(240 days)
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.
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.
The CRYOcheck Chromogenic Factor VIII device is intended for the quantitative determination of Factor VIII (FVIII) activity in human plasma to identify FVIII deficiency and aid in Hemophilia A management.
Here's an analysis based on the provided document:
1. Table of Acceptance Criteria and Reported Device Performance
The acceptance criteria are generally implied by the successful outcomes of various studies, aligning with CLSI guidelines and demonstrating performance comparable to the predicate device. Specific numerical acceptance criteria were not explicitly stated as pass/fail thresholds in the document, but the reported performance values are presented as the fulfillment of these criteria.
| Performance Characteristic | Acceptance Criteria (Implied by Study Design & Comparator Performance) | Reported Device Performance (CRYOcheck Chromogenic Factor VIII) |
|---|---|---|
| Precision | CV < 10% for FVIII > 1%; SD $\leq$ 0.1% for very low FVIII | Pooled precision: <10% CV for FVIII > 1%; $\leq$0.1% SD for very low FVIII plasma (Table in document) |
| Reproducibility | CV < 10% for FVIII > 1%; SD $\leq$ 0.1% for very low FVIII | Pooled reproducibility: <10% CV for FVIII > 1%; $\leq$0.1% SD for very low FVIII plasma (Table in document) |
| Linearity Range | Clinically relevant range (e.g., 0-200% FVIII activity) | 0 to 200% FVIII activity |
| Reference Interval | Established from healthy population | 43.2-159.3% FVIII activity (95% CI) |
| Shelf-Life Stability | At least 12 months at recommended storage conditions | At least 12 months at $\leq$-70°C (study completed up to 13 months) |
| In-Use Stability | Defined operational stability durations | 8 hours on-board instrument; 5 days at 2-8°C; 1 month refrozen storage at $\leq$-70°C (if refrozen within 4 hours of initial thaw) |
| Limit of Detection (LoD) | Clinically relevant low detection limit | 0.5% FVIII activity |
| Limit of Blank (LoB) | Clinically relevant lower blank limit | 0.4% FVIII activity |
| Limit of Quantitation (LoQ) | Clinically relevant low quantitation limit | 0.5% FVIII activity |
| Interferences | No significant interference from common substances | No interference from tested: Hemoglobin $\leq$ 500 mg/dL, Intralipid $\leq$ 500 mg/dL, Bilirubin (unconjugated) $\leq$ 29 mg/dL, vWF $\leq$ 20 µg/mL, Unfractionated heparin $\leq$ 2 IU/mL, Low molecular weight heparin $\leq$ 2 IU/mL, Fondaparinux $\leq$ 1.25 mg/L, Lupus Anticoagulant $\leq$ 1.8 dRVVT ratio. Interference from Rivaroxaban and Dabigatran noted. |
| Method Comparison | Equivalent performance to predicate device | Passing-Bablok regression: Slope ~1, Intercept ~0, Pearson Correlation Coefficient > 0.99 (Overall: Slope 1.038, Intercept 0.473, r=0.994) |
| Sample Integrity | Defined sample stability durations | 2 hours at room temperature; 3 months at $\leq$-70°C (including up to two freeze-thaw cycles) |
2. Sample Sizes Used for the Test Set and Data Provenance
- Precision Study: One normal and two abnormal reference controls, and five patient plasma samples (representing very low, mid, normal, and high FVIII activity). Each sample was measured in duplicate, twice a day for 20 days (total 80 replicates per sample per lot) with 3 lots of the device. Data provenance is internal (as it's an internal study).
- Reproducibility Study: One normal and two abnormal reference controls, and three patient plasma samples (representing very low, normal, and high FVIII activity). Each sample was measured in triplicate, twice a day for 5 days at each of 3 sites with 3 lots of the device. Data provenance includes one internal and two external sites, but specific countries are not mentioned beyond "Canada" for the submitter. This appears to be prospective data collection for the study.
- Linearity/Assay Reportable Range Study: Fifteen sample dilutions created by combining high FVIII plasma (260%) with congenital FVIII deficient plasma (0%). Each level was tested in quadruplicate. Data provenance is internal (as it implies an internal study).
- Reference Interval Study: One hundred and twenty ostensibly healthy individuals $\geq$ 18 years. Data provenance is not specified beyond "citrated plasma samples collected from". This appears to be prospective data collection.
- Shelf-Life Stability Study: Six plasma samples representing low to normal FVIII activity levels. Three lots of the device were tested at various time points up to 37 months (13 months completed). Data provenance is internal.
- In-Use Stability Study: Six plasma samples representing low to normal FVIII activity levels. Three lots of the device were tested at various time points. Data provenance is internal.
- Detection Limit (LoB/LoD/LoQ) Studies:
- LoB: Four blank plasma samples from individuals with severe congenital hemophilia A.
- LoD: Four plasma samples with low FVIII activity from congenital hemophilia A donors.
- LoQ: Aliquots of four plasma samples with low FVIII activity from congenital hemophilia A donors.
All samples for LoB/LoD were measured in triplicate over five days using three lots of the device. For LoQ, samples were tested in triplicate on five different days at an external laboratory. Data provenance for LoB/LoD/LoQ samples is from individuals with hemophilia A.
- Interference Studies: Plasma samples spiked with possible interferents. Ten replicates of each spiked sample and 10 replicates of corresponding blank matrix control were tested. Data provenance is internal.
- Method Comparison Studies: Three hundred and eighteen human plasma samples from normal individuals, patients with congenital or acquired hemophilia A, and various types of von Willebrand disease. Samples were distributed across three sites. Data provenance spans "normal ostensibly healthy individuals and from patients". This appears to be prospective data collection.
- Sample Integrity Study: Forty-six plasma samples. Data provenance is not further specified. This appears to be prospective data collection at two external sites.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts
For this in vitro diagnostic device, "ground truth" is typically established by reference methods or accepted clinical classifications rather than expert consensus on individual images or cases.
- For studies like Precision, Reproducibility, Linearity, Stability, and Detection Limits, the "ground truth" for the FVIII activity levels in controls and calibrators would be established by the manufacturer's internal characterization methods, traceable to international standards if applicable (though not explicitly stated as such for FVIII activity). The expertise would lie in clinical chemistry, hematology, and laboratory medicine for the development and validation of these reference materials and methods. No specific number or qualification of experts is mentioned for these.
- For the Reference Interval Study, the ground truth is derived from the statistical distribution of FVIII activity in a population of ostensibly healthy individuals. This doesn't involve individual expert ground-truthing but rather statistical analysis.
- For the Method Comparison Study, the ground truth for FVIII activity was established by a comparator device, Coatest SP FVIII (K042576), used at a central reference laboratory. The "experts" in this context are the trained laboratory personnel performing the reference method.
- For sample selection in studies involving "congenital hemophilia A donors" or "patients with congenital or acquired hemophilia A and various types of von Willebrand disease," the diagnosis (which forms part of the "ground truth" for classifying these samples) would have been made by medical professionals (e.g., hematologists) based on standard clinical and laboratory diagnostic criteria. No specific number or qualifications are given.
4. Adjudication Method for the Test Set
Adjudication methods like "2+1" or "3+1" are typically used for subjective diagnostic tasks, often involving image interpretation or clinical reviews. Since the CRYOcheck Chromogenic Factor VIII is a quantitative assay for FVIII activity, the "adjudication method" is the quantitative measurement itself, often with replicates and statistical analysis, as detailed in the study designs (e.g., "in duplicate," "in triplicate"). No expert adjudication in the traditional sense is described or expected for this type of device.
5. If a Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study Was Done, and the Effect Size
No, a multi-reader multi-case (MRMC) comparative effectiveness study was not done. MRMC studies are primarily for evaluating the impact of a new diagnostic tool on the diagnostic performance of human readers, typically with subjective assessments. This device is a quantitative in vitro diagnostic, not a tool for human reading in the conventional sense.
6. If a Standalone (i.e., algorithm only without human-in-the-loop performance) Was Done
Yes, the device operates as a standalone algorithm (assay) measuring FVIII activity. The performance studies described (Precision, Reproducibility, Linearity, Reference Interval, Stability, Detection Limits, Interferences, Method Comparison, Sample Integrity) all represent the performance of the device itself, without human-in-the-loop diagnostic interpretation as part of its core function, other than trained laboratory personnel operating the instrument and interpreting numerical output.
7. The Type of Ground Truth Used
The ground truth used for these studies varies by the specific test:
- Reference materials and controls: For precision, linearity, and detection limit studies, the ground truth for FVIII activity levels is based on established values of these materials, likely traceable to international standards or well-characterized internal standards.
- Healthy population data: For the reference interval, the ground truth for "normal" FVIII activity is derived from a statistically representative healthy population.
- Comparator device: For the method comparison study, the ground truth was established by the predicate device, Coatest SP FVIII (K042576).
- Clinical diagnosis: For studies involving patient samples (e.g., hemophilia A, von Willebrand disease), the ground truth for their disease status would be based on clinical diagnosis by medical professionals.
8. The Sample Size for the Training Set
The document describes performance studies (validation tests) for the device. It does not provide information on a "training set" in the context of machine learning. This is because the device is a chemical assay, not an AI/ML-based diagnostic system that would require a training set to develop its model.
9. How the Ground Truth for the Training Set Was Established
As no training set is mentioned or applicable for this type of chemical assay device, this question is not relevant based on the provided information.
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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
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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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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'sInformation | Precision BioLogic Inc.140 Eileen Stubbs Ave.Dartmouth, Nova Scotia B3B 0A9Canada | ||
|---|---|---|---|
| Contact Person | Karen M. Black, VP of Compliance & Product DevelopmentPhone: 902-468-6422, ext. 226, or 902-706-3125E-mail: kblack@precisionbiologic.com | ||
| Preparation Date | 9 July 2020 | ||
| Device TradeName | CRYOCheck™ Chromogenic Factor VIII | ||
| RegulatoryInformation | Regulation Number andDescription | 21 CFR 864.7290Factor Deficiency Test | |
| Classification | Class II | ||
| Product Code | GGP; Test, Qualitative and QuantitativeFactor 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 thequantitative determination of factor VIII activity in 3.2% citrated humanplasma. It is intended to be used in identifying factor VIII deficiency andas an aid in the management of hemophilia A in individuals aged 2 yearsand older. For in vitro diagnostic use. | ||
| DeviceDescription | CRYOcheck Chromogenic Factor VIII is used for determination of FVIIIactivity and contains the following four components, packaged in glassvials and provided frozen to preserve the integrity of the components:Reagent 1: Bovine FX and a fibrin polymerization inhibitor, withactivators and stabilizers.Reagent 2: Human FIIa, human FIXa, calcium chloride andphospholipids.Reagent 3: FXa substrate containing EDTA and a thrombin inhibitor.Diluent Buffer: Tris buffer solution containing 1% BSA and a heparinantagonist. | ||
| Comparison to Predicate | |||
| Item | Predicate | New Device | |
| Proprietary andEstablished 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 | GGPTest, Qualitative andQuantitative Factor Deficiency | GGPTest, Qualitative and QuantitativeFactor Deficiency | |
| Regulation Section | 21 CFR 864.7290 | 21 CFR 864.7290 | |
| Factor Deficiency TestClass II | Factor Deficiency TestClass II | ||
| Classification | 81 (Haematology) | 81 (Haematology) | |
| Panel | Coatest SP FVIII is intended forthe photometric determination offactor VIII activity in citratedplasma. | CRYOCheck Chromogenic Factor VIIIis for clinical laboratory use in thequantitative determination of factorVIII activity in 3.2% citrated humanplasma. It is intended to be used inidentifying factor VIII deficiency andas an aid in the management ofhemophilia A in individuals aged 2years and older. For in vitrodiagnostic use. | |
| Intended Use | Quantitative (chromogenicmeasurement of FVIII) | Quantitative (chromogenicmeasurement of FVIII) | |
| Assay Type | Coatest SP FVIII is a modifiedversion of Coatest Factor VIII(K833892) reformulated toEuropean PharmacopoeiaStandards. Coatest SP FVIII is aphotometric assay containing achromogenic substrate, S-2765,with EDTA added as apreservative, lyophilized bovinefactors IXa and X with bovinealbumin added as a stabilizingagent. The device also containscalcium chloride, Tris bufferstock solution containing sodiumchloride, bovine serum albuminwith added antimicrobial inaddition to a mixture of highlypurified synthetic phospholipids. | CRYOCheck Chromogenic Factor VIIIis used for determination of FVIIIactivity and contains the followingfour components, packaged in glassvials and provided frozen to preservethe integrity of the components:Reagent 1: Bovine FX and a fibrinpolymerization inhibitor, withactivators and stabilizers.Reagent 2: Human Flla, humanFIXa, calcium chloride andphospholipids.Reagent 3: FXa substrate containingEDTA and a thrombin inhibitor.Diluent Buffer: Tris buffer solutioncontaining 1% BSA and a heparinantagonist. | |
| Device Description | In the presence of calcium andphospholipids, factor X isactivated to factor Xa by factorIXa. This generation is greatlystimulated by factor VIII, whichmay be considered as a cofactorin this reaction. By using optimalamounts of Ca2+ andphospholipids and an excess offactors IXa and X, the rate ofactivation of factor X is solelydependent on the amount offactor VIII. Factor Xa hydrolysesthe chromogenic substrate S-2765 thus liberating thechromophoric group, pNA. Thecolor is then readphotometrically at 405 nm. Thegenerated factor Xa and thusthe intensity of color are | In the first stage of the chromogenicassay, test plasma (containing anunknown amount of functional FVIII)is added to a reaction mixturecomprised of calcium, phospholipids,purified human thrombin and FIXa,and purified bovine FX (Reagent 1and Reagent 2). This mixture swiftlyactivates FVIII to FVIIIa, which worksin concert with FIXa to activate FX.When the reaction is stopped, FXaproduction is assumed to beproportional to the amount offunctional FVIII present in thesample. The second stage of theassay is to measure FXa throughcleavage of an FXa-specific peptidenitroanilide substrate (FXaSubstrate). P-nitroaniline isproduced, giving a color that can be | |
| Methodology | |||
| proportional to the factor VIIIactivity in the sample. Hydrolysisof S-2765 by thrombin formed isprevented by the addition of thesynthetic thrombin inhibitor, I-2581, together with thesubstrate. | measured spectrophotometrically byabsorbance at 405 nm. | ||
| Expression ofresults | Quantitative; results areexpressed as percent activityinterpreted relative to acalibration curve. | Quantitative; results are expressedas percent activity interpreted relativeto a calibration curve. | |
| Differences | |||
| Instrument(s) | Manual method, IL ACL 9000 | IL ACL TOP CTS Series and IL ACLTOP 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% (instrumentapplication) | 43.2-159.3% FVIII activity | |
| Limit of Detection | 1% FVIII activity | 0.5% FVIII activity | |
| In Use Stability | Working Factor ReagentStability (phospholipids + factorIXa + factor X reagent): 12hours on ice.S-2765 + I-2581: Reconstitutedsubstrate is stable 3 months at2-8°CCaCl2: Opened vial is stable 3months at 2-8ºCBuffer, stock solution: Openedvial is stable 3 months at 2-8ºCPhospholipid: Opened vial isstable for 3 months at 2-8ºCFactor Reagent (IXa + X):Aliquoted for -20ºC for 3months. | 8 hours on-board instrument5 days at 2-8ºC1 month refrozen storage at ≤-70 °Cif refrozen within 4 hours of the initialthaw. Previously refrozen reagentscan be thawed and used once for upto four hours on board theinstrument. | |
| Interferences | Manual method:Triglycerides up to 700 mg/dLBilirubin up to 20 mg/dLHemoglobin up to 100 mg/dLUnfractionated Heparin up to 1.0IU/mLAutomated Method:Triglycerides up to 900 mg/dLBilirubin up to 20 mg/dLHemoglobin up to 50 mg/dL | Hemoglobin: ≤ 500 mg/dLIntralipid: ≤ 500 mg/dLBilirubin (unconjugated): ≤ 29 mg/dLvWF: ≤ 20 µg/mLUnfractionated heparin: ≤ 2 IU/mLLow molecular weight heparin: ≤2 IU/mLFondaparinux: ≤ 1.25 mg/LLupus Anticoagulant: ≤ 1.8 dRVVTratioRivaroxaban and dabigatraninterfered with the quantification ofFVIII activity. | |
| Unfractionated Heparin up to 1.0IU/mL | The potential interference effects ofconjugated bilirubin on this devicehave not been evaluated. | ||
| Due to the high dilutions usedthere is no underestimation ofFVIII activity in samplescontaining lupus anticoagulant | The performance of this device hasnot been established in individualswith von Willebrand disease Type2M. | ||
| The performance of this device hasnot been established in evaluatingthe potency of FVIII concentrates. | |||
| Sample Stability | None specified.Package insert indicates "Referto NCCLS document H21-A4 forfurther instructions on specimencollection, handling andstorage." | 2 hours at room temperature and 3months at ≤-70 °C, including up totwo freeze thaw cycles |
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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 <10% CV and ≤0.1% SD for the Very Low FVIII plasma sample.
| Aggregated Data (Lots 1, 2 and 3) | ||||
|---|---|---|---|---|
| Sample | Mean FVIII (%) | Within-Laboratory | ||
| SD | %CV | |||
| CRYOcheck Reference Control Normal | 80.8 | 4.0 | 5.0 | |
| CRYOcheck Abnormal 1 Reference Control | 26.1 | 1.9 | 7.1 | |
| CRYOcheck Abnormal 2 Reference Control | 7.8 | 0.8 | 9.9 | |
| Very Low FVIII Plasma Sample | 1.0 | 0.1 | NA | |
| Low FVIII Plasma Sample | 5.4 | 0.4 | 7.4 | |
| Mid FVIII Plasma Sample | 26.0 | 1.7 | 6.7 | |
| Normal FVIII Plasma Sample | 85.3 | 4.3 | 5.1 | |
| High FVIII Plasma Sample | 152.1 | 5.7 | 3.8 |
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Multi-Reagent Lot Site to Site Reproducibility
Reproducibility studies were conducted at three sites (one internal and two external) by different operators on an IL ACL TOP CTS (K160276) and two different IL ACL TOP 750 (K150877) analyzers using three lots of cRYOcheck Chromogenic Factor VIII in accordance with CLSI EP05-A3. The study quantified one normal and two abnormal reference controls and three patient plasma samples representing very low, normal and high levels of FVIII activity. Each sample was measured in triplicate, twice a day for 5 days at each site. The data across three sites demonstrated a pooled reproducibility of <10% CV for the controls and plasma samples >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 1Reference 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 2Reference 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
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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.
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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 CorrelationCoefficient | |||
|---|---|---|---|---|---|---|
| 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.
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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.
§ 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).