Immunoassay for the in vitro quantitative determination of N terminal pro Brain natriuretic peptide in human serum and plasma. This assay is used as an aid in the diagnosis of individuals suspected of having heart failure. The test is further indicated for the risk stratification of patients with acute coronary syndrome and heart failure. The test may also serve as an aid in the assessment of increased risk of cardiovascular events and mortality in patients at risk for heart failure who have stable coronary artery disease.

The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

Device Description

Elecsys proBNP II (updated assay) is a second-generation assay by Roche Diagnostics for the in vitro quantitative determination of N-terminal pro-Brain natriuretic peptide (NT-proBNP) in human serum and plasma with increased biotin tolerance. The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

The cobas e family of analyzers employs the electrochemiluminescence immunoassay "ECLIA" technology. The assays are an 18-minute (Elecsys proBNP II) and 9 minute (Elecsys proBNP II STAT) application following a sandwich principle using two monoclonal antibodies which are specifically directed against NT-proBNP.

AI/ML Overview

Acceptance Criteria and Device Performance Study for Elecsys proBNP II and Elecsys proBNP II STAT Assays

The document describes the analytical performance studies conducted for the Elecsys proBNP II and Elecsys proBNP II STAT assays, which are in vitro diagnostic devices. These assays are intended for the quantitative determination of N-terminal pro-Brain natriuretic peptide (NT-proBNP) in human serum and plasma, aiding in the diagnosis of heart failure, risk stratification, and assessment of cardiovascular event risk. The assays have been modified to include increased biotin tolerance.

The studies aim to demonstrate that the updated assays meet predetermined acceptance criteria for various analytical parameters, ensuring their safety and effectiveness and substantial equivalence to their predicate devices.

1. Table of Acceptance Criteria and Reported Device Performance

The provided document doesn't explicitly list "acceptance criteria" alongside "reported device performance" in a single, dedicated table for all parameters. However, the "Conclusion" sections for each analytical study implicitly state whether the acceptance criteria were met. Based on the provided text, a table can be constructed as follows:

Acceptance Criteria Category	Specific Metric	Predetermined Acceptance Criterion (Implicitly Met)	Reported Device Performance (Elecsys proBNP II)	Reported Device Performance (Elecsys proBNP II STAT)
Analytical Sensitivity	Limit of Blank (LoB)	≤ 8 pg/mL	All lots met ≤ 8 pg/mL	All lots met ≤ 8 pg/mL
	Limit of Detection (LoD)	≤ 10 pg/mL	All lots met ≤ 10 pg/mL	All lots met ≤ 10 pg/mL
	Limit of Quantitation (LoQ) (Intermediate precision 20% CV)	≤ 36 pg/mL (Inferred from reported LoQ values)	32.7 - 35.7 pg/mL	7.28 - 13.6 pg/mL
Precision	Repeatability (Within-run precision)	Low CV% (Specific numerical thresholds not stated)	See tables on pages 8-9 for detailed CV	See tables on pages 9-10 for detailed CV
	Intermediate Precision (Within-laboratory precision)	Low CV% (Specific numerical thresholds not stated)	See tables on pages 8-9 for detailed CV	See tables on pages 9-10 for detailed CV
	Inter-Instrument Variability (Inter-laboratory precision)	Low CV% (Specific numerical thresholds not stated)	See table on page 10 for detailed CV	See table on page 11 for detailed CV
Linearity/Reportable Range	Measurements across claimed measuring range are linear	Not explicitly quantified, but demonstrated	Linearity data on page 15	Linearity data on page 15
Interference	Bilirubin (Conjugated & Unconjugated)	No interference up to 25.0 mg/dL	No interference up to 25.0 mg/dL	No interference up to 25.0 mg/dL
	Hemoglobin	No interference up to 1000 mg/dL	No interference up to 1000 mg/dL	No interference up to 1000 mg/dL
	Lipemia	No interference up to 1500 mg/dL	No interference up to 1500 mg/dL	No interference up to 1500 mg/dL
	Biotin	No interference up to 3500 ng/mL	No interference up to 5000 ng/mL	No interference up to 5000 ng/mL
	Rheumatoid Factor	No interference up to 1500 IU/mL	No interference demonstrated	No interference demonstrated
	Albumin	No interference up to 7 g/dL	No interference demonstrated	No interference demonstrated
Cross-Reactivity	Absence of significant cross-reactivity with various substances	Recovery % close to 100% (Implied)	See tables on pages 19-20	See tables on pages 19-20
Exogenous Interference	No interference with listed common and special therapeutic drugs	No significant interference (Implied)	No interference seen with tested drugs	No interference seen with tested drugs
Matrix Comparisons	Equivalence between Serum, Li-Heparin, and K2-EDTA plasma	Slope close to 1, Intercept close to 0, High 'r'	Slope 0.990-1.01, Intercept -0.985-0.898, r≥0.998	Not explicitly detailed for STAT
Method Comparison	Substantial equivalence to predicate device	High Pearson's r, Passing-Bablok Slope close to 1, Intercept close to 0	Pearson's r ≥ 0.999, Slope 0.98, Intercept -2.88	Pearson's r ≥ 0.999, Slope 1.01, Intercept -1.60

2. Sample Sizes and Data Provenance

Test Set Sample Sizes:
- Precision (Repeatability & Intermediate): 8 human serum samples and 2 controls (n=10 total) for each assay, measured in 84 determinations over 21 operating days. The exact number of individual patient samples contained within the "8 human serum samples" is not specified but appears to be 8 distinct pools/matrices.
- Precision (Inter-Instrument): 8 native human serum sample pools and 2 quality control levels (n=10 total) for each assay, with 75 determinations per sample/QC level (due to 5 days, 3 laboratories, 25 determinations/site, 5x5=25, 3 sites x 25 = 75 total reported).
- Analytical Sensitivity (LoB): 60 determinations of an analyte-free sample.
- Analytical Sensitivity (LoD): 5 low-level human serum samples, with 60 determinations per reagent lot.
- Analytical Sensitivity (LoQ): 9 native, unaltered serum samples for Elecsys proBNP II and 10 native, unaltered serum samples for Elecsys proBNP II STAT, each tested with 25 measured values.
- Linearity/Assay Reportable Range: One high analyte human, native serum sample diluted to 11 concentrations.
- Endogenous Interference (Bilirubin, Hemoglobin, Lipemia, Biotin, Rheumatoid Factor, Albumin): Three different analyte concentration levels (low, medium, high) in human native serum samples. Specific number of samples at each level not explicitly stated but implied to be several.
- Cross-Reactivity: Two human native serum samples (low and high analyte levels) spiked with potential cross-reactants.
- Exogenous Interference (Drugs): Two human native serum samples (low and high analyte concentrations).
- Matrix Comparisons: Single donor samples drawn into Serum (reference), Li-Heparin (98 pairs), and K2-EDTA plasma (111 pairs).
- Method Comparison: 1928 subjects for Elecsys proBNP II STAT and 1940 subjects for Elecsys proBNP II.
Data Provenance: The document generally refers to "human serum samples" and "native human serum samples" without specifying the country of origin. The studies are described as internal (e.g., "one internal site" for precision). There is no explicit mention of the data being retrospective or prospective, but the nature of in vitro diagnostic device performance studies (analytical validation) typically involves prospective testing of samples under controlled laboratory conditions, simulating diagnostic use.

3. Number of Experts and Qualifications for Ground Truth

The document does not describe the use of human experts to establish "ground truth" for the test set in the context of diagnostic interpretation (e.g., radiologists, cardiologists). This document details the analytical performance of an in vitro diagnostic assay, not an AI/imaging diagnostic device that requires expert adjudication of images. The "ground truth" in this context refers to the true analytical concentration of NT-proBNP in the samples, established through well-defined laboratory methodologies and traceable standards, or comparison to a previously cleared predicate device.

4. Adjudication Method for the Test Set

Not applicable. As this is an analytical validation of an in vitro diagnostic assay, there is no "adjudication method" in the sense of multiple human readers or experts resolving discrepancies in diagnostic interpretation. The methods involve quantitative analytical measurements of biochemical markers.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

Not applicable. This is not an imaging or AI-assisted diagnostic device that would typically undergo an MRMC study. The study focuses on the analytical performance of a laboratory immunoassay.

6. Standalone (Algorithm Only) Performance

Not applicable in the typical sense for medical imaging AI. The "algorithm" here is the chemical reaction and measurement process of the immunoassay itself. The analytical performance metrics (precision, sensitivity, linearity, interference, matrix comparison) are effectively the "standalone performance" of the device. The method comparison study directly compares the performance of the updated device against its predicate (older version) without human intervention in the result generation.

7. Type of Ground Truth Used

The ground truth used for these analytical studies is primarily measured analytical concentration, established through:

Reference materials: Calibrators and controls with known concentrations.
Spiking experiments: Adding known amounts of analyte or interfering substances to samples.
Dilutions: Creating samples with predictable concentrations.
Comparison to predicate device: The method comparison studies compare the new device's results against the results from the previously cleared Elecsys proBNP II and Elecsys proBNP II STAT (older versions) as the reference.
Consensus laboratory methods/standards: Studies like LoB, LoD, LoQ, and precision follow CLSI (Clinical and Laboratory Standards Institute) guidelines, which are established consensus standards for analytical validation.

There is no mention of pathology, clinical outcomes data, or expert consensus interpretation of results as a primary ground truth in this analytical performance section.

8. Sample Size for the Training Set

Not applicable. This document describes the analytical validation of laboratory assays, not a machine learning model that requires a "training set" in the same sense. The assay works based on established biochemical principles (electrochemiluminescence immunoassay, ECLIA, utilizing specific antibodies) and wet-lab procedures, not on learned patterns from a large dataset. Reagent formulation and process optimization might involve internal development data, but it's not a "training set" in the AI/ML context.

9. How the Ground Truth for the Training Set was Established

Not applicable, as there is no "training set" for an AI/ML model in this context. The "ground truth" for the development of the assay itself would be based on fundamental analytical chemistry principles, extensive laboratory testing, control materials, and established reference methods for quantifying NT-proBNP.

Summary

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Image /page/0/Picture/0 description: The image shows the logo of the U.S. Food and Drug Administration (FDA). The logo consists of two parts: a symbol on the left and the FDA name on the right. The symbol on the left is a stylized image of a human figure, while the FDA name on the right is written in blue letters. The words "U.S. FOOD & DRUG ADMINISTRATION" are written in a clear, sans-serif font.

March 31, 2022

Roche Diagnostics Jane Phillips Sr. Regulatory Program Manager 9115 Hague Road Indianapolis, Indiana 46250

Re: K210546

Trade/Device Name: Elecsys proBNP II, Elecsys proBNP II STAT Regulation Number: 21 CFR 862.1117 Regulation Name: B-Type Natriuretic Peptide Test System Regulatory Class: Class II Product Code: NBC Dated: December 8, 2021 Received: December 8, 2021

Dear Jane Phillips:

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.

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Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 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.

Marianela Perez-Torres, Ph.D. Deputy Director Division of Chemistry and Toxicology 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) K210546

Device Name Elecsys proBNP II

Indications for Use (Describe)

The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

Type of Use (Select one or both, as applicable)
X Prescription Use (Part 21 CFR 801 Subpart D)	Over-The-Counter Use (21 CFR 801 Subpart C)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.

DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

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"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

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Indications for Use

510(k) Number (if known) K210546

Device Name Elecsys proBNP II STAT

Indications for Use (Describe)

Immunoassay for the in vitro quantitative determination of Nterminal proBrain natruretic peptide in human serum and plasma. This assay is used as an aid in the diagnosis of individuals suspected of having heart failure. The test is further indicated for the risk stratification of patients with acute coronary syndrome and heart failure. The test may also serve as an aid in the assessment of increased risk of cardiovascular events and mortality in patients at risk for heart failure who have stable coronary artery disease.

The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

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)

CONTINUE ON A SEPARATE PAGE IF NEEDED.

This section applies only to requirements of the Paperwork Reduction Act of 1995.

DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.

Department of Health and Human Services Food and Drug Administration Office of Chief Information Officer Paperwork Reduction Act (PRA) Staff PRAStaff(@fda.hhs.gov

"An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number."

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510(k) Summary

March 28th, 2022

U.S. Food and Drug Administration Center for Devices and Radiological Heath Document Mail Center - WO66 Room 0609 10903 New Hampshire Avenue Silver Spring, MD 20993-0002

Purpose In accordance with 21 CFR 807.87, Roche Diagnostics Corporation hereby submits official notification as required by Section 510(k) of the Federal Food, Drug and Cosmetics Act of our intention to market the device described in this Premarket Notification [510(k)].

510(k) Summary K210546

Device Name	Proprietary name:	Elecsys proBNP II and Elecsys proBNP II STAT
	Common name:	proBNP II and proBNP II STAT

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Owner	Roche Diagnostics9115 Hague RoadIndianapolis, IN 46250Phone: 317-521-2000Fax: 317-521-1413
Contact	Jane Phillips, PhD
Date	March 28th, 2022

Panel	ProductCode	Classification Name	Regulationn Citation
Clinical Chemistry	NBC	B-Type NatriureticPeptide Test System	862.1117

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Substantial The Elecsys proBNP II (updated assay) is substantially Equivalence equivalent to the Elecsys proBNP II (old assay, K072437) The Elecsys proBNP II (updated assay) STAT is substantially equivalent to the Elecsys proBNP II STAT (old assav, K092649) Intended

Use

Elecsys proBNP II STAT

Immunoassay for the in vitro quantitative determination of N-terminal proBrain natriuretic peptide in human serum and plasma. This assay is used as an aid in the diagnosis of individuals suspected of having heart failure. The test is further indicated for the risk stratification of patients with acute coronary syndrome and heart failure. The test may also serve as an aid in the assessment of increased risk of cardiovascular events and mortality in patients at risk for heart failure who have stable coronary artery disease. The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

Elecsvs proBNP II

Immunoassay for the in vitro quantitative determination of N-terminal pro-Brain natriuretic peptide in human serum and plasma. This assay is used as an aid in the diagnosis of individuals suspected of having heart failure. The test is further indicated for the risk stratification of patients with acute coronary syndrome and heart failure. The test may also serve as an aid in the assessment of increased risk of cardiovascular events and mortality in patients at risk for heart failure who have stable coronary artery disease. The electrochemiluminescence immunoassay "ECLIA" is intended for use on cobas e immunoassay analyzers.

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ltem	Elecsys proBNP II(old assay design)	Elecsys proBNP II(updated assay design)	Change description
Proprietary name	Elecsys proBNP II	Elecsys proBNP II	None
Technology	ECLIA	ECLIA	None
Test format	Sandwich	Sandwich	None
Test type	Quantitative	Quantitative	None
Assay protocol	R1 + R2 + sample, incubation, +beads, incubation	R1 + R2 + sample, incubation, +beads, incubation	None
Measuring Range	5-35000 pg/ml	36-35000 pg/ml	Changing the lowerend of MR from LoD toLoQ
Biotin Tolerance	Up to 30 ng/mL	Up to 3500 ng/mL	Increase of biotintolerance

Table 1: Similarities and Differences between the Elecsys proBNP II assays

Table 2: Similarities and Differences between the Elecsys proBNP STAT assays

Item	Elecsys proBNP II(old assay design)	Elecsys proBNP II(updated assay design)	Change description
Proprietary name	Elecsys proBNP II STAT	Elecsys proBNP II STAT	None
Technology	ECLIA	ECLIA	None
Test format	Sandwich	Sandwich	None
Test type	Quantitative	Quantitative	None
Assay protocol	R1 + R2 + sample+ beads,incubation	R1 + R2 + sample+ beads,incubation	None
Measuring Range	5-35000 pg/ml	36-35000 pg/ml	Changing the lowerend of MR from LoD toLoQ
Biotin Tolerance	Up to 30 ng/mL	Up to 3500 ng/mL	Increase of biotintolerance

Device Description

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Changes to Reagent Composition

For the neutralization of free biotin in serum and plasma, Roche developed an antibody, which binds to free biotin. The antibodies are specific for free biotin and do not bind to or interact with the biotin-linker conjugates.

Analytical Performance

Precision/Reproducibility

Repeatability and Intermediate Precision

Precision measurements were conducted to evaluate repeatability (within-run precision) and the intermediate precision (within-laboratory precision) according to the CLSI guideline EP5-A3.

Methods:

The precision of the Elecsys proBNP II assay was evaluated on the cobas e 411 and proBNP II STAT assay was evaluated on one cobas e 601 analyzer at one internal site with three reagent lots over 21 days.

The protocol consisted of testing the eight serum samples and two controls in single determinations in four separate aliquots (divided into two runs per day) for 21 operating days (n=84). Repeatability and Intermediate Precision were calculated according to EP05-A3.

Results:

cobas e 411 analyzer
	Repeatability				Intermediate precision
Sample(Serum)	Meanpg/mL	SDpg/mL	SD 95%UCLpg/mL	CV%	CV 95%UCL%	SDpg/mL	SD95%UCLpg/mL	CV%	CV 95%UCL%
Human serum 1	55.9	2.62	3.20	4.7	5.7	4.35	5.35	7.8	9.6
Human serum 2	129	3.07	3.76	2.4	2.9	7.40	9.61	5.7	7.5
Human serum 3	423	8.91	10.9	2.1	2.6	18.0	22.3	4.3	5.3
Human serum 4	925	23.0	28.1	2.5	3.0	44.3	55.6	4.8	6.0

Lot 391671: Elecsys proBNP II

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cobas e 411 analyzer
	Repeatability				Intermediate precision
Sample(Serum)	Meanpg/mL	SDpg/mL	SD 95%UCLpg/mL	CV%	CV 95%UCL%	SDpg/mL	SD95%UCLpg/mL	CV%	CV 95%UCL%
Human serum 5	1924	43.8	53.5	2.3	2.8	88.8	110	4.6	5.7
Human serum 6	15620	248	303	1.6	1.9	662	844	4.2	5.4
Human serum 7	33526	778	950	2.3	2.8	1591	2010	4.7	6.0
Human serum 8	337	5.11	6.24	1.5	1.8	11.4	14.4	3.4	4.3
PreciControlCardiac II 1	132	3.29	4.02	2.5	3.1	5.97	7.38	4.5	5.6
PreciControlCardiac II 2	4477	135	165	3.0	3.7	216	267	4.8	6.0

Lot 391674: Elecsys proBNP II STAT

cobas e 601 analyzer
	Repeatability				Intermediate precision
Sample(Serum)	Meanpg/mL	SDpg/mL	SD 95%UCL	CV%	CV 95%UCL	SDpg/mL	SD95%UCL	CV%	CV 95%UCL
Human serum 1	63.9	2.42	2.96	3.8	4.6	4.39	5.25	6.9	8.2
Human serum 2	144	3.36	4.10	2.3	2.8	5.87	7.08	4.1	4.9
Human serum 3	482	12.7	15.5	2.6	3.2	18.5	22.4	3.8	4.6
Human serum 4	1060	24.6	30.1	2.3	2.8	34.4	40.9	3.2	3.9
Human serum 5	2219	51.4	62.8	2.3	2.8	77.2	94.1	3.5	4.2
Human serum 6	18371	405	494	2.2	2.7	654	801	3.6	4.4
Human serum 7	33967	912	1114	2.7	3.3	1338	1608	3.9	4.7
Human serum 8	331	5.01	6.12	1.5	1.8	12.7	16.1	3.8	4.9
PreciControlCardiac II 1	155	4.06	4.97	2.6	3.2	6.08	7.27	3.9	4.7
PreciControlCardiac II 2	5660	116	142	2.1	2.5	194	237	3.4	4.2

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Inter-Instrument Variability (CLSI EP5-A3)

Methods (Inter-Instrument):

This was done using the Elecsys proBNP II (updated assay) on three cobas e 411 analyzers and using the Elecsys proBNP II STAT (updated assay) on three cobas e 601 analyzers according to the precision model described in CLSI EP5-A3 guideline. One reagent lot of the updated assay was measured on 5 days in three laboratory sites.

A precision experiment according to the CLSI EP05-A3 setup (5-day model, 25 determinations total per sample pool, reagent lot, and site) was conducted at all 3 laboratories, for each of the assays. The sample panel was identical for both and consisted of 8 native human serum sample pools, and 2 concentration levels made of quality control material (PreciControl Cardiac II Level 1 and 2). The samples were not spiked.

Results:

Sample	N	Mean	SD	SD UCL	CV[%]	CV[%]UCL
HS 1	75	55.2	1.02	2.06	1.8	3.7
HS 2	75	131	3.46	5.84	2.6	4.5
HS 3	75	332	5.50	10.3	1.7	3.1
HS 4	75	471	10.3	18.0	2.2	3.8
HS 5	75	963	13.2	27.1	1.4	2.8
HS 6	75	1773	20.0	41.1	1.1	2.3
HS 7	75	18,719	0.00	N/A	0.00	N/A
HS 8	75	31,425	0.00	N/A	0.00	N/A
PC 1	75	144	1.89	3.79	1.3	2.6
PC 2	75	4799	0.00	N/A	0.00	N/A

Inter-Instrument Precision, proBNP II on the e411. Units = pg/mL

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Sample	N	Mean	SD	SD UCL	CV[%]	CV[%] UCL
HS 1	75	52.1	3.91	6.45	7.5	12.4
HS 2	75	128	9.22	15.2	7.2	11.8
HS 3	75	317	23.8	38.9	7.5	12.3
HS 4	75	465	34.6	56.5	7.4	12.1
HS 5	75	961	74.3	122.7	7.7	12.7
HS 6	75	1845	134	220	7.3	11.9
HS 7	75	18,477	1466	2396	7.9	13.0
HS 8	75	30,089	2394	3914	8.0	13.0
PC 1	75	145	10.9	17.9	7.5	12.3
PC 2	75	5328	399	652	7.5	12.2

Inter-Instrument Precision, proBNP II STAT on the e601. Units = pg/mL

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Analytical Sensitivity

The analytical studies to establish the limit of blank (LoB), limit of detection (LoD) and limit of quantitation (LoQ) were conducted according to the experimental design described in CLSI EP-17-A2

Limit of Blank (LoB) (CLSI EP17-A2)

LoB of the Elecsys proBNP II (updated assay) on the cobas e 411 analyzer and of Elecsys proBNP II STAT (updated assay) on the cobas e 601 was determined according to CLSI EP17-A2. Limit of Blank determines the highest observed measurement values for samples free of analyte. The Limit of Blank was determined as the 95th percentile of measurements of blank samples.

Methods:

In total 60 determinations of an analyte-free sample were obtained on one instrument over ≥ three days in 6 runs with 10-fold determination per run. Three lots of reagent were used in the experimental design.

As the analyzer does not report negative sample concentrations, the data set was truncated and the data were evaluated as the linear interpolation of the 57th ranked observation.

Conclusion:

All lots met the predetermined acceptance criterion of ≤ 8 pg/mL.

Limit of Detection (LoD) (CLSI EP17-A2)

LoD of the Elecsys proBNP II (updated assay) on the cobas e 411 analyzer and of Elecsys proBNP II STAT (updated assay) on the cobas e 601 analyzer was determined according to CLSI EP17-A2. The LoD determines the lower limit for samples with analyte. The LoD was determined as the lowest amount of analyte in a sample that can be detected with a 95% probability.

Methods:

Five low-level human serum samples were measured on one instrument over ≥ three days in 6 runs with a two-fold determination per run. In total sixty determinations per reagent lot. The experiment was conducted using three reagent lots.

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A pooled estimate of the precision (SD total) for the 5 low level samples was calculated.

LoD was calculated according to EP17-A2, chapter 5.3.3.2 as:

LoD = LoB + 1.653 x SD total (of low analyte samples)

Conclusion:

All lots met the predetermined acceptance criterion of ≤ 10 pg/mL. The LoD claim in the labeling will be set to ≤ 10 pg/mL.

Limit of Quantitation (CLSI EP17-A2)

The LoQ of the Elecsys proBNP II (updated assay) on the cobas e 411 analyzer and of the Elecsys proBNP II STAT (updated assay) on the cobas e 601 analyzer was determined according to CLSI Guideline EP17-A2.

The LoQ was determined as the lowest concentration of analyte which can be reproducibly measured with an intermediate precision of 20% CV.

Methods:

A low level sample set 9 native, unaltered serum samples using the Elecsys proBNP II on the cobas e 411 analyzer and 10 native, unaltered serum samples using the Elecsys proBNP II STAT on the cobas e 601 analyzer of known measurand concentration was tested in 5 replicates (aliquots) on one instrument in singleton per day, over 5 days with 1 run per day. Three lots of reagent were used in the LoQ experiment. A total of n=25 measured values were obtained for each sample. The mean value and the intermediate precision as coefficient of variation (CV) and standard deviation (SD) were calculated for each LoQ sample.

LoQ is defined as the mean value of that sample which is the first that fulfills the specification for the intermediate precision and for which no sample with higher concentration exists that exceeds this specification.

Data are summarized in the following tables, below.

Reagent Lot	LoQ [pg/mL]
1 391671	33.7
2 391672	32.7
3 391673	35.7

LoQ Results for Elecsys proBNP II

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LoQ Results for proBNP II STAT

Reagent Lot	LoQ [pg/mL]
1	391674	8.98
2	391675	13.6
3	391676	7.28

Linearity/Assav Reportable Range (CLSI EP06-Ed2)

The linearity study was conducted to demonstrate that measurements across the claimed measuring range for each parameter are linear. The study was performed according to CLSI guideline EP06-Ed2.

Methods:

One high analyte human, native serum sample above measuring rage was diluted with analyte free serum. 11 concentrations were prepared throughout the entire measuring range. Samples were assayed in 3-fold determination within a single run. The linearity data were analyzed as described below.

For each sample and its dilution levels a weighted least square regression by pooled variance is performed.

As one high analyte human, native serum sample above measuring range was diluted with analyte free serum (CLSI Design A1), a regression without intercept is chosen.

As variance increases with concentration, weighted linear regression is used.

As only 3 replicates are available at each dilution step, the weights are computed based on the pooled variance including also the replicates of the next higher as well as the next lower dilution step.

The result table is composed for each experiment, i.e. for the proBNP II (18min) on e411 and the proBNP II STAT on e601. The resulting linearity statistics and deviation results are given in the tables below (rounded data).

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Level	Rel Conc	Mean Conc	ExpectedConc.	Predicted Conc.	Deviation	Deviation (%)
1	0	8.417	9.911	8.424	-0.007	-0.084
2	0.001	22.792	25.371	21.567	1.225	5.681
3	0.002	55.455	63.428	53.916	1.538	2.853
4	0.004	132.273	158.569	134.791	-2.518	-1.868
5	0.01	310.973	396.423	336.977	-26.004	-7.717
6	0.026	780.863	1014.843	862.660	-81.798	-9.482
7	0.064	1939.433	2537.108	2156.651	-217.217	-10.072
8	0.16	4823.735	6342.77	5391.626	-567.891	-10.533
9	0.4	13053.629	15856.925	13479.066	-425.437	-3.156
10	0.7	24009.672	27749.618	23588.365	421.307	1.786
11	1	39642.311	39642.311	33697.665	5944.647	17.641

Linearity for Elecsys proBNP II

Linearity for Elecsys proBNP II STAT

Level	Rel Conc	Mean Conc	ExpectedConc.	Predicted Conc.	Deviation	Deviation (%)
1	0	8.515	10.024	9.720	-1.206	-12.403
2	0.001	22.355	25.662	24.884	-2.529	-10.164
3	0.002	55.493	64.156	62.210	-6.717	-10.797
4	0.004	137.301	160.39	155.524	-18.224	-11.717
5	0.01	328.36	400.975	388.810	-60.450	-15.547
6	0.026	828.107	1026.495	995.354	-167.248	-16.803
7	0.064	2066.792	2566.238	2488.386	-421.593	-16.942
8	0.16	5351.93	6415.594	6220.964	-869.034	-13.969
9	0.4	14054.029	16038.985	15552.410	-1498.381	-9.634
10	0.7	25238.171	28068.223	27216.718	-1978.547	-7.270
11	1	40097.462	40097.462	38881.026	1216.436	3.129

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Endogenous Interference Studies

The purpose of these studies was to evaluate endogenous substances for potential interference with the parameters measured on the cobas e 411 for Elecsys proBNP II (updated assay) and on the cobas e 601 for Elecsys proBNP II STAT (updated assay).

Method for Bilirubin, Hemoglobin and Lipemia:

The effect on quantitation of analyte in the presence of endogenous interfering substances using the Elecsys proBNP II was determined on the cobas e 411 and the Elecsys proBNP II STAT was determined on the cobas e 601.

Endogenous interferences were determined by testing three different analyte concentration levels (low about 130 pg/mL, medium about 900 pg/mL, high about 20000 pg/mL) in human native serum samples. The high concentrations were spiked with recombinant human proBNP for the bilirubin and lipemia testing.

One aliquot of each serum sample was spiked with the interfering substance (= interference pool) and another aliquot was spiked (if applicable) with the same volume of the solvent of the interfering substance (= dilution pool). The interfering pool was then dilution pool in 10 % increments. The recovery for each sample was calculated by comparison to the reference (unspiked sample).

Interfering substance	No interference up to
Conjugated Bilirubin	25.0 mg/dL
Unconjugated Bilirubin	25.0 mg/dL
Hemoglobin	1000 mg/dL
Lipemia	1500 mg/dL

Results/Conclusion for both proBNP II and proBNP II STAT:

Method for Biotin:

The effect on quantitation of analyte in the presence of Biotin using the Elecsys proBNP II was determined on the cobas e 411 and the Elecsys proBNP II STAT was determined on the cobas e 601.

Biotin interferences were determined by testing three different analyte concentration levels (low about 125 pg/mL, medium about 800 pg/mL, high about 18000 pg/mL) in human native serum samples.

One aliquot of each serum sample was spiked with 15000 ng/mL Biotin (= interference pool) and another aliquot was spiked (if applicable) with the same volume of the solvent of the interfering substance (= dilution pool). The interfering pool was then diluted into the dilution

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pool in 11 steps. The recovery for each sample was calculated by comparison to the reference (unspiked sample).

Results:

Volume (Relation)		Measuredconcentrationof analytepg/mL	Expectedconcentrationof analytepg/mL	Concentrationof interferentng/mL	Recovery%
Sample 1a	Sample 1b
100	0.0000	123	123	0.0000	100
96.7	3.33	124	123	500	101
93.3	6.67	127	123	1000	103
90.0	10.0	126	123	1500	102
86.7	13.3	126	123	2000	102
83.3	16.7	127	123	2500	103
80.0	20.0	127	123	3000	103
76.0	24.0	125	123	3600	102
66.7	33.3	116	123	5000	93.9
33.3	66.7	85.1	123	10000	69.1
0.0000	100	46.9	123	15000	38.1

Biotin: Elecsys proBNP II

Conclusion:

No interference was seen up to 5000 ng/mL biotin. All predetermined acceptance criteria were met. The claimed Biotin concentration at which no interference is observed is 3500 ng/mL.

Method for Rheumatoid Factor:

The effect on quantitation of analyte in the presence of Rheumatoid Factors using the Elecsys proBNP II was determined on the cobas e 411 and the Elecsys proBNP II STAT was determined on the cobas e 601.

Rheumatoid Factors interferences were determined by testing three different analyte concentration levels (low about 130 pg/mL, medium about 900 pg/mL, high about 20000 pg/mL) in human native serum samples. The high sample was spiked with recombinant human NTproBNP.

One aliquot of each serum sample was spiked with 1500 IU/mL Rheumatoid Factors

(= interference pool) and another aliquot was spiked (if applicable) with the same volume of the solvent of the interfering substance (= dilution pool). The interfering pool was then diluted into the dilution pool in 10 % increments. The recovery for each sample was calculated by comparison to the reference (unspiked sample).

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The claimed Rheumatoid factors concentration is 1500 IU/mL.

Method for Albumin:

The effect on quantitation of analyte in the presence of Albumin using the Elecsys proBNP II was determined on the cobas e 411 and the Elecsys proBNP II STAT was determined on the cobas e 601.

Albumin interferences were determined by testing three different analyte concentration levels (low about 140 pg/mL, medium about 1000 pg/mL, high about 23000 pg/mL) in human native serum samples. The high sample was spiked with human recombinant proBNP.

One aliquot of each serum sample was spiked with 7 g/dL Albumin (= interference pool) and another aliquot was spiked (if applicable) with the same volume of the solvent of the interfering substance (= dilution pool). The interfering pool was then dilution pool in 10 % increments. The recovery for each sample was calculated by comparison to the reference (unspiked sample).

The claimed Albumin concentration is 7 g/dL.

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Cross-Reactivity

This study was conducted to evaluate the Elecsys proBNP II and proBNP II STAT assay on the cobas e 411 and 601, respectively, for potential cross-reactivity.

Methods

To determine the analytical specificity of the Elecsys proBNP II (updated assay) and Elecsys proBNP II STAT (updated assay), two human native serum samples with low (about 150 pg/mL) and high (about 2500 pg/mL) analyte levels were aliquoted and spiked with potential crossreactants. One aliquot was left unspiked to serve as a reference. Samples were measured on the cobas e 411 analyzer for Elecsys proBNP II and on the cobas e 601 analyzer for Elecsys proBNP II STAT and cross reactivity was calculated according to the formula:

x percent cross reaction =

100 x simulated analyte conc. conc. of cross-reactant spiked

	Concentrationreactant	Low analyte level			High analyte level
Cross-reactant		Measuredanalyteconcentrationwithout Crossreactant[pg/mL]	MeasuredAnalyteconcentrationwith Crossreactant[pg/mL]	Recoverywith x-reactant[%]	Measuredanalyteconcentrationwithout Crossreactant[pg/mL]	MeasuredAnalyteconcentrationwith Crossreactant[pg/mL]	Recoverywith x-reactant[%]
Adrenomedullin	1.0 ng/mL	143	130	91	2655	2573	97
Angiotensin I	0.6 ng/mL	145	144	99	2640	2615	99
Angiotensin II	0.6 ng/mL	147	147	100	2657	2603	98
Angiotensin III	1.0 ng/mL	147	147	100	2692	2617	97
Arg-Vasopressin	1.0 ng/mL	139	139	100	2659	2647	100
Endothelin	20 pg/mL	148	148	100	2680	2593	97
Aldosterone	0.6 ng/mL	151	146	97	2655	2541	96
Renin	50 ng/mL	144	136	94	2671	2498	94
BNP32	3.5 µg/mL	140	140	100	2655	2646	100
CNP22	2.2 µg/mL	143	130	91	2579	2467	96
Urodilatin	3.5 µg/mL	141	142	100	2682	2668	99
ANP 1-28	3.1 µg/mL	141	142	101	2668	2642	99

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	Concentrationreactant	Low analyte level			High analyte level
Cross-reactant		Measuredanalyteconcentrationwithout Crossreactant[pg/mL]	MeasuredAnalyteconcentrationwith Crossreactant[pg/mL]	Recoverywith x-reactant[%]	Measuredanalyteconcentrationwithout Crossreactant[pg/mL]	MeasuredAnalyteconcentrationwith Crossreactant[pg/mL]	Recoverywith x-reactant[%]
NT-proANP(1-30)[preproANP (26-55)]	3.5 µg/mL	146	144	99	2677	2568	96
NT-proANP (31-67) [preproANP56-92]	1.0 ng/mL	146	141	97	2694	2622	97
NT-proANP (79-98) [preproANP104-123]	1.0 ng/mL	146	146	100	3077	3181	103

Exogenous Interference - Drugs

The purpose of this study was to evaluate drugs for potential interference with the parameters measured on the cobas e 411 analyzer for Elecsys proBNP II (updated assay) and on the cobas e 601 analyzer for Elecsys proBNP II STAT (updated assay).

Methods:

Two human native serum samples with approximately 125 pg/mL and 2000 pg/mL were analyzed using the cobas e 411 and cobas e 601 analyzer. These samples were divided into an appropriate number of aliquots. One aliquot of each serum sample was spiked with the respective amount (volume) of the drug (= interference sample) and another aliquot was spiked (if applicable) with the same volume of the solvent of the respective drug (= reference sample). The recovery of the interference sample was calculated as percent recovery compared to the reference sample.

In total. 18 common and 33 special pharmaceutic compounds were analyzed and measured.

Potentially Interfering Drugs and Test Concentrations

Common therapeutic drugs	Drug concentration[mg/L]
Acetylcystein	150
Ampicillin-Na	1000
Ascorbic acid	300
Cefoxitin	2500

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Common therapeutic drugs	Drug concentration
	[mg/L]
Heparin	5000 IU/L
Levodopa	20
Methyldopa	20
Metronidazole	200
Doxycycline	50
Acetylsalicylic Acid	1000
Rifampicin	60
Cyclosporine	5
Phenylbutazone	400
Acetaminophen	200
Ibuprofen	500
Theophylline	100
Intralipid	10000
Ca-Dobesilate	200

Special therapeutic drugs	Drug concentration[mg/L]
Carvedilol	150
Clopidogrel	75
Digoxin	0.5
Epinephrine	0.37
Insulin	0.84
Lidocaine	100
Lisinopril	40
Methylprednisolone	80
Metoprolol	15
Nifedipine	60
Marcumar	6
Propafenone	900
Reteplase	1.12
Simvastin	40
Spirolactone	400
Tolbutamide	3000
Torasemide	200

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Special therapeutic drugs	Drug concentration[mg/L]
Verapamil	120
Propanolol	0.32
Enalapril	40
Captopril	50
Gentamycin	500
Lovostatin	80
Pravastatin	40
Bisoprolol	10
Glycerol nitrate	192
Molsidormin	24
Nicardipin	90
Streptokinase	300 IE/mL
Urokinase	600 IE/mL
Digitoxin	0.3
Sotalol	320
Low molecular weight heparin	18

No interference was seen with the drugs tested.

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Matrix Comparisons

Methods:

The effect on quantitation of analyte in the presence of anticoagulants with the Elecsys proBNP II (updated assay) was determined on the cobas e 411 and with the Elecsys proBNP II STAT (updated assay) was determined on the cobas e 601 by comparing values obtained from native samples (single donors) drawn into Serum, Li-Heparin and K2-EDTA plasma.

Results:

Sample Matrix Comparison for Elecsys proBNP II

Elecsys proBNP II	Li Heparin	K2-EDTA
Slope[95% LCL/UCL]	0.990[0.981/0.997]	1.01[0.994/1.01]
Intercept (pg/mL)[95% LCL/UCL]	0.898[-0.981/3.45]	-0.985[-4.23/2.21]
Correlation coefficientPearson's r	0.998	0.999
Absolute Bias at 125 pg/mL	-0.391	-0.189
% Bias at 125 pg/mL[95% LCL/UCL]	-0.3[-1.5/1.3]	-0.2[-2.3/2.1]
Serum/plasma pairs	98	111

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Method comparison

We performed a method comparison study with 1928 subjects with the biotin remediated Elecsys proBNP II STAT assay, 09315276160 (y) and the Elecsys proBNP II STAT assay, 05390109160 (x).

Image /page/24/Figure/2 description: This image is a scatter plot that compares two proBNP II STAT measurements, labeled (09315276160) and (05390109160). The plot includes a Passing-Bablok regression fit with the equation -1.60 + 1.01 * x, where n=1928. An identity line is also plotted for reference. The Pearson's r correlation coefficient is 0.999, indicating a strong positive correlation between the two measurements.

Passing Bablok Regression Fit

Correlation	Estimate
Kendall'stau	0.99
Pearson's r	1.00

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Passing-Bablok Regression				Predicted Relative Bias (%)
TestMethod	Comparison Method	N	Intercept(95% CI)	Slope (95%CI)	125 pg/mL(95% CI)	300 pg/mL(95% CI)	450 pg/mL(95% CI)	900 pg/mL(95% CI)	1800pg/mL(95% CI)
Biotin-remediatedElecsysproBNP IISTAT(09315276160)	ElecsysproBNP IISTAT(05390109160)	1,928	-1.60 (-2.09,-1.16)	1.01(1.00,1.01)	-0.8 (-1.0,-0.5)	-0.0 (-0.2,0.2)	0.2 (-0.1,0.4)	0.3(0.1,0.6)	0.4(0.2,0.7)

Regression results and relative bias at selected NT-proBNP concentration levels (all, STAT)

We performed a method comparison study with 1940 subjects with the biotin remediated Elecsys proBNP II assay, 09315268160 (y) and the Elecsys proBNP II assay, 04842464160 (x).

Image /page/25/Figure/3 description: This image is a scatter plot that compares two proBNP II measurements, labeled as (09315268160) on the y-axis and (04842464160) on the x-axis. The plot includes a Passing-Bablok regression fit, represented by the equation -2.88 + 0.98 * x, with n=1940 data points. An identity line is also plotted for reference. The Pearson's r correlation coefficient is 0.999, indicating a strong positive correlation between the two measurements.

Passing Bablok Regression Fit

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Correlation	Estimate
Kendall'stau	0.99
Pearson's r	1.00

Regression results and relative bias at selected NT-proBNP concentration levels (all, 18min)

Passing-Bablok Regression				Predicted Relative Bias (%)
TestMethod	ComparisonMethod	N	Intercept(95% CI)	Slope (95%CI)	125 pg/mL(95% CI)	300 pg/mL(95% CI)	450 pg/mL(95% CI)	900 pg/mL(95% CI)	1800pg/mL(95% CI)
Biotin-remediatedElecsysproBNP II(09315268160)	ElecsysproBNP II(04842464160)	1,940	-2.88 (-3.20,-2.46)	0.98(0.98,0.98)	-4.2 (-4.4,-3.9)	-2.8 (-3.0,-2.7)	-2.5 (-2.6,-2.4)	-2.2 (-2.3,-2.1)	-2.0 (-2.2,-1.9)

Conclusion: Testing demonstrated that the Elecsys proBNP II and Elecsys proBNP II STAT assays are safe and effective and are substantially equivalent to the assays cleared in K072437 and K092649. Biotin tolerance has been improved for patient safety.

Regulation Number and Section

§ 862.1117 B-type natriuretic peptide test system.

(a)
Identification. The B-type natriuretic peptide (BNP) test system is an in vitro diagnostic device intended to measure BNP in whole blood and plasma. Measurements of BNP are used as an aid in the diagnosis of patients with congestive heart failure.(b)
Classification. Class II (special controls). The special control is “Class II Special Control Guidance Document for B-Type Natriuretic Peptide Premarket Notifications; Final Guidance for Industry and FDA Reviewers.”