(305 days)
CRYOcheck Hex LA is for clinical laboratory use as a qualitative test kit intended to aid in the detection of lupus anticoagulants (LA) in 3.2% citrated human plasma by the application of hexagonal phase phospholipids. CRYOcheck Hex LA should be used as an integrated test for lupus anticoagulant detection. For in vitro diagnostic use. The performance of this device has not been established in neonate and pediatric patient populations.
CRYOcheck Hex LA is comprised of three reagents supplied in a frozen format as follows:
LA Start: Pooled normal plasma with buffer and a heparin neutralizer.
LA Correct: Pooled normal plasma with buffer, a heparin neutralizer, and inverted hexagonal phase phospholipid.
LA APTT: Silica-based lupus sensitive APTT reagent with stabilizer.
1. Acceptance Criteria and Device Performance:
| Acceptance Criteria (Internal Precision) | Reported Device Performance |
|---|---|
| Pooled precision of < 5% CV for LA Start | Pooled precision of < 5% CV for LA Start (e.g., 2.6%-4.2%) |
| Pooled precision of < 8% CV for LA Correct | Pooled precision of < 8% CV for LA Correct (e.g., 4.2%-7.6%) |
| Acceptance Criteria (Reproducibility) | Reported Device Performance |
| Pooled reproducibility of < 5% CV for LA Start (across 3 sites) | Pooled reproducibility of < 5% CV for LA Start (e.g., 3.0%-4.8%) |
| Pooled reproducibility of ≤ 8% CV for LA Correct (across 3 sites) | Pooled reproducibility of ≤ 8% CV for LA Correct (e.g., 4.8%-8.0%) |
| Acceptance Criteria (Normal Range / Cut-off Establishment) | Reported Device Performance |
| Cut-off for assay delta correction determined as mean + 4 SD from normal range study. | Cut-off established as mean + 4 SD, resulting in: |
| < 6.0 seconds = LA Negative | |
| ≥ 6.0 seconds = LA Positive | |
| Acceptance Criteria (Shelf Life Stability) | Reported Device Performance |
| Supports a shelf-life stability claim of at least 12 months at -70°C or colder. | Supported for at least 12 months at -70°C or colder. |
| Acceptance Criteria (In-Use Stability) | Reported Device Performance |
| Supports a 4-hour in-use stability at room temperature. | Supported for 4 hours at room temperature. |
| Supports an 8-hour in-use stability when stored on-board the instrument. | Supported for 8 hours when stored on-board the instrument. |
| Acceptance Criteria (Method Comparison - Qualitative Agreement vs. Predicate) | Reported Device Performance |
| Positive Percent Agreement (PPA) | 95.6% (95% CI, 91%-98%) |
| Negative Percent Agreement (NPA) | 95.2% (95% CI, 92%-97%) |
| Overall Agreement | 95.3% (95% CI, 93%-97%) |
| Acceptance Criteria (Sample Integrity - Fresh Sample Stability) | Reported Device Performance |
| Supports fresh sample stability of 4 hours at room temperature. | Supported for 4 hours at room temperature. |
| Acceptance Criteria (Sample Integrity - Frozen Storage & Freeze-Thaw) | Reported Device Performance |
| Supports frozen storage claim of 2 months at ≤ -70°C, including one freeze-thaw cycle. | Supported for 2 months at ≤ -70°C, including one freeze-thaw cycle. |
| Acceptance Criteria (Interferences - No Interference) | Reported Device Performance (no interference up to indicated concentrations) |
| Hemoglobin ≤ 500 mg/dL | Hemoglobin: ≤ 500 mg/dL |
| Bilirubin (unconjugated) ≤ 20 mg/dL | Bilirubin (unconjugated): ≤ 20 mg/dL |
| Bilirubin (conjugated) ≤ 2 mg/dL | Bilirubin (conjugated): ≤ 2 mg/dL |
| Intralipid ≤ 500 mg/dL | Intralipid: ≤ 500 mg/dL |
| Unfractionated Heparin ≤ 2 IU/mL | Unfractionated Heparin: ≤ 2 IU/mL |
| Low Molecular Weight Heparin ≤ 2 IU/mL | Low Molecular Weight Heparin: ≤ 2 IU/mL |
| Dabigatran, rivaroxaban, fondaparinux (no interpretation interference) | No interpretation interference observed |
| Elevated Factor VIII activity (up to 180%) (no interference) | No interference observed |
| Elevated fibrinogen concentrations (no interpretation interference) | No interpretation interference observed |
| C-reactive protein (no interpretation interference) | No interpretation interference observed |
| Factor VIII inhibitor antibodies (no interpretation interference) | No interpretation interference observed |
| Plasma samples with elevated INR (up to 4.5) (no interpretation interference) | No interpretation interference observed |
| High platelet counts (>10,000 platelets/µL) should show interference | Showed interference |
| Abnormally low factor II activities (below 50%) (may interfere, potentially false negative) | May interfere, potentially resulting in false negative results |
| Factor VII and factor IX deficiencies (no interference) | No interference observed |
| Abnormally low factor X activities (below 50%) (no interpretation interference) | No interpretation interference observed |
2. Sample Sizes and Data Provenance:
- Precision Study:
- 3 control plasmas and 5 plasmas with varying LA positivity.
- Tested in duplicate, twice a day for 20 days per lot of reagent (3 lots used).
- Reproducibility Study:
- 3 control plasmas and 5 plasmas with varying LA positivity.
- Each sample tested in triplicate, twice a day for 5 days for each of the 3 lots of reagent.
- Normal Range and Assay Cut-off Study:
- Normal samples: 137 on Analyzer A, 126 on Analyzer B.
- Each sample tested using 3 lots of CRYOcheck Hex LA.
- Shelf Life Stability Study:
- 3 lots of CRYOcheck Hex LA.
- 10 replicates of 3 controls tested at time = 0 and regular intervals up to 37 months.
- One additional plasma sample close to the cut-off for one lot.
- In-Use Stability Study:
- 3 lots of CRYOcheck Hex LA.
- 5 replicates of 3 control plasmas and 4 test plasmas with varying levels of LA.
- Tested at 0, 2, 4, 6, 7, 8, and 9 hours.
- Interference Studies:
- Patient plasma samples spiked with possible interferents.
- 20 replicates of spiked samples tested alongside 20 replicates of corresponding blank matrix control.
- Single lot of CRYOcheck Hex LA used.
- Method Comparison Studies (Test Set):
- Total samples: 446
- 124 known (previously characterized) LA positive samples.
- 75 normal (presumed LA negative) samples from individuals with other medical conditions including autoimmune disorders.
- 220 LA target screening population samples.
- Data Provenance: The document does not explicitly state the country of origin for the patient samples. The study involved one internal site and three external sites, suggesting a multi-center study. It is a retrospective study since samples were "known (previously characterized) LA positive" or "presumed LA negative."
- Total samples: 446
- Sample Integrity Study:
- 64 samples.
3. Number of Experts and Qualifications for Ground Truth:
- This device is an in vitro diagnostic (IVD) test, not an AI/imaging device requiring expert interpretation for ground truth.
- The ground truth for the method comparison study was established by comparing the CRYOcheck Hex LA results against a legally marketed predicate device, Staclot® LA (K923731).
- The determination of "known (previously characterized)" LA positive samples and "presumed LA negative" samples would implicitly rely on established clinical or laboratory diagnostic procedures, which are performed by qualified laboratory personnel, not typically "experts" in the sense of a radiology reader study.
4. Adjudication Method for the Test Set:
- Not applicable as this is a comparison study against a predicate device's results, not a human reader study needing adjudication. The "ground truth" for the test set was the result obtained from the predicate device (Staclot® LA).
5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study:
- No, an MRMC comparative effectiveness study was not done. This is an IVD device measuring a biomarker, not an imaging AI device assisting human readers. Therefore, the concept of "effect size" of how human readers improve with AI vs. without AI assistance is not relevant here.
6. Standalone Performance:
- Yes, the performance data presented (Precision, Reproducibility, Normal Range, Stability, Interferences) are essentially standalone (algorithm-only) performance characteristics of the CRYOcheck Hex LA device in a laboratory setting.
- The method comparison study also evaluated the device's performance independently against a predicate, effectively demonstrating its standalone performance in identifying LA status.
7. Type of Ground Truth Used:
- For the method comparison study (test set), the ground truth was comparison to a legally marketed predicate device (Staclot® LA).
- For the known LA positive/negative samples, the ground truth was "previously characterized" LA status, likely established through a combination of clinical diagnosis and existing laboratory methods.
8. Sample Size for the Training Set:
- This document describes a 510(k) submission for an in vitro diagnostic (IVD) device, not an AI/machine learning device that typically involves a "training set" in the same computational sense.
- The development and optimization of such an IVD assay would involve internal development samples and studies, but these are not referred to as "training sets" in the context of this regulatory filing. The provided information focuses on analytical and clinical performance validation.
9. How the Ground Truth for the Training Set Was Established:
- Not applicable due to the nature of the device as explained in point 8. The device's "training" or development would involve laboratory optimization and calibration using reference materials and characterized samples, but not "ground truth" in the AI sense for a dedicated "training set."
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October 23, 2020
Precision BioLogic Karen Black VP of Compliance and Product Development 140 Eileen Stubbs Avenue Dartmouth, Nova Scotia B3B 0A9 Canada
Re: K193556
Trade/Device Name: Cryocheck Hex LA Regulation Number: 21 CFR 864.7925 Regulation Name: Partial Thromboplastin Time Tests Regulatory Class: Class II Product Code: GFO Dated: December 20, 2019 Received: December 23, 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 requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part
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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 (QS) 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 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,
Takeesha Taylor-Bell Chief Division of Immunology and Hematology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health
Enclosure
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Indications for Use
510(k) Number (if known) K193556
Device Name Cryocheck Hex LA
Indications for Use (Describe)
Cryocheck Hex LA is for clinical laboratory use as a qualitative test kit intended to aid in the detection of lupus anticoagulants (LA) in 3.2% citrated human plasma by the application of hexagonal phase phospholipids. Cryocheck Hex LA should be used as an integrated test for lupus anticoagulant detection. For in vitro diagnostic use. The performance of this device has not been established in neonate and pediatric patient populations.
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
510(k) Summary cryocheck™ Hex LA™
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 K193556
| Submitter'sInformation | Precision BioLogic Inc.140 Eileen Stubbs Ave.Dartmouth, Nova Scotia B3B 0A9Canada | Similarities | ||||
|---|---|---|---|---|---|---|
| Contact Person | Karen M. Black, VP of Compliance & Product DevelopmentPhone: 902-468-6422 ext. 226, or 902-706-3125E-mail: kblack@precisionbiologic.com | Staclot LA | CRYOcheck Hex LA | |||
| Preparation Date | 14 October 2020 | Measurand | lupus anticoagulant | lupus anticoagulant | ||
| Device Trade Name | CRYOcheck ™ Hex LA | Product Code | GFO | GFO | ||
| RegulatoryInformation | Regulation Number andDescription | 21 CFR 864.7925Partial thromboplastin time test | Partial thromboplastin time tests | Partial thromboplastin time tests | ||
| Classification | Class II | Regulation Section | 21 CFR 864.7925 | 21 CFR 864.7925 | ||
| Product Code | GFO; Partial thromboplastin time test; 21CFR 864.7290 | Partial thromboplastin time tests | Partial thromboplastin time test | |||
| Classification Panel | Hematology | Classification | Class II | Class II | ||
| Panel | 81 (Haematology) | 81 (Haematology) | ||||
| Predicate Device | Staclot LA (K923731) | Intended Use | The Staclot LA test kit is a reagentsystem designed for the qualitativedetection of lupus anticoagulants(LA) in plasma by the use ofhexagonal HII phase phospholipidmolecules. (In the USA thisprocedure has been assigned tothe high complexity category perCLIA 1988 - CDC Analyte Code3728; CDC Test System Code13285). | CRYOcheck Hex LA is for clinicallaboratory use as a qualitativetest kit intended to aid in thedetection of lupus anticoagulants(LA) in 3.2% citrated humanplasma by the application ofhexagonal phase phospholipids.CRYOcheck Hex LA should beused as an integrated test forlupus anticoagulant detection.For in vitro diagnostic use. Theperformance of this device hasnot been established in neonateand pediatric patientpopulations. | ||
| Indication for Use/Intended Use | CRYOcheck Hex LA is for clinical laboratory use as a qualitative test kitintended to aid in the detection of lupus anticoagulants (LA) in 3.2%citrated human plasma by the application of hexagonal phasephospholipids. CRYOcheck Hex LA should be used as an integratedtest for lupus anticoagulant detection. For in vitro diagnostic use. Theperformance of this device has not been established in neonate andpediatric patient populations. | Assay Type | Qualitative; hexagonal phaseneutralization test | Qualitative; hexagonal phaseneutralization test | ||
| Device Description | CRYOcheck Hex LA is comprised of three reagents supplied in a frozenformat as follows:LA Start: Pooled normal plasma with buffer and a heparin neutralizer.LA Correct: Pooled normal plasma with buffer, a heparin neutralizer,and inverted hexagonal phase phospholipid.LA APTT: Silica-based lupus sensitive APTT reagent with stabilizer. | Methodology | The Staclot LA test procedure isbased on the following principle:the test plasma that is suspected tocontain LA is first allowed toincubate at 37°C with (Tube 2) andwithout (Tube 1) hexagonal phasephosphatidylethanolamine (HPE)(Reagent 2); next, an APTT isperformed on both tubes using anLA sensitive reagent (Reagent 4); ifLA were present in the test plasma,they would be neutralized by HPEin tube 2, and this would result in ashortening of the clotting time oftube 2 compared with that of tube1. By comparing the differencebetween the two clotting times, thepresence of LA antibodies in thetest plasma can be identified.The Reagent 3 contains a heparininhibitor which makes the testsystem insensitive to heparin levelsup to 1 IU/mL. Furthermore, theStaclot LA procedure calls for theaddition of a normal plasma(Reagent 3) to the test system to | CRYOcheck Hex LA is ahexagonal-phase phospholipidneutralization test (HPNT),which is an integrated test thatcombines screening,confirmatory and mixing testprocedures into a single assay.CRYOcheck LA works on theprinciple that LA are neutralizedby hexagonal phasephospholipids that are present inthe assay's confirmatoryreaction mixture and not thescreening reaction mixture. Thepresence of LA in plasmasamples is confirmed by thecorrection of APTT clot times inthe presence of a reactionmixture containing hexagonalphase phospholipids.In the CRYOcheck Hex LA assay,the test plasma suspected tocontain LA is incubated in tworeaction cuvettes, both of whichentail dilution with pooled normal | ||
| Comparison to Predicate | ||||||
| Item | Predicate | New Device | ||||
| Proprietary andEstablished Names | Staclot LA | CRYOcheck Hex LA | ||||
| Manufacturer | American Bioproducts Inc (originalapplicant); Diagnostica Stago(current manufacturer) | Precision BioLogic Inc |
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| Similarities | ||
|---|---|---|
| Staclot LA | CRYOcheck Hex LA | |
| clotting time due to factordeficiencies that might be present.If the Staclot LA does not producea shortening of the clotting time,then the presence of anti-factorantibodies should be suspected; inthis case, use an appropriate testfor anti-factor antibodies.Compare the clotting time of tube 1(CT1) with that of tube 2 (CT2). Ashortening of clotting time of 8seconds or more of tube 2compared with that of tube 1 issignificant of a neutralization ofanti-phospholipid antibodies (this 8-second cut-off in clotting times hasbeen determined with the ST4/STart® instrument - DiagnosticaStago). | neutralizer), thus satisfying themixing test requirement. In thefirst cuvette, the screening testreaction is performed by mixingthe test plasma with the LA Startreagent (pooled normal plasma).In the second cuvette, theconfirmatory reaction isperformed by mixing the testplasma with the LA Correctreagent (pooled normal plasmawith hexagonal phasephospholipid). The LA APTTreagent is then added to eachcuvette, followed by 0.025 MCaCl2 to activate clotting via theintrinsic pathway. Clot times arerecorded for the screeningreaction mixture containing LAStart and the confirmatoryreaction mixture containing LACorrect. The result is reportedas the difference in clot time inseconds ("delta correction")between LA Start and LACorrect cuvettes.delta correction = (CT LA Start)- (CT LA Correct)The result is then compared toan established cut-off. A resultgreater than or equal to theestablished cut-off is consideredLA positive, while a result lessthan the established cut-off isconsidered LA negative. | |
| Expression of results | Qualitative; results are reported asclot time delta (seconds) and areinterpreted as positive or negativerelative to an established cut-offvalue. | Qualitative; results are reportedas clot time delta (seconds) andare interpreted as positive ornegative relative to anestablished cut-off value. |
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| Differences | ||||||||
|---|---|---|---|---|---|---|---|---|
| Staclot LA | CRYOcheck Hex LA | |||||||
| Format | Staclot LA is comprised of three lyophilized reagents and two reconstitution liquids as follows:Reagent 1 : ready-for-use bufferReagent 2 : lyophilized hexagonal phase phosphatidylethanolamineReagent 3 : lyophilized normal human plasma containing a heparin inhibitorReagent 4 : lyophilized PTT-LS reagent consisting of cephalin prepared from rabbit cerebral tissues and a particulate siliceous activatorReagent 5 : solvent for reconstitution of Reagent 4. | CRYOCheck Hex LA is comprised of three reagents supplied in a frozen format as follows:LA Start : Pooled normal plasma with buffer and a heparin neutralizer.LA Correct : Pooled normal plasma with buffer, a heparin neutralizer, and inverted hexagonal phase phospholipid.LA APTT : Silica-based LA sensitive APTT reagent with stabilizer. | ||||||
| Storage | 2-8°C until expiration | ≤-70°C until expiration | ||||||
| Instrument | Manual (ST4/ST art®) | STA-R Evolution® | ||||||
| Associated Controls | STA® - Control LA 1 + 2 | CRYOCheck Lupus Negative ControlCRYOCheck Weak Lupus Positive ControlCRYOCheck Lupus Positive Control | ||||||
| Cut-off | A shortening of clotting time of 8 seconds or more of tube 2 compared with that of tube 1 is significant of a neutralization of anti-phospholipid antibodies (this 8-second cut-off in clotting times has been determined with the ST4/ST art® instrument - Diagnostica Stago).Each laboratory should verify this 8-second cut-off by testing the plasma of at least 20 normal individuals, using its own methodology to obtain the mean delta T + 4 SD. | The cut-off for the assay delta correction was determined using pooled data from a normal range study conducted on Stago STA-R Evolution® analyzers and calculating the mean + 4 SD, with the following results:Delta Correction Interpretation < 6.0 seconds LA Negative ≥ 6.0 seconds LA Positive The results were obtained using specific lots of reagent. The cut-off is calculated as the mean of the delta correction + 4 SD, consistent with accepted methods for hexagonal phase neutralization tests. This method of establishing cut-off is different than that indicated for confirmatory tests in Pengo et al., 2009.1 Each laboratory should verify its own cut-off, by |
1 Pengo V, Tripodi A, Reber G, Rand JH, Ortel TL, Galli M, deGroot PG. Update of the guidelines for lupus anticoagulant detection. J. Thromb. Haemost. 2009;7(10):1737-1740.
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| Differences | ||
|---|---|---|
| Staclot LA | CRYOcheck Hex LA | |
| testing the plasma of at least 20normal individuals. | ||
| Heparin Interference | No interference up to 1 IU/mL | Unfractionated heparin: nointerference up to 2 IU/mLLow molecular weight heparin:no interference up to 2 IU/mL |
| Direct Thrombin andXa InhibitorInterference | Thrombin inhibitors (e.g., hirudin,argatroban...) present in thesample to be tested may interferein the test and lead to falselypositive results. | Dabigatran, rivaroxaban, andfondaparinux do not interferewith the interpretation ofCRYOcheck Hex LA results butmay increase the deltacorrection of LA positivesamples. |
| Warfarin Interference | The Staclot® LA procedure wasused to test plasmas fromstabilized coumadin patients (n =29). All plasmas gave negativeresults with Staclot® LA. | Plasma samples with elevatedINR (up to 4.5) do not interferewith the interpretation ofCRYOcheck Hex LA results |
| Factor VIII InhibitorAntibody Interference | The presence of anti-factorantibodies does not normallyproduce a correction in clottingtime with the Staclot LA testprocedure. However consideringthe heterogeneity of theseantibodies, some may interfere inthe test. Consequently, when theseare suspected, use an appropriatetest for anti-factor antibodies | Factor VIII inhibitor antibodiesdo not interfere with theinterpretation of CRYOcheck HexLA results, but at titers above 15BU/mL may increase the deltacorrection of LA positivesamples. |
| Factor DeficiencyInterference | A total of 21 factor deficientplasmas, comprising deficiencies ofF. VIII (n = 8), F. VIII with thepresence of anti-F. VIII-antibodies(n = 5), F. IX (n = 4), F. XI (n = 2),F. XII (n = 1) and F. II (n = 1), weretested with the Staclot® LAprocedure. The observed CT1-CT2was found in all cases < 8seconds. | Factor VII and factor IXdeficiencies do not interfere withCRYOcheck Hex LA.Abnormally low factor Xactivities (below 50%) do notinterfere with the interpretationof CRYOcheck Hex LA results butmay increase the deltacorrection for LA positivesamples.Abnormally low factor II activities(below 50%) may interfere withthe interpretation of CRYOcheckHex LA, potentially resulting infalse negative results for weaklyLA positive plasmas. |
| HIL interference | Unknown | Hemoglobin: ≤ 500 mg/dLBilirubin (unconjugated): ≤ 20mg/dLBilirubin (conjugated): ≤ 2 mg/dLIntralipid: ≤ 500 mg/dL |
| Differences | ||
| Staclot LA | CRYOcheck Hex LA | |
| C-reactive proteininterference | Unknown | C-reactive protein does notinterfere with the interpretationof CRYOcheck Hex LA results butat concentrations above 15µg/mL may increase the deltacorrection of LA positivesamples. |
| Elevated factorinterference | Unknown | Elevated factor VIII activity (upto 180%) does not interfere withCRYOcheck Hex LA.Elevated fibrinogenconcentrations do not interferewith the interpretation ofCRYOcheck Hex LA results butmay increase the deltacorrection of LA positivesamples. |
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Performance Summary:
All studies were performed using CRYOcheck Hex LA on Diagnostica Stago STA-R Evolution® instrument(s).
Precision
An internal precision study was performed using three different lots of CRYOcheck Hex LA on a STA-R Evolution instrument in accordance with CLSI EP05-A3. Three lot numbers of CRYOcheck Hex LA were used to test three control plasmas and five plasmas with varying LA positivity, in duplicate, twice a day for 20 days. The results demonstrated a pooled precision of < 5% CV for LA Start and < 8% CV for LA Correct.
| Sample | Within Laboratory PrecisionLA Start | Within Laboratory PrecisionLA Correct | ||||
|---|---|---|---|---|---|---|
| MeanClot Time (s) | SD | %CV | MeanClot Time (s) | SD | %CV | |
| CRYOCheck Lupus Negative Control | 53.0 | 1.6 | 3.0 | 52.8 | 2.8 | 5.3 |
| CRYOCheck Weak Lupus Positive Control | 87.3 | 3.2 | 3.7 | 65.4 | 2.8 | 4.2 |
| CRYOCheck Lupus Positive Control | 125.4 | 5.2 | 4.2 | 79.8 | 4.5 | 5.7 |
| LA Negative Plasma Sample | 55.9 | 1.7 | 3.1 | 55.1 | 2.5 | 4.5 |
| LA Near Cut-Off Plasma Sample | 67.6 | 2.5 | 3.8 | 58.4 | 2.6 | 4.5 |
| LA Weak Positive Plasma Sample | 89.8 | 3.3 | 3.7 | 66.4 | 3.0 | 4.6 |
| LA Moderate Positive Plasma Sample | 146.5 | 6.0 | 4.1 | 85.9 | 5.8 | 6.7 |
| LA Strong Positive Plasma Sample | 270.7 | 9.6 | 3.6 | 118.0 | 9.0 | 7.6 |
Reproducibility
Reproducibility studies were conducted at three sites (one internal and two external) using three lots of CRYOcheck Hex LA in accordance with CLSI EP05-A3. The study tested three control plasmas as well as five plasmas with varying LA positivity. Each sample was tested in triplicate, twice a day for 5 days for each of the 3 lots of CRYOcheck Hex LA. The data across three sites demonstrated a pooled reproducibility of <5% CV for LA Start and ≤8 % CV for LA Correct as summarized in the reproducibility tables below.
| Reproducibility: LA Start | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Sample | MeanClotTime (s) | Within-Run(Repeatability) | Between-Run | Between-Day | Between-Site | Reproducibility | |||||
| SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV | ||
| CRYOcheck LupusNegative Control | 52.8 | 1.4 | 2.7 | 0.3 | 0.6 | 0 | 0 | 0.4 | 0.7 | 1.6 | 3.0 |
| CRYOcheck WeakLupus PositiveControl | 85.6 | 3.3 | 3.9 | 0.5 | 0.6 | 0.5 | 0.6 | 1.9 | 2.2 | 3.9 | 4.6 |
| CRYOcheck LupusPositive Control | 123.6 | 5.0 | 4.0 | 0 | 0 | 1.5 | 1.3 | 2.1 | 1.7 | 5.7 | 4.6 |
| LA NegativePlasma Sample | 55.8 | 1.5 | 2.6 | 0.1 | 0.2 | 0.3 | 0.5 | 0.7 | 1.3 | 1.8 | 3.2 |
| LA Near Cut-OffPlasma Sample | 66.9 | 2.3 | 3.5 | 0.4 | 0.6 | 0.8 | 1.2 | 0.8 | 1.2 | 2.7 | 4.0 |
| LA Weak PositivePlasma Sample | 88.3 | 3.7 | 4.1 | 0 | 0 | 1.1 | 1.3 | 1.8 | 2.0 | 4.3 | 4.8 |
| LA StrongPositive PlasmaSample | 264.9 | 6.9 | 2.6 | 0.3 | 0.1 | 2.3 | 0.9 | 4.5 | 1.7 | 10.3 | 3.9 |
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| Reproducibility: LA Correct | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|
| Mean | Within-Run(Repeatability) | Between-Run | Between-Day | Between-Site | Reproducibility | ||||||
| Sample | ClotTime (s) | SD | %CV | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| CRYOcheck LupusNegative Control | 53.7 | 1.7 | 3.1 | 0.8 | 1.6 | 0 | 0 | 0 | 0 | 3.1 | 5.8 |
| CRYOcheck WeakLupus PositiveControl | 65.7 | 2.4 | 3.6 | 1.0 | 1.5 | 0.7 | 1.0 | 0.7 | 1.1 | 3.1 | 4.8 |
| CRYOcheck LupusPositive Control | 80.2 | 3.2 | 4.0 | 1.5 | 1.8 | 1.4 | 1.7 | 1.8 | 2.2 | 4.8 | 5.9 |
| LA NegativePlasma Sample | 56.0 | 1.7 | 3.0 | 0.6 | 1.1 | 0.2 | 0.4 | 0 | 0 | 2.9 | 5.2 |
| LA Near Cut-OffPlasma Sample | 59.2 | 2.2 | 3.7 | 1.2 | 2.0 | 0.7 | 1.1 | 0.3 | 0.6 | 3.0 | 5.0 |
| LA Weak PositivePlasma Sample | 66.9 | 2.7 | 4.0 | 1.3 | 2.0 | 1.2 | 1.8 | 2.2 | 3.3 | 4.0 | 6.0 |
| LA StrongPositive PlasmaSample | 117.4 | 4.5 | 3.9 | 3.1 | 2.6 | 2.0 | 1.7 | 2.6 | 2.2 | 9.4 | 8.0 |
Normal Range and Assay Cut-off
A normal range study was performed in-house using on two analyzers using normal samples (Analyzer A, n = 137; Analyzer B, n = 126) according to CLSI EP28: A3c. Each sample was tested using three lots of CRYOcheck Hex LA. A pooled mean ±2 SD range was determined for delta correction results and is shown in the table below:
| Normal Range | |
|---|---|
| Lower Range (s) | Upper Range (s) |
| -5.9 | 2.0 |
The cut-off for the assay delta correction was determined using pooled data from the normal range study and calculating the mean + 4 SD, with the following results:
| Delta Correction | Interpretation |
|---|---|
| < 6.0 seconds | LA Negative |
| ≥ 6.0 seconds | LA Positive |
The cut-off results were obtained using specific lots of reagent. The cut-off is calculated as the mean of the delta correction + 4 SD, consistent with accepted methods for hexagonal phase neutralization tests. This method of establishing cut-off is different than that indicated for confirmatory tests in Pengo et al., 2009.1 Each laboratory should verify its own cut-off, by testing the plasma of at least 20 normal individuals.
Stability
Shelf Life Stability
A shelf life stability study was conducted in accordance with CLSI EP25-A. Three lots of CRYOcheck Hex LA were stored -70 ℃ (-66 to -72 ℃) and -80 ℃ (-76 to -82 ℃) and tested at time = 0 and regular intervals up to 37 months. At each timepoint, 10 replicates of three controls were tested. For one lot, an additional plasma sample close to the assay cut-off was also tested. The study has been completed up to 12 months and supports a shelf-life stability claim of at least 12 months when the product is stored at -70 ℃ or colder.
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In-Use Stability
An in-use stability study was conducted in accordance with CLSI EP25-A. Three lots of CRYOcheck Hex LA were maintained at room temperature (18–25 °C) or on-board the instrument and used to test five replicates of three control plasmas as well as four test plasmas with varying levels of LA at 0. 2. 4. 6. 7. 8 and 9 hours. The data support a 4-hour in-use stability of the product when maintained at room temperature or 8 hours when stored on-board the instrument.
Interferences
Interference studies were conducted according to CLSI EP07, 3d ed. using a single lot of CRYOcheck Hex LA. Patient plasma samples were spiked with possible interferents and 20 replicates were tested alongside 20 replicates of the corresponding blank matrix control. The following substances showed no interference up to the concentrations indicated:
| Substance Tested | Test Concentration |
|---|---|
| Hemoglobin | ≤ 500 mg/dL |
| Bilirubin (unconjugated) | ≤ 20 mg/dL |
| Bilirubin (conjugated) | ≤ 2 mg/dL |
| Intralipid | ≤ 500 mg/dL |
| Unfractionated Heparin | ≤ 2 IU/mL |
| Low Molecular Weight Heparin | ≤ 2 IU/mL |
- . Dabigatran, rivaroxaban, and fondaparinux do not interfere with the interpretation of cryocheck Hex LA results but may increase the delta correction of LA positive samples.
- . Elevated factor VIII activity (up to 180%) does not interfere with CRYOcheck Hex LA.
- Elevated fibrinogen concentrations do not interfere with the interpretation of CRYOcheck Hex ● LA results but may increase the delta correction of LA positive samples.
- . C-reactive protein does not interfere with the interpretation of CRYOcheck Hex LA results but at concentrations above 15 uq/mL may increase the delta correction of LA positive samples.
- Factor VIII inhibitor antibodies do not interfere with the interpretation of CRYOcheck Hex LA results, but at titers above 15 BU/mL may increase the delta correction of LA positive samples.
- Plasma samples with elevated INR (up to 4.5) do not interfere with the interpretation of . cryocheck Hex LA results.
- High platelet counts (>10,000 platelets/μL) showed interference with cRYOcheck Hex LA . results when compared with platelet poor (<10.000/ µL, single centrifuged) or platelet free (double centrifuged) plasma samples from the same donors.
- . Abnormally low factor II activities (below 50%) may interfere with the interpretation of CRYOcheck Hex LA, potentially resulting in false negative results for weakly LA positive plasmas.
- . Factor VII and factor IX deficiencies do not interfere with CRYOcheck Hex LA.
- Abnormally low factor X activities (below 50%) do not interfere with the interpretation of ● CRYOcheck Hex LA results but may increase the delta correction for LA positive samples.
Method Comparison Studies
A method comparison study was conducted to assess the efficacy of CRYOcheck Hex LA in the qualitative detection of LA relative to a comparator assay, Staclot® LA. A total of 446 samples were included in the study: 124 known (previously characterized) LA positive samples, 75 normal (presumed LA negative) samples from individuals with other medical conditions including autoimmune disorders and 220 LA target screening population samples. The study was conducted at one internal and three external sites. Each site performed the investigational device assay on their assigned portion of the samples using a single lot of CRYOcheck Hex LA. One external site, acting as the central laboratory, performed the comparator device testing on all 446 samples using the Staclot LA assay on a STA-R Evolution. The data demonstrated positive
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percent agreement of 95.6% (95% Cl, 91-98%), negative percent agreement of 95.2% (95% Cl, 92%-97%), and overall agreement of 95.3% (95% Cl, 93%-97%) as summarized below.
| CRYOcheck Hex LA results | ||||
|---|---|---|---|---|
| Comparator device results | Negative | Positive | Total | |
| Comparator device results | Negative | 295 | 15 | 310 |
| Positive | 6 | 130 | 136 | |
| Total | 301 | 145 | 446 | |
| Agreement | Point Estimate (95% Confidence Interval) | |||
| Positive Percent Agreement | 95.6% (91% - 98%) | |||
| Negative Percent Agreement | 95.2% (92% - 97%) |
Sample Integrity
95.3% (93% - 97%)
A sample integrity study was conducted to assess sample stability of fresh samples at room temperature, when stored at ≤ -70 ℃ and after up to two freeze-thaw cycles. Sixty-four samples were measured with a single lot of CRYOcheck Hex LA. Results were compared using regression analysis and support a fresh sample stability claim of 4 hours at room temperature and a frozen storage claim of 2 months at ≤ -70 ℃, including one freeze-thaw cycle.
Conclusion
The performance testing results demonstrate that CRYOcheck Hex LA is substantially equivalent to the predicate device, Staclot LA (K923731), and that the assay is effective for its labeled intended use.
Overall Agreement
§ 864.7925 Partial thromboplastin time tests.
(a)
Identification. A partial thromboplastin time test is a device used for primary screening for coagulation abnormalities, for evaluation of the effect of therapy on procoagulant disorders, and as an assay for coagulation factor deficiencies of the intrinsic coagulation pathway.(b)
Classification. Class II (performance standards).