(399 days)
Not Found
No
The summary describes a genetic test kit and associated software for analyzing a specific gene mutation. There is no mention of AI, ML, or any technology that would typically involve learning from data or complex pattern recognition beyond standard genetic analysis. The software is likely for data processing and reporting, not AI/ML inference.
No
The device is an in vitro diagnostic test for detecting and genotyping a specific gene mutation, which is used for diagnostic purposes rather than for treating or curing a disease.
Yes
The "Intended Use / Indications for Use" section explicitly states that "The Invader® MTHFR 677 test is an in vitro diagnostic test".
No
The device description explicitly lists multiple physical components (Oligo Mix, Buffers, Enzymes, Controls) in addition to the software. This indicates it is a kit with both hardware (reagents) and software components, not a software-only device.
Yes, this device is an IVD (In Vitro Diagnostic).
The "Intended Use / Indications for Use" section explicitly states: "The Invader® MTHFR 677 test is an in vitro diagnostic test intended for the detection and genotyping of a single point mutation...". This directly identifies the device as an IVD.
N/A
Intended Use / Indications for Use
The Invader® MTHFR 677 test is an in vitro diagnostic test intended for the detection and genotyping of a single point mutation (C to T at position 677) of the human 5.10-methylenetetrahydrofolate reductase (MTHFR) gene in isolated genomic DNA obtained from whole blood Potassium EDTA samples from patients with suspected thrombophilia.
Product codes
OMM
Device Description
The Invader MTHFR 677 test consists of the following components:
MTHFR 677 Oligo Mix
Universal Buffer
Universal Enzyme Mix
No DNA Control
MTHFR 677 Wild Type Control
MTHFR 677 Heterozygous Control
MTHFR 677 Mutant Control
Invader Call Reporter™ Software
Invader® MTHFR 677 Software
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
Not Found
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
Not Found
Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
1. Analytical performance:
- Precision/Reproducibility:
- External Reproducibility (Study #1): Two operators from three (3) different sites (2 external, 1 internal) tested a panel of nine (9) whole blood samples (3 genotypes) in duplicate over five (5) non-consecutive days (ten-day period). Total 540 samples tested. Final % Agreement: 100%.
- Lot-to-Lot Reproducibility (Study #9): Nine (9) genomic DNA samples (3 wild type, 3 heterozygous, 3 mutants) were tested in quadruplicate using three (3) different kit lots. Total 108 samples tested. Percent agreement between Invader® MTHFR 677 test and sequencing was 100%.
- Linearity/assay reportable range: Refer to paragraph D below.
- Traceability, Stability, Expected values (controls, calibrators, or methods):
- Real-Time Stability Study (Study #5): Three (3) lots of product stored at -30° to -15°C and +2° to +8°C. Functional testing performed with samples representing all 3 genotypes in quadruplicate at each time point. Demonstrated 7 months stability.
- Reagent Freeze-Thaw Stability Study (Study #6): Product subjected to 15 freeze-thaw cycles. Functional testing performed using genomic DNA isolated from cell lines, representing all possible genotypes. Percent agreement between sequencing and Invader® MTHFR 677 test was 100%.
- Detection limit/Analytical Sensitivity and Normal Range (Study #3): Three (3) genomic DNA samples (WT, HET, MUT) extracted from whole blood were diluted to eight concentrations (0.5, 5, 20, 40, 80, 200, 400, 800 ng/uL) and tested in forty (40) replicates. The recommended range of the assay was determined to be between 5-80 ng/uL of input gDNA, based on 100% concordance of all tested replicates with bi-directional sequencing.
- Analytical specificity (Interfering Substances) (Study #4): Tested the effect of Heparin, Cholesterol, Bilirubin, Hemoglobin, Potassium EDTA, and Ethanol-based Wash Buffer on nine (9) whole blood samples (3 WT, 3 HET, 3 MUT). Test performance was not affected, with 100% agreement with sequencing genotype and untreated samples.
- Pre-Analytical Equivalency Study/Genomic DNA Extraction Reproducibility (Study #7): Thirty (30) human whole blood samples and ten (10) leukocyte depleted whole blood spiked with cell lines were extracted using four (4) commercially available DNA extraction methods. 160 extracted DNAs were analyzed in singlicate. Percent agreement between the Invader® MTHFR 677 test for each extraction method and bi-directional sequencing was 100%.
- Instrument Equivalency (Study #8): Twenty-nine (29) human whole blood samples and ten (10) leukocyte depleted whole blood samples spiked with cell lines were extracted using two (2) extraction methods. Extracts tested with Invader® MTHFR 677 using three (3) thermal cyclers and raw fluorescent data acquired on three (3) fluorometers. Results showed 100% concordance with bi-directional sequencing.
2. Comparison studies:
- Method comparison: Bi-directional Sequencing (Study #2): Human whole blood samples (n = 361) underwent DNA extraction and bi-directional DNA sequence analysis, then analyzed using Invader® MTHFR 677 test. Observed agreement was 100% (359/359). First run agreement with bi-directional sequencing was 99.45% (359/361). Two samples failed to generate valid results on the first run.
3. External Reproducibility studies:
- Clinical Sensitivity: please refer to section 1d above.
- Clinical specificity: please refer to section 1e above.
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
- External Reproducibility (Study #1): Final % Agreement: 100%
- Lot-to-Lot Reproducibility (Study #9): Percent agreement between Invader® MTHFR 677 test and sequencing was 100%.
- Detection Anlaysis (Study #3): 100% concordance with bi-directional sequencing for gDNA concentrations between 5-80 ng/uL.
- Analytical Specificity (Study #4): 100% agreement with sequencing genotype and untreated samples when interfering substances were added.
- Pre-Analytical Equivalency (Study #7): Percent agreement between the Invader® MTHFR 677 test for each extraction method and bi-directional sequencing was 100%.
- Instrument Equivalency (Study #8): 100% concordance for all thermal cycler and fluorometer combinations.
- Method Comparison (Study #2):
- Observed agreement between the Invader® MTHFR 677 test and bi-directional DNA sequencing was 100% (359/359).
- First run agreement with bi-directional sequencing was 99.45% (359/361).
- First Pass Agreement: Homozygous Wild Type (GG) - 98.89% (178/180); Heterozygous (GA) - 100% (104/104); Homozygous Mutant (AA) - 100% (77/77).
- Final Agreement (after accounting for no-calls): Homozygous Wild Type (GG) - 99.45%; Heterozygous (GA) - 100%; Homozygous Mutant (AA) - 100%.
Predicate Device(s)
Nanosphere, K070597
Reference Device(s)
Not Found
Predetermined Change Control Plan (PCCP) - All Relevant Information
Not Found
§ 864.7280 Factor V Leiden DNA mutation detection systems.
(a)
Identification. Factor V Leiden deoxyribonucleic acid (DNA) mutation detection systems are devices that consist of different reagents and instruments which include polymerase chain reaction (PCR) primers, hybridization matrices, thermal cyclers, imagers, and software packages. The detection of the Factor V Leiden mutation aids in the diagnosis of patients with suspected thrombophilia.(b)
Classification. Class II (special controls). The special control is FDA's guidance entitled “Class II Special Controls Guidance Document: Factor V Leiden DNA Mutation Detection Systems.” (See § 864.1(d) for the availability of this guidance document.)
0
Invader® MTHFR 677 510(k) SUMMARY
-
A. 510(k) Number: K100987
: -
B. Purpose for Submission: New Device
-
C. Measurand: MTHFR 677
-
D. Type of Test:
Qualitative genotyping test for single nucleotide polymorphism detection.
E. Applicant:
Hologic Inc. Third Wave Technologies 250 Campus Drive Marlborough, MA 01752 508-263-8912 Contact Person: Randall J. Covill, Manager, Regulatory Affairs Date of Submission: April 2010
- F. Proprietary and Established Names: Invader® MTHFR 677
G. Regulatory Information:
-
- Regulation Sections: 21 CFR 864.7280
-
- Classification: Class II
-
- Product Code: OMM: Test 5,10-Methylenetetrahydrofolate Reductase Mutations, Genomic DNA PCR
-
- Panel:
Hematology (81)
H. Intended Use:
-
- Intended Use(s):
The Invader® MTHFR 677 test is an in vitro diagnostic test intended for the detection and genotyping of a single point mutation (C to T at position 677) of the human 5.10-methylenetetrahydrofolate reductase (MTHFR) gene in isolated genomic DNA obtained from whole blood Potassium EDTA samples from patients with suspected thrombophilia.
- Intended Use(s):
-
Indication(s) for use: 2.
Same as Intended Use -
Special Conditions for use statements(s): ల For prescription use only
-
Special instrument requirements: 4. None
I. Device Description:
The Invader MTHFR 677 test consists of the following components:
MTHFR 677 Oligo Mix
- Universal Buffer
Universal Enzyme Mix
No DNA Control
MTHFR 677 Wild Type Control
MTHFR 677 Heterozygous Control
MTHFR 677 Mutant Control
Invader Call Reporter™ Software
1
· Invader® MTHFR 677 Software
J. Substantial Equivalence Information:
-
- Predicate device name(s):
... .
- Predicate device name(s):
-
Verigene® MTHFR Nucleic Acid Test (Nanosphere, K070597)
- Predicate 510(k) number(s):
- Nanosphere, K070597
-
- Comparison with predicate:
| Table 1: Comparison with Predicate Device | Use | ||||
|---|---|---|---|---|---|
| Predicate Device | Proposed Device | Target | |||
| Population | Patients with suspected | ||||
| thrombophilia | Same as predicate | ||||
| Product Name | |||||
| (Manufacturer, | |||||
| Submission) | Verigene® MTHFR Nucleic Acid | ||||
| Test | |||||
| (Nanosphere, K070597) | Invader® MTHFR 677 | ||||
| (Hologic, Inc. K100987) | Chemistry | SNP discrimination via | |||
| oligonucleotide probes; detection | |||||
| via evanescent wave light scatter | |||||
| with nanoparticles. | PCR followed by Invader® achieving | ||||
| SNP discrimination via | |||||
| oligonucleotide probes. | |||||
| Intended Use | "The Verigene MTHFR Nucleic | ||||
| Acid Test is an in vitro diagnostic | |||||
| for the detection and genotyping of | |||||
| a single point mutation (C to T at | |||||
| position 677) of the human 5, 10 | |||||
| methylene-tetrahydrofolate | |||||
| reductase gene (MTHFR) in | |||||
| patients with suspected | |||||
| thrombophilia, from isolated | |||||
| genomic DNA obtained from | |||||
| whole blood samples. The test is | |||||
| intended to be used on the | |||||
| Verigene System." |
The Verigene System is a bench-
top molecular diagnostic
workstation that automates the in
vitro diagnostic analysis and
detection of nucleic acids using
gold nanoparticle probe
technology. The Verigene System
is intended to be used by
experienced laboratory
professionals with training on basic
laboratory techniques and on the
use of the system components. | The Invader® MTHFR 677 test is an
in vitro diagnostic test intended for
the detection and genotyping of a
single point mutation (C to T at
position 677) of the human 5,10-
methylenetetrahydrofolate reductase
(MTHFR) gene in isolated genomic
DNA obtained from whole blood
Potassium EDTA samples from
patients with suspected
thrombophilia. | Hardware | The Verigene System consists of
two instruments, the Verigene
Processor and the Verigene Reader,
and utilizes single-use, disposable
Test Cartridges to process and
genotype multiple genes in a DNA
samples in approximately 1.5
hours. | Non-specified, third-party
fluorometer and thermal cycler. |
| Specimen Type | Purified DNA isolated from human
whole peripheral blood | Same as predicate | Software
Interface | Embedded software in closed
system, integrated graphical user
interface. | Java-based software installed on a
standalone PC capable of converting
raw fluorescence data into genotype
calls. |
| Indications for | Same as Intended Use | Same as Intended Use | Detection Method | Single-image sensor where
nanoparticles are illuminated using
a fixed-wavelength light source. | PCR and Fluorescence Resonance
Energy Transfer (FRET) chemistry
for signal reporting. |
| Sample Size | 25µL | 20µl reaction containing 0.25-4ng/ul
gDNA extracted from human
peripheral whole blood. | | | |
| Detection
Procedure | Single-image sensor where
nanoparticles are illuminated using
a fixed-wavelength light source. | Multi-well fluorometer to detect raw
fluorescence. | | | |
| Detection
Chemistry | Detection via evanescent wave
light scatter with nanoparticles. | PCR and Invader® using
Fluorescence Resonance Energy
Transfer (FRET) chemistry for signal
reporting. | | | |
| Analysis Time | 90 min. processing with 2 min
analysis time. | ~90 min. amplification followed by 1
min signal detection. Software
analysis post signal detection. | | | |
2
.
3
K. Standard/Guidance Document Referenced (if applicable):
- Guidance for Industry and FDA Staff Class II Special Controls Guidance Document: Factor V Leiden DNA Mutation Detection Systems issued on March 16, 2004
- Guidance for Industry and FDA Staff Guidance for the Content of . Premarket Submissions for Software Contained in Medical Devices issued May 11, 2005
- Guidance for Industry and FDA Staff Format for Traditional and . Abbreviated 510(k)s issued on August 12, 2005
L. Test Principle:
The Invader® MTHFR 677 test utilizes the Invader Plus® chemistry with DNA isolated from human whole blood, for the detection of the targeted sequence polymorphism. Specifically, the Invader Plus® chemistry utilizes a single-tube, two phase reaction, including target amplification and signal generation (mediated by Invader chemistry). Invader Plus® reaction mixes are assembled by combining the MTHFR 677 Oligo Mix, Universal Enzyme Mix, and Universal Buffer. In a 96-well plate, reaction mix is combined with purified genomic DNA samples, as well as four (4) controls included with the test. The No DNA Control is used by the interpretive software to set the "noise" component of the run for "signal-to-noise" calculations. The genotype-specific controls (WT, HET, MUT) ensure reagents were assembled correctly and perform according to the specifications. The 96-well plate is transferred to an appropriately programmed thermal cycler for target amplification and signal generation. In the target amplification phase of the reaction, amplification is carried out using "two-step" cycling conditions (i.e. denaturation & annealing/extension). Following amplification, Taq polymerase is inactivated by a 10 minute incubation at 99°C, after which the thermal cycler proceeds to 63°C to initiate the signal generation (Invader ) phase of the reaction (see Figure 1).
4
Image /page/4/Figure/0 description: The image shows a comparison of two structure formation processes, one for a wildtype specific primary probe and one for a mutation specific primary probe. Both processes involve three steps: structure formation, structure recognition and cleavage, and a secondary reaction. The secondary reaction for the wildtype probe involves a FRET cassette (FAM) and results in fluorescence 1 (FAM), while the secondary reaction for the mutation specific probe involves a FRET cassette (RED) and results in fluorescence 2 (RED).
Figure 1. Invader® Signal Generation Phase
During the signal generation phase, a discriminatory Probe transiently hybridizes to the amplified target sequence along with an Invader® oligonucleotide, to form an overlapping structure. The 5-end of the Primary Probe includes a 5-flap that does not hybridize to the target DNA. The 3'-nucleotide of the bound Invader oligonucleotide overlaps the Primary Probe, and does not hybridize to the target DNA. The Cleavase enzyme recognizes this overlapping structure and cleaves off the unpaired 5'-flap of the Primary Probe, releasing it as a target-specific product. The Primary Probe is designed to have a melting temperature aligned with the Invader® reaction temperature so that under the isothermal reaction conditions (~63°C) the Primary Probes cycle on and off the target DNA. This allows for multiple rounds of Primary Probe cleavage for each DNA target resulting in an accumulation of the number of released 5'-flaps. The released 5'-flap transiently hybridizes with a corresponding FRET cassette forming an overlapping structure that is recognized and the fluorophore is cleaved from the FRET cassette by the Cleavase enzyme. The 5'-flap is designed to have a melting temperature aligned with the Invader reaction temperature, so that the 5'-flaps cycle on and off of the corresponding FRET cassettes. This allows for multiple rounds of FRET cassette cleavage for each 5'flap, and an accumulation of released fluorophore. When the FRET cassette is cleaved, a fluorophore and quencher are separated, generating detectable fluorescence signal. The format uses two different discriminatory Primary Probes, one for the mutant allele and one for the wild type allele (Figure 1). Each Primary Probe is assigned a unique 5-flap, and distinct FRET cassette, with a spectrally distinct fluorophore. By design, the released 5flaps will bind only to their respective FRET cassettes to generate a target-specific signal, linking the wild type allele with one fluorophore (Fluorescence 1: FAM) and the mutant allele with the second fluorophore (Fluorescence 2: RED).
5
The Invader® MTHFR 677 software, in combination with Invader Call Reporter™ software, is a data analysis software package developed by Hologic for use with the Invader® MTHFR 677 test. The software package provides a working template for the setup of reaction mixes and sample placement, and following the import of fluorescence data, it determines results and validity for controls and samples. A summary of the Invader Call Reporter™- Invader® MTHFR 677 package workflow is shown in Figure 2.
Image /page/5/Figure/1 description: This image is a flowchart that describes an assay selection process. The process starts with assay selection, where the operator enters the run ID, selects MTHFR 677, and enters the number of samples. The next step is mix preparation, where the master kit lot number and expiration date are entered, as well as the component lot numbers and expiration dates, and the reaction mix amounts are calculated. The next step is sample placement, where the sample IDs and comments are entered, followed by the results, where fluorescence data is imported, Invader MTHFR 677 results and data for controls and samples are displayed for valid plates, results are saved as PDF, and results are exported to spreadsheet and CSV, and finally the summary, where Invader MTHFR 677 results are displayed for controls and samples, the summary is saved as PDF, and the assay is finished.
Figure 2. Invader Call Reporter™. Invader® MTHFR 677 Package Workflow
Page 6 of 13
6
M. Performance Characteristics (if/when applicable):
-
- Analytical performance:
- a. Precision/Reproducibility:
External Reproducibility (Study #1): Two operators each from three (3) different sites (2 external sites and 1 internal site) performed the testing, in duplicate, over five (5) non-consecutive days for a ten (10) day period using the same testing materials including a panel of nine (9) whole blood samples specific for each of the three (3) possible genotypes (i.e. wild type, heterozygous, homozygous mutant).
| Table 2: Inter-laboratory Reproducibility of Invader® 677 Test | ||||||||
|---|---|---|---|---|---|---|---|---|
| Site | Operator | Samples | ||||||
| tested | Correct | |||||||
| Calls | First Pass | |||||||
| No Calls | ||||||||
| (Invalid, EQ) | Miscalls | Correct | ||||||
| Calls | Final | |||||||
| No Calls | ||||||||
| (Invalid, EQ) | Final % | |||||||
| Agreement | ||||||||
| Final Correct Calls | ||||||||
| Samples Tested | ||||||||
| Site | ||||||||
| 001 | 1 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| Site | ||||||||
| 001 | 2 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| Site | ||||||||
| 002 | 1 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| Site | ||||||||
| 002 | 2 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| Site | ||||||||
| 003 | 1 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| Site | ||||||||
| 003 | 2 | 90 | 90 | 0 | 0 | 90 | 0 | 100% |
| All | All | 540 | 540 | 0 | 0 | 540 | 0 | 100% |
Lot-to-Lot Reproducibility (Study #9): A total of nine (9) genomic DNA samples (three (3) wild type , three (3) heterozygous and three (3) mutants) were tested in quadruplicate using three (3) different kit lots of the Invader® MTHFR 677 test. The percent agreement between Invader® MTHFR 677 test and sequencing was 100% (n=108).
| Table 3: Lot to Lot Reproducibility | ||||||
|---|---|---|---|---|---|---|
| Lot | # | |||||
| Samples | ||||||
| Tested | First | |||||
| Pass | ||||||
| Correct | ||||||
| Calls | First | |||||
| Pass No | ||||||
| Calls | Miscalls | Final | ||||
| Correct | ||||||
| Calls | Final | |||||
| Agreement | ||||||
| % | ||||||
| 1 | 36 | 36 | 0 | 0 | 36 | 100 |
| 2 | 36 | 36 | 0 | 0 | 36 | 100 |
| 3 | 36 | 36 | 0 | 0 | 36 | 100 |
| Total | 108 | 108 | 0 | 0 | 108 | 100 |
- b. Linearity/assay reportable range: Refer to paragraph D below.
- Traceability, Stability, Expected values (controls, calibrators, or methods): C. Real-Time Stability Study (Study #5): Three (3) lots of product in the final configuration are being stored under recommended conditions: (1) -30° to -15°C (Standard Storage of intermediate components) as well as (2) +2° to +8°C (Standard Storage of Genotype-Specific Controls). Functional testing is performed with samples representing all 3 genotypes in quadruplicate at each time point. The interim test results have demonstrated 7 months stability for the device.
7
| Sample/
Control | Sequencing/
Expected
MTHER 677
Genotype | T₀ Result | | | T₁ Result | | | T₂ Result | | |
|--------------------|--------------------------------------------------|-----------|-------|-------|-----------|-------|-------|-----------|-------|-------|
| | | Lot 1 | Lot 2 | Lot 3 | Lot 1 | Lot 2 | Lot 3 | Lot 1 | Lot 2 | Lot 3 |
| Control 1 | WT | WT | WT | WT | WT | WT | WT | WT | WT | WT |
| Control 2 | HET | HET | HET | HET | HET | HET | HET | HET | HET | HET |
| Control 3 | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT |
| gDNA 1 | WT | WT | WT | WT | WT | WT | WT | WT | WT | WT |
| gDNA 2 | WT | WT | WT | WT | WT | WT | WT | WT | WT | WT |
| gDNA 3 | HET | HET | HET | HET | HET | HET | HET | HET | HET | HET |
| gDNA 4 | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT | MUT |
| Percent Agreement | | 100 | 100 | 100 | 100 | 100 | 100 | 100 | 100 | 100 |
Reagent Freeze-Thaw Stability Study (Study #6): Product in the final configuration was subject to 15 freeze-thaw cycles prior to the final thaw at the time of testing. Functional testing was performed using genomic DNA isolated from cell lines, representing all possible genotypes. The percent agreement between the sequencing result and the Invader® MTHFR 677 test were 100%, therefore demonstrating stability for up to fifteen (15) freeze/thaw cycles.
| Table 5: Freeze/Thaw Stability of Invader® MTHFR 677 | |||||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 1 | |||||||||||||||||
| Sample 1 1 11 | َ َ َ َ َ َ َ َ | 3 3 8 8 8 9 1 8 8 9 1 10 2 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 | |||||||||||||||
| Control 1 | |||||||||||||||||
| (WINE KEEK | ﻢ ﺍﻟﻤﺮﺍﺟﻊ | ﻢ ﺍﻟﻤﺴﺘﻘﻠﺔ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟ | ਪ੍ | 3 | ਤੇ | 3 | 3 | ﺔ | 3 | ﻓ | ﻢ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤﺴﺘﻮﻯ ﺍﻟﻤ | 3 | ব | 3 | વર્તર | 100 | |
| Control 2; 4 | |||||||||||||||||
| CHET) THET | 3 | 7 | ﻢ ﺍﻟﻤﺮﺍﺟﻊ ﺍﻟﻤﺮﺍﺟﻊ | 3 | 3 | ತೆ | ನ | 3 | 3 | 3 | ಗೆ | 8 | ತಿ | 3 | 3 | વે રે | 100 |
| Control 3 | |||||||||||||||||
| EMUT | ਪ੍ | ਪ੍ | ﻢ | 3 | 3 | 3 | 3 | ಗ | 3 | 3 | ਪੈ | 3 | 3 | 3 | વર્સ | 100 | |
| ISEDNA | |||||||||||||||||
| TWT) : | 6 | 6 | 12 | 1 | ત | 6 | 本 | * | 6 | 36 | 100 | ||||||
| gDNA | |||||||||||||||||
| (HET) | ನ್ನ | 8 | 8 | 18 | * | * | 8 | 8 | 本 | * | 8 | 48 | 100 | ||||
| gDNA® | |||||||||||||||||
| : "(MUT) | 6 | 6 | ત | ત્ | 6 | * | * | 6 | 36 | 100 | |||||||
| Total | 29 | 9 | 29 | Q | 29 | ರ | 0 | 9 | ਉ | 29 | 0 | 29 | ರ | ರಿ | 29 | 255 | 100 |
| * Testing with oDNA samples did not occur at this testing point. |
*Testing with gDNA samples did not occur at this testing point.
- Detection limit/Analytical Sensitivity and Normal Range (Study #3): d. Three (3) genomic DNA samples with different genotypes (i.e. WT, HET, MUT) were extracted from whole blood collected in potassium EDTA. Each sample was diluted to eight different concentrations 0.5, 5, 20, 40, 80, 200, 400, 800 ng/ uL and tested in replicates of forty (40). The recommend range of the assay was
8
determined to be between 5-80 ng/uL of input gDNA, based on 100% concordance of all tested replicates with bi-directional sequencing.
| Table 6: Percent Agreement Between Replicates | |||
|---|---|---|---|
| Sample ID (Genotype based on Sequencing) | |||
| Input Sample | |||
| Concentration | 03-4520 (MUT) | 03-4525 (HET) | 03-4524 (WT) |
| 0.5 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 5 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 20 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 40 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 80 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 200 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 400 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
| 800 ng/µl | 100% (40/40) | 100% (40/40) | 100% (40/40) |
-
Analytical specificity (Interfering Substances) (Study #4): e.
Test performance was not affected by addition of the following substances to nine (9) whole blood samples of different genotype (3 WT, 3 HET, 3 MUT) prior to extraction: -
Heparin (1500 U/dL human whole blood) ●
-
Cholesterol (300 mg/dL human whole blood) .
-
. Bilirubin (10 mg/dL human whole blood)
-
Hemoglobin (up to 0.2% in whole blood) .
-
Potassium EDTA (1.8 mg/mL human whole blood) .
-
Ethanol-based Wash Buffer (5% in DNA sample) .
9
| Table 7: Summary, Comparison of Invader MTHFR 677 Interfering Substance Results to Sequencing | ||||
|---|---|---|---|---|
| Interfering | ||||
| Substance | ||||
| Code | Substance Concentration / | |||
| (in blood or DNA sample) | % Agreement | |||
| with | ||||
| Sequencing | ||||
| Genotype | % Agreement | |||
| with | ||||
| Untreated | ||||
| Sample | ||||
| Invader® | ||||
| MTHFR 677 | ||||
| Genotype | PASS / | |||
| FAIL | ||||
| A | No Addition Control | |||
| (Untreated) | 100% (18 of 18) | N/A | PASS | |
| B | Bilirubin 10mg/dl (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| C | Cholesterol 300mg/dl (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| D | K2EDTA 1.8mg/ml (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| E | Heparin 1500 U/dl (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| F1 | Hemoglobin 0.2% (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| F2 | Hemoglobin 0.1% (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| F3 | Hemoglobin 0.05% (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| F4 | Hemoglobin 0.025% (blood) | 100% (18 of 18) | 100% (18 of 18) | PASS |
| G | Buffer AW2 5% (DNA) | 100% (18 of 18) | 100% (18 of 18) | PASS |
61677 Summers Ecomparison of Invodor Max 2010 September 10, 2016 Conservering Substance Count
- Pre-Analytical Equivalency Study/Genomic DNA Extraction Reproducibility f (Study #7): Thirty (30) human whole blood samples and ten (10) leukocyte depleted whole blood spiked with cell lines were divided and extracted using four (4), commercially available DNA extraction methods (A. Qiagen QIAamp® 96 DNA Blood Kit, B. Qiagen QIAamp® DNA Blood Mini Kit, C. Gentra Generation® Capture Column Kit (Oiagen), D. Roche MagNA Pure LC DNA Isolation Kit I). The 160 extracted DNAs were analyzed in singlicate with one (1) lot of the device. The percent agreement between the Invader® MTHFR 677 test for each extraction method and bi-directional sequencing was 100% (n=40).
10
| Table 8a: Pre-Analytical Equivalency | ||||||
|---|---|---|---|---|---|---|
| Extraction | ||||||
| Method | # | |||||
| Samples | ||||||
| Tested | First | |||||
| Pass | ||||||
| Correct | ||||||
| Calls | First | |||||
| Pass | ||||||
| No. | ||||||
| Calls | Miscalls | Final | ||||
| Correct | ||||||
| Calls | Final | |||||
| Agreement | ||||||
| % | ||||||
| A | 40 | 40 | 0 | 0 | 40 | 100 |
| B | 40 | 39 | 1* | 0 | 39* | 100* |
| C | 40 | 40 | 0 | 0 | 40 | 100 |
| D | 40 | 40 | 0 | 0 | 40 | 100 |
| Total | 160 | 159 | 1 | 0 | 159 | 100 |
| *Sample was removed from study due to loss of traceability of the sample | ||||||
| identification. |
| Table 8b: Genotype Distribution Across Sample Type | |||||
|---|---|---|---|---|---|
| Sample Type Type WT WT MET MET HET Total | |||||
| Human Whole Blood | 0 * | 10 | |||
| LDWB Spiked with Cell Lines** | 10 | ||||
| Total | 0* | રું તેમ | |||
| *Sample was removed from study due to loss of traceability of the sample identification. | |||||
| **LDWB=Leukocyte Depleted Whole Blood |
- Instrument Equivalency (Study #8): Twenty-nine (29) human whole blood g. samples and ten (10) leukocyte depleted whole blood samples spiked with cell lines were extracted using two (2) commonly used extraction methods. The extracts were tested with the Invader® MTHFR 677 test using three (3) commercially available thermal cyclers (1. ABI GeneAmp® PCR System 9700 with 96-well gold block, 2. ABI Veriti™ and 3. MJ Research PTC-100) and the raw fluorescent data acquired on three (3) commercially available fluorometers (A. Tecan Infinite®, B. Tecan Genios® and C. BioTek®, FLx800). Results from the three (3) fluorometers were transferred into the interpretive software and genotype calls compared to bi-directional sequencing.
| Table 9: Concordance by Instrument | |||
|---|---|---|---|
| Fluorometer | Thermal Cycler | ||
| 1 | 2 | 3 | |
| A | 78 of 78 = 100% | 78 of 78 = 100% | 78 of 78 = 100% |
| B | 78 of 78 = 100% | 78 of 78 = 100% | 78 of 78 = 100% |
| C | 78 of 78 = 100% | 78 of 78 = 100% | 78 of 78 = 100% |
2. Comparison studies:
a. Method comparison: Bi-directional Sequencing (Study #2):
- Human whole blood samples (n = 361) underwent DNA extraction and subsequent bi-directional DNA sequence analysis. The same DNA samples were then analyzed using the Invader® MTHFR 677 test. The observed agreement between the Invader® MTHFR 677 test and bidirectional DNA sequencing was 100% (359/359). The first run agreement with bi-directional sequencing was 99.45% (359/361).
11
| MTHFR 677
Genotype* | Number
tested | Number of
valid results
on 1st run | Number of
Correct
genotype
calls on First
Run | First Run
Agreement |
|----------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------|------------------------------------------|-----------------------------------------------------------|------------------------|
| Homozygous
Wild Type
(GG) | 180 | 178** | 178** | 98.89% |
| Heterozygous
(GA) | 104 | 104 | 104 | 100% |
| Homozygous
Mutant
(AA) | 77 | 77 | 77 | 100% |
| Total | 361 | 359** | 359** | 99.45% |
| * Genotype determined through bi-directional DNA sequencing | | | | |
| ** Two samples failed to generate valid results. These samples were reported as invalid (EO) and no genotype calls were assigned by the interpretive software. | | | | |
The EQ result was used to determine the First Run Agreement.
| Table 10b: Agreement between the Invader® MTHFR 677 Test and | |||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Bi-directional DNA Sequencing | |||||||||||||||
| Genotype* | First Pass Results | Final Results | |||||||||||||
| WT | HET | MUT | No-Call | Mis-Call | % Agreement | 95% LCB** | WT | HET | MUT | No-Call | Mis-Call | % Agreement | 95% LCB | ||
| WT | 178 | 0 | 0 | 2 | 0 | 99.45 | 96.04 | 178 | 0 | 0 | 2 | 0 | 99.45 | 96.04 | |
| HET | 0 | 104 | 0 | 0 | 0 | 100.00 | 97.16 | 0 | 104 | 0 | 0 | 0 | 100.00 | 97.16 | |
| MUT | 0 | 0 | 77 | 0 | 0 | 100.00 | 96.18 | 0 | 0 | 77 | 0 | 0 | 100.00 | 96.18 | |
| *Genotype determined through bi-directional DNA sequencing. **Lower boundary of the 95% confidence interval for each genotype. †Calculated across all samples. |
- External Reproducibility studies:
Clinical Sensitivity: please refer to section 1d above. a.
Clinical specificity: please refer to section 1e above. b.
-
- Expected values/Reference range: (Prevalence) MTHFR 677: 30-40% Caucasians
N. System Descriptions:
- Expected values/Reference range: (Prevalence) MTHFR 677: 30-40% Caucasians
.
-
- Modes of Operation: · Closed System
12
-
- Software:
FDA has reviewed applicant's Hazard Analysis and software development processes for this line of product type. Yes X
- Software:
-
- Specimen Identification: Manual Labeling
-
- Specimen Sampling and Handling:
DNA should be extracted using a validated DNA extraction method that generates DNA concentration range of greater than 5ng/ul.
-
- Quality Control:
Each test contains positive and negative controls to assure proper functioning of the system: Failure of any controls will be indicated as "Invalid" in the test results section of the report. The genotyping test result will not be reported for any sample for which a positive or negative control failure occurs. Positive Control: The genotype controls (WT, HET, MUT) ensure reagents were assembled correctly and perform according to the specifications. Negative Control: The No DNA Control is used by the interpretive software to set the "noise" component of the run for "signal-to-noise" calculations.
- Quality Control:
Hardware and Software Controls:
The genotyping test result will not be reported for any sample for which a positive or negative control failure occurs.
O. Proposed Labeling:
The labeling is sufficient and satisfies the requirements of 21 CFR Part 809.10.
- P. Conclusion:
The submitted information in this 510 (k) notification is complete and supports a substantial equivalence decision.
13
Image /page/13/Picture/1 description: The image shows the logo for the U.S. Department of Health and Human Services. The logo features a stylized caduceus, which is a symbol of medicine, with three wavy lines representing the serpent entwined around the staff. The logo is encircled by the text "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA" in a circular arrangement.
Food and Drug Administration 10903 New Hampshire Avenue Document Mail Center - WO66-0609 Silver Spring, MD 20993-0002
Hologic Inc. Third Wave Technologies c/o Mr. Randall J. Covill Senior Specialist Regulatory Affairs 502 South Rosa Road Madison, WI 53719
MAY 1 3 2011
Re: K100987
Trade/Device Name: Regulation Number: Regulation Name: Regulatory Class: Product Code: Dated: March 31, 2011 Received: April 7, 2011 Invader® MTHFR 677 21 CFR §864.7280 Factor V Leiden DNA Mutation Detection Systems Class II OMM
Dear Mr. Covill:
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.
If your device is classified (see above) into class II (Special Controls), it may be subject to such additional controls. Existing major regulations affecting your device can be found in Title 21, Code of Federal Regulations (CFR), Parts 800 to 895. 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); and good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820). This letter
14
Page 2 - Mr. Randall J. Covill
will allow you to begin marketing your device as described in your Section 510(k) premarket notification. The FDA finding of substantial equivalence of your device to a legally marketed predicate device results in a classification for your device and thus, permits your device to proceed to the market.
If you desire specific advice for your device on our labeling regulation (21 CFR Parts 801 and 809), please contact the Office of In Vitro Diagnostic Device Evaluation and Safety at (301) 796-5450. 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 Small Manufacturers, International and Consumer Assistance at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address http://www.fda.gov/cdrh/industry/support/index.html.
Sincerely yours,
Thava In Chen
Maria M. Chan, Ph.D Director Division of Immunology and Hematology Devices Office of In Vitro Diagnostic Device Evaluation and Safety Center for Devices and Radiological Health
Enclosure
15
Indication for Use Statement
510(k) Number (if known): K100987
Invader MTHFR 677 test Device Name:
Indication for Use:
The Invader® MTHFR 677 test is an in vitro diagnostic test intended for the detection and genotyping of a single point mutation (C to T at position 677) of the human 5,10methylenetetrahydrofolate reductase (MTHFR) gene in isolated genomic DNA obtained from whole blood potassium EDTA samples from patients with suspected thrombophilia.
Prescription Use x (Part 21 CFR 801 Subpart D) AND/OR
Over-The-Counter Use (21 CFR 801 Subpart C)
(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE OF NEEDED)
Concurrence of CDRH, Office of In Vitro Diagnostic Devices (OIVD)
Division Sign-Off
Office of In Vitro Diagnostic Device Evaluation and Safety
510(k) K100987