalent to 959 oocysts detected per test), and 1.676 PZs/mL (equivalent to 34 PZs detected per test). For fecal matrix/Cary Blair, the LoD for the assay was determined to be 34,155 cysts/mL of feces for Giardia (equivalent to 427 cysts detected per test), 99,456 oocysts/mL of feces for Cryptosporidium (equivalent to 1243 occysts detected per test), and 4655 PZs/mL (equivalent to 58 PZs detected per test). For fecal matrix/C&S, the LoD for the assay was determined to be 37.095 cysts/mL of feces for Giardia (equivalent to 464 cysts detected per test), 122,299 occysts/mL of feces for Cryptosporidium (equivalent to 1529 ocysts detected per test), and 3948 PZs/mL (equivalent to 49 PZs detected per test).
Because the TRI-COMBO PARASITE SCREEN test detects soluble antigen in fecal specimens in addition to cysts, oocysts, and trophozoites, this LOD study represents an estimate of analytical sensitivity.
Analytical Specificity (Cross Reactivity)
The TRI-COMBO PARASITE SCREEN test was evaluated for cross-reactivity with the bacterial and viral strains listed below. None of the strains were shown to interfere with the performance of the TRI-COMBO PARASITE SCREEN test.
Aeromonas hydrophila Bacteroides fragilis Campylobacter jejuni Clostridium difficile Escherichia coli 0157:H7 Escherichia coli ETEC Shigella dysenteriae Staphylococcus aureus Vibrio parahaemolyticus
Bacillus cereus Campylobacter coli Candida albicans Enterococcus faecalis Escherichia coli EIEC Klebsiella pneumonia Shigella flexneri Staphylococcus aureus (Cowan's) Yersinia enterocolitica
Bacillus subtilis Campylobacter fetus Clostridium bifermentans Escherichia coli Escherichia coli EPEC Salmonella typhimurium Shigella sonnei Staphylococcus epidermidis
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Cytomegalovirus Echovirus 11. 18. 33 Calicivirus Human Adenovirus 1, 2, 3, 5, 40, 41 Human Coronavirus Human Coxsackievirus B2, B3, B4, B5 Human Echovirus 9 Human Enterovirus 68, 69, 70, 71 Human parechovirus 1 [Echovirus 22] Human Rotavirus
Additionally, the TRI-COMBO PARASITE SCREEN test was run on fecal specimens documented to be positive for other parasites by microscopy. The number in parentheses is the quantity of each organism found in the clinical specimens. No cross-reactivity was seen with the following organisms.
| Ascaris lumbricoides and with eggs (22) | Entamoeba coli (16) | Trichuris trichiura eggs (12) |
|---|---|---|
| Blastocystis hominis (11) | Entamoeba moshkovskii (3) | |
| Entamoeba bangladeshi (3) | Iodamoeba bütschlii (10) |
Cross reactivity with Norovirus is unknown because it was not tested in analytical studies. However, Norovirus GI/GII was identified in 34 clinical specimens and Enterotoxigenic E. Coli - ETEC LT/ST was identified in 107 clinical specimens, using an FDA cleared multiplex NAAT assay during clinical testing and no cross reactivity was found using the TRI-COMBO PARASITE SCREEN in those samples.
Strain Specific Study
Due to the similarity in morphology between pathogenic and non-pathogenic Entamoeba species, 3 specimens identified by PCR as positive for non-pathogenic Entamoeba moshkovskii and 3 positive for non-pathogenic Entamoeba bangladeshi were evaluated using the TRI-COMBO PARASITE SCREEN test. These 6 specimens tested negative in the TRI-COMBO PARASITE SCREEN test.
Interfering Substances (U.S. Formulation)
The following substances had no effect on positive or negative TRI-COMBO PARASITE SCREEN test results analyzed at the concentrations indicated:
Barium sulfate (5% w/v), Benzalkonium Chloride (1% w/v), Ciprofloxacin (0.25% w/v), Ethanol (1% w/v), Hog gastric mucin (3.5% w/v), Human blood (40% v/v), Hydrocortisone (1% w/v), Imodium® (5% v/v), Kaopectate® (5% v/v), Leukocytes (0.05% w/v), Maalox® Advanced (5% v/v), Mesalazine (10% w/v), Metronidazole (0.25% w/v), Mineral Oil (10% w/v), Mylanta® (4.2 mg/mL), Naproxen Sodium (5% w/v), Nonoxynol-9 (1% w/v), Nystatin (1% w/v), Palmitic Acid/Fecal Fat (40% w/v), Pepto-Bismol® (5% v/v), Phenylephrine (1% w/v), Polyethylene glycol 3350 (10% w/v ), Prilosec OTC® (5 µg/mL), Sennosides (1% w/v), Simethicone (10% w/v), Stearic Acid/Fecal Fat (40% w/v), Tagamet® (5 µg/mL), TUMS (50 µg/mL), Human Urine (5% v/v), and Vancomycin (0.25% w/v).
Precision - Intra-assay
For the determination of intra-assay performance. 24 fecal samples were analyzed by the TRI-COMBO PARASITE SCREEN test. The samples included 2 negative, 2 high negative, 2 low positive, and 2 moderate positive samples for each analyte. Each specimen was assayed a total of five times using two different kit lots. Positive specimens consistently tested positive and negative specimens consistently tested negative. High negative samples tested within 95% agreement of each other for all three analytes.
Precision - Inter-assay
For the determination of inter-assay performance, 24 fecal samples were analyzed by the TRI-COMBO PARASITE SCREEN test. The samples included 2 negative, 2 high negative, 2 low positive, and 2 moderate positive samples for each analyte. The samples were tested twice a day by multiple
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technicians over a 12-day period using 2 different kit lots. All positive samples remained positive and all negative samples remained negative.
Fresh Versus Frozen Samples
The effect of long term frozen specimen storage on antigen stability was evaluated. For the analysis, a total of 39 fecal samples were tested with the TRI-COMBO PARASITE SCREEN test. The samples consisted of negative fecal samples, high negative fecal samples, low positive fecal samples, moderate positive fecal samples, and high positive fecal samples. Samples were prepared by spiking a negative fecal pool with Giardia cysts, Cryptosporidium oocysts, or E. histolytica pathogenic zymodemes at respective concentrations and stored at ≤ -10°C. These samples were tested at 0, 1, 4 and 8 weeks. No conversion of positive-to-negative-to-positive was observed in any of the samples at the specified time points.
Prozone
To ensure that a high concentration of analyte does not interfere with a positive reaction in TRI-COMBO PARASITE SCREEN test, high samples were prepared by spiking a negative fecal pool with Giardia cysts, Cryptosporidium oocysts, or E. histolytica pathogenic zymodemes, and then tested. A total of 5 different dilutions of each, up to and including the clinically observed high concentration, were prepared and tested in triplicate. The results demonstrated that there was no overall prozone affect, that elevated levels of analyte did not affect the detection of each organism.
Conclusion
The conclusions drawn from the nonclinical and clinical tests demonstrate that the TRI-COMBO PARASITE SCREEN test is safe and effective for the detection of Giardia spp., Cryptosporidium spp., and E. histolytica in human fecal specimens. The information submitted in this premarket notification is complete and supports a substantial equivalence decision.