(240 days)
No
The summary describes a chromogenic assay kit for determining factor VIII activity, which is a chemical-based diagnostic method. There is no mention of AI, ML, image processing, or any computational analysis that would suggest the use of these technologies. The performance studies focus on analytical performance characteristics typical of laboratory assays.
No
The device is an in vitro diagnostic intended for quantitative determination of factor VIII activity in human plasma, used to identify factor VIII deficiency and aid in hemophilia A management. It does not provide direct therapy or treatment to a patient.
Yes
Explanation: The "Intended Use / Indications for Use" section explicitly states that the device is for "quantitative determination of factor VIII activity" and is "intended to be used in identifying factor VIII deficiency and as an aid in the management of hemophilia A." These activities fall under the definition of diagnosing or aiding in the diagnosis of a medical condition.
No
The device description clearly states it contains four components packaged in glass vials and provided frozen, which are physical reagents, not software.
Yes, this device is an IVD (In Vitro Diagnostic).
The "Intended Use / Indications for Use" section explicitly states: "For in vitro diagnostic use."
N/A
Intended Use / Indications for Use
CRYOcheck Chromogenic Factor VIII is for clinical laboratory use in the quantitative determination of factor VIII activity in 3.2% citrated human plasma. It is intended to be used in identifying factor VIII deficiency and as an aid in the management of hemophilia A in individuals aged 2 years and older. For in vitro diagnostic use.
Product codes
GGP
Device Description
CRYOcheck Chromogenic Factor VIII is used for determination of FVIII activity and contains the following four components, packaged in glass vials and provided frozen to preserve the integrity of the components: Reagent 1: Bovine FX and a fibrin polymerization inhibitor, with activators and stabilizers. Reagent 2: Human FIIa, human FIXa, calcium chloride and phospholipids. Reagent 3: FXa substrate containing EDTA and a thrombin inhibitor. Diluent Buffer: Tris buffer solution containing 1% BSA and a heparin antagonist.
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
individuals aged 2 years and older.
Intended User / Care Setting
clinical laboratory use
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
Multi-Reagent Lot Precision:
- Study type: Internal precision study
- Sample size: one normal and two abnormal reference controls and five patient plasma samples (Very Low, Low, Mid, Normal, High FVIII levels) with 80 replicates per sample per lot.
- Key results: Pooled precision of 1% FVIII and ≤0.1% SD for the Very Low FVIII plasma sample.
Multi-Reagent Lot Site to Site Reproducibility:
- Study type: Reproducibility study
- Sample size: one normal and two abnormal reference controls and three patient plasma samples (very low, normal, and high FVIII activity), measured in triplicate, twice a day for 5 days at each of three sites.
- Key results: Pooled reproducibility of 1% FVIII; and ≤0.1% SD for the Very Low FVIII plasma sample.
Linearity/Assay Reportable Range:
- Study type: Linearity study
- Sample size: Three lots of CRYOcheck Chromogenic Factor VIII, fifteen sample dilutions (0 to 260% FVIII estimated activity), each level tested in quadruplicate.
- Key results: Linearity Range: 0 to 200% FVIII activity.
Reference Interval:
- Study type: Reference interval study
- Sample size: Citrated plasma samples from one hundred and twenty ostensibly healthy individuals ≥ 18 years.
- Key results: Reference Interval: 43.2-159.3% FVIII activity.
Stability:
- Shelf Life Stability:
- Study type: Shelf life stability study
- Sample size: Three lots of CRYOcheck Chromogenic Factor VIII, five replicates of six plasma samples representing low to normal FVIII activity levels.
- Key results: Support a shelf-life stability claim of at least 12 months when the product is stored at ≤-70°C.
- In-Use Stability:
- Study type: In-use stability study
- Sample size: Three lots of CRYOcheck Chromogenic Factor VIII, five replicates of six plasma samples representing low to normal FVIII activity levels.
- Key results: Support a stability claim of 8 hours on board the instrument and 120 hours (5 days) at 2-8 °C. Also, 1 month refrozen storage at ≤-70 °C if refrozen within 4 hours of the initial thaw, with previously refrozen reagents usable once for up to four hours on board the instrument.
Detection Limit:
- Limit of Blank (LoB):
- Study type: LoB determination
- Sample size: Four blank plasma samples, measured in triplicate over five days using three lots.
- Key results: LoB was determined to be 0.4% FVIII activity.
- Limit of Detection (LoD):
- Study type: LoD determination
- Sample size: Four plasma samples with low FVIII activity, measured in triplicate over five days using three lots.
- Key results: LoD was determined to be 0.5% FVIII activity.
- Limit of Quantitation (LoQ):
- Study type: LoQ determination
- Sample size: Aliquots of four plasma samples with low FVIII activity, tested in three replicates on five different days at an external laboratory.
- Key results: LoQ was determined to be 0.5% FVIII activity.
Interferences:
- Study type: Interference study
- Sample size: Plasma samples spiked with possible interferents and 10 replicates of corresponding blank matrix control.
- Key results: No interference up to indicated concentrations for Hemoglobin (≤ 500 mg/dL), Intralipid (≤ 500 mg/dL), Bilirubin (unconjugated) (≤ 29 mg/dL), vWF (≤ 20 µg/mL), Unfractionated heparin (≤ 2 IU/mL), Low molecular weight heparin (≤ 2 IU/mL), Fondaparinux (≤ 1.25 mg/L), Lupus Anticoagulant (≤ 1.8 dRVVT ratio). Rivaroxaban and dabigatran interfered.
Method Comparison Studies:
- Study type: Method comparison study (CLSI EP09c)
- Sample size: Three hundred and eighteen human plasma samples from normal individuals and patients (congenital/acquired hemophilia A, Type 1/2A/2B/2N von Willebrand disease).
- Key results: Performed equivalently to the comparator method. Overall Pearson Correlation Coefficient: 0.994 (r2=0.987).
Sample Integrity:
- Study type: Sample integrity study
- Sample size: Forty-six plasma samples.
- Key results: Support a fresh sample stability claim of 2 hours at room temperature and a frozen storage claim of 3 months at ≤-70 °C, including up to two freeze thaw cycles.
Key Metrics
- Pooled precision: 1% FVIII; and ≤0.1% SD for Very Low FVIII plasma sample.
- Linearity Range: 0 to 200% FVIII activity.
- Reference Interval: 43.2-159.3% FVIII activity.
- Limit of blank (LoB): 0.4% FVIII activity.
- Limit of detection (LoD): 0.5% FVIII activity.
- Limit of quantitation (LoQ): 0.5% FVIII activity.
- Method Comparison (Overall): Slope 1.038, Intercept 0.473, Pearson Correlation 0.994 (r2=0.987).
- Predicted Bias at FVIII activity (%):
- 1%: -1.11%
- 5%: -0.81%
- 45%: 2.20%
- 50%: 2.57%
- 100%: 6.33%
- 150%: 10.09%
Predicate Device(s)
Reference Device(s)
Predetermined Change Control Plan (PCCP) - All Relevant Information
Not Found
§ 864.7290 Factor deficiency test.
(a)
Identification. A factor deficiency test is a device used to diagnose specific coagulation defects, to monitor certain types of therapy, to detect coagulation inhibitors, and to detect a carrier state (a person carrying both a recessive gene for a coagulation factor deficiency such as hemophilia and the corresponding normal gene).(b)
Classification. Class II (performance standards).
0
Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). On the left 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.
July 17, 2020
Precision BioLogic Karen Black VP of Compliance & Product Development 140 Eileen Stubbs Avenue Dartmouth, Nova Scotia B3B 0A9 Canada
Re: K193204
Trade/Device Name: CRYOcheck Chromogenic Factor VIII Regulation Number: 21 CFR 864.7290 Regulation Name: Factor deficiency test Regulatory Class: Class II Product Code: GGP Dated: November 19, 2019 Received: November 20, 2019
Dear Karen Black:
We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database located at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.
If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.
Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's
1
requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.
Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.
For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).
Sincerely,
for
Takeesha Taylor-Bell Chief Division of Immunology and Hematology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Ouality Center for Devices and Radiological Health
Enclosure
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Indications for Use
510(k) Number (if known) K193204
Device Name CRYOcheck Chromogenic Factor VIII
Indications for Use (Describe)
CRYOcheck Chromogenic Factor VIII is for clinical laboratory use in the quantitative determination of factor VIII activity in 3.2% citrated human plasma. It is intended to be used in identifying factor VIII deficiency and as an aid in the management of hemophilia A in individuals aged 2 years and older. For in vitro diagnostic use.
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
cryocheck™ Chromogenic Factor VIII
This summary of 510(k) safety and effectiveness information is being submitted in accordance with the requirements of SMDA 1990 and 21 CFR 807.92.
The assigned 510(k) number is K193204
| Submitter's
Information | Precision BioLogic Inc.
140 Eileen Stubbs Ave.
Dartmouth, Nova Scotia B3B 0A9
Canada | | |
|--------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|--|
| Contact Person | Karen M. Black, VP of Compliance & Product Development
Phone: 902-468-6422, ext. 226, or 902-706-3125
E-mail: kblack@precisionbiologic.com | | |
| Preparation Date | 9 July 2020 | | |
| Device Trade
Name | CRYOCheck™ Chromogenic Factor VIII | | |
| Regulatory
Information | Regulation Number and
Description | 21 CFR 864.7290
Factor Deficiency Test | |
| | Classification | Class II | |
| | Product Code | GGP; Test, Qualitative and Quantitative
Factor Deficiency; 21 CFR 864.7290 | |
| Predicate Device | Coatest SP FVIII (K042576) | | |
| Indication for Use/
Intended Use | CRYOcheck Chromogenic Factor VIII is for clinical laboratory use in the
quantitative determination of factor VIII activity in 3.2% citrated human
plasma. It is intended to be used in identifying factor VIII deficiency and
as an aid in the management of hemophilia A in individuals aged 2 years
and older. For in vitro diagnostic use. | | |
| Device
Description | CRYOcheck Chromogenic Factor VIII is used for determination of FVIII
activity and contains the following four components, packaged in glass
vials and provided frozen to preserve the integrity of the components:
Reagent 1: Bovine FX and a fibrin polymerization inhibitor, with
activators and stabilizers.
Reagent 2: Human FIIa, human FIXa, calcium chloride and
phospholipids.
Reagent 3: FXa substrate containing EDTA and a thrombin inhibitor.
Diluent Buffer: Tris buffer solution containing 1% BSA and a heparin
antagonist. | | |
| Comparison to Predicate | | | |
| Item | Predicate | New Device | |
| Proprietary and
Established Names | Coatest SP FVIII | CRYOCheck Chromogenic Factor VIII | |
| Manufacturer | Instrumentation Laboratory | Precision BioLogic | |
| Similarities | | | |
| Item | Predicate | New Device | |
| Measurand | Human Factor VIII | Human Factor VIII | |
| Product Code | GGP
Test, Qualitative and
Quantitative Factor Deficiency | GGP
Test, Qualitative and Quantitative
Factor Deficiency | |
| Regulation Section | 21 CFR 864.7290 | 21 CFR 864.7290 | |
| | Factor Deficiency Test
Class II | Factor Deficiency Test
Class II | |
| Classification | 81 (Haematology) | 81 (Haematology) | |
| Panel | Coatest SP FVIII is intended for
the photometric determination of
factor VIII activity in citrated
plasma. | CRYOCheck Chromogenic Factor VIII
is for clinical laboratory use in the
quantitative determination of factor
VIII activity in 3.2% citrated human
plasma. It is intended to be used in
identifying factor VIII deficiency and
as an aid in the management of
hemophilia A in individuals aged 2
years and older. For in vitro
diagnostic use. | |
| Intended Use | Quantitative (chromogenic
measurement of FVIII) | Quantitative (chromogenic
measurement of FVIII) | |
| Assay Type | Coatest SP FVIII is a modified
version of Coatest Factor VIII
(K833892) reformulated to
European Pharmacopoeia
Standards. Coatest SP FVIII is a
photometric assay containing a
chromogenic substrate, S-2765,
with EDTA added as a
preservative, lyophilized bovine
factors IXa and X with bovine
albumin added as a stabilizing
agent. The device also contains
calcium chloride, Tris buffer
stock solution containing sodium
chloride, bovine serum albumin
with added antimicrobial in
addition to a mixture of highly
purified synthetic phospholipids. | CRYOCheck Chromogenic Factor VIII
is used for determination of FVIII
activity and contains the following
four components, packaged in glass
vials and provided frozen to preserve
the integrity of the components:
Reagent 1: Bovine FX and a fibrin
polymerization inhibitor, with
activators and stabilizers.
Reagent 2: Human Flla, human
FIXa, calcium chloride and
phospholipids.
Reagent 3: FXa substrate containing
EDTA and a thrombin inhibitor.
Diluent Buffer: Tris buffer solution
containing 1% BSA and a heparin
antagonist. | |
| Device Description | In the presence of calcium and
phospholipids, factor X is
activated to factor Xa by factor
IXa. This generation is greatly
stimulated by factor VIII, which
may be considered as a cofactor
in this reaction. By using optimal
amounts of Ca2+ and
phospholipids and an excess of
factors IXa and X, the rate of
activation of factor X is solely
dependent on the amount of
factor VIII. Factor Xa hydrolyses
the chromogenic substrate S-
2765 thus liberating the
chromophoric group, pNA. The
color is then read
photometrically at 405 nm. The
generated factor Xa and thus
the intensity of color are | In the first stage of the chromogenic
assay, test plasma (containing an
unknown amount of functional FVIII)
is added to a reaction mixture
comprised of calcium, phospholipids,
purified human thrombin and FIXa,
and purified bovine FX (Reagent 1
and Reagent 2). This mixture swiftly
activates FVIII to FVIIIa, which works
in concert with FIXa to activate FX.
When the reaction is stopped, FXa
production is assumed to be
proportional to the amount of
functional FVIII present in the
sample. The second stage of the
assay is to measure FXa through
cleavage of an FXa-specific peptide
nitroanilide substrate (FXa
Substrate). P-nitroaniline is
produced, giving a color that can be | |
| Methodology | | | |
| | proportional to the factor VIII
activity in the sample. Hydrolysis
of S-2765 by thrombin formed is
prevented by the addition of the
synthetic thrombin inhibitor, I-
2581, together with the
substrate. | measured spectrophotometrically by
absorbance at 405 nm. | |
| Expression of
results | Quantitative; results are
expressed as percent activity
interpreted relative to a
calibration curve. | Quantitative; results are expressed
as percent activity interpreted relative
to a calibration curve. | |
| | Differences | | |
| Instrument(s) | Manual method, IL ACL 9000 | IL ACL TOP CTS Series and IL ACL
TOP 50 CTS Series | |
| Storage | 2-8 °C until expiration | ≤-70°C until expiration | |
| Linearity Range | 0-150% FVIII activity | 0-200% FVIII activity | |
| Reference Range | 48.6- 126% (manual method)
55.4-148.9% (instrument
application) | 43.2-159.3% FVIII activity | |
| Limit of Detection | 1% FVIII activity | 0.5% FVIII activity | |
| In Use Stability | Working Factor Reagent
Stability (phospholipids + factor
IXa + factor X reagent): 12
hours on ice.
S-2765 + I-2581: Reconstituted
substrate is stable 3 months at
2-8°C
CaCl2: Opened vial is stable 3
months at 2-8ºC
Buffer, stock solution: Opened
vial is stable 3 months at 2-8ºC
Phospholipid: Opened vial is
stable for 3 months at 2-8ºC
Factor Reagent (IXa + X):
Aliquoted for -20ºC for 3
months. | 8 hours on-board instrument
5 days at 2-8ºC
1 month refrozen storage at ≤-70 °C
if refrozen within 4 hours of the initial
thaw. Previously refrozen reagents
can be thawed and used once for up
to four hours on board the
instrument. | |
| Interferences | Manual method:
Triglycerides up to 700 mg/dL
Bilirubin up to 20 mg/dL
Hemoglobin up to 100 mg/dL
Unfractionated Heparin up to 1.0
IU/mL
Automated Method:
Triglycerides up to 900 mg/dL
Bilirubin up to 20 mg/dL
Hemoglobin up to 50 mg/dL | Hemoglobin: ≤ 500 mg/dL
Intralipid: ≤ 500 mg/dL
Bilirubin (unconjugated): ≤ 29 mg/dL
vWF: ≤ 20 µg/mL
Unfractionated heparin: ≤ 2 IU/mL
Low molecular weight heparin: ≤
2 IU/mL
Fondaparinux: ≤ 1.25 mg/L
Lupus Anticoagulant: ≤ 1.8 dRVVT
ratio
Rivaroxaban and dabigatran
interfered with the quantification of
FVIII activity. | |
| | Unfractionated Heparin up to 1.0
IU/mL | The potential interference effects of
conjugated bilirubin on this device
have not been evaluated. | |
| | Due to the high dilutions used
there is no underestimation of
FVIII activity in samples
containing lupus anticoagulant | The performance of this device has
not been established in individuals
with von Willebrand disease Type
2M. | |
| | | The performance of this device has
not been established in evaluating
the potency of FVIII concentrates. | |
| Sample Stability | None specified.
Package insert indicates "Refer
to NCCLS document H21-A4 for
further instructions on specimen
collection, handling and
storage." | 2 hours at room temperature and 3
months at ≤-70 °C, including up to
two freeze thaw cycles | |
4
5
6
Performance Summary:
All studies were performed using CRYOcheck Chromogenic Factor VIII on Instrumentation Laboratories' ACL TOP Series or TOP 50 Series Instruments; the specific instrument(s) used for each study are indicated in the summary reports below.
Multi-Reagent Lot Precision
An internal precision study was performed using three (3) lots of CRYOcheck Chromogenic Factor VIII by one operator on an IL ACL TOP CTS analyzer (K160276) in accordance with CLSI EP05-A3. The study quantified one normal and two abnormal reference controls and five patient plasma samples representing very low, mid, normal and high levels of FVIII activity. Each sample was measured with each product lot in duplicate, twice a day for 20 days for a total of 80 replicates per sample per lot. The results demonstrated a pooled precision of 1% FVIII; and ≤0.1% SD for the Very Low FVIII plasma sample.
| Pooled 3-Site Data | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Sample | Mean (%) | Within-Run | Between- Run | Between-Day | Between-Site | Across-Site | |||||
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | ||
| CRYOcheck Reference Control Normal | 80.3 | 3.6 | 4.5 | 0.2 | 0.2 | 0.5 | 0.7 | 0.0 | 0.0 | 4.1 | 5.1 |
| CRYOcheck Abnormal 1 | |||||||||||
| Reference Control | 25.4 | 1.3 | 5.2 | 0.2 | 0.6 | 0.3 | 1.2 | 1.5 | 5.7 | 2.1 | 8.2 |
| CRYOcheck Abnormal 2 | |||||||||||
| Reference Control | 7.7 | 0.5 | 7.0 | 0.0 | 0.4 | 0.1 | 1.6 | 0.4 | 4.9 | 0.7 | 9.6 |
| Very Low FVIII Plasma Sample | 1.1 | 0.1 | NA | 0.0 | NA | 0.0 | NA | 0.0 | NA | 0.2 | NA |
| Normal FVIII Plasma Sample | 85.4 | 4.3 | 5.1 | 1.1 | 1.3 | 0.0 | 0.0 | 0.0 | 0.0 | 5.0 | 5.8 |
| High FVIII Plasma Sample | 156.9 | 7.8 | 5.0 | 0.9 | 0.5 | 0.0 | 0.0 | 6.1 | 3.9 | 11.4 | 7.3 |
Linearity/Assav Reportable Range
A linearity study was conducted in accordance with CLSI EP6-A using three lots of CRYOcheck Chromogenic Factor VIII on an IL ACL TOP CTS instrument (K160276). A high FVIII (260%) plasma was combined with congenital FVIII deficient plasma (0%) to create fifteen sample dilutions with estimated FVIII activities in the range of 0 to 260% FVIII. Each level was tested in quadruplicate. The results support the linearity claim described below.
Linearity Range: 0 to 200% FVIII activity
Reference Interval
A reference interval study was conducted by multiple operators in accordance with CLSI EP28-A3c using three lots of cRYocheck Chromogenic Factor VIII on an IL ACL TOP CTS instrument (K160276). Citrated plasma samples were collected from one hundred and twenty ostensibly healthy individuals ≥ 18 years. The reference interval was established by calculating the nonparametric 95% confidence interval (2.5th to 97.5th percentiles).
Reference Interval: 43.2-159.3% FVIII activity
Stability
Shelf Life Stability
A shelf life stability study was conducted in accordance with CLSI EP25-A using an IL ACL TOP CTS instrument (K160276). Three lots of CRYOcheck Chromogenic Factor VIII were stored at and at ≤-70°C (monitored condition -76 to -82°C) and tested at t=0 and regular intervals defined by the lot-specific pull schedule up to 37 months. At each timepoint, five replicates of six plasma samples representing low to normal FVIII activity levels were quantified. The study has been
8
completed up to 13 months and supports a shelf-life stability claim of at least 12 months when the product is stored at ≤-70°C.
In-Use Stability
An in-use stability study was conducted in accordance with CLSI EP25-A using an IL ACL TOP CTS instrument (K160276). Three lots of CRYOcheck Chromogenic Factor VIII were maintained on board the analyzer (12–18 °C) for up to 9 hours and in a refrigerator (2–8 °C) for up to 121 hours. Each lot was used to quantify five replicates of six plasma samples representing low to normal FVIII activity levels at each storage condition at defined timepoints. The data support a stability claim of 8 hours on board the instrument and 120 hours (5 days) at 2-8 °C.
Three lots of CRYOcheck Chromoqenic Factor VIII were maintained on board an analyzer for 4 hours, then subsequently refrozen at ≤-70 ℃ for up to 2 months. Each lot was used to quantify five replicates of six plasma samples representing low to normal FVIII activity levels at defined timepoints. The data support a stability claim of 1 month refrozen storage at ≤-70 °C if refrozen within 4 hours of the initial thaw. Previously refrozen reagents can be thawed and used once for up to four hours on board the instrument.
Detection Limit
The limit of blank (LoB) was determined following the CLSI EP17-A2 quideline by measuring four blank plasma samples obtained from individuals with severe congenital hemophilia A. Samples were measured in triplicate on an IL ACL TOP CTS instrument (K160276) using three lots of CRYOcheck Chromogenic Factor VIII over five days. The LoB was determined to be 0.4% FVIII activity.
The limit of detection (LoD) was determined following the CLSI EP17-A2 guideline by measuring four plasma samples with low FVIII activity obtained from congenital hemophilia A donors. Samples were measured in triplicate on an IL ACL TOP CTS instrument (K160276) using three lots of CRYOcheck Chromogenic Factor VIII over five days. The LoD was determined to be 0.5% FVIII activity.
The limit of quantitation (LoQ) was determined according to the CLSI EP17-A2 quideline. Aliquots of four plasma samples with low FVIII activity obtained from congenital hemophilia A donors were sent to an external laboratory for testing in three replicates on five different days on an IL ACL TOP 700 instrument (K160276) to determine assigned values using Coatest SP FVIII. The LoQ was determined to be 0.5% FVIII activity.
Interferences
Interference studies were conducted according to CLSI EP7-A3 using a single lot of CRYOcheck Chromogenic Factor VIII on an IL ACL TOP CTS instrument (K160276). Plasma samples were spiked with possible interferents and 10 replicates were tested alongside 10 replicates of the corresponding blank matrix control. The following substances showed no interference up to the concentrations indicated:
| Possible Interferent | Concentration |
|---|---|
| Hemoglobin | ≤ 500 mg/dL |
| Intralipid | ≤ 500 mg/dL |
| Bilirubin (unconjugated) | ≤ 29 mg/dL |
| vWF | ≤ 20 µg/mL |
| Unfractionated heparin | ≤ 2 IU/mL |
| Low molecular weight heparin | ≤ 2 IU/mL |
| Fondaparinux | ≤ 1.25 mg/L |
| Lupus Anticoagulant | ≤ 1.8 dRVVT ratio |
Rivaroxaban and dabigatran interfered with the quantification of FVIII activity.
9
The potential interference effects of conjugated bilirubin on this device have not been evaluated.
The performance of this device has not been established in individuals with yon Willebrand disease Type 2M.
The performance of this device has not been established in evaluating the potency of FVIII concentrates.
Method Comparison Studies
A method comparison study was conducted at three sites (one internal and two external) according to CLSI EP09c to compare the accuracy of CRYOcheck Chromogenic Factor VIII relative to a comparator device. Three hundred and eighteen human plasma samples from normal ostensibly healthy individuals and from patients with congenital or acquired hemophilia A and Type 1, Type 2A, Type 2B and Type 2N von Willebrand disease were distributed across three sites and tested for FVIII activity using a single lot of CRYOcheck Chromogenic Factor VIII on an IL ACL TOP CTS (K160276) and two different IL ACL TOP 750 (K150877) analyzers. A second aliquot of each sample was tested at a central reference laboratory using Coatest SP FVIII on an IL ACL TOP 700 instrument (K160276).
Results were compared by Passing-Bablok regression statistics show that CRYOcheck Chromogenic Factor VIII performed equivalently to the comparator method.
| | N | Slope | | Intercept | | Pearson Correlation
Coefficient |
|---------|-----|-------|--------------|-----------|---------------|------------------------------------|
| | | Value | 95% CI | Value | 95% CI | |
| Site 1 | 133 | 1.041 | 1.027, 1.058 | 0.720 | 0.252, 1.205 | 0.997 (r2=0.993) |
| Site 2 | 53 | 1.138 | 1.109, 1.168 | 0.252 | 0.001, 0.409 | 0.998 (r2=0.996) |
| Site 3 | 132 | 1.012 | 0.989, 1.045 | -0.140 | -1.768, 0.404 | 0.991 (r2=0.982) |
| Overall | 318 | 1.038 | 1.022, 1.051 | 0.473 | 0.265, 0.594 | 0.994 (r2=0.987) |
Absolute predicted biases are reported below.
| FVIII activity (%) | Predicted Bias (%) | Lower CI (%) | Upper CI (%) |
|---|---|---|---|
| 1 | -1.11 | -1.87 | -0.35 |
| 5 | -0.81 | -1.53 | -0.08 |
| 45 | 2.20 | 1.71 | 2.69 |
| 50 | 2.57 | 2.09 | 3.06 |
| 100 | 6.33 | 5.51 | 7.15 |
| 150 | 10.09 | 8.71 | 11.47 |
Sample Integrity
A sample integrity study was conducted at two external sites to assess sample stability of fresh samples at room temperature, when stored frozen at ≤-70 ℃ and after up to two freeze thaw cycles. The FVIII activity of forty-six plasma samples was measured using a single lot of CRYocheck Chromogenic Factor VIII on an IL ACL TOP 300 and IL ACL TOP 700 (K160276) analyzer. Results were compared using Passing Bablok regression analysis and support a fresh sample stability claim of 2 hours at room temperature and a frozen storage claim of 3 months at ≤-70 °C, including up to two freeze thaw cycles.
10
Conclusion
The performance testing results demonstrate that cRYOcheck Chromogenic Factor VIII is substantially equivalent to the predicate device, Coatest SP FVIII (K042576), and that the assay
is effective for its labeled intended use.