(87 days)
Not Found
No
The summary describes a standard real-time PCR in vitro diagnostic test and does not mention any AI or ML components in the device description, intended use, or performance studies.
No
The device is an in vitro diagnostic test intended for the qualitative detection and differentiation of parainfluenza viruses to aid in diagnosis, not to treat or alleviate a disease.
Yes
The "Intended Use / Indications for Use" section explicitly states that the Panther Fusion Paraflu assay is an "in vitro diagnostic " and "is intended to aid in the differential diagnosis" of HPIV infections.
No
The device is an in vitro diagnostic test that involves physical steps like sample lysis, nucleic acid capture and elution, and multiplex RT-PCR, all performed on the Panther Fusion system, which is a hardware platform. While software is undoubtedly involved in controlling the system and analyzing results, the device itself is not solely software.
Yes, this device is an IVD (In Vitro Diagnostic).
The "Intended Use / Indications for Use" section explicitly states: "The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test..."
The "Device Description" also refers to it as an "in vitro diagnostic test".
These statements clearly identify the device as an IVD.
N/A
Intended Use / Indications for Use
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test for the rapid and qualitative detection and differentiation of parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus and parainfluenza 4 virus (HPIV-1, HPIV-2, HPIV-3, and HPIV-4). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection.
This assay is intended to aid in the differential diagnosis of HPIV-1, HPIV-2, HPIV-3, and HPIV-4 infections in humans. Negative results do not preclude HPIV-1, HPIV-2, HPIV-3, and HPIV-4 infections and should not be used as the sole basis for treatment or other management decisions. This assay is designed for use on the Panther Fusion system.
Product codes (comma separated list FDA assigned to the subject device)
OOU, OOI
Device Description
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test developed for use on the fully automated Panther Fusion system to detect and differentiate parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus, and parainfluenza 4 virus directly from nasopharyngeal swab specimens.
The Panther Fusion Paraflu assay involves the following steps: Sample lysis, nucleic acid capture and elution, and multiplex RT-PCR where analytes (when present) are simultaneously amplified, detected and differentiated. Nucleic acid capture and elution takes place in a single tube on the Panther Fusion system. The eluate is transferred to the Panther Fusion system reaction tube containing the assay reagents. Multiplex RT-PCR is then performed for the eluted nucleic acid on the Panther Fusion system.
Nucleic acid capture and elution: Prior to processing and testing on the Panther Fusion system, specimens are transferred to a tube containing specimen transport media (STM) that lyses the cells, releases target nucleic acid and protects them from degradation during storage. The Internal Control-S (IC-S) is added to each test specimen and controls via the working Panther Fusion Capture Reagent-S (wFCR-S). The IC-S in the reagent is used to monitor specimen processing, amplification and detection. Magnetic particles with covalently bound oligonucleotides mediate the nucleic acid capture. Capture oligonucleotides hybridize to total nucleic acid in the test specimen. Hybridized nucleic acid is then separated from the lysed specimen in a magnetic field. Wash and aspiration steps remove extraneous components debris from the reaction tube. The elution step elutes purified nucleic acid.
Elution transfer and RT-PCR: During the elution transfer step, eluted nucleic acid is transferred to a Panther Fusion tube already containing oil and reconstituted mastermix. Target amplification occurs via RT-PCR. A reverse transcriptase generates a DNA copy of the target sequence. Target specific forward and reverse primers and probes then amplify targets while simultaneously detecting and discriminating multiple target types via multiplex RT-PCR. The Panther Fusion system compares the fluorescence signal to a predetermined cut-off to produce a qualitative result for the presence or absence of the analyte. The positive result for each analyte will be accompanied by the cycle threshold (Ct) value.
Mentions image processing
Not Found
Mentions AI, DNN, or ML
Not Found
Input Imaging Modality
Not Found
Anatomical Site
nasopharyngeal (NP) swab specimens
Indicated Patient Age Range
all ages
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
Description of Clinical Performance Study:
This study was performed to demonstrate clinical performance characteristics for the Panther Fusion Paraflu assay. A prospective multicenter study was conducted with leftover, remnant nasopharyngeal (NP) swab specimens from male and female individuals of all ages exhibiting signs and/or symptoms of a respiratory tract infection. Four participating US pediatric/ adolescent, private, and/or university hospitals obtained 2961 leftover, remnant NP swab specimens. The samples were tested with the Panther Fusion Paraflu assay, with reference viral culture followed by direct fluorescent antibody (DFA) identification (for HPIV-1, and HPIV-3), and with 2 unique and independently developed reverse transcriptase PCR assays followed by bi-directional sequencing (PCR/sequencing, for HPIV-4). A validated PCR assay was used for discordant resolution testing for HPIV-1, and HPIV-3. No discordant resolution testing was performed for HPIV-4 since reference assay was based on composite result of two different reverse transcriptase PCR assays followed by by-directional sequencing as stated above. Performance characteristics were estimated relative to valid culture/DFA or PCR/sequencing results for each sample. Sensitivity and specificity (for HPIV-1, and HPIV-3) and positive and negative percent agreement (for HPIV-4) were estimated with corresponding 2-sided 95% Score CIs. Analyses were performed separately for each target analyte (HPIV-1, HPIV-2, HPIV-3, and HPIV-4).
Sample size:
2961 leftover, remnant NP swab specimens were obtained. Of the 2961 specimens, 31 specimens/samples were withdrawn, 2930 samples were processed in valid Panther Fusion Paraflu runs, 2877 (98.2%) had final valid results, and 53 (1.8%) had final invalid results. Of the 2877 samples with valid Panther Fusion results, 1359 samples were from females and 1518 samples were from males. 2870 samples were evaluable for analyses for each analyte.
Data Source:
US pediatric/adolescent, private, and/or university hospitals.
Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)
Study Type: Analytical (Non-Clinical) Studies.
Analytical Sensitivity and Limit of Detection (LoD)
The LoD was determined by testing dilution panels for HPIV-1, HPIV-2, HPIV-3 and HPIV-4 made by spiking them into pooled negative NP swab clinical specimens. At least 36 replicates were tested using three reagent lots on three instruments per test concentration, per each virus types. The LoD was based on results from the lowest concentration with > 95% positive rate. The LoD values for each virus type were verified by testing newly prepared panel for at least 20 replicates.
LoD results:
- HPIV-1: 1x10^-2.0 TCID50/mL
- HPIV-2: 1x10^2.0 TCID50/mL
- HPIV-3: 1x10^1.0 TCID50/mL
- HPIV-4: 1x10^0.5 TCID50/mL
Interference Study
Mucin, whole blood and other potentially interfering substances (medications and over-thecounter or OTC products) that may be present in the samples were evaluated. Clinically relevant amount of the potentially interfering substances were added to simulated clinical matrix and tested unspiked with cultured HPIV-1, HPIV-2, HPIV-3 and HPIV-4 at their respective 3X LoD concentrations. No interference in performance of the Panther Fusion Paraflu assay was observed.
Competitive Interference Study
Competitive Interference was evaluated using a simulated clinical matrix with pairs of target viruses at two different concentrations: near the Limit of Detection (3 - 5X LoD) and high (1000X LoD). The presence of two viruses at varying concentrations in a single sample had no effect on the analytical sensitivity (100% detection for both targets) at the concentration tested.
Analytical Specificity Study
The analytical specificity was evaluated by testing a panel of 58 organisms (31 viral, 26 bacterial, and 1 yeast strain) representing common respiratory pathogens or flora. Bacteria and yeast were tested at concentrations of 10^5 to 10^7 CFU/mL or IFU/mL, except where noted. Viruses were tested at concentrations of 10^3 to 10^7 TCID50/mL. Analytical specificity of the Panther Fusion Paraflu assay was 100% for HPIV-1, HPIV-2, HPIV-3 and HPIV-4.
Carry-Over/Contamination Study
The study was performed with negative samples alternately placed between high HPIV positive samples (HPIV-2 at 10^6 TCID50/mL, over 10,000X LoD). A total of nine separate runs with 450 positive and 450 negative samples were tested across three different instruments. The carry-over rate was 0.0%.
Assay Precision Study
Evaluated with a 9-member panel tested by three operators on two separate runs per day, using three reagent lots on three Panther Fusion systems over 45 days.
Sample size: 160-162 replicates per panel member.
Key results:
- HPIV-1: 3X LoD (100.0% positive), 1X LoD (100.0% positive), 0.01X LoD (3.1% positive), Negative (0.0% positive).
- HPIV-2: 3X LoD (100.0% positive), 1X LoD (100.0% positive), 0.01X LoD (27.8% positive), Negative (0.0% positive).
- HPIV-3: 3X LoD (100.0% positive), 1X LoD (97.5% positive), 0.01X LoD (4.9% positive), Negative (0.6% positive).
- HPIV-4: 3X LoD (100.0% positive), 1X LoD (98.1% positive), 0.01X LoD (4.3% positive), Negative (0.0% positive).
Overall (total) %CV for Ct values ranged from 1.0% to 8.5% for different targets and concentrations.
Clinical Performance Study
Study type: Prospective multicenter study.
Sample size: 2870 evaluable samples.
Key results:
- HPIV-1: 1.5% positivity.
- HPIV-2: 1.3% positivity.
- HPIV-3: 2.8% positivity.
- HPIV-4: 1.2% positivity.
Reproducibility Study
Evaluated at three US sites using nine panel members, one lot of assay reagents, and six operators (two at each site). Testing was performed for at least five days, with three replicates of each panel member per run.
Key results:
- Agreement with expected results was 100% in the negative and moderate positive panel members.
- Agreement was >=96.6% in low-positive panel members for HPIV-1, HPIV-2, HPIV-3, and HPIV-4.
- Total HPIV-1, HPIV-2, HPIV-3, and HPIV-4 signal variability measured as %CV ranged from 1.11% to 5.88% in low and moderate positive panel members.
- Signal variability as measured as %CV was 99.9)
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
§ 866.3980 Respiratory viral panel multiplex nucleic acid assay.
(a)
Identification. A respiratory viral panel multiplex nucleic acid assay is a qualitative in vitro diagnostic device intended to simultaneously detect and identify multiple viral nucleic acids extracted from human respiratory specimens or viral culture. The detection and identification of a specific viral nucleic acid from individuals exhibiting signs and symptoms of respiratory infection aids in the diagnosis of respiratory viral infection when used in conjunction with other clinical and laboratory findings. The device is intended for detection and identification of a combination of the following viruses:(1) Influenza A and Influenza B;
(2) Influenza A subtype H1 and Influenza A subtype H3;
(3) Respiratory Syncytial Virus subtype A and Respiratory Syncytial Virus subtype B;
(4) Parainfluenza 1, Parainfluenza 2, and Parainfluenza 3 virus;
(5) Human Metapneumovirus;
(6) Rhinovirus; and
(7) Adenovirus.
(b)
Classification. Class II (special controls). The special controls are:(1) FDA's guidance document entitled “Class II Special Controls Guidance Document: Respiratory Viral Panel Multiplex Nucleic Acid Assay;”
(2) For a device that detects and identifies Human Metapneumovirus, FDA's guidance document entitled “Class II Special Controls Guidance Document: Testing for Human Metapneumovirus (hMPV) Using Nucleic Acid Assays;” and
(3) For a device that detects and differentiates Influenza A subtype H1 and subtype H3, FDA's guidance document entitled “Class II Special Controls Guidance Document: Testing for Detection and Differentiation of Influenza A Virus Subtypes Using Multiplex Nucleic Acid Assays.” See § 866.1(e) for the availability of these guidance documents.
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Public Health Service
Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002
October 23, 2017
Hologic, Inc. Jeffrey Hergesheimer Regulatory Affairs Specialist 10210 Genetic Center Drive San Diego CA 92121
Re: K172282
Trade/Device Name: Panther Fusion Paraflu Assay Regulation Number: 21 CFR 866.3980 Regulation Name: Respiratory viral panel multiplex nucleic acid assay Regulatory Class: II Product Code: OOU, OOI Dated: July 28, 2017 Received: July 28, 2017
Dear Mr. Hergesheimer:
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,
Steven R. Gitterman -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
2
Indications for Use
510(k) Number (if known) K172282
Device Name Panther Fusion Paraflu Assay
Indications for Use (Describe)
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test for the rapid and qualitative detection and differentiation of parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus and parainfluenza 4 virus (HPIV-1, HPIV-2, HPIV-3, and HPIV-4). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection.
This assay is intended to aid in the differential diagnosis of HPIV-1, HPIV-2, HPIV-3, and HPIV-4 infections in humans. Negative results do not preclude HPIV-1, HPIV-3, and HPIV-4 infections and should not be used as the sole basis for treatment or other management decisions. This assay is designed for use on the Panther Fusion system.
| 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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3
510(k) SUMMARY Panther Fusion Paraflu Assay
I. SUBMITTER
Hologic, Inc. 10210 Genetic Center Drive San Diego, CA 92121
| Contact Person: | Jeffrey Hergesheimer, MS, RAC
Regulatory Affairs Specialist
Jeffrey.Hergesheimer@Hologic.com
Phone: (858) 410-8536
Fax: (858) 410-5557 |
| ----------------- | ---------------------------------------------------------------------------------------------------------------------------------------------------- |
|---|
Date Prepared: July 24, 2017
II. DEVICE
| Proprietary Name of Device: | Panther Fusion Paraflu Assay | |
|---|---|---|
| Classification Name: | Respiratory viral panel multiplex nucleic acid assay | |
| Regulation Number: | 21 CFR 866.3980 and 862.2570 | |
| Regulatory Class: | Class II | |
| Product Code: | OOU and OOI |
III. PREDICATE DEVICE
The predicate device is the Prodesse ProParaFlu+ Assay (K153223; cleared December 9, 2015, Hologic, San Diego, CA).
IV. DEVICE DESCRIPTION
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test developed for use on the fully automated Panther Fusion system to detect and differentiate parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus, and parainfluenza 4 virus directly from nasopharyngeal swab specimens.
4
The Panther Fusion Paraflu assay involves the following steps: Sample lysis, nucleic acid capture and elution, and multiplex RT-PCR where analytes (when present) are simultaneously amplified, detected and differentiated. Nucleic acid capture and elution takes place in a single tube on the Panther Fusion system. The eluate is transferred to the Panther Fusion system reaction tube containing the assay reagents. Multiplex RT-PCR is then performed for the eluted nucleic acid on the Panther Fusion system.
Nucleic acid capture and elution: Prior to processing and testing on the Panther Fusion system, specimens are transferred to a tube containing specimen transport media (STM) that lyses the cells, releases target nucleic acid and protects them from degradation during storage. The Internal Control-S (IC-S) is added to each test specimen and controls via the working Panther Fusion Capture Reagent-S (wFCR-S). The IC-S in the reagent is used to monitor specimen processing, amplification and detection. Magnetic particles with covalently bound oligonucleotides mediate the nucleic acid capture. Capture oligonucleotides hybridize to total nucleic acid in the test specimen. Hybridized nucleic acid is then separated from the lysed specimen in a magnetic field. Wash and aspiration steps remove extraneous components debris from the reaction tube. The elution step elutes purified nucleic acid.
Elution transfer and RT-PCR: During the elution transfer step, eluted nucleic acid is transferred to a Panther Fusion tube already containing oil and reconstituted mastermix. Target amplification occurs via RT-PCR. A reverse transcriptase generates a DNA copy of the target sequence. Target specific forward and reverse primers and probes then amplify targets while simultaneously detecting and discriminating multiple target types via multiplex RT-PCR. The Panther Fusion system compares the fluorescence signal to a predetermined cut-off to produce a qualitative result for the presence or absence of the analyte. The positive result for each analyte will be accompanied by the cycle threshold (Ct) value. The analytes and the channel used for their detection on the Panther Fusion system is summarized in the table below.
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| Analyte | Gene Targeted | Instrument Channel |
|---|---|---|
| HPIV-1 | Hemagglutinin neuraminidase | FAM |
| HPIV-2 | Hemagglutinin neuraminidase | HEX |
| HPIV-3 | Hemagglutinin neuraminidase | ROX |
| HPIV-4 | Nucleocapsid | RED647 |
| Internal Control | Not applicable | RED677 |
Assay Components
The reagents required to perform the Panther Fusion Paraflu assay are packaged and sold separately. There are 7 boxes containing 9 reagents which are required for sample processing. A description of the components that are required to perform the Panther Fusion Paraflu assay are detailed in Table 1. In addition, there is one ancillary kit, Panther Fusion Specimen Lysis Tubes, which is required for processing of specimens prior to testing on the Panther Fusion system.
| Box | Components Description | |
|---|---|---|
| 1 | Panther Fusion Paraflu Assay Cartridges | |
| 2 | Panther Fusion Extraction Reagent-S Box Contains: Panther Fusion Capture Reagent-S Panther Fusion Enhancer Reagent-S | |
| 3 | Panther Fusion Internal Control-S | |
| 4 | Panther Fusion Reconstitution Buffer I | |
| 5 | Panther Fusion Elution Buffer | |
| 6 | Panther Fusion Oil | |
| 7 | Panther Fusion Paraflu Assay Controls Box Contains: Panther Fusion Paraflu Positive Control Panther Fusion Negative Control |
Table 1: Reagents Required to Perform the Panther Fusion Paraflu Assay
In addition, select components can also be ordered in the following bundles:
- Panther Fusion Universal Fluids Kit: (contains Panther Fusion Oil and Panther Fusion . Elution Buffer).
- Panther Fusion Assay Fluids I-S: (contains Panther Fusion Extraction Reagents-S, . Panther Fusion Internal Control-S, and Panther Fusion Reconstitution Buffer I).
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Instrumentation
The Panther Fusion Paraflu assay has been designed for and validated on the Panther Fusion system. The Panther Fusion system is an integrated hardware and software system that together with the Panther Fusion Paraflu assay fully automates all the steps necessary to perform the assay.
The Panther Fusion system integrates Hologic's commercialized Panther instrument system with an add-on sidecar, the Panther Fusion module, which extends the functionality of the Panther system by increasing the assay processing capabilities to include multiplex real-time RT-PCR. The Panther Fusion module includes instrument hardware and can be installed on existing Panther instruments or ordered with new Panther instruments.
The Panther Fusion system employs non-specific target capture (NSTC) for the purification of RNA and DNA from the sample, followed by nucleic acid amplification and real-time fluorescent detection. The process involves sample loading and preparation (i.e. nucleic acid extraction) on the Panther instrument using the same workflow and processing steps as for other commercialized Hologic Aptima TMA assays. The extracted nucleic acid for each sample is transferred to the Panther Fusion module where PCR amplification and detection occurs.
V. INDICATIONS FOR USE
Intended Use
The Panther Fusion Paraflu assay is a multiplex real-time PCR (RT-PCR) in vitro diagnostic test for the rapid and qualitative detection and differentiation of parainfluenza 1 virus, parainfluenza 2 virus, parainfluenza 3 virus and parainfluenza 4 virus (HPIV-1, HPIV-2, HPIV-3, and HPIV-4). Nucleic acids are isolated and purified from nasopharyngeal (NP) swab specimens obtained from individuals exhibiting signs and symptoms of a respiratory tract infection.
This assay is intended to aid in the differential diagnosis of HPIV-1, HPIV-3, and HPIV-4 infections in humans. Negative results do not preclude HPIV-1, HPIV-2, HPIV-3, and
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HPIV-4 infections and should not be used as the sole basis for treatment or other management decisions. This assay is designed for use on the Panther Fusion system.
VI. COMPARISON OF TECHNOLOGICAL CHARACTERISTICS WITH THE PREDICATE DEVICE
A comparison of the Panther Fusion Paraflu assay to the predicate Prodesse ProParaFlu+ (K153223) is summarized in Table 2 (similarities) and Table 3 (differences).
| Item | Prodesse ProParaFlu+ Assay
(Predicate Device) | Panther Fusion Paraflu Assay
(Subject Device) |
|--------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Technology Principle
of Operation | Multiplex Real Time RT-PCR | Same |
| Analyte | Viral RNA | Same |
| Patient Population | Male and female patients with
signs/symptoms of respiratory
infection | Same |
| Specimen Types | Nasopharyngeal (NP) swab specimens | Same |
| Assay Controls | Internal control in each sample.
External control processed at periodic
intervals. | Same |
| Item | Prodesse ProParaFlu+ Assay
(Predicate Device) | Panther Fusion Paraflu Assay
(Subject Device) |
| Organisms Detected | Human Parainfluenza Virus (HPIV)
Types 1, 2, and 3 | Same.
Plus HPIV-4. |
| Platform | Manual multiplex real-time RT-PCR
platform.
Uses Roche MagNA Pure LC System
or bioMerieux NucliSENS easyMAG
for nucleic acid extraction and the
Cepheid SmartCycler II system for real
time RT-PCR | Automated multiplex real-time RT-
PCR platform.
Uses Panther Fusion system for all
steps including nucleic acid extraction,
amplification, detection and result
processing. |
| Intended Use | The Prodesse ProParaflu+ Assay is a
multiplex Real-Time PCR (RT-PCR)
in vitro diagnostic test for the
qualitative detection and
discrimination of Parainfluenza 1
Virus, Parainfluenza 2 Virus and
Parainfluenza 3 Virus (HPIV-1, HPIV-
2 and HPIV-3) nucleic acids isolated
and purified from nasopharyngeal (NP)
swab specimens obtained from
individuals exhibiting signs and
symptoms of respiratory tract
infections. This Assay targets the
conserved regions of the
Hemagglutinin-Neuraminidase (HN)
gene of HPIV-1, HPIV-2 and HPIV-3,
respectively. The detection and
discrimination of HPIV-1, HPIV-2 and
HPIV-3 nucleic acids from
symptomatic patients aid in the
diagnosis of human respiratory tract
parainfluenza infections if used in
conjunction with other clinical and
laboratory findings. This test is not
intended to detect Parainfluenza 4a or
Parainfluenza 4b Viruses.
Negative test results are presumptive
and should be confirmed by cell
culture. Negative results do not
preclude Parainfluenza 1, 2 or 3 virus
infections and should not be used as
the sole basis for treatment or other
management decisions. | The Panther Fusion Paraflu assay is a
multiplex real-time PCR (RT-PCR) in
vitro diagnostic test for the rapid and
qualitative detection and
differentiation of parainfluenza 1 virus,
parainfluenza 2 virus, parainfluenza 3
virus and parainfluenza 4 virus (HPIV-
1, HPIV-2, HPIV-3, and HPIV-4).
Nucleic acids are isolated and purified
from nasopharyngeal (NP) swab
specimens obtained from individuals
exhibiting signs and symptoms of a
respiratory tract infection.
This assay is intended to aid in the
differential diagnosis of HPIV-1,
HPIV-2, HPIV-3, and HPIV-4
infections in humans. Negative results
do not preclude HPIV-1, HPIV-2,
HPIV-3, and HPIV-4 infections and
should not be used as the sole basis for
treatment or other management
decisions. This assay is designed for
use on the Panther Fusion system. |
| Time to Obtain Test
Results | Approximately 4 hours | Approximately 2.5 hours |
Table 2: Similarities Between Panther Fusion Paraflu Assay and Predicate Device
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Table 3: Differences Between Panther Fusion Paraflu Assay and Predicate Device
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VII. PERFORMANCE DATA
The following performance data were provided in support of the substantial equivalence determination.
Brief Description of Analytical (Non-Clinical) Studies
The following analytical studies (non-clinical) were conducted to support the clearance of the Panther Fusion Paraflu Assay on the Panther Fusion System.
Analytical Sensitivity and Limit of Detection (LoD) of Nasopharyngeal Swab Specimens
The LoD was determined by testing dilution panels for HPIV-1, HPIV-2, HPIV-3 and HPIV-4 made by spiking them into pooled negative NP swab clinical specimens. At least 36 replicates were tested using three reagent lots on three instruments per test concentration, per each virus types. The LoD was based on results from the lowest concentration with > 95% positive rate. The LoD values for each virus type were verified by testing newly prepared panel for at least 20 replicates. Verified LOD concentrations are shown in Table 4.
| Viral Strain | LoD Concentration |
|---|---|
| HPIV-1 | $1x10^{-2.0}$ TCID50/mL |
| HPIV-2 | $1x10^{2.0}$ TCID50/mL |
| HPIV-3 | $1x10^{1.0}$ TCID50/mL |
| HPIV-4 | $1x10^{0.5}$ TCID50/mL |
Table 4: LoD Determination Panel Description
Interference
Mucin, whole blood and other potentially interfering substances (medications and over-thecounter or OTC products) that may be present in the samples were evaluated in the Panther Fusion Paraflu assay. Clinically relevant amount of the potentially interfering substances were added to simulated clinical matrix and tested unspiked with cultured HPIV-1, HPIV-2, HPIV-3 and HPIV-4 at their respective 3X LoD concentrations. The substances consisted of nasal sprays (liquid and powder), ingestible pills, lozenges, injectable and endogenous substances. No interference in performance of the Panther Fusion Paraflu assay was observed
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in the presence of a representative brand of the following potentially interfering substances at the concentrations stated in Table 5.
| Type | Substance Name | Active Ingredient(s) | Concentration |
|---|---|---|---|
| Endogenous | Mucin | Purified mucin protein | 60 µg/mL |
| Nasal sprays or drops | Human blood | Blood | 2% v/v |
| Neo-Synephrine® | Phenylephrine | 15% v/v | |
| Anefrin | Oxymetazoline | 15% v/v | |
| Saline | Sodium chloride | 15% v/v | |
| Ventolin® HFA | Albuterol | 15% v/v | |
| Nasal | |||
| corticosteroids | QVAR®, Beconase AQ | Beclomethasone | 5% v/v |
| Dexacort | Dexamethasone | 5% v/v | |
| AEROSPAN® | Flunisolide | 5% v/v | |
| Nasacort | Triamcinolone | 5% v/v | |
| Rhinocort | Budesonide | 5% v/v | |
| Nasonex | Mometasone | 5% v/v | |
| Flonase | Fluticasone | 5% v/v | |
| Nasal gel | Zicam® (Allergy Relief) | Luffa opperculata, Galphimia, | |
| Glauca, Histaminum | |||
| hydrochloricum, Sulfur | 5% v/v | ||
| Throat lozenges | Chloraseptic Throat | ||
| Lozenges | Benzocaine | ||
| Menthol | 0.63 mg/mL | ||
| Anti-viral drugs | Relenza® | Zanamivir | |
| TamiFlu | Oseltamivir | 25 mg/mL | |
| Rebitol | Ribavirin | 20 mg/mL | |
| Antibiotic, nasal | |||
| ointment | Bactroban cream | Mupirocin | 10 mg/mL |
| Antibiotic, systemic | Tobramycin | Tobramycin | 4.0 µg/mL |
Table 5: Potentially Interfering Substances
Competitive Interference
Competitive Interference of the Panther Fusion Paraflu assay was evaluated using a simulated clinical matrix with pairs of target viruses at two different concentrations. One of the concentrations was near the Limit of Detection (3 - 5X LoD) while the other concentration was high (1000X LoD). The presence of two viruses at varying concentrations in a single sample had no effect on the analytical sensitivity (100% detection for both targets) at the concentration tested (see Table 6).
11
| Target 1 | Target 2 | Result | ||||||
|---|---|---|---|---|---|---|---|---|
| Condition | Description | Concentration | Description | Concentration | HPIV-1 | HPIV-2 | HPIV-3 | HPIV-4 |
| 1 | HPIV-1 | 3X LoD | HPIV-2 | 1000X LoD | + | + | - | - |
| 2 | HPIV-1 | 3X LoD | HPIV-3 | 1000X LoD | + | - | + | - |
| 3 | HPIV-1 | 5X LoD | HPIV-4 | 1000X LoD | + | - | - | + |
| 4 | HPIV-2 | 3X LoD | HPIV-1 | 1000X LoD | + | + | - | - |
| 5 | HPIV-2 | 3X LoD | HPIV-3 | 1000X LoD | - | + | + | - |
| 6 | HPIV-2 | 3X LoD | HPIV-4 | 1000X LoD | - | + | - | + |
| 7 | HPIV-3 | 3X LoD | HPIV-1 | 1000X LoD | + | - | - | - |
| 8 | HPIV-3 | 3X LoD | HPIV-2 | 1000X LoD | - | + | + | - |
| 9 | HPIV-3 | 3X LoD | HPIV-4 | 1000X LoD | - | - | + | + |
| 10 | HPIV-4 | 3X LoD | HPIV-1 | 1000X LoD | + | - | - | + |
| 11 | HPIV-4 | 3X LoD | HPIV-2 | 1000X LoD | - | + | - | + |
| 12 | HPIV-4 | 3X LoD | HPIV-3 | 1000X LoD | - | - | + | + |
Table 6: Co-Infection Concentrations and Results
Analytical Specificity
The analytical specificity of the Panther Fusion Paraflu assay was evaluated by testing a panel of 58 organisms, consisting of 31 viral, 26 bacterial, and 1 yeast strain representing common respiratory pathogens or flora commonly present in respiratory tract. Bacteria and yeast were tested at concentrations of 105 to10° CFU/mL or IFU/mL, except where noted. Viruses were tested at concentrations of 103 to 10' TCID50/mL. Analytical specificity of the Panther Fusion Paraflu assay was 100% for HPIV-1, HPIV-2, HPIV-3 and HPIV-4 (see Table 7).
| Organism | Concentration | Result | |||
|---|---|---|---|---|---|
| HPIV-1 | HPIV-2 | HPIV-3 | HPIV-4 | ||
| Adenovirus 1 | 1x105 TCID50/mL | - | - | - | - |
| Adenovirus 7a | 1x105 TCID50/mL | - | - | - | - |
| Bordetella bronchiseptica | 1x107 CFU/mL | - | - | - | - |
| Bordetella pertussis | 1x108 CFU/mL | - | - | - | - |
| Candida albicans | 1x107 CFU/mL | - | - | - | - |
| Chlamydia trachomatis | 1x105 CFU/mL | - | - | - | - |
| Chlamydophila pneumoniae | |||||
| (formerly Chlamydia pneumoniae) | 1x105 IFU/mL | - | - | - | - |
| CMV Strain AD 169 | 1x104 TCID50/mL | - | - | - | - |
| Coronavirus 229E | 1x104 TCID50/mL | - | - | - | - |
| Corynebacterium diphtheria | 1x107 CFU/mL | - | - | - | - |
| Result | |||||
| Organism | Concentration | HPIV-1 | HPIV-2 | HPIV-3 | HPIV-4 |
| Coxsackie B4 | 1x106 TCID50/mL | - | - | - | - |
| Coxsackie B5/10/2006 | 1x105 TCID50/mL | - | - | - | - |
| E. coli | 1x107 CFU/mL | - | - | - | - |
| EBV | 1x107 TCID50/mL | - | - | - | - |
| Echovirus 11 | 1x105 TCID50/mL | - | - | - | - |
| Echovirus 2 | 1x104 TCID50/mL | - | - | - | - |
| Echovirus 3 | 1x105 TCID50/mL | - | - | - | - |
| Echovirus 6 | 1x104 TCID50/mL | - | - | - | - |
| Enterovirus 68 | 1x105 TCID50/mL | - | - | - | - |
| Enterovirus 70 | 1x104 TCID50/mL | - | - | - | - |
| Haemophilus Influenzae | 1x107 CFU/mL | - | - | - | - |
| HPIV-1, C35 | 1x102 TCID50/mL | + | - | - | - |
| HPIV-2, Greer | 1x102 TCID50/mL | - | + | - | - |
| HPIV-3, C243 | 1x102 TCID50/mL | - | - | + | - |
| HPIV-4b, CH19503 | 1x102 TCID50/mL | - | - | - | + |
| hMPV Subtype A2 | 1x106 TCID50/mL | - | - | - | - |
| HSV-1 Macinytre Strain | 1x105 TCID50/mL | - | - | - | - |
| HSV-2 Type 2G Strain | 1x105 TCID50/mL | - | - | - | - |
| Influenza A (H1N1) | 1x104 TCID50/mL | - | - | - | - |
| Influenza A (H3N2) | 1x104 TCID50/mL | - | - | - | - |
| Influenza B | 1x104 TCID50/mL | - | - | - | - |
| Klebsiella pneumonia | 1x107 CFU/mL | - | - | - | - |
| Lactobacillus plantarum | 1x107 CFU/mL | - | - | - | - |
| Legionella pneumophila | 1x107 CFU/mL | - | - | - | - |
| Measles/7/2000 | 1x105 TCID50/mL | - | - | - | - |
| Moraxella catarrhalis | 1x106 CFU/mL | - | - | - | - |
| Mumps virus | 1x104 TCID50/mL | - | - | - | - |
| Mycobacterium intracellulare | 1x1010 rRNA | ||||
| Copies/mL | - | - | - | - | |
| Mycobacterium tuberculosis | 1x1010 rRNA | ||||
| Copies/mL | - | - | - | - | |
| Mycoplasma pneumoniae | 1x106 CFU/mL | - | - | - | - |
| Neisseria gonorrhea | 1x107 CFU/mL | - | - | - | - |
| Neisseria meningitides | 1x107 CFU/mL | - | - | - | - |
| Neisseria mucosa | 1x107 CFU/mL | - | - | - | - |
| Polio virus | 1x106 TCID50/mL | - | - | - | - |
| Proteus mirabilis | 1x107 CFU/mL | - | - | - | - |
| Proteus vulgaris | 1x107 CFU/mL | - | - | - | - |
| Pseudomonas aeruginosa | 1x107 CFU/mL | - | - | - | - |
| Rhinovirus 1A | 1x105 TCID50/mL | - | - | - | - |
| Organism | Concentration | Result | |||
| HPIV-1 | HPIV-2 | HPIV-3 | HPIV-4 | ||
| RSV A | 1x104 TCID50/mL | - | - | - | - |
| RSV B | 1x104 TCID50/mL | - | - | - | - |
| Staphlycoccus aureus | 1x107 CFU/mL | - | - | - | - |
| Staphlycoccus epidermidis | 1x107 CFU/mL | - | - | - | - |
| Streptococcus pneumoniae | 1x106 CFU/mL | - | - | - | - |
| Streptococcus pyogenes | 1x107 CFU/mL | - | - | - | - |
| Streptococcus salivarius | 1x106 CFU/mL | - | - | - | - |
| Tatlockia micdadei (formerly | |||||
| Legionella micdadei) | 1x107 CFU/mL | - | - | - | - |
| Varicella Zoster Virus | 1x103 TCID50/mL | - | - | - | - |
Table 7: Specificity Results
12
13
Carry-Over/Contamination
The carry-over/cross-contamination study was performed with negative samples alternately placed between high HPIV positive samples and tested. High positive samples were prepared by spiking HPIV-2 at 10° TCID50/mL (over 10,000X LoD). A total of nine separate runs with negative samples and positive samples placed in a checkerboard pattern were tested over three different instruments for a combined total of 450 positive and 450 negative samples. The carry-over rate was 0.0%.
Assay Precision
Panther Fusion Paraflu assay precision was evaluated with a 9-member panel. The panel was tested by three operators on two separate runs per day, using three reagent lots on three Panther Fusion systems over 45 days. The panel members, along with a summary of the agreement with expected results for each target is presented in Table 8. The mean and variability analysis between instruments, between reagent lots, between operators, between days, between runs and within runs, and overall (total) for Ct are also presented in Table 9.
14
| Target | Panel Description | % Positive | % Agreement (95% CI) |
|---|---|---|---|
| HPIV-1 | 3X LoD | 100.0% (162/162) | 100.0% (97.7 - 100%) |
| 1X LoD | 100.0% (160/160) | 100.0% (97.7 - 100%) | |
| 0.01X LoD | 3.1% (5/161) | 96.9% (92.9 - 98.7%) | |
| Negative | 0.0% (0/162) | 100.0% (97.7 - 100%) | |
| HPIV-2 | 3X LoD | 100.0% (162/162) | 100.0% (97.7 - 100%) |
| 1X LoD | 100.0% (162/162) | 100.0% (97.7 - 100%) | |
| 0.01X LoD | 27.8% (45/162) | 72.2% (64.9 - 78.5%) | |
| Negative | 0.0% (0/162) | 100.0% (97.7 - 100%) | |
| HPIV-3 | 3X LoD | 100.0% (162/162) | 100.0% (97.7 - 100%) |
| 1X LoD | 97.5% (158/162) | 97.5% (93.8 - 99.0%) | |
| 0.01X LoD | 4.9% (8/162) | 95.1% (90.6 - 97.5%) | |
| Negative | 0.6% (1/162) | 99.4% (96.6 - 99.9%) | |
| HPIV-4 | 3X LoD | 100.0% (161/161) | 100.0% (97.7 - 100%) |
| 1X LoD | 98.1% (159/162) | 98.1% (94.7 - 99.4%) | |
| 0.01X LoD | 4.3% (7/162) | 95.7% (91.4 - 97.9%) | |
| Negative | 0.0% (0/162) | 100.0% (97.7 - 100%) |
Table 8: Percent Positive and Agreement
15
| Target | Panel
Member | Mean | Between
Instruments | | Between
Reagent Lots | | Between
Operators | | Between Days | | Between Runs | | Within Run | | Total | |
|--------|-----------------|------|------------------------|-----------|-------------------------|-----------|----------------------|-----------|--------------|-----------|--------------|-----------|------------|-----------|-------|-----------|
| | | | SD | CV
(%) | SD | CV
(%) | SD | CV
(%) | SD | CV
(%) | SD | CV
(%) | SD | CV
(%) | SD | CV
(%) |
| HPIV-1 | 3X LoD | 35.2 | 0.0 | 0.0 | 0.1 | 0.2 | 0.0 | 0.0 | 0.1 | 0.3 | 0.0 | 0.0 | 0.4 | 1.1 | 0.4 | 1.2 |
| HPIV-1 | 1X LoD | 37.0 | 0.0 | 0.0 | 0.1 | 0.4 | 0.0 | 0.0 | 0.0 | 0.2 | 0.0 | 0.0 | 0.6 | 1.7 | 0.6 | 1.8 |
| HPIV-1 | 0.01X LoD | 42.3 | 0.3 | 0.9 | 0.4 | 1.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.4 | 1.0 | 0.7 | 1.7 |
| HPIV-2 | 3X LoD | 32.8 | 0.0 | 0.0 | 0.0 | 0.1 | 0.0 | 0.1 | 0.0 | 0.0 | 0.1 | 0.3 | 0.3 | 0.9 | 0.3 | 1.0 |
| HPIV-2 | 1X LoD | 34.3 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.1 | 0.0 | 0.0 | 0.0 | 0.0 | 0.5 | 1.5 | 0.5 | 1.5 |
| HPIV-2 | 0.01X LoD | 40.7 | 0.1 | 0.3 | 0.0 | 0.1 | 0.0 | 0.0 | 0.3 | 0.8 | 0.0 | 0.0 | 1.1 | 2.8 | 1.2 | 3.0 |
| HPIV-3 | 3X LoD | 35.5 | 0.5 | 1.4 | 0.0 | 0.0 | 0.0 | 0.0 | 0.2 | 0.7 | 0.0 | 0.0 | 1.5 | 4.4 | 1.6 | 4.7 |
| HPIV-3 | 1X LoD | 37.5 | 0.2 | 0.6 | 0.4 | 1.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.3 | 1.0 | 2.0 | 5.4 | 2.1 | 5.7 |
| HPIV-3 | 0.01X LoD | 40.1 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 0.0 | 3.3 | 8.3 | 0.7 | 1.7 | 3.4 | 8.5 |
| HPIV-4 | 3X LoD | 36.2 | 0.0 | 0.0 | 0.0 | 0.0 | 0.3 | 0.9 | 0.0 | 0.0 | 0.5 | 1.4 | 1.5 | 4.3 | 1.6 | 4.6 |
| HPIV-4 | 1X LoD | 38.1 | 0.0 | 0.0 | 0.0 | 0.0 | 0.2 | 0.7 | 0.0 | 0.0 | 0.0 | 0.0 | 1.9 | 5.0 | 1.9 | 5.1 |
| HPIV-4 | 0.01X LoD | 42.5 | 0.0 | 0.0 | 1.1 | 2.6 | 0.8 | 1.9 | 0.0 | 0.0 | 0.0 | 0.0 | 0.7 | 1.8 | 1.6 | 3.7 |
| IC | Negative | 32.1 | 0.0 | 0.0 | 0.0 | 0.1 | 0.0 | 0.2 | 0.0 | 0.0 | 0.1 | 0.5 | 0.4 | 1.2 | 0.4 | 1.4 |
Table 9: Signal Variability Analysis Results
16
Brief Description of Clinical Studies
Clinical testing of the Panther Fusion Paraflu assay on the Panther Fusion system included performance and reproducibility testing. Substantial equivalence is based in part on the performance study.
Clinical Performance Study
This study was performed to demonstrate clinical performance characteristics for the Panther Fusion Paraflu assay. A prospective multicenter study was conducted with leftover, remnant nasopharyngeal (NP) swab specimens from male and female individuals of all ages exhibiting signs and/or symptoms of a respiratory tract infection. Four participating US pediatric/ adolescent, private, and/or university hospitals obtained 2961 leftover, remnant NP swab specimens. The samples were tested with the Panther Fusion Paraflu assay, with reference viral culture followed by direct fluorescent antibody (DFA) identification (for HPIV-1, and HPIV-3), and with 2 unique and independently developed reverse transcriptase PCR assays followed by bi-directional sequencing (PCR/sequencing, for HPIV-4). A validated PCR assay was used for discordant resolution testing for HPIV-1, and HPIV-3. No discordant resolution testing was performed for HPIV-4 since reference assay was based on composite result of two different reverse transcriptase PCR assays followed by by-directional sequencing as stated above. Performance characteristics were estimated relative to valid culture/DFA or PCR/sequencing results for each sample. Sensitivity and specificity (for HPIV-1, and HPIV-3) and positive and negative percent agreement (for HPIV-4) were estimated with corresponding 2-sided 95% Score CIs. Analyses were performed separately for each target analyte (HPIV-1, HPIV-2, HPIV-3, and HPIV-4).
Of the 2961 specimens, 31 specimens/samples were withdrawn, 2930 samples were processed in valid Panther Fusion Paraflu runs, 2877 (98.2%) had final valid results, and 53 (1.8%) had final invalid results. Of the 2877 samples with valid Panther Fusion results, 1359 samples were from females and 1518 samples were from males (see Table 10). The positivity of each analyte by age group is presented in Table 11.
17
| N (%) | |
|---|---|
| Total | 2877 (100) |
| Sex | |
| Female | 1359 (47.2) |
| Male | 1518 (52.8) |
| Age Group | |
| 0 to 28 days | 82 (2.9) |
| 29 days to 65 years | 420 (14.6) |
Table 10: Summary of Subject Demographics for Prospective Samples in the Panther Fusion Paraflu Assay Evaluation
| Table 11: Panther Fusion Paraflu Positivity by Analyte and Age Group | ||||||||
|---|---|---|---|---|---|---|---|---|
| -- | -- | -- | -- | -- | -- | -- | -- | ---------------------------------------------------------------------- |
| % Positivity (n/N) | ||||
|---|---|---|---|---|
| Analyte | HPIV-1 | HPIV-2 | HPIV-3 | HPIV-4 |
| All | 1.5% | 1.3% | 2.8% | 1.2% |
| 0 to 28 days | 0.0% (0/82) | 0.0% (0/82) | 1.2% (1/82) | 0.0% (0/82) |
| 29 days to 99.9) | ||||
| FN=false negative. FP=false positive. NPA=negative percent agreement. PPA=positive percent agreement. TP=true positive. TN=true negative |
1 Study prevalence reported, "Score Confidence Interval, "PPA and NPA apply to HPIV-4
8/10 false positive results were confirmed positive and 1/1 false negative result was confirmed negative for HPIV-1 by PCR 14/15 false positive results were confirmed positive and 2/2 false negative results were confirmed negative for HPV-2 by PCR
26/28 false positive results were confirmed positive and 2/2 false negative results were confirmed negative for HPV-4 by PCR No discordant resolution testing were performed for the 5 false positive and 1 false negative results for HPV-4
Reproducibility
Panther Fusion Paraflu assay reproducibility was evaluated at three US sites using nine panel members. Testing was performed using one lot of assay reagents and six operators (two at each site). At each site, testing was performed for at least five days. Each run had three replicates of each panel member.
A negative panel member was created using a matrix of simulated nasal swab specimen in viral transport medium (VTM). Positive panel members were created by spiking 1-2X LoD (low
19
positive) or 2-3X LoD (moderate-positive) concentrations of the target analyte into a matrix of simulated nasal swab specimen, composed of cultured human cells suspended in VTM.
The agreement with expected results was 100% in the negative and moderate positive panel members and ≥96.6% in low-positive panel members for HPIV-1, HPIV-2, HPIV-3, and HPIV-4 as shown in Table 13.
| | Expected Results
HPIV- | | | | Agreement with Expected Results
HPIV-1 | | HPIV-2 | | HPIV-3 | | HPIV-4 | | | |
|-------------------|---------------------------|----------------------|---|---|-------------------------------------------|---|--------|-------------------|--------|-------------------|--------|---------------------|-------|---------------------|
| Description | Comp. | Conc.
(TCID50/mL) | 1 | 2 | 3 | 4 | N | %
(95% CI) | N | %
(95% CI) | N | %
(95% CI) | N | %
(95% CI) |
| HPIV-1
Low Pos | 1-2X LoD | 1.00E-02 | + | - | - | - | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) |
| HPIV-1
Mod Pos | 2-3X LoD | 3.00E-02 | + | - | - | - | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) |
| HPIV-2
Low Pos | 1-2X LoD | 1.00E+02 | - | + | - | - | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) |
| HPIV-2
Mod Pos | 2-3X LoD | 3.00E+02 | - | + | - | - | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) |
| HPIV-3
Low Pos | 1-2X LoD | 1.00E+01 | - | - | + | - | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 86/87 | 98.9
(93.8-99.8) | 87/87 | 100
(95.8-100) |
| HPIV-3
Mod Pos | 2-3X LoD | 3.00E+01 | - | - | + | - | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) | 89/89 | 100
(95.9-100) |
| HPIV-4
Low Pos | 1-2X LoD | 3.16E+00 | - | - | - | + | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 84/87 | 96.6
(90.3-98.8) |
| HPIV-4
Mod Pos | 2-3X LoD | 9.49E+00 | - | - | - | + | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) | 88/88 | 100
(95.8-100) |
| Neg | N/A | N/A | - | - | - | - | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) | 87/87 | 100
(95.8-100) |
Table 13: Agreement of Panther Fusion Paraflu Assay Results With Expected Results
Comp.=composition, Conc.=concentration, Cl=Score confidence interval, Mod=moderate, Neg=negative, Pos=positive, TCID50/mL=50% tissue culture infective dose (measure of virus titer)
The total HPIV-1, HPIV-2, HPIV-3, and HPIV-4 signal variability measured as %CV ranged from 1.11% to 5.88% in low and moderate positive panel members. For the sources of variation except the 'within-run' factor, %CV values were ≤1.40% as shown in Table 14.
20
| | | | Between
Sites | | Between
Operators | | Between
Days | | Between
Runs | | Within
Runs | | Total | |
|-------------------|----|------------|------------------|--------|----------------------|--------|-----------------|--------|-----------------|--------|----------------|--------|-------|--------|
| Panel Description | N | Mean
Ct | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) | SD | CV (%) |
| HPIV-1 Low Pos | 88 | 37.2 | 0.0 | 0.0 |