Predicate Devices

Reference Devices

AI/ML (Artificial Intelligence / Machine Learning)

No
The device description and performance studies focus on a standard immunoassay technology and statistical analysis of the results, with no mention of AI or ML algorithms.

Therapeutic

No
The device is an in vitro diagnostic assay used to quantify NT-proBNP levels, which aids in the diagnosis of heart failure. It does not provide any treatment or therapeutic intervention.

Diagnostic

Yes

The intended use explicitly states that the device is "used as an aid in the diagnosis of heart failure (HF)".

SaMD (Software as a Medical Device)

No

The device is a chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination of NT-proBNP in human serum and plasma. This involves physical reagents and laboratory equipment, not just software.

IVD (In Vitro Diagnostic)

Yes, this device is an IVD (In Vitro Diagnostic).

Here's why:

Intended Use / Indications for Use: The document explicitly states the assay is used for the "in vitro quantitative determination" of NT-proBNP in human serum and plasma. "In vitro" means "in glass" or "outside the body," which is a key characteristic of IVDs. It also states its use as an "aid in the diagnosis of heart failure," which is a diagnostic purpose.
Device Description: The description details a laboratory-based assay using human serum and plasma samples, involving chemical reactions and measurement of light units. This is consistent with the nature of an IVD.
Clinical Performance Studies: The document describes studies using human samples (serum and plasma) to evaluate the performance of the assay in a clinical setting (emergency department) for diagnosing heart failure. This is a standard part of the validation process for IVDs.

The entire context of the document, from the intended use to the performance studies, clearly indicates that this device is designed to be used outside of the human body to analyze biological samples for diagnostic purposes, which is the definition of an In Vitro Diagnostic device.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

The Alere NT-proBNP for Alinity i assay is a chemiluminescent microparticle immunoassay (CMIA) used for the in vitro quantitative determination of N-terminal pro B-type natriuretic peptide (NT-proBNP) in human serum and plasma on the Alinity i system.

In the emergency department, measurements of NT-proBNP are used as an aid in the diagnosis of heart failure (HF) in patients with clinical suspicion of new onset or worsening HF.

Product codes

NBC

Device Description

The Alere NT-proBNP for Alinity i assay is an automated, two-step immunoassay for the in vitro quantitative determination of NT-proBNP in human serum and plasma using chemiluminescent microparticle immunoassay (CMIA) technology.

Sample and anti-NT-proBNP coated paramagnetic microparticles are combined and incubated. The NT-proBNP present in the sample binds to the anti-NT-proBNP coated microparticles. The mixture is washed. Anti-NT-proBNP acridinium-labeled conjugate is added to create a reaction mixture and incubated. Following a wash cycle, Pre-Trigger and Trigger Solutions are added. The resulting chemiluminescent reaction is measured as a relative light unit (RLU). There is a direct relationship between the amount of NT-proBNP in the sample and the RLU detected by the system optics.

Mentions image processing

Not Found

Mentions AI, DNN, or ML

Not Found

Input Imaging Modality

Not Found

Anatomical Site

Not Found

Indicated Patient Age Range

18 years and older

Intended User / Care Setting

Emergency department

Description of the training set, sample size, data source, and annotation protocol

Not Found

Description of the test set, sample size, data source, and annotation protocol

A multi-center prospective study including 17 collection sites across the US was conducted to establish the performance characteristics of the Alere NT-proBNP for Alinity i assay. Subjects 18 years and older presenting to the ED with signs and symptoms consistent with a clinical suspicion of new onset or acute exacerbation of HF were included in the study. Subjects with renal insufficiency requiring dialysis or known estimated glomerular filtration rate (eGFR) 75 years old: Female 75 years).
Age-independent negative cutoff: 300 pg/mL.

All Subjects (N=2127)

Positive Predictive Value (Posttest Probability of HF for Positive result): 75.2% (95% CI: 72.3, 77.8) for all ages.
Negative Predictive Value (Posttest Probability of Non-HF for Negative result): 94.0% (95% CI: 92.2, 95.4) for all ages.
Grayzone: Posttest probability of HF: 35.6% (95% CI: 30.7, 40.8). Posttest probability of Non-HF: 64.4% (95% CI: 59.2, 69.3).
Likelihood Ratio (HF): Positive: 4.29; Grayzone: 0.78; Negative: 0.09.

Subgroup Analysis:

History of HF vs. No History of HF: Patients with a history of HF had a higher false positive rate (36.4%) compared to those without (12.3%).
**eGFR

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.”

Summary

0

Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). On the left is the Department of Health & Human Services logo. To the right of that is the FDA logo in blue, with the words "U.S. FOOD & DRUG" stacked on top of the word "ADMINISTRATION".

January 16, 2025

Axis-Shield Diagnostics Limited Claire Dora Associate Director, Regulatory Affairs 17 Luna Place, The Technology Park Dundee. Scotland DD 1XA United Kingdom

Re: K241176

Trade/Device Name: Alere NT-proBNP for Alinity i Reagent Kit Regulation Number: 21 CFR 862.1117 Regulation Name: B-Type Natriuretic Peptide Test System Regulatory Class: Class II Product Code: NBC Dated: December 11, 2024 Received: December 11, 2024

Dear Claire Dora:

We have reviewed your section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (the Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database available at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Additional information about changes that may require a new premarket notification are provided in the FDA guidance documents entitled "Deciding When to Submit a 510(k) for a Change to an Existing Device"

1

(https://www.fda.gov/media/99812/download) and "Deciding When to Submit a 510(k) for a Software Change to an Existing Device" (https://www.fda.gov/media/99785/download).

Your device is also subject to, among other requirements, the Quality System (QS) regulation (21 CFR Part 820), which includes, but is not limited to, 21 CFR 820.30. Design controls; 21 CFR 820.90. Nonconforming product; and 21 CFR 820.100, Corrective and preventive action. Please note that regardless of whether a change requires premarket review, the OS regulation requires device manufacturers to review and approve changes to device design and production (21 CFR 820.30 and 21 CFR 820.70) and document changes and approvals in the device master record (21 CFR 820.181).

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR Part 803) for devices or postmarketing safety reporting (21 CFR Part 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safetyreporting-combination-products); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR Part 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR Parts 1000-1050.

All medical devices, including Class I and unclassified devices and combination product device constituent parts are required to be in compliance with the final Unique Device Identification System rule ("UDI Rue"). The UDI Rule requires, among other things, that a device bear a unique device identifier (UDI) on its label and package (21 CFR 801.20(a)) unless an exception or alternative applies (21 CFR 801.20(b)) and that the dates on the device label be formatted in accordance with 21 CFR 801.18. The UDI Rule (21 CFR 830.300(a) and 830.320(b)) also requires that certain information be submitted to the Global Unique Device Identification Database (GUDID) (21 CFR Part 830 Subpart E). For additional information on these requirements, please see the UDI System webpage at https://www.fda.gov/medical-device-advicecomprehensive-regulatory-assistance/unique-device-identification-system-udi-system.

Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems.

2

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,

Paula V. Caposino -S

Paula Caposino, Ph.D. Division Deputy Director Division of Chemistry and Toxicology Devices OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

3

Indications for Use

510(k) Number (if known) K241176

Device Name Alere NT-proBNP for Alinity i

The Alere NT-proBNP for Alinity i assay is a chemiluminescent microparticle immunoassay (CMIA) used for the in vitro quantitative determination of N-terminal pro B-type natriuretic peptide (NT-proBNP) in human serum and plasma on the Alinity i system.

In the emergency departments of NT-proBNP are used as an aid in the diagnosis of heart failure (HF) in patients with clinical suspicion of new onset or worsening HF.

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.

The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to:

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."

Indications for Use (Describe)

4

Administrative Documentation - 510(k) Summary

This summary of the 510(k) safety and effectiveness information is being submitted in accordance with the requirements of SMDA 1990 and 21 CFR §807.92.

I. 510(k) Number

K241176

II. Applicant Name

Axis-Shield Diagnostics Limited Luna Place, The Technology Park, Dundee DD2 1XA United Kingdom

Primary contact:

Claire Dora, Associate Director, Regulatory Affairs Axis-Shield Diagnostics Ltd Phone: +44(0)790994098 Email: claire.dora@abbott.com

Date Summary Prepared: January 16, 2025

III. Device Name

Trade Name: Alere NT-proBNP for Alinity i

Alere NT-proBNP for Alinity i Device Classification: Class II Classification Name: BNP Test System Governing Regulation: 21 CFR Part 862.1117 Product Code: NBC

5

IV. Predicate Device

Roche Elecsys proBNP II (K092649)

V. Description of Device

A. Reagents

The various configurations of the Alere NT-proBNP for Alinity i Reagent Kit are described below.

	List Number
	04S7921	04S7931
Tests per cartridge	100	500
Number of cartridges per kit	2	2
Tests per kit	200	1000
Microparticle volume	6.7 mL	27.0 mL
Conjugate volume	6.1 mL	26.5 mL

Microparticles: Anti-NT-proBNP (sheep, monoclonal) coated microparticles in Bis-TRIS buffer with protein (bovine) stabilizer, non-specific binding blocking agents, and surfactant. Minimum concentration: 0.05% solids. Preservative: sodium azide.

Conjugate: Anti-NT-proBNP (mouse, monoclonal) acridinium-labeled conjugate in MES buffer with protein (bovine) stabilizer and surfactant. Minimum concentration: 0.12 ug/mL. Preservatives: antimicrobial agents.

B. Biological Principles of the Procedure

The Alere NT-proBNP for Alinity i assay is an automated, two-step immunoassay for the in vitro quantitative determination of NT-proBNP in human serum and plasma using chemiluminescent microparticle immunoassay (CMIA) technology.

Sample and anti-NT-proBNP coated paramagnetic microparticles are combined and incubated. The NT-proBNP present in the sample binds to the anti-NT-proBNP coated microparticles. The mixture is washed. Anti-NT-proBNP acridinium-labeled conjugate is added to create a reaction mixture and incubated. Following a wash cycle,

Pre-Trigger and Trigger Solutions are added.

6

The resulting chemiluminescent reaction is measured as a relative light unit (RLU). There is a direct relationship between the amount of NT-proBNP in the sample and the RLU detected by the system optics.

Assay Standardization

There is currently no international recognized reference method or reference material for standardization. The Alere NT-proBNP for Alinity i Calibrators are traceable to internal reference standards at every concentration level that were value assigned on and made traceable to the predicate device by transference, as described by Clinical and Laboratory Standards Institute (CLSI) Document EP32-R.*

C. Interpretation of Results

Emergency Department

For patients presenting to the emergency department (ED) with clinical suspicion of heart failure (HF), the results should be interpreted as indicated in the table below.

| Age Group

(Years)	NT-proBNP
	(pg/mL)	(pmol/L)	Interpretation
All	75	≥ 300.0 to 75	≥ 1800.0

" Clinical and Laboratory Standards Institute (CLSI). Metrological Traceability and Its Implementation; A Report. CLSI document EP32-R. Wayne, PA: CLSI; 2006.

7

VI. Intended Use of the Device

The Alere NT-proBNP for Alinity i assay is a chemiluminescent microparticle immunoassay (CMIA) used for the in vitro quantitative determination of N-terminal pro B-type natriuretic peptide (NT-proBNP) in human serum and plasma on the Alinity i system.

In the emergency department, measurements of NT-proBNP are used as an aid in the diagnosis of heart failure (HF) in patients with clinical suspicion of new onset or worsening HF.

VII. Comparison with the Predicate Device

The similarities and differences between the subject device and the predicate device are presented in the table below.

| | Subject Device:
Alere NT-proBNP for Alinity i | Predicate Device:
Roche Elecsys proBNP II (K092649) |
|----------------------------------------|----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| | General Device Characteristic Similarities | |
| Intended
Use/Indications
for Use | Immunoassay used for the in vitro
quantitative determination of N-terminal
pro B-type natriuretic peptide
(NT-proBNP) in human serum and
plasma.
In the emergency department,
measurements of NT-proBNP are used
as an aid in the diagnosis of heart failure
(HF) in patients with clinical suspicion
of new onset or worsening HF. | 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 congestive
heart failure.
The test is further indicated for the risk
stratification of patients with acute coronary
syndrome and congestive 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. |
| Specimen Type | Human serum and plasma | Same |
| Test Principle | Sandwich immunoassay | Same |
| Measurement
Type | Quantitative | Same |
| Analyte | NT-proBNP | Same |

8

| | Subject Device:
Alere NT-proBNP for Alinity i | Predicate Device:
Roche Elecsys proBNP II (K092649) |
|----------------------------|------------------------------------------------------------------------------------------------------------------------------|---------------------------------------------------------------|
| Detection
Technology | Chemiluminescence | Same |
| General Device Differences | | |
| Principle of
Operation | CMIA | Electro chemiluminescent microparticle
immunoassay (ECLIA) |
| Intended Use
Setting | Emergency department | Not specified |
| Measuring
Interval | Analytical Measuring Interval (AMI):
15.8 to 35,000.0 pg/mL
(1.9 to 4130.0 pmoL/L) | 5 to 35,000 pg/mL |
| Hook Effect | No hook effect up to 372,620 pg/mL | No hook effect up to 300,000 pg/mL |
| Results
Interpretation | Negative: HF unlikely for all patients:
75 ≥ 300.0 to 75 ≥ 1800.0 | |

9

VIII. Summary of Nonclinical Performance

A. Reportable Interval

Based on representative data for the limit of quantitation (LoQ), the ranges over which results can be reported are provided below.

	pg/mL	pmol/L
Analytical Measuring
Interval (AMI)a	15.8 - 35,000.0	1.9 - 4130.0
Extended Measuring
Interval (EMI)b	35,000.0 - 350,000.0	4130.0 - 41,300.0
Reportable Intervalc	15.8 - 350,000.0	1.9 - 41,300.0

ª AMI: The AMI extends from the LoQ to the upper limit of quantitation (ULoQ). This is determined by the range of values in pg/mL (pmol/L) that demonstrated acceptable performance for linearity, imprecision, and bias.

b EMI: The EMI extends from the ULoQ to the ULoQ × dilution factor. The value reflects a 1:10 dilution factor.

^ The reportable interval extends from the LoQ to the upper limit of the EMI.

B. Within-Laboratory Precision (20-Day)

A study was performed based on guidance from CLSI EP05-A3. * Testing was conducted using 3 lots of the Alere NT-proBNP for Alinity i reagents, 1 lot of the Alere NT-proBNP for Alinity i Calibrators, 1 lot of the Alere NT-proBNP for Alinity i Controls, and 1 instrument. Three controls and 8 human plasma panels (7 native panels and 1 panel supplemented with NT-proBNP analyte) were tested in 2 replicates at 2 separate times per day on 20 days.

Clinical and Laboratory Standards Institute (CLS). Evaluation of Quantitative Measurement Procedures; Approved Guideline—Third Edition. CLSI Document EP05-A3. Wayne, PA: CLSI; 2014.

10

| | | Mean | Repeatability | | Within-Laboratorya | | Between-Lot | | Overall Within-
Laboratoryb | |
|---------------------------|-----|----------|---------------|-----|--------------------|-----|-------------|-----|--------------------------------|------|
| Sample | n | (pg/mL) | SD | %CV | SD | %CV | SD | %CV | SD | %CV |
| Low Control | 240 | 143.0 | 6.40 | 4.5 | 7.63 | 5.3 | 4.42 | 3.1 | 8.82 | 6.2 |
| Medium Control | 240 | 503.4 | 16.26 | 3.2 | 18.21 | 3.6 | 9.29 | 1.8 | 20.44 | 4.1 |
| High Control | 240 | 5144.2 | 177.02 | 3.4 | 202.18 | 3.9 | 21.47 | 0.4 | 203.32 | 4.0 |
| Panel A | 240 | 18.8 | 1.39 | 7.4 | 1.72 | 9.2 | 0.75 | 4.0 | 1.88 | 10.0 |
| Panel B | 240 | 55.6 | 2.64 | 4.7 | 3.02 | 5.4 | 1.60 | 2.9 | 3.41 | 6.1 |
| Panel C | 240 | 130.4 | 4.28 | 3.3 | 5.44 | 4.2 | 4.87 | 3.7 | 7.30 | 5.6 |
| Panel D | 240 | 449.1 | 13.10 | 2.9 | 18.68 | 4.2 | 14.02 | 3.1 | 23.36 | 5.2 |
| Panel E | 240 | 1011.4 | 28.31 | 2.8 | 34.49 | 3.4 | 21.92 | 2.2 | 40.87 | 4.0 |
| Panel F | 240 | 1938.0 | 54.02 | 2.8 | 65.28 | 3.4 | 22.56 | 1.2 | 69.07 | 3.6 |
| Panel G | 240 | 14,939.8 | 417.56 | 2.8 | 518.88 | 3.5 | 296.61 | 2.0 | 597.67 | 4.0 |
| Panel H
(Supplemented) | 240 | 31,711.0 | 989.02 | 3.1 | 1185.11 | 3.7 | 2118.19 | 6.7 | 2427.19 | 7.7 |

ª Includes repeatability (within-run), between-run, and between-day variability.

b Includes repeatability (within-run), between-run, between-day, and between-lot variability.

11

C. Linearity

A study was performed based on guidance from CLSI EP06, 2nd ed. This assay is linear across the AMI of 15.8 to 35,000.0 pg/mL (1.9 to 4130.0 pmol/L).

D. Lower Limits of Measurement

A study was performed based on guidance from CLSI EP17-A2. T Testing was conducted using 3 lots of the Alere NT-proBNP for Alinity i reagents on each of 2 instruments over a minimum of 3 days. The maximum observed limit of blank (LoB), limit of detection (LoD), and limit of quantitation (LoQ) values are summarized below.

	pg/mL	pmol/L
LoBa	0.1	0.0
LoDb	3.6	0.4
LoQc	15.8	1.9

a The LoB represents the 95th percentile from n ≥ 60 replicates of zero-analyte samples.

b The LoD represents the lowest concentration at which the analyte can be detected with 95% probability based on n ≥ 60 replicates of low-analyte level samples.

6 The LoQ is defined as the lowest concentration at which a maximum allowable precision of 20 %CV was met and was determined from n ≥ 60 replicates of low-analyte level samples.

E. Analytical Specificity

1. Potentially Interfering Endogenous Substances

A study was performed based on guidance from CLSI EP07, 3rd ed. + Each substance was tested at 2 levels of the analyte (approximately 125 pg/mL and 2000 pg/mL).

Clinical and Laboratory Standards Institute (CLSI). Evaluation of the Linearity of Quantitative Measurement Procedures. 2nd ed. CLSI Guideline EP06. Wayne, PA: CLSI; 2020.

T Clinical and Laboratory Standards Institute (CLSI). Evaluation of Detection Capability for Clinical Measurements; Approved Guideline-Second Edition. CLSI Document EP17-A2. Wayne, PA: CLSI; 2012.

Clinical and Laboratory Standards Institute (CLSI). Interference Testing in Clinical Chemistry. 3rd ed. CLSI Guideline EP07. Wayne, PA: CLSI; 2018.

12

No significant interference (interference within ± 10.0%) was observed at the following concentrations.

No Significant Interference (Interference within ± 10.0%)
Potentially Interfering Substance	Interferent Level
Bilirubin (conjugated)	60 mg/dL
Bilirubin (unconjugated)	60 mg/dL
Biotin	4250 ng/mL
Cholesterol	700 mg/dL
Human Anti-mouse Antibodies (HAMA)	1500 ng/mL
Hemoglobin	1 g/dL
IgG	6 g/dL
Intralipid	3000 mg/dL
Rheumatoid Factor (RF)	600 IU/mL
Total Protein	12.6 g/dL

Interference beyond ± 10.0% (based on 95% confidence interval [CI]) was

observed at the concentrations shown below for the following substances.

Interference beyond ± 10.0% (based on 95% CI)
Potentially Interfering
Substance	Interferent Level	Analyte Level	% Interference
(95% CI)
RF	1520 IU/mL	125 pg/mL	-8.9%
(-10.4%, -7.5%)
	1520 IU/mL	2000 pg/mL	-11.4%
(-12.4%, -10.4%)
Total Protein	15.2 g/dL	125 pg/mL	-12.7%
(-14.7%, -10.7%)
	15.5 g/dL	2000 pg/mL	-9.9%
(-11.4%, -8.5%)

13

2. Potentially Interfering Drugs

A study was performed based on guidance from CLSI EP07, 3rd ed. * Each substance was tested at 2 levels of the analyte (approximately 125 pg/mL and 2000 pg/mL).

No significant interference (interference within ± 10.0%) was observed at the following concentrations.

No Significant Interference (Interference within ± 10.0%)
Potentially Interfering
Drug	Interferent Level	Potentially Interfering
Drug	Interferent Level
Acetaminophen	15.6 mg/dL	Lidocaine	8.0 mg/dL
Acetylsalicylic Acid	60 mg/dL	Lisinopril	1.6 mg/dL
Allopurinol	6.0 mg/dL	Lithium	4.20 mg/dL
Amikacin	15 mg/dL	Losartan potassium	5.99 mg/dL
Amiodarone	4.2 mg/dL	Lovastatin	2.0 mg/dL
Amlodipine Besylate	0.4 mg/dL	L-Thyroxine	0.06 mg/dL
Ampicillin	7.5 mg/dL	Methyldopa	2.5 mg/dL
Ascorbic acid	5.25 mg/dL	Methylprednisolone	0.75 mg/dL
Atenolol	1.0 mg/dL	Metoprolol	1.5 mg/dL
Atorvastatin	32 mg/dL	Milrinone	0.183 mg/dL
Caffeine	10.8 mg/dL	Naproxen	49.97 mg/dL
Captopril	5.0 mg/dL	Nicotine	0.1 mg/dL
Carbamazepine	4.5 mg/dL	Nicotinic acid	4.0 mg/dL
Carvedilol	3.75 mg/dL	Nifedipine	6.0 mg/dL
Chloramphenicol	7.8 mg/dL	Nitrofurantoin	4.0 mg/dL
Chlordiazepoxide	1.0 mg/dL	Nitroglycerin	0.016 mg/dL
Chlorpromazine	0.33 mg/dL	Oxazepam	1.2 mg/dL
Cimetidine	3.0 mg/dL	Oxytetracycline	10 mg/dL
No Significant Interference (Interference within ± 10.0%)
Potentially Interfering
Drug	Interferent Level	Potentially Interfering
Drug	Interferent Level
Cinnarizine	3.1 mg/dL	Penicillin G	25 U/mL
Clopidogrel bisulfate	7.5 mg/dL	Pentobarbital	12.6 mg/dL
Creatinine	30 mg/dL	Phenobarbitol	69 mg/dL
Cyclosporine A	0.4 mg/dL	Phenprocoumon
(Marcumar)	1.53 mg/dL
Dextran 40	6000 mg/dL	Phenytoin	6.0 mg/dL
Diazepam	3.0 mg/dL	Primidone	5.7 mg/dL
Diclofenac	6.0 mg/dL	Probenecid	60 mg/dL
Digoxin	0.025 mg/dL	Procainamide	4.8 mg/dL
Diltiazem	12 mg/dL	Propafenone	30 mg/dL
Dipyridamole	8.0 mg/dL	Propanolol	0.2 mg/dL
Disopyramide	4.0 mg/dL	Propoxyphene	0.321 mg/dL
Dobutamine	10 mg/dL	Quinidine	2.0 mg/dL
Dopamine	65 mg/dL	Ramipril	0.6 mg/dL
Enalapril Maleate	1.6 mg/dL	Retaplase	3.33 mg/dL
Epinephrine	0.05 mg/dL	Simvastatin	3.2 mg/dL
Erythromycin	13.8 mg/dL	Spironolactone	7.5 mg/dL
Ethanol	600 mg/dL	Sulfamethoxazole	112 mg/dL
Ethosuximide	30 mg/dL	Theophylline	6.0 mg/dL
Fenofibrate	4.5 mg/dL	Tolbutamide	150 mg/dL
Furosemide	6.0 mg/dL	Torasemide	1.5 mg/dL
Gentamicin	12 mg/dL	Trandolapril	4.0 mg/dL
Heparin	8 U/mL	Trasylol/Aprotinin	501.8 KIE/mL
Hydralazine	2.0 mg/dL	Trimethoprim	6.4 mg/dL
Hydrochlorothiazide	2.0 mg/dL	Uric Acid	23.5 mg/dL
Insulin	0.16 mg/dL	Valproic Acid	50 mg/dL
Ibuprofen	50 mg/dL	Verapamil	24 mg/dL
No Significant Interference (Interference within ± 10.0%)
Potentially Interfering
Drug	Interferent Level	Potentially Interfering
Drug	Interferent Level
Indomethacin	3.6 mg/dL	Warfarin	7.5 mg/dL
Isosorbide dinitrate	6.0 mg/dL

Clinical and Laboratory Standards Institute (CLSI). Interference Testing in Clinical Chemistry. 3rd ed. CLSI Guideline EP07. Wayne, PA: CLSI; 2018.

14

15

3. Cross-Reactants

A study was performed based on guidance from CLSI EP07, 3rd ed. * Samples with NT-proBNP target concentrations of 125 pg/mL and 2000 pg/mL containing the cross-reactants at the concentration listed below were tested with the Alere NT-proBNP for Alinity i assay on the Alinity i system. For each cross-reactant, the % recovery was calculated as: (NT-proBNP concentration with cross-reactant) / (NT-proBNP concentration without cross-reactant) × 100%. The observed % recovery of NT-proBNP was within 100% ± 10% for all cross-reactants evaluated at each analyte level.

Potential Cross-Reactant	Cross-Reactant Concentration
Adrenomedullin	1000 pg/mL
Aldosterone	600 pg/mL
Angiotensin I	600 pg/mL
Angiotensin II	600 pg/mL
Angiotensin III	1000 pg/mL
ANP 28	3100 ng/mL
Arg-Vasopressin	1000 pg/mL
BNP 32	3500 ng/mL
CNP 22	2200 ng/mL
Endothelin	20 pg/mL
NT-proANP 1-30 (preproANP26-55)	3500 ng/mL

Clinical and Laboratory Standards Institute (CLSI). Interference Testing in Clinical Chemistry. 3rd ed. CLSI Guideline EP07. Wayne, PA: CLSI; 2018.

16

Potential Cross-Reactant	Cross-Reactant Concentration
NT-proANP 31-67 (preproANP56-92)	1000 pg/mL
NT-proANP 79-98 (preproANP104-123)	1000 pg/mL
Renin	50,000 pg/mL
Urodilatin	3500 ng/mL

F. High Dose Hook

High dose hook effect was not observed on samples up to 372,620 pg/mL.

G. Expected Values (Reference Interval)

A reference interval study was performed based on guidance from CLSI EP28-A3c. Testing was performed with a US-based general population from apparently healthy individuals (≥ 18 years old). The specimens were tested with the Alere NT-proBNP for Alinity i assay on the Alinity i system. Based on the results, the 95% reference interval of an apparently healthy population for each sex and age range was determined to be as follows:

				Reference Interval
			Median	(2.5th to 97.5th Percentile)
Age Range	Sex	n	(pg/mL)	(pg/mL)
18 to 75 years old	Female	127	81.4	* Clinical and Laboratory Standards Institute (CLSI). Evaluation of Quantitative Measurement Procedures; Approved Guideline-Third Edition. CLSI Document EP05-A3. Wayne, PA: CLSI; 2014.

18

| Sample | n | Mean
(pg/mL) | Repeatability
SD | Repeatability
%CV | Within-
Laboratorya
SD | Within-
Laboratorya
%CV | Between-Site
SD | Between-Site
%CV | Between-Lot
SD | Between-Lot
%CV | Overall
Reproducibilityb
SD | Overall
Reproducibilityb
%CV |
|---------------------------|-----|-----------------|---------------------|----------------------|------------------------------|-------------------------------|--------------------|---------------------|-------------------|--------------------|-----------------------------------|------------------------------------|
| Low Control | 360 | 136.6 | 4.75 | 3.5 | 5.75 | 4.2 | 2.63 | 1.9 | 1.45 | 1.1 | 6.49 | 4.7 |
| Medium Control | 360 | 477.1 | 12.24 | 2.6 | 15.35 | 3.2 | 14.20 | 3.0 | 8.88 | 1.9 | 22.72 | 4.8 |
| High Control | 359 | 4932.2 | 201.01 | 4.1 | 311.08 | 6.3 | 105.17 | 2.1 | 0.00 | 0.0 | 328.38 | 6.7 |
| Panel 1 | 359 | 14.5 | 2.10c | 14.5 | 2.74c | 18.9 | 0.00 | 0.0 | 0.00 | 0.0 | 2.74c | 18.9 |
| Panel 2 | 360 | 51.9 | 2.14 | 4.1 | 3.07 | 5.9 | 0.49 | 0.9 | 0.00 | 0.0 | 3.11 | 6.0 |
| Panel 3 | 360 | 127.1 | 4.67 | 3.7 | 5.95 | 4.7 | 1.36 | 1.1 | 0.78 | 0.6 | 6.15 | 4.8 |
| Panel 4 | 360 | 436.9 | 10.46 | 2.4 | 18.67 | 4.3 | 3.18 | 0.7 | 0.00 | 0.0 | 18.94 | 4.3 |
| Panel 5 | 360 | 996.5 | 21.86 | 2.2 | 43.45 | 4.4 | 17.12 | 1.7 | 0.00 | 0.0 | 46.70 | 4.7 |
| Panel 6 | 360 | 1891.1 | 33.18 | 1.8 | 79.86 | 4.2 | 35.96 | 1.9 | 0.00 | 0.0 | 87.58 | 4.6 |
| Panel 7
(Supplemented) | 359 | 15,664.6 | 319.91 | 2.0 | 974.61 | 6.2 | 281.81 | 1.8 | 217.28 | 1.4 | 1037.54 | 6.6 |
| Panel 8
(Supplemented) | 360 | 25,630.5 | 601.42 | 2.3 | 1654.86 | 6.5 | 220.39 | 0.9 | 807.19 | 3.1 | 1854.37 | 7.2 |

Includes repeatability (within-run), between-run, and between-day variability. a

b Includes repeatability (within-run), between-run, between-site, and between-lot variability.

င An outlying run was observed. Based on guidance from CLSI EPO5-A3, a replacement run was performed and the results are shown in the preceding table. Without the replacement run, the repeatability SD was 9.5 pg/mL (1.12 pmol2), within-laboratory precision SD was 9.64 pg/mL (1.14 pmoVL), and the overall reproducibility SD was 9.64 pg/mL (1.14 pmol/L).

19

B. Clinical Performance

A multi-center prospective study including 17 collection sites across the US was conducted to establish the performance characteristics of the Alere NT-proBNP for Alinity i assay. Subjects 18 years and older presenting to the ED with signs and symptoms consistent with a clinical suspicion of new onset or acute exacerbation of HF were included in the study. Subjects with renal insufficiency requiring dialysis or known estimated glomerular filtration rate (eGFR) 75 | All | 75 | All |
| All Subjects | | | | | | | | |
| N | 160 | 553 | 167 | 880 | 300 | 799 | 148 | 1247 |
| Mean | 4354.5 | 6092.9 | 6891.3 | 5928.3 | 437.9 | 760.4 | 1738.8 | 798.9 |
| SD | 5454.61 | 8144.69 | 6860.32 | 7521.45 | 1686.57 | 1833.48 | 3494.96 | 2100.93 |
| Median | 2664.9 | 3297.2 | 4787.6 | 3420.7 | 70.1 | 198.9 | 565.9 | 181.8 |

20

| Adjudicated

Diagnosis	HF				Non-HF
Age Group
(Years)	75	All	75	All
Minimum	22.8	19.8	158.9	19.8	0.0	0.0	4.2	0.0
Maximum	38,880.9	60,716.9	38,938.4	60,716.9	24,737.1	17,988.9	31,154.0	31,154.0
Female Subjects
N	55	236	85	376	165	407	82	654
Mean	4034.7	5370.1	6814.2	5501.2	273.5	614.8	1320.8	617.2
SD	6302.82	7732.10	7042.32	7417.01	732.64	1543.13	2403.63	1556.92
Median	1851.5	2541.7	4787.6	2786.4	73.1	174.9	499.7	161.2
Minimum	22.8	24.9	158.9	22.8	0.0	0.9	25.7	0.0
Maximum	38,880.9	47,385.1	38,938.4	47,385.1	5633.9	14,849.5	18,827.7	18,827.7
Male Subjects
N	105	317	82	504	135	392	66	593
Mean	4522.0	6631.0	6971.2	6247.0	638.8	911.6	2258.0	999.4
SD	4977.78	8410.52	6708.84	7590.10	2369.85	2084.06	4464.35	2557.26
Median	2780.1	4066.5	4773.9	3904.3	64.1	221.0	701.8	203.2
Minimum	58.3	19.8	186.9	19.8	0.0	0.0	4.2	0.0
Maximum	26,986.3	60,716.9	32,286.4	60,716.9	24,737.1	17,988.9	31,154.0	31,154.0

The pretest probability of adjudicated HF (prevalence of adjudicated HF in the study), posttest probabilities, and likelihood ratios of the Alere NT-proBNP for Alinity i result vs adjudicated diagnosis (along with the 95% CIs) were determined based on guidance from CLSI EP12, 3rd ed., * for all subjects and by sex using the age-dependent positive cutoffs (450 pg/mL for subjects 18 to 75 years of age) and age-independent negative cutoff (300 pg/mL).

Clinical and Laboratory Standards Institute (CLSI). Evaluation of Qualitative, Binary Output Examination Performance. 3rd ed. CLSI Guideline EP12. Wayne, PA: CLSI; 2023.

21

			All Subjects
NT-proBNP
Result			Adjudicated
Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF	Total
Age Group: 18 to 75 Years

Pretest Probability of HF (Prevalence of HF in Study): 53.02% (167/315)

22

				All Subjects
	Adjudicated Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
NT-proBNP
Result	HF	Non-HF	Total
Positive	131	38	169	77.5
(131/169)
(70.6, 83.2)	-	3.06
(2.30, 4.06)
Grayzone	34	59	93	36.6
(34/93)
(27.5, 46.7)	63.4
(59/93)
(53.3, 72.5)	0.51
(0.36, 0.73)
Negative	2	51	53	-	96.2
(51/53)
(87.2, 99.0)	0.03
(0.01, 0.14)
Total	167	148	315
All Subjectsc
Pretest Probability of HF (Prevalence of HF in Study): 41.37% (880/2127)
Positive	708	234	942	75.2
(708/942)
(72.3, 77.8)	-	4.29
(3.80, 4.83)
Grayzone	121	219	340	35.6
(121/340)
(30.7, 40.8)	64.4
(219/340)
(59.2, 69.3)	0.78
(0.64, 0.96)
Negative	51	794	845	-	94.0
(794/845)
(92.2, 95.4)	0.09
(0.07, 0.12)
Total	880	1247	2127

The results for female subjects are presented in the following table.

23

			Female Subjects
NT-proBNP
Result	Adjudicated
Diagnosis		Total	Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF
Age Group: 18 to 75 Years
	Pretest Probability of HF (Prevalence of HF in Study): 50.90% (85/167)
Positive	64	19	83	77.1
(64/83)
(67.0, 84.8)	-	3.25
(2.15, 4.91)
Female Subjects
	Adjudicated
Diagnosis
NT-proBNP
Result	HF	Non-HF	Total	Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
Grayzone	20	34	54	37.0
(20/54)
(25.4, 50.4)	63.0
(34/54)
(49.6, 74.6)	0.57
(0.36, 0.90)
Negative	1	29	30	-	96.7
(29/30)
(83.3, 99.4)	0.03
(0.00, 0.24)
Total	85	82	167
All Female Subjectsc
Pretest Probability of HF (Prevalence of HF in Study): 36.50% (376/1030)
Positive	282	95	377	74.8
(282/377)
(70.2, 78.9)	-	5.16
(4.25, 6.27)
Grayzone	58	117	175	33.1
(58/175)
(26.6, 40.4)	66.9
(117/175)
(59.6, 73.4)	0.86
(0.65, 1.15)
Negative	36	442	478	-	92.5
(442/478)
(89.7, 94.5)	0.14
(0.10, 0.19)
Total	376	654	1030

Administrative Documentation – 510(k) Summary
Administrative Documentation – 510(k) Summary
Page 20 of 42

24

The results for male subjects are presented in the following table.

25

	Male Subjects
NT-proBNP
Result	Adjudicated
Diagnosis			Posttest
Probability of	Posttest
Probability of
Non-HF %	Likelihood
	HF	Non-
HF	Total	HF % (n/N)
(95% CI)a	(n/N)
(95% CI)a	Ratio (HF)
(95% CI)b
Age Group: 18 to 75 Years
				Pretest Probability of HF (Prevalence of HF in Study): 55.41% (82/148)
Positive	67	19	86	77.9
(67/86)	-	2.84
(1.92, 4.20)

(68.1, 85.4)

26

Male Subjects
NT-proBNP
Result	Adjudicated Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF	Total
Grayzone	14	25	39	35.9
(14/39)
(22.7, 51.6)	64.1
(25/39)
(48.4, 77.3)	0.45
(0.26, 0.80)
Negative	1	22	23	-	95.7
(22/23)
(79.0, 99.2)	0.04
(0.01, 0.26)
Total	82	66	148
All Male Subjectsc
Pretest Probability of HF (Prevalence of HF in Study): 45.94% (504/1097)
Positive	426	139	565	75.4
(426/565)
(71.7, 78.8)	-	3.61
(3.10, 4.19)
Grayzone	63	102	165	38.2
(63/165)
(31.1, 45.8)	61.8
(102/165)
(54.2, 68.9)	0.73
(0.54, 0.97)
Negative	15	352	367	-	95.9
(352/367)
(93.4, 97.5)	0.05
(0.03, 0.08)
Total	504	593	1097

The pretest probability of adjudicated HF (prevalence of adjudicated HF in the study), posttest probabilities, and likelihood ratios of the Alere NT-proBNP for Alinity i result vs adjudicated diagnosis (along with the 95% CIs) were also determined for relevant clinical subgroups using the age-dependent positive cutoffs and age-independent negative cutoff.

The results for subjects with a history of HF are presented in the following table.

27

			Subjects With History of HF
NT-proBNP
Result		Adjudicated
Diagnosis		Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF	Total
Age Group: 18 to 75 Years
	Pretest Probability of HF (Prevalence of HF in Study): 69.31% (131/189)
Positive	107	22	129	82.9	-	2.15
	Subjects With History of HF
		Adjudicated
Diagnosis		Posttest
Probability of	Posttest
Probability of
Non-HF %	Likelihood
NT-proBNP
Result	HF	Non-
HF	Total	HF % (n/N)
(95% CI)a	(n/N)
(95% CI)a	Ratio (HF)
(95% CI)b
Grayzone	24	21	45	53.3
(24/45)
(39.1, 67.1)	46.7
(21/45)
(32.9, 60.9)	0.51
(0.31, 0.83)
Negative	0	15	15	-	100.0
(15/15)
(79.6, 100.0)	0.01
(0.00, 0.24)
Total	131	58	189
All Subjects With History of HFc
Pretest Probability of HF (Prevalence of HF in Study): 66.93% (672/1004)
Positive	554	121	675	82.1
(554/675)
(79.0, 84.8)	-	2.26
(1.95, 2.62)
Grayzone	87	69	156	55.8
(87/156)
(47.9, 63.3)	44.2
(69/156)
(36.7, 52.1)	0.62
(0.47, 0.83)
Negative	31	142	173	-	82.1
(142/173)
(75.7, 87.1)	0.11
(0.07, 0.16)
Total	672	332	1004

(75.5, 88.5)

28

The results for subjects without a history of HF are presented in the following table.

29

			Subjects Without History of HF
NT-proBNP Result			Adjudicated Diagnosis	Total	Posttest Probability of HF % (n/N) (95% CI)a	Posttest Probability of Non-HF % (n/N) (95% CI)a	Likelihood Ratio (HF) (95% CI)b
Age Group: 18 to 75 Years
Pretest Probability of HF (Prevalence of HF in Study): 28.57% (36/126)
Positive	24	16	40	60.0
(24/40)	-	3.75
(2.27, 6.19)
Subjects Without History of HF
NT-proBNP Result	Adjudicated Diagnosis			Posttest Probability of HF % (n/N) (95% CI)a	Posttest Probability of Non-HF % (n/N) (95% CI)a	Likelihood Ratio (HF) (95% CI)b
	HF	Non-HF	Total
Grayzone	10	38	48	20.8 (10/48) (11.7, 34.3)	79.2 (38/48) (65.7, 88.3)	0.66 (0.37, 1.17)
Negative	2	36	38	-	94.7 (36/38) (82.7, 98.5)	0.14 (0.04, 0.55)
Total	36	90	126
All Subjects Without History of HFc
Pretest Probability of HF (Prevalence of HF in Study): 18.52% (208/1123)
Positive	154	113	267	57.7 (154/267) (51.7, 63.5)	-	6.00 (4.96, 7.25)
Grayzone	34	150	184	18.5 (34/184) (13.5, 24.7)	81.5 (150/184) (75.3, 86.5)	1.00 (0.71, 1.40)
Negative	20	652	672		97.0 (652/672) (95.4, 98.1)	0.13 (0.09, 0.21)
Total	208	915	1123

(44.6, 73.7)

30

Patients with a history of HF had a higher rate of false positives compared to patients without a history of HF. Of the 332 total patients with a history of HF and adjudicated as non-HF, 121 (36.4%) had NT-proBNP concentrations greater than or equal to agedependent cutoffs. Of the 915 total patients with no history of HF and adjudicated as non-HF, 113 (12.3%) had NT-proBNP concentrations greater than or equal to agedependent cutoffs. This difference is likely due to long-standing chronic elevation of NT-proBNP in patients with a previous diagnosis of HF. Results should always be

31

assessed in conjunction with patient's medical history, clinical examination, and other findings.

The results for subjects with an eGFR less than 60 mL/min/1.73 m² are presented in the following table. This group was comprised of 548 patients with an eGFR between 30 and less than 60 mL/min/1.73 m² and 80 patients with an eGFR less than 30 mL/min/1.73 m².

| | | | | Subjects With eGFR 75 Years
Pretest Probability of HF (Prevalence of HF in Study): 59.12% (94/159) | | | | | | |
| Positive | 78 | 22 | 100 | 78.0
(78/100)
(68.9, 85.0) | - | 2.45
(1.72, 3.49) |
| Grayzone | 16 | 22 | 38 | 42.1
(16/38)
(27.9, 57.8) | 57.9
(22/38)
(42.2, 72.1) | 0.50
(0.29, 0.88) |
| Negative | 0 | 21 | 21 | - | 100.0
(21/21)
(84.5, 100.0) | 0.02
(0.00, 0.26) |
| Total | 94 | 65 | 159 | | | |
| All Subjects With eGFR 75 Years

Pretest Probability of HF (Prevalence of HF in Study): 46.45% (72/155)

34

Patients with an eGFR less than 60 mL/min/1.73 m² had a higher rate of false positives compared to patients with an eGFR greater than or equal to 60 mL/min/1.73 m². Of the 258 total patients with an eGFR less than 60 mL/min/1.73 m² and adjudicated as non-HF, 85 (32.9%) had NT-proBNP concentrations greater than or equal to agedependent cutoffs. Of the 981 total patients with an eGFR greater than or equal to

35

60 mL/min/1.73 m² and adjudicated as non-HF, 149 (15.2%) had NT-proBNP concentrations greater than or equal to age-dependent cutoffs. Eight patients adjudicated as non-HF (0.6%) had an unknown eGFR. Patients with an eGFR less than 60 mL/min/1.73 m² had a higher rate of false negatives (9.0% [10/111]) compared to patients with an eGFR greater than or equal to 60 mL/min/1.73 m² (5.6% [41/726]). All 10 of the false negatives in the eGFR less than 60 mL/min/1.73 m² group were from patients with a BMI greater than or equal to 30 kg/m2. In patients with eGFR less than 60 mL/min/1.73 m², caution should be used when interpreting NT-proBNP results. Results should always be assessed in conjunction with the patient's medical history, clinical examination, and other findings.

The results for subjects with a BMI greater than or equal to 30 kg/m² are presented in the following table.

				Subjects With BMI ≥ 30 kg/m²
NT-proBNP
Result	Adjudicated Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF	Total
Age Group: 18 to 75 Years
Pretest Probability of HF (Prevalence of HF in Study): 59.81% (64/107)
Positive	48	5	53	90.6
(48 / 53)
(79.7, 95.9)	-	6.45
(2.80, 14.88)
Grayzone	16	19	35	45.7
(16 / 35)
(30.5, 61.8)	54.3
(19 / 35)
(38.2, 69.5)	0.57
(0.33, 0.97)
Negative	0	19	19	-	100.0
(19/19)
(83.2, 100.0)	0.02
(0.00, 0.28)
Total	64	43	107
All Subjects With BMI ≥ 30 kg/m2 c
Pretest Probability of HF (Prevalence of HF in Study): 46.69% (500/1071)

Age Group: 50 to 75 Years

Pretest Probability of HF (Prevalence of HF in Study): 46.92% (328/699)

36

Positive	369	85	454	81.3	4.96
				(369/454)	(4.05, 6.07)
				(77.4, 84.6)

37

| NT-proBNP

Result	Subjects With BMI ≥ 30 kg/m²
	Adjudicated
Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
Grayzone	HF	90	Non-HF	91	Total	181	49.7
(90/181)
(42.5, 56.9)	50.3
(91/181)
(43.1, 57.5)	1.13
(0.87, 1.47)
Negative	HF	41	Non-HF	395	Total	436	-

                        | 90.6

(395/436)
(87.5, 93.0) | 0.12
(0.09, 0.16) |
| Total | HF | 500 | Non-HF | 571 | Total | 1071 | | | |

The results for subjects with a BMI less than 30 kg/m² are presented in the following table.

| | Subjects With BMI 75 Years | | | | | | | | |
| | Pretest Probability of HF (Prevalence of HF in Study): 51.08% (95/186) | | | | | | | |
| Positive | 76 | 31 | 107 | 71.0
(76/107)
(61.8, 78.8) | - | 2.35
(1.73, 3.18) | | |
| Grayzone | 17 | 32 | 49 | 34.7
(17/49)
(22.9, 48.7) | 65.3
(32/49)
(51.3, 77.1) | 0.51
(0.30, 0.85) | | |
| Negative | 2 | 28 | 30 | - | 93.3
(28/30)
(78.7, 98.2) | 0.07
(0.02, 0.28) | | |
| Total | 95 | 91 | 186 | | | | | |
| All Subjects With BMI 97.8
(307/314)
(95.5, 98.9) | 0.04
(0.02, 0.08) | | |
| Total | 338 | 539 | 877 | | | | | |
| | NT-proBNP
Result | Adjudicated Diagnosis | | | Subjects With Comorbidities | | | |
| | | HF | Non-HF | Total | Posttest
Probability of
HF % (n/N)
(95% CI)a | Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a | Likelihood
Ratio (HF)
(95% CI)b | |
| Age Group: 18 to 75 Years | | | | | | | | |

38

(66.1, 74.7)

39

Patients with a BMI greater than or equal to 30 kg/m2 had a higher rate of false negatives compared to patients with a BMI less than 30 kg/m². Of the total observed false negatives (51/880), 41 (80.4%) were from patients with a BMI greater than or equal to 30 kg/m², 7 (13.7%) were from patients with a BMI less than 30 kg/m², and 3 (5.9%) were from patients with an unknown BMI. In patients with a BMI less than 30 kg/m2, the age greater than 75 years group had a higher rate of false negatives (6.7% [2/30]) than the other age groups. Results should always be assessed in conjunction with the patient's medical history, clinical examination, and other findings. In patients with obesity, natriuretic peptides should be interpreted with caution.

The results for subjects with comorbidities are presented in the following table. Comorbidities include at least one of the following: diabetes, renal dysfunction (eGFR less than 60 mL/min/1.73 m²), hypertension, and/or chronic obstructive pulmonary disease (COPD). Patients with these comorbidities had a higher rate of false positives compared to patients without these comorbidities.

40

Positive	128	36	164	78.0	-	2.88
				(128/164)		(2.15, 3.85)
				(71.1, 83.7)

41

Subjects With Comorbidities
NT-proBNP
Result	Adjudicated
Diagnosis			Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood
Ratio (HF)
(95% CI)b
	HF	Non-HF	Total
Grayzone	33	48	81	40.7
(33/81)
(30.7, 51.6)	59.3
(48/81)
(48.4, 69.3)	0.56
(0.38, 0.81)
Negative	2	48	50	-	96.0
(48/50)
(86.5, 98.9)	0.03
(0.01, 0.14)
Total	163	132	295
All Subjects With Comorbiditiesc
Pretest Probability of HF (Prevalence of HF in Study): 45.26% (821/1814)
Positive	658	206	864	76.2
(658/864)
(73.2, 78.9)	-	3.86
(3.41, 4.38)
Grayzone	118	191	309	38.2
(118/309)
(32.9, 43.7)	61.8
(191/309)
(56.3, 67.1)	0.75
(0.61, 0.92)
Negative	45	596	641	-	93.0
(596/641)
(90.7, 94.7)	0.09
(0.07, 0.12)
Total	821	993	1814
Subjects Without Comorbidities
NT-proBNP Result	Adjudicated Diagnosis
HF	Non-HF	Total	Posttest Probability of HF % (n/N)
(95% CI)a	Posttest Probability of Non-HF % (n/N)
(95% CI)a	Likelihood Ratio (HF)
(95% CI)b
Age Group: 18 to 75 Years
Pretest Probability of HF (Prevalence of HF in Study): 20.00% (4/20)
Positive	3	2	5	60.0
(3/5)
(23.1, 88.2)	-	6.00
(1.46, 24.69)

The results for subjects without comorbidities are presented in the following table.

42

43

	Subjects Without Comorbidities
NT-proBNP
Result		Adjudicated
Diagnosis		Posttest
Probability of
HF % (n/N)
(95% CI)a	Posttest
Probability of
Non-HF %
(n/N)
(95% CI)a	Likelihood Ratio
(HF)
(95% CI)b
	HF	Non-HF	Total
Grayzone	1	11	12	8.3
(1/12)
(1.5, 35.4)	91.7
(11/12)
(64.6, 98.5)	0.36
(0.06, 2.05)
Negative	0	3	3	-

                                                              | 100.0

(3/3)
(43.9, 100.0) | 0.52
(0.03, 8.39) | |
| Total | 4 | 16 | 20 | | | | |
| All Subjects Without Comorbiditiesc | | | | Pretest Probability of HF (Prevalence of HF in Study): 18.85% (59/313) | | | |
| Positive | 50 | 28 | 78 | 64.1
(50/78)
(53.0, 73.9) | - | 7.69
(5.33, 11.08) | |
| Grayzone | 3 | 28 | 31 | 9.7
(3/31)
(3.3, 24.9) | 90.3
(28/31)
(75.1, 96.7) | 0.46
(0.15, 1.47) | |
| Negative | 6 | 198 | 204 | -
| 97.1
(198/204)
| 0.13
(0.06, 0.28) | |

a The Wilson score method was used to calculate 95% CI for posttest probability.

313

b The log method was used to calculate 95% CI for likelihood ratio.

254

રેત્રે રહે

Total

c The analysis across age groups used the pooled result based on age-dependent positive cutoff and age-independent rule-out cutoff.

The formulas below were used to calculate the pretest probabilities, posttest probabilities, and likelihood ratios in the tables above.

	Adjudicated Diagnosis
Alere NT-proBNP for Alinity i Result	HF	Non-HF
Positive (≥ age-dependent cutoff)	A	B
Grayzone (≥ 300 pg/mL to