(746 days)
Not Found
No
The device is a chemical solution and collection tube for specimen stabilization and inactivation, with no mention of computational analysis or algorithms.
No
The device is described as a solution and collection tube intended for the stabilization, transportation, and inactivation of upper respiratory specimens containing SARS-CoV-2. Its purpose is for diagnostic sample preparation, not for treating or preventing disease.
No
Explanation: The device is intended for the stabilization, transportation, and inactivation of specimens, making them suitable for use with legally marketed molecular diagnostic devices. It does not perform the diagnostic analysis itself.
No
The device description clearly states it is a "virus-inactivating and lysis solution" provided in a "polypropylene flat bottom collection tube with a high-density polyethylene (HDPE) cap". This describes a physical product (solution and tube), not a software-only device.
Based on the provided information, yes, this device is an IVD (In Vitro Diagnostic).
Here's why:
- Intended Use: The intended use explicitly states that InActiv Blue is for the "stabilization, transportation and inactivation of upper respiratory specimens suspected of containing SARS-CoV-2." It also states that specimens collected and stored in the tube are "suitable for use with legally marketed molecular diagnostic devices." This clearly indicates its role in the pre-analytical phase of an in vitro diagnostic test for SARS-CoV-2.
- Device Description: The description details a solution designed to preserve the integrity of RNA from collected specimens for subsequent diagnostic testing.
- Performance Studies: The performance studies described (Limit of Detection, Sample Stability, Viral Inactivation) are all relevant to evaluating the performance of a device used in the context of an in vitro diagnostic workflow for detecting SARS-CoV-2.
- Predicate Device: The mention of a predicate device (K202641 – DNA/RNA Shield™ Collection Tube) which is also a specimen collection and transport device for molecular diagnostics, further supports its classification as an IVD.
While the device itself doesn't perform the diagnostic test, it is an essential component in the process of collecting, transporting, and preparing a specimen for an in vitro diagnostic test. This function falls squarely within the definition of an IVD device.
N/A
Intended Use / Indications for Use
InActiv Blue is intended for the stabilization and inactivation of upper respiratory specimens suspected of containing SARS-CoV-2. These devices can be used for collection, transport and storage of specimens at refrigerated (2-8°C) or ambient temperatures (20-25°C). SARS-CoV-2 RNA in the specimens is stabilized for 7 days at these specified temperatures. Specimens collected and stored in an InActiv Blue collection tube are suitable for use with legally marketed molecular diagnostic devices.
Product codes
QBD
Device Description
InActiv Blue is a virus-inactivating and lysis solution that abrogates the infectious potential of the collected material without affecting the integrity of the RNA. Two milliliters of InActiv Blue solution are provided in a polypropylene flat bottom collection tube with a high-density polyethylene (HDPE) cap (product code IB TUB). The device is non-sterile, single use. Secondary Packaging for the collection tube configuration includes racks of 50 collection tubes and cardboard boxes with either 400 or 1200 collection tubes. Sample types intended for collection with the device include upper respiratory swab samples. Swabs are not provided with the InActiv Blue collection tubes. The active components of the InActiv Blue solution are guanidine thiocyanate, Nlauroylsarcosine sodium salt and EDTA. These components are formulated in a sodium phosphate buffered solution containing methylene blue.
Mentions image processing
Not Found
Mentions AI, DNN, or ML
Not Found
Input Imaging Modality
Not Found
Anatomical Site
Upper respiratory specimens
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)
Shelf-Life Stability:
- Study Type: Real-time stability study.
- Sample Size: At least eight lots of InActiv Blue.
- Key Results: The product met all stability specifications for the duration of the study, supporting 21 months of shelf life. Parameters assessed were appearance by visual inspection, pH (6+/- 0.6), osmolarity (mOsm/kg 4200+/-420), and evaporation.
Limit of Detection (LOD):
- Study Type: Preliminary and Confirmatory LOD studies.
- Key Results: The LOD for nasopharyngeal swab specimen in InActiv Blue was determined to be 45 particles/mL of SARS-CoV-2 using the Roche cobas SARS-CoV-2 assay on the cobas 6800 system.
- Preliminary LOD: Found to be within the range of 5-45 particles/mL. At 15 particles/mL and 45 particles/mL, 100% positivity was observed in triplicate.
- Confirmatory LOD: At 45 particles/mL, 100% of 21 replicates were positive. At 15 particles/mL, 90% of 21 replicates (19 positive, 2 negative) were positive.
Sample Stability Study:
- Study Type: Stability of SARS-CoV-2 in nasopharyngeal specimens stored in InActiv Blue.
- Sample Size: Six replicate samples per lot, spiked with 135 particles/mL (= 3x LOD).
- Key Results: All samples collected in the four lots (newly manufactured, middle aged and near/recently expired lots) and stored at both temperatures (2-8°C and 20-25°C) remained within 3 Ct of the initial reading up to 7 days, supporting the sample stability claim.
Viral Inactivation:
- Study Type: Evaluation of residual infective SARS-CoV-2 viral particles.
- Sample Size: Three lots of InActiv Blue (approximately 21 months old).
- Key Results: The log infectivity reduction factor was calculated to be at least 3.89 logs for the virus exposed to InActiv Blue medium for exposure times of 1, 2.5, and 5 minutes. Inactivation performance after 1 minute exposure was comparable to 2.5 and 5 minutes.
Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)
LOD (Limit of Detection): 45 particles/mL of SARS-CoV-2 (for nasopharyngeal swab specimen in InActiv Blue).
Viral Inactivation: The log infectivity reduction factor was calculated to be at least 3.89 logs.
Predicate Device(s)
Reference Device(s)
Not Found
Predetermined Change Control Plan (PCCP) - All Relevant Information
Not Found
§ 866.2950 Microbial nucleic acid storage and stabilization device.
(a)
Identification. A microbial nucleic acid storage and stabilization device is a device that consists of a container and reagents intended to stabilize microbial nucleic acids in human specimens for subsequent isolation and purification of nucleic acids for further molecular testing. The device is not intended for preserving morphology or viability of microorganisms.(b)
Classification. Class II (special controls). The special controls for this device are:(1) The intended use for the labeling required under § 809.10 of this chapter must include a detailed description of microorganisms and types of human specimens intended to be preserved.
(2) The labeling required under § 809.10(b) of this chapter must include the following:
(i) A detailed device description, including all device components;
(ii) Performance characteristics from applicable analytical studies, including nucleic acid stability and microorganism inactivation;
(iii) A limiting statement that erroneous results may occur when the transport device is not compatible with molecular testing; and
(iv) A limiting statement that the device has only been validated to preserve the representative microorganisms used in the analytical studies.
(3) Design verification and validation must include the following:
(i) Overall device design, including all device components and all control elements incorporated into the analytical validation procedures;
(ii) Thorough description of the microorganisms and methodology used in the validation of the device including, extraction platforms and assays used for the detection of preserved nucleic acids; and
(iii) The limit of detection (LoD) of the molecular test used to establish microorganism nucleic acid stability.
0
June 11, 2024
Image /page/0/Picture/1 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). On the left, there is the Department of Health & Human Services logo. To the right of that is the FDA logo, which is a blue square with the letters "FDA" in white. To the right of the blue square is the text "U.S. FOOD & DRUG ADMINISTRATION" in blue.
FertiPro NV % Gregor Kleinert Official Correspondent Ducks in a Row Compliance LLC 1447 Bentley Drive Warrington, Pennsylvania 18976
Re: K221547
Trade/Device Name: InActiv Blue Regulation Number: 21 CFR 866.2950 Regulation Name: Microbial nucleic acid storage and stabilization device Regulatory Class: Class II Product Code: QBD Dated: May 26, 2022 Received: May 27, 2022
Dear Gregor Kleinert:
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 (the Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database available at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.
If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.
Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device" (https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).
1
Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30, Design controls; 21 CFR 820.90, Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review. the OS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).
Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safetyreporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.
Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.
For comprehensive regulatory information about medical devices and radiation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).
Sincerely, Noel J. Gerald -S
Noel J. Gerald, Ph.D. Branch Chief Bacterial Respiratory and Medical Countermeasures Branch Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Quality Center for Devices and Radiological Health
2
Indications for Use
510(k) Number (if known) K221547
Device Name InActiv Blue
Indications for Use (Describe)
InActiv Blue is intended for the stabilization and inactivation of upper respiratory specimens suspected of containing SARS-CoV-2. These devices can be used for collection, transport and storage of specimens at refrigerated (2-8°C) or ambient temperatures (20-25°C). SARS-CoV-2 RNA in the specimens is stabilized for 7 days at these specified temperatures. Specimens collected and stored in an InActiv Blue collection tube are suitable for use with legally marketed molecular diagnostic devices.
| Type of Use (Select one or both, as applicable) | |
|---|---|
| Prescription Use (Part 21 CFR 801 Subpart D) | Over-The-Counter Use (21 CFR 801 Subpart C) |
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3
510(k) Summary - K221547
| Applicant: | FertiPro NV
Industriepark Noord 32
8730 Beernem
Belgium |
|--------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Establishment Registration No.: | 3003272500 |
| Primary correspondent: | Gregory P. Kleinert
Ducks in a Row Compliance LLC.
1447 Bentley Drive
Warrington, PA 18976
Phone: 215-858-2429
Email: DucksComplianceLLC@gmail.com |
| Summary Date: | 14 March 2024 |
| Device Proprietary Name: | InActiv Blue |
| Classification Name: | Microbial Nucleic Acid Storage
and Stabilization Device |
| Regulation:
Class:
Panel:
Product code: | 866.2950
Class II
Microbiology
QBD |
| Predicate Device: | K202641 – DNA/RNA Shield™ Collection
Tube (Manufacturer: Zymo Research) |
Intended Use/Indication for Use:
InActiv Blue is intended for the stabilization, transportation and inactivation of upper respiratory specimens suspected of containing SARS-CoV-2. These devices can be used for collection transport and storage of specimens at refrigerated (2-8°C) or ambient temperatures (20-25°C). SARS-CoV-2 RNA in the specimens is stabilized for 7 days at these specified temperatures. Specimens collected and stored in an InActiv Blue collection tube are suitable for use with legally marketed molecular diagnostic devices.
Device Description:
InActiv Blue is a virus-inactivating and lysis solution that abrogates the infectious potential of the collected material without affecting the integrity of the RNA. Two milliliters of InActiv Blue solution are provided in a polypropylene flat bottom collection tube with a high-density polyethylene (HDPE) cap (product code IB TUB). The device is non-sterile, single use. Secondary Packaging for the collection tube configuration includes racks of 50 collection tubes and cardboard boxes with either 400 or 1200 collection tubes.
4
Sample types intended for collection with the device include upper respiratory swab samples. Swabs are not provided with the InActiv Blue collection tubes.
The active components of the InActiv Blue solution are guanidine thiocyanate, Nlauroylsarcosine sodium salt and EDTA. These components are formulated in a sodium phosphate buffered solution containing methylene blue.
Principles of Operation:
InActiv Blue main active components are guanidine thiocyanate, N-lauroylsarcosine and EDTA.
Guanidine thiocyanate is a chaotropic salt acting as a denaturing agent for macromolecules leading to virus inactivation. Additionally, guanidine acts as a ribonuclease (RNase) inhibitor, therefore preventing RNA degradation in samples and so keeping the virus RNA detectable for a longer period of time.
N-lauroylsarcosine (LS) is an anionic surfactant capable of reducing viral infection by interfering with the initial binding of the virus to the cells. Inactivation of SARS-CoV-2, is accomplished by disrupting the viral envelope, making it unable for the virus to enter cells. Additionally, LS is able to inhibit RNase and so protecting RNA from degradation, keeping the virus RNA detectable for a longer period of time.
EDTA is a chelating agent and nuclease inhibitor. Due to the high affinity of EDTA to divalent cations, such as Ca2+, Mn2+ and Mg2+, which act as cofactors for nucleases it could inhibit the degradation of RNA by RNases.
Comparison to Predicate Device
InActiv Blue has similar intended use, design and technological characteristics as the predicate device, DNA/RNA Shield Collection Tube (K202641). Below is a comparison table showing similarities and differences between InActiv Blue and predicate device K202641:
| Device & Predicate Device(s): | Device: K221547 | Predicate: K202641 |
|---|---|---|
| Device Trade Name | InActiv Blue | DNA/RNA Shield Collection |
| Tube | ||
| General Device | ||
| Characteristic Similarities |
5
| Intended Use/Indications For
Use | InActiv Blue is intended for the
stabilization, transportation and
inactivation of upper respiratory
specimens suspected of
containing SARS-CoV-2. These
devices can be used for
collection transport and storage
of specimens at refrigerated (2-
8°C) or ambient temperatures
(20-25°C). SARS-CoV-2 RNA
in the specimens is stabilized for
7 days at these specified
temperatures. Specimens
collected and stored in an
InActiv Blue collection tube are
suitable for use with legally
marketed molecular diagnostic
devices. | The DNA/RNA Shield
collection tube is intended for
the stabilization and inactivation
of upper and lower respiratory
human specimens suspected of
containing SARS-CoV-2. These
devices can be used for
collection transport and storage
of specimens at ambient
temperatures (20-25°C).
Specimens collected and stored
in a DNA/RNA Shield collection
tube are suitable for use with
legally marketed molecular
diagnostic devices. |
|----------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Inactivation | RNA virus particles | Same |
| Analyte | RNA | Same |
| General Device
Characteristic Differences | | |
| Sample Source | Human upper respiratory | Human lower and upper
respiratory |
| Sample storage Temperature | 7 days at refrigerated (2-8°C) or
ambient temperatures (20-25°C) | 28 days at 20-25°C |
Performance Characteristics:
Shelf-Life Stability:
Shelf-life stability of InActiv Blue was evaluated to ensure product integrity over the complete shelf-life of the product. Completed real-time stability studies included at least eight lots of InActiv Blue which have been stored at room temperature for different periods of time, with performance testing conducted at regularly spaced timepoints to support a 21-month shelf-life stability claim.
At each time points, appearance by visual inspection pH (6+/- 0.6), osmolarity (mOsm/kg 4200+/-420) and evaporation were assessed. The product met all stability specifications for the duration of the study, supporting 21 months of shelf life. The results are summarized below (Table 1).
Table 1: Shelf-life Stability of InActiv Blue when stored at 15-25°C
6
| InActiv
Blue lots
stored at
15-25°C | Lot
number | Sample age | pH ^ | Osmolality
(mOsm/kg)
^^ | Appearance * | Evaporation** |
|----------------------------------------------|--------------------|----------------|----------------|-------------------------------|--------------|---------------|
| Recently
produced | FP23IB06 | 38 days | 6.02 | 435 | PASS | PASS |
| Intermediate aged | FP23IB03 | 12 months | 6.03 | 438 | PASS | PASS |
| | FP23IB02 | 12 months | 6.06 | 449 | PASS | PASS |
| | FP23IB01 | 12 months | 6.07 | 448 | PASS | PASS |
| | Expired
batches | FP21IB06 | > 21
months | 5.97 | 460 | PASS |
| FP21IB04 | | > 21
months | 5.90 | 459 | PASS | PASS |
| FP21IB03 | | > 21
months | 5.95 | 461 | PASS | PASS |
| FP20IB06 | | > 21
months | 6.02 | 452 | PASS | PASS |
^ Mean of three determinations. At production, pH values range from 5.98 to 6.03
^^ Mean of three determinations. At production osmolarity values range from 396 to 441 mOsm/kg. Determinations were made on product diluted 1:10
- Criteria: labeling does not show decreased quality and adheres well to the tube, and liquid is clear ** Criterium: >1.5 mL of InActiv Blue
Limit of Detection:
The limit of detection (LOD) of SARS-CoV-2 in InActiv Blue was established through preliminary and confirmatory LOD studies. The LOD was determined for InActiv Blue with nasopharyngeal swab samples, using the Roche cobas SARS-CoV-2 assay (EUA200009) on the cobas 6800 system.
The preliminary LOD was determined by spiking InActiv Blue with nasopharyngeal clinical matrix and SARS-CoV-2 at concentrations of 5, 15, 45, 135, and 405 particles/mL, with each determination performed in triplicate. The LOD was found to be within the range of 5-45 particles/mL.
| Table 1. Preliminary LOD Study Results | ||||
|---|---|---|---|---|
| -- | ---------------------------------------- | -- | -- | -- |
| SARS-CoV-2
(particles/mL) | Number of
Positive
Replicates | Number of
Negative
Replicates | % Positive |
|------------------------------|-------------------------------------|-------------------------------------|------------|
| 5 | 1 | 2 | 33% |
| 15 | 3 | 0 | 100% |
| 45 | 3 | 0 | 100% |
7
| 135 | 3 | 0 | 100% |
|---|---|---|---|
| 405 | 3 | 0 | 100% |
The confirmatory LOD was established by spiking a swab with negative clinical nasopharyngeal matrix containing either 15 or 45 SARS-CoV-2 particles/mL, into InActiv Blue media. A total of 20 replicates were evaluated for each concentration. The nasopharyngeal swabs in InActiv Blue, containing 45 particles/mL, showed that 100% of the replicates were positive, and at a concentration of 15 particles/mL 90% of the replicates were positive.
| Table 2. Confirmatory LOD Study Results | |
|---|---|
| ----------------------------------------- | -- |
| SARS-CoV-2
(particles/mL) | Number
of
Positive
Samples | Number
of
Negative
Samples | % Positive |
|------------------------------|-------------------------------------|-------------------------------------|------------|
| 15 | 19 | 2 | 90% |
| 45 | 21 | 0 | 100% |
The LOD for nasopharyngeal swab specimen in InActiv Blue was determined to be 45 particles/mL of SARS-CoV-2.
Sample Stability Study:
Stability of SARS-CoV-2 in nasopharyngeal specimens stored InActiv Blue was evaluated using product lots of different ages to ensure that there is no decrease in the performance of the product over its claimed shelf-life. Lots of InActiv Blue from different ages (newly manufactured, middle aged and near/recently expired lots), spanning the claimed shelf-life were evaluated. Six replicate samples containing nasopharyngeal matrix were collected and stored in each of the lots of InActiv Blue. Each replicate was spiked with 135 particles/mL (= 3x LOD). Three of the replicates from each lot were stored at 2-8°C while the other 3 replicates were stored at 20-25°C for at least 7 days. Each sample was evaluated using the cobas SARS-CoV-2 assay (EUA200009) on the 6800 system at day 0, day 4 and day 7. All samples collected in the four lots and stored at both temperatures remained within 3 Ct of the initial reading, supporting the sample stability claim for InActiv Blue of 7 days at a temperature between 2-8 and 20-25°C. The following table summarizes the results obtained.
8
| Storage
Temperature | Lot
manufacturing
age | Lot # | Target 1 (ORF 1Ab)
Delta Ct's | | Target 2 (E gene)
Delta Ct's | |
|------------------------|-----------------------------|----------|----------------------------------|-----------------------|---------------------------------|-----------------------|
| | | | Ct Day4 - Ct
Day 0 | Ct Day7 - Ct
Day 0 | Ct Day4 - Ct
Day 0 | Ct Day7 - Ct
Day 0 |
| 4°C | new | FP23IB01 | -0.12 | 0.00 | 0.74 | 0.80 |
| | middle aged | FP21IB04 | 0.09 | 0.20 | 0.80 | 0.37 |
| | after expiry date | FP21IB01 | -0.36 | 0.05 | 0.13 | 0.12 |
| | stressed | FP21IB01 | -0.30 | 0.84 | 0.42 | 0.97 |
| 20°C | new | FP23IB01 | -0.55 | -0.22 | -0.87 | -0.49 |
| | middle aged | FP21IB04 | -0.23 | 0.47 | 0.33 | 0.45 |
| | after expiry date | FP21IB01 | 0.02 | -0.41 | 1.10 | -0.42 |
| | stressed | FP21IB01 | -0.27 | -0.37 | 0.63 | -0.05 |
Table 3. SARS-CoV-2 RNA stability after 4 and 7 days of incubation at 2-8°C and 20-25°C
Viral Inactivation:
Inactivation was evaluated by measuring the presence of residual infective SARS-CoV-2 viral particles spiked into InActiv Blue. Inactivation studies were conducted in two phases. First a 1:1000 dilution of InActiv Blue was established to eliminate the cytotoxicity of the medium. Higher concentrations of medium were found to render the cultured cells nonviable and unusable for the viral inactivation study. The second phase of viral inactivation evaluated cvtotoxicity of SARS-COV-2 spiked into a 1:1000 dilution of the transport media for 1, 2.5 and 5 minutes after inoculation.
The second phase of the study used a high concentration of infectious viral particles (TCID50mL=10 75 or 10 739). Briefly, 100 uL of virus stock was mixed 1:1 with InActiv Blue and incubated for 1, 2.5 and 5 minutes. Negative and positive controls were prepared similarly. The virus and InActiv Blue (as well as positive and negative control samples) were then diluted 1:1000 before each replicate was added to cell culture plates containing adherent Vero E6 cells to determine the minimal incubation time required to inactivate virus. Cultures were incubated for several minutes and observed for presence or absence of cytopathic effect and TCID50 was calculated by the Reed and Muench method. Virus inactivation was evaluated using 3 lots near the expiration of InActiv Blue (approximately 21 months old). Inactivation performance of InActiv Blue after 1 minute exposure was comparable to 2.5 and 5 minutes of exposure for each of the three batches of InActiv Blue evaluated.
Since the starting viral concentration was 7.5 logs (TCIDsomL=10 7.5) for two of the lots examined and 7.39 logs (TCID50mL=10 7.39) for the third lot, the log infectivity reduction factor was calculated to be at least 3.89 logs for the virus exposed to InActiv Blue medium. Negative
9
control (without virus) demonstrated minimal cellular toxicity and positive control (virus in DMEM) showed no meaningful loss of viral recovery (data not shown). The study results are summarized in Table 4 below.
| Lot | exposure time | log (TCID50 control) - log
(TCID50 test) | virus log reduction factor |
|-----|---------------|---------------------------------------------|----------------------------|
| 1 | 1 min | 7.50-3.50 | 4 |
| 1 | 2.5 min | 7.50-3.50 | 4 |
| 1 | 5 min | 7.50-3.50 | 4 |
| 2 | 1 min | 7.50-3.50 | 4 |
| 2 | 2.5 min | 7.50-3.50 | 4 |
| 2 | 5 min | 7.50-3.50 | 4 |
| 3 | 1 min | 7.39-3.50 | 3.89 |
| 3 | 2.5 min | 7.39-3.50 | 3.89 |
| 3 | 5 min | 7.39-3.50 | 3.89 |
Table 4: SARS-CoV-2 Viral Inactivation when exposed to InActiv Blue