The ADVIA Centaur® Erythropoietin (EPO) assay is for in the quantitative measurement of exythropoitin in pediatric and adult human serum or plasma (K2-EDTA, lithium heparin) using the ADVIA Centaur XP system. Measurement of erythropoietin is used as an aid in the diagnosis of anemias and polycythemias.

Device Description

The ADVIA Centaur EPO assay is a fully automated, one-step sandwich immunoassay using direct chemiluminescent technology. The assay utilizes an acridinium-ester-labeled monoclonal mouse anti-EPO antibody in the Lite Reagent. The Solid Phase consists of mouse anti-EPO monoclonal antibody-coated paramagnetic microparticles.

AI/ML Overview

Here's an analysis of the provided text to extract the acceptance criteria and study details for the ADVIA Centaur Erythropoietin (EPO) assay:

1. Table of Acceptance Criteria and Reported Device Performance

This table compiles information primarily from the "Summary of Non-Clinical Performance" and "Summary of Clinical Performance" sections.

Acceptance Criterion (Implicit)	Reported Device Performance (ADVIA Centaur EPO assay)
Linearity (range over which results are proportional to actual concentration)	Linear from 0.83–750.00 mIU/mL.
Dilution Recovery (accuracy after dilution)	Observed percent recovery for individual samples ranged from 76 - 111% when diluted 1:10.
Measuring Interval (reportable range)	0.83 - 750.00 mIU/mL.
Limit of Blank (LoB)	0.46 mIU/mL.
Limit of Detection (LoD) (lowest concentration detectable with 95% probability)	0.75 mIU/mL.
Limit of Quantitation (LoQ) (lowest concentration detectable at total error of 30%)	0.83 mIU/mL. (Results below LoQ should be reported as < 0.83 mIU/mL).
High Dose Hook Effect (no paradoxical decrease at high concentrations)	Patient samples with EPO levels as high as 114,500 mIU/mL will assay greater than 750.00 mIU/mL (i.e., no high-dose hook effect within the measuring interval).
Cross-reactivity (minimal interference from related substances and normal plasma proteins)	Showed minimal cross-reactivity with normal human alpha-2-macroglobulin, transferrin (iron-saturated and non-saturated), rh Thrombopoietin, alpha-1-antitrypsin, alpha- and beta-globulins, Gamma Globulins, and alpha-1-acid glycoprotein. Cross-reactivity with Epoetin alfa and Dabepoetin alfa was also quantified (e.g., Epoetin alfa at 250 mIU/mL showed 27.13% cross-reactivity, Dabepoetin alfa at 2075 mIU/mL showed 7.14% cross-reactivity).
Interference (minimal effect from common endogenous and exogenous interfering substances)	Designed to be ≤ 10% change in EPO values at approximately 4-6 mIU/mL and 25-35 mIU/mL. Insignificant effect from hemolyzed samples (up to 500 mg/dL hemoglobin), icteric (up to 60 mg/dL unconjugated bilirubin, 40 mg/dL conjugated bilirubin), and lipemic (up to 3000 mg/dL Intralipid). No significant interference from Acetaminophen, Acetylsalicylic acid, Biotin, Cholesterol, EPO Soluble Receptor, Heparin, Human Gamma Globulins, Ibuprofen, Multivitamin, Protein Albumin (human), Rheumatoid Factor, Silwet L720, Total Protein, and Triglycerides at specified concentrations (e.g., Acetaminophen >18 mg/dL caused >10% change at 4-6 mIU/mL EPO; Albumin >6.8 g/dL caused >10% change at 4-6 mIU/mL EPO; EPO soluble receptor >31.25 ng/dL caused >10% change at 4-6 mIU/mL EPO; Human gamma globulins (IgG) 6.7 g/dL caused >10% change at 25-35 mIU/mL EPO).
Precision (reproducibility and repeatability)	Coefficients of Variation (CV%) for Repeatability (Within-Run) ranged from 1.6% to 4.8%. CV% for Within-Lab (Total) ranged from 2.6% to 8.4% across 7 samples with EPO concentrations from 1.69 to 579.41 mIU/mL.
Specimen Collection Comparison (equivalence across different tube types)	Correlation coefficient (r) ≥ 0.95, a slope of 0.90-1.10, and an intercept ± 1.00 mIU/mL for alternate tube types (y) versus human serum (x). Demonstrated r values of 0.99-1.00, slopes of 0.97-1.02 and intercepts of -0.33 to -0.20 for K2-EDTA, Lithium Heparin, Sodium Heparin, Plasma Separator Tube, and Serum Separator Tube compared to human serum.
Method Comparison (Agreement with a legally marketed predicate device)	Passing-Bablok regression: ADVIA Centaur EPO (y) = 0.99 (x) + 0.81 mIU/mL (intercept), r = 0.99 (1st study). ADVIA Centaur EPO (y) = 1.07 (x) + 0.00 mIU/mL (intercept), r = 1.00 (2nd study). ADVIA Centaur EPO (y) = 1.01 (x) + 0.36 mIU/mL (intercept), r = 0.99 (3rd multi-site study).
Expected Values (establishment of reference ranges for adult and pediatric populations)	Established 95% Reference Range for combined adult male and female: 5.44 - 26.25 mIU/mL. Established pediatric ranges for Male Child (2-12): 4.13-25.52 mIU/mL; Male Adolescent (13-21): 4.15-26.15 mIU/mL; Female Child (2-12): 4.94-24.47 mIU/mL; Female Adolescent (13-21): 4.07-40.30 mIU/mL.
Standardization (traceability to international standards)	Traceable to WHO 2nd International Reference Preparation for Erythropoietin (human, urinary derived); NIBSC code: 67/343, and WHO 3rd International Standard for Erythropoietin, recombinant, for bioassay; NIBSC code: 11/170.
Substantial Equivalence (Overall conclusion based on studies showing similar performance to predicate)	The ADVIA Centaur EPO assay demonstrated substantially equivalent performance to the Beckman Coulter Access EPO assay.

2. Sample Size Used for the Test Set and Data Provenance

Linearity: Not specified, but involved three high EPO samples mixed with low EPO human serum.
Dilution Recovery: 10 samples (containing high EPO levels: 618.63-986.07 mIU/mL).
Detection Capability (LoD): 323 determinations using 10 low-level samples.
Cross-reactivity: Not explicitly stated as a "sample size," but involved numerous cross-reactants (e.g., various plasma proteins, epoetin alfa, darbepoetin alfa).
Interference: Not explicitly stated as a specific "sample size" for each interferent, but involved various substances tested at different concentrations.
Precision: 7 pooled serum samples. For each sample, there were 80 observations (replicates of 2, in 2 runs/day, over 20 days).
Specimen Collection Comparison: 65 samples (serum EPO values ranging from 4.39 - 707.81 mIU/mL).
Method Comparison:
- Study 1: 216 human serum samples (range: 3.29 – 691.60 mIU/mL).
- Study 2: 100 human serum samples from US population (range: 4.45 - 407.74 mIU/mL).
- Study 3 (Multi-site): 327 human serum samples (range: 3.55 - 596.81 mIU/mL), with ≥ 100 samples per site.
Expected Values (Adult): 251 apparently healthy subjects (128 males, 123 females), older than 21 years of age.
Expected Values (Pediatric): 266 apparently healthy children (2 to <13 years) and adolescents (13 to <22 years).

Data Provenance:
The document does not explicitly state the country of origin for all data sets, but mentions:

Method Comparison Study 2: "100 human serum samples from US population".
Method Comparison Study 3: "3 sites, 2 within Europe and 1 within the US".
Most studies imply retrospective (samples collected before the study) or prospective (samples collected during the study for specific evaluation), but this is not consistently specified for each study. For establishing expected values, samples were collected prospectively following specific criteria (e.g., healthy subjects, specific age ranges, exclusion criteria).

3. Number of Experts Used to Establish the Ground Truth for the Test Set and Their Qualifications

This device is an in vitro diagnostic assay that measures erythropoietin levels. The "ground truth" for such assays typically comes from the actual concentration of the analyte in the biological sample, often confirmed by a reference method or standardized preparations.
The document describes traceability to World Health Organization (WHO) International Reference Preparations and Standards (NIBSC codes 67/343 and 11/170). These international standards are established through collaborative studies involving multiple expert laboratories and scientists, but the immediate "ground truth" for the test samples in this submission would be their assigned values based on these standards or the results from the predicate device.
No "experts" in the sense of radiologists or other clinicians interpreting images or assessments are directly involved in establishing the ground truth for individual test samples for this type of device. The ground truth is biochemical measurement.

4. Adjudication Method for the Test Set

Not applicable. This is a quantitative diagnostic assay. "Adjudication" typically refers to resolving discrepancies between multiple human readers in diagnostic imaging or clinical assessment studies. For an immunoassay, the "ground truth" is a measured concentration.

5. If a Multi Reader Multi Case (MRMC) Comparative Effectiveness Study Was Done, and the Effect Size of How Much Human Readers Improve with AI vs Without AI Assistance

Not applicable. This is an immunoassay, not an AI-assisted diagnostic imaging or clinical decision support device involving human readers.

6. If a Standalone (i.e. algorithm only without human-in-the-loop performance) Was Done

Yes, the performance studies described (Linearity, Dilution Recovery, Measuring Interval, Detection Capability, High Dose Hook, Cross-reactivity, Interference, Precision, Specimen Collection Comparison, Method Comparison, Expected Values) represent the standalone performance of the ADVIA Centaur Erythropoietin (EPO) assay itself, meaning the algorithm/instrument's measurement capabilities without direct human interpretation of the final measurement result for diagnostic purposes. The device directly produces a quantitative EPO value.

7. The Type of Ground Truth Used

Reference materials/standards: The assay is traceable to WHO International Reference Preparations/Standards for Erythropoietin.
Predicate device comparison: For method comparison studies, the results from the legally marketed predicate device (Beckman Coulter Access EPO Assay) serve as a comparative "ground truth" to demonstrate substantial equivalence.
Expert consensus (indirectly): The establishment of reference intervals (expected values) involved defining "healthy" populations based on standard medical criteria, which would implicitly rely on shared medical understanding and the consensus of medical professionals. The CLSI protocols referenced (e.g., EP28-A3) also represent a form of expert consensus on how to establish such values.
Biochemical measurement: The fundamental ground truth is the actual concentration of EPO in the samples, determined through rigorous biochemical methods and validated against international standards.

8. The Sample Size for the Training Set

For an immunoassay like this, there isn't a "training set" in the machine learning sense. The device is not learning from data in the same way an AI algorithm would. Instead, the assay's reagents, calibration curves, and analytical procedures are developed and optimized through extensive R&D and internal validation studies. The studies described in the 510(k) are primarily for verification and validation (V&V) of the final device's performance characteristics. Therefore, a specific "training set sample size" as might be provided for an AI/ML device is not applicable or provided.

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

As above, the concept of a "training set" and its "ground truth" in the context of machine learning is not directly applicable to an immunoassay. The development of the assay involves extensive characterization of reagents, optimization of reaction conditions, and establishment of calibration curves using purified EPO standards of known concentrations, which are themselves traceable to international standards. The "ground truth" during this development phase would be the known concentrations of these standards.

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, which is a blue square with the letters "FDA" in white. To the right of the blue square is the text "U.S. FOOD & DRUG ADMINISTRATION" in blue.

August 2, 2019

Axis-Shield Diagnostics Limited Claire Dora Regulatory Affairs Manager Luna Place, The Technology Park Dundee DD2 1XA, Scotland, UK

Re: K183088

Trade/Device Name: ADVIA Centaur Erythropoietin (EPO) assay Regulation Number: 21 CFR 864.7250 Regulation Name: Erythropoietin Assay Regulatory Class: Class II Product Code: GGT Dated: July 1, 2019 Received: July 3, 2019

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 (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database located at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

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

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's

{1}------------------------------------------------

requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

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

For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100).

Sincerely,

Takeesha Taylor-Bell Chief Division of Immunology and Hematology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

{2}------------------------------------------------

Indications for Use

510(k) Number (if known)

K183088

Device Name ADVIA Centaur® Erythropoietin (EPO) assay

Indications for Use (Describe)

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.

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."

{3}------------------------------------------------

ADVIA Centaur Erythropoietin (EPO) Assay 510(k) Summary

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

A. 510(k) number is: K183088

B. Date of Preparation: 08-02-2019

C. Submission correspondent:

Dr. Claire Dora Regulatory Affairs Manager Axis-Shield Diagnostics Ltd. The Technology Park Dundee DD2 1XA, Scotland, UK

D. Device Name:

ADVIA Centaur® Erythropoietin (EPO) Assay

E. Regulatory Information:

Classification Name: Assay, Erythropoietin Trade Name: ADVIA Centaur® Erythropoietin (EPO) Assay Common Name: Erythropoietin Governing Regulation: 864.7250 Device Classification: Class II Classification Panel: Hematology Product Code: GGT

F. Legally marketed device to which equivalency is claimed:

Beckman Coulter Access EPO Assay (K052223)

{4}------------------------------------------------

G. Intended Use of Device:

The ADVIA Centaur® Erythropoietin (EPO) assay is for in vitro diagnostic use in the quantitative measurement of erythropoietin in pediatric and adult human serum or plasma (K2-EDTA, lithium heparin, sodium heparin) using the ADVIA Centaur XP system. Measurement of erythropoietin is used as an aid in the diagnosis of anemias and polycythemias.

H. Device Description:

Principles of the Device:

The ADVIA Centaur EPO reagent kit contains the following:

ADVIA Centaur EPO ReadyPack® primary reagent pack;

Lite Reagent (10.0 mL/reagent pack): monoclonal mouse anti-EPO antibody (0.5 µg/mL) labeled with DMAE acridinium conjugate reagent in buffer with bovine serum albumin, surfactant, and sodium azide (< 0.1%)

Solid Phase Reagent (24.0 mL/reagent pack): anti-EPO mouse monoclonal antibody coated streptavidin microparticles (0.30 mg/mL) in buffer with bovine serum albumin, surfactant, and preservatives

ADVIA Centaur EPO Calibrator (2.0 mL/vial): recombinant human EPO, calf serum, and preservative.

I. Comparison of Technological Characteristics:

The ADVIA Centaur EPO assay and the Beckman Coulter Access EPO Assay are both automated immunoassays for the quantitative measurement of erythropoietin in human serum and plasma.

The ADVIA Centaur System and Beckman Access share similar detection methods both utilizing chemiluminescent microparticle immunoassay (CMIA) technology. Both assays also demonstrated substantial equivalence as described in the table below:

{5}------------------------------------------------

Comparison of the subject device with the predicate device:

Similarities:

Parameter	New DeviceADVIA Centaur Erythropoietin (EPO)	Predicate DeviceBeckman Access EPO
Intended use	The ADVIA Centaur® Erythropoietin(EPO) assay is for in vitro diagnostic usein the quantitative measurement oferythropoietin in human serum andplasma using the ADVIA Centaursystems. Measurement of erythropoietinis used as an aid in the diagnosis ofanemias and polycythemias.	The Access EPO assay is a paramagneticparticle, chemiluminescent immunoassay forthe quantitative determination oferythropoietin levels in human serum andplasma (heparin) using the AccessImmunoassay Systems. This assay is intendedas an aid in the diagnosis of anemias andpolycythemias. With the advent of theadministration of recombinant erythropoietinas a biologic therapy to increase red blood cellmass, an erythropoietin assay may be usedalso to aid in the prediction and monitoring ofresponse to recombinant erythropoietintreatment of anemias.
Assay technology	Chemiluminescent microparticleimmunoassay (CMIA)	Chemiluminescent microparticleimmunoassay (CMIA)
Calibration range	Up to 750 mIU/mL	Up to 750 mIU/mL
Storage conditions	Reagent, calibrator and control pack mustbe stored at intended storage (2-8°C)	Reagent and calibrator pack must be stored atintended storage (2-10°C)
Open stability	Calibrators are stable opened for 90 days	Calibrators are stable at 2-10°C for 90 daysafter initial use.

Differences:

Parameter	New DeviceADVIA Centaur Erythropoietin (EPO)	Predicate DeviceBeckman Access EPO
Substrate / signalgeneration	Acridinium tracer	Alkaline phosphatase
Conjugate antibody	Monoclonal mouse anti-EPO antibody	Polyclonal chicken anti-recombinant mouse EPO antibody
Calibration	2-point calibration using 2 level adjustors	6 level calibration
Calibration frequency	14 days	28 days
Specimen type	Human Serum and plasma (K2-EDTA, lithium heparin, sodium heparin)	Serum, Plasma (Heparin)
Expected Values –Adults (>21yrs)	5.44 – 26.25 mIU/mL	2.59 – 18.50 mIU/mL
Expected Values –Paediatrics (< 22 yrs)	Male Child (2-12yrs) : 4.13 – 25.52Male Adolescent (13-21yrs) : 4.15 – 26.15Female Child (2-12yrs) : 4.94 – 24.47Female Adolescent (13-21yrs): 4.07 – 40.30	Not reported
Sensitivity	Limit of Quantitation is 0.83 mIU/mL	Analytical Sensitivity, lowest limit ofDetection: 0.6 mIU/mL

{6}------------------------------------------------

Parameter	New Device	Predicate Device
	ADVIA Centaur Erythropoietin (EPO)	Beckman Access EPO
Linearity	0.83 – 750.00 mIU/mL	3.2 – 557.2 mIU/mL
Measureable range	0.83 – 750.00 mIU/mL	0.6 – 750 mIU/mL
Assay dilutionprotocol	1:10 On-board auto dilute with ADVIACentaur Multi-Diluent 13	On-board 1:5 with Access Sample Diluent AManual 1:5 or 1:10 with Access EPOCalibrator S0 or Access Sample Diluent A.
On-board stability	Reagents can be stored on-board for 28days.Calibrators are stable on-board for 24 hours	Reagents: On-board not stated.Calibrators - On-board not stated.

Summary of Non-Clinical Performance: J.

The ADVIA Centaur EPO assay demonstrated substantially equivalent performance to the Beckman Coulter Access EPO assay. A summary of the non-clinical performance data included in this 510(k) submission has been presented.

Linearity

Linearity was evaluated according to the CLSI protocol EP06-A.

Three samples containing high levels of EPO were mixed with low EPO human serum. The resulting sample mixtures were assayed for EPO. The ADVIA Centaur EPO assay is linear from 0.83–750.00 mIU/mL.

Dilution Recovery

Ten samples containing high levels of EPO (618.63-986.07 mIU/mL) were diluted 1:10 (1 part sample plus 9 parts diluent) with ADVIA Centaur Multi-Diluent 13 and assayed for recovery correcting the diluted sample by the dilution factor.

In a representative study the observed percent recovery for individual samples ranged from 76 - 111%.

Measuring Interval

The ADVIA Centaur EPO assay measures erythropoietin concentrations from 0.83 -750.00 mIU/mL.

{7}------------------------------------------------

Detection Capability

The limit of blank (LoB), limit of detection (LoD), and the limit of quantitation (LoQ) were determined as described in CLSI Document EP17-A2. The LoB is defined as the highest measurement result that is likely to be observed for a blank sample. The ADVIA Centaur EPO assay has an LoB of 0.46 mIU/mL. The LoD is defined as the lowest concentration of EPO that can be detected with 95% probability. The ADVIA Centaur EPO assay has an LoD of 0.75 mIU/mL based on 323 determinations using 10 low level samples, and an LoB of 0.46 mIU/mL. The LoQ is defined as the lowest concentration of EPO that can be detected at a total error of 30%. The ADVIA Centaur EPO assay has an LoQ of 0.83 mIU/mL. Report results below the LoQ as < 0.83 mIU/mL.

High Dose Hook

Patient samples with high EPO levels can cause a paradoxical decrease in the Relative Light Units (RLUs) (high-dose hook effect). In the ADVIA Centaur EPO assay, patient samples EPO levels as high as 114,500 mIU/mL will assay greater with than 750.00 mIU/mL.

{8}------------------------------------------------

Cross-reactivity

rh Thrombopoietin

α-1-antitrypsin α- and β-globulins

Gamma Globulins

α-1-acid glycoprotein

Cross-reactivity was tested in the presence of EPO at concentrations of approximately 4 - 6 mIU/mL of EPO according to CLSI EP07-A2. Testing was performed with normal human plasma proteins, and with recombinant erythropoiesis-stimulating agents (ESAs).

Cross-reactant	Concentration	% Cross reactivity
α-2-macroglobulin	400 mg/dL	- 0.01
Transferrin (iron-saturated)	200 mg/dL	0.00
Transferrin (non-saturated)	200 mg/dL	0.00

10,000 ng/mL

80 mg/dL

200 mg/dL

5 g/dL

6 g/dL

0.10

0.01

0.00

0.000

The ADVIA Centaur EPO assay showed minimal cross-reactivity with normal human

The ESAs, epoetin alfa and darbepoetin alfa were also tested. The concentration of darbepoetin was converted using the following formula: 1ng/mL = 166 mIU/mL.4

Note; The cross-reactivity results of the ESAs are provided for information only and should not be used to make a therapeutic decision.

Cross-reactant	Concentration	EPO (mIU/mL)	% Cross-reactivity
Epoetin alfa	1000 mIU/mL	>750	N/A
	250 mIU/mL	72.35	27.13
	125 mIU/mL	27.59	18.46
	65 mIU/mL	18.28	21.17
Dabepoetin alfa	2075 mIU/mL (12.5 ng/mL)	152.82	7.14
	518.75 mIU/mL (3.125 ng/mL)	32.26	5.31
	259.29 mIU/mL (1.562 ng/mL)	20.84	6.22
	129.65 mIU/mL (0.781 ng/mL)	13.5	6.78

1 Conversion from ng/mL to mIU/mL used, ng/mL x 166, this was taken from Owen & Roberts Clinica Acta. 412 (2011) 480-482

{9}------------------------------------------------

Interference

Potential interference in the ADVIA Centaur EPO assay from hemoglobin, bilirubin, and lipemia, is designed to be ≤ 10% at EPO concentrations of approximately 4 - 6 mIU/mL and 25 - 35 mIU/mL.

Interfering substances at the levels indicated in the table below were tested as described in CLSI Document EP07-A227 using the ADVIA Centaur EPO assay.

Serum specimens that are...	Have an insignificant effect on the assay up to...
hemolyzed	500 mg/dL (0.31 mmol/L) of haemoglobin
icteric	60 mg/dL (1026 $ \mu $ mol/L) of unconjugated bilirubin
icteric	40 mg/dL (475 $ \mu $ mol/L) of conjugated bilirubin
lipemic	3000 mg/dL (34.0 mmol/L) of lipemia (Intralipid)

Two levels of EPO were tested with each of the following substances at the levels indicated, and caused no significant interference in the ADVIA Centaur EPO assay at EPO concentrations of approximately 4 - 6 mIU/mL and 25 - 35 mIU/mL.

Substances	Concentrations
Acetaminophen	14 mg/dL (927 $\mu$ mol/L) a
Acetylsalicyclic acid	50 mg/dL (2.8 mmol/L)
Biotin	100 mg/dL (4.1 mmol/L)
Cholesterol	500 mg/dL (12.95 mmol/L)
EPO Soluble Receptor	15 ng/mLa
Heparin	8000 U/dL
Human Gamma Globulins	4.9 g/dLb
Ibuprofen	40 mg/dL (1942 $\mu$ mol/L)
Multivitamin	0.2% (therapeutic level)
Protein Albumin (human)	6 g/dL (60 g/L)a
Rheumatoid Factor	200 IU/mL
Silwet L720	0.2 mg/dL
Total Protein	12 g/dL (120 g/L)
Triglycerides	1000 mg/dL (11.3 mmol/L)

4 concentration-response curve performed to determine non-interfering levels (See detailed study below) bconcentration determined by spiking and recovery method

{10}------------------------------------------------

When targeted concentrations of interfering substances caused >10% change in EPO values, further investigation was performed.

Substances	EPOConcentrationsTested	Level ≤10% change wasobserved	Level >10% change wasobserved
Acetaminophen	4-6 mIU/mL	< 14 mg/dL (927µmol/L)	> 18 mg/dL (1192µmol/L)
	25 - 35 mIU/mL	20 mg/dL (1324 µmol/L)	--
Albumin (human)	4-6 mIU/mL	< 6.0 g/dL (60 g/L)	> 6.8 g/dL (68 g/L)
	25 – 35 mIU/mL	65.2 mg/mL	--
EPO soluble receptor	4-6 mIU/mL	< 25.00 ng/dL	> 31.25 ng/dL
	25 - 35 mIU/mL	< 15.61 ng/mL	> 18.75 ng/mL
Human gammaglobulins (IgG)	4-6 mIU/mL	7.2 g/dL	--
	25 – 35 mIU/mL	4.9 g/dL	6.7 g/dL

Assay results obtained at individual laboratories may vary from the data presented.

{11}------------------------------------------------

Precision

Seven pooled serum samples were prepared with EPO concentrations spanning the measuring interval. Samples were tested in replicates of 2, in 2 runs per day, over 20 days, yielding 80 observations per sample. One ADVIA Centaur XP system and 3 reagent lots were used.

			Repeatability (Within-Run)			Within-Lab (Total)
Specimen	N	Mean (mIU/mL)	SD (mIU/mL)	CV (%)	SD (mIU/mL)	CV (%)
Sample 1	80	1.69	0.08	4.8	0.14	8.4
Sample 2	80	4.51	0.13	2.9	0.23	5.0
Sample 3	80	9.30	0.25	2.6	0.33	3.6
Sample 4	80	25.16	0.54	2.2	1.00	4.0
Sample 5	80	94.30	1.82	1.9	3.22	3.4
Sample 6	80	220.25	3.69	1.7	7.03	3.2
Sample 7	80	579.41	9.20	1.6	15.17	2.6

Representative data from the study is shown in the following table:

Specimen Collection Comparison

The ADVIA Centaur EPO assay was evaluated using different human serum and plasma matrices. The assay is designed to have a correlation coefficient (r) ≥ 0.95, a slope of 0.90-1.10, and an intercept ± 1.00 mIU/mL for alternate tube types (y) versus human serum (x). A specimen collection study was performed with serum EPO values ranging from 4.39 - 707.81 mIU/mL.

Passing-Bablok regression and a Pearson coefficient analysis were performed and no significant difference between tube types was observed. The following results were obtained:

Human Serum (x) Vs	n	Slope	Intercept (mIU/mL)	r
K2-EDTA (y)	65	0.97	-0.30	1.00
Lithium Heparin (y)	65	1.00	-0.27	0.99
Sodium Heparin	65	0.98	-0.33	0.99
Plasma Separator Tube	65	0.99	-0.33	1.00
Serum Separator Tube	65	1.02	-0.20	0.99

{12}------------------------------------------------

K. Summary of Clinical Performance:

The ADVIA Centaur EPO assay demonstrated substantially equivalent performance to the predicate as indicated by reference intervals (expected values) and a method comparison.

Method Comparison

A total of 216 human serum samples in the range of 3.29 – 691.60 mIU/mL were tested on the Centaur XP system vs. the predicate following CLSI EP09-A3.

The relationship of the ADVIA Centaur EPO assay (y) and a comparator EPO assay (x) is described using Passing-Bablok regression and a Pearson coefficient using the ADVIA Centaur XP system. Data from the study is shown below:

ADVIA Centaur EPO (y) = 0.99 (x) + 0.81 mIU/mL (intercept), r = 0.99.

A second method comparison was performed with a total of 100 human serum samples from US population in the range of 4.45 - 407.74 mIU/mL were tested on the Centaur EPO assay and a comparator EPO assay following CLSI EP09-A3.

The relationship of the ADVIA Centaur EPO assay (y) and the comparator EPO assay (x) is described using Passing-Bablok regression and a Pearson coefficient using the ADVIA Centaur XP system. Data from the study is shown below:

ADVIA Centaur EPO (y) = 1.07 (x) + 0.00 mIU/mL (intercept), r = 1.00.

A third mutli-site study was performed 3 sites, 2 within Europe and 1 within the US tested a total of 327 human serum samples (≥ 100 samples per site) in the range of 3.55 - 596.81 mIU/mL were tested on the Centaur EPO assay and a comparator EPO assay.

ADVIA Centaur EPO (y) = 1.01 (x) + 0.36 mIU/mL (intercept), r = 0.99.

{13}------------------------------------------------

Expected Values

The ADVIA Centaur EPO assay results were obtained on 251 apparently healthy subjects, males (n=128) and females (n=123), older than 21 years of age using the ADVIA Centaur XP system. Samples were collected between 7:30 am and 12 noon from individuals with normal hematocrit and hemoglobin levels. Smokers, pregnant women, people living at high altitude or having donated blood within 60 days were excluded.

The reference intervals in the following table were calculated non-parametrically according to EP28-A3.

Gender	N	Mean(mIU/mL)	Median(mIU/mL)	95% CI of Median(mIU/mL)	95% Reference Range(mIU/mL)
CombinedMale and Female	251	11.20	10.08	9.43 - 10.52	5.44 -26.25

ADVIA Centaur EPO Reference Ranges

ADVIA Centaur EPO Pediatric Ranges

The ADVIA Centaur EPO assay results were obtained on 266 apparently healthy children (2 to <13 years) and adolescents (13 to <22 years) using the ADVIA Centaur XP system. Samples were collected 6:00 am and 12 noon from individuals who were of normal weight and height for their age, and were without known diseases.

The reference intervals are presented in the table below and were calculated by means of robust biweight estimators.2

Gender	Age	N	Mean(mIU/mL)	Median(mIU/mL)	95% CI of Median(mIU/mL)	95% Reference Range(mIU/mL)
Male Child	2-12	72	9.80	9.21	8.14-9.89	4.13-25.52
Male Adolescent	13-21	60	10.44	9.30	8.18-10.40	4.15-26.15
Female Child	2-12	74	10.62	9.02	8.20-9.97	4.94-24.47
Female Adolescent	13-21	60	11.96	8.50	8.04-10.78	4.07-40.30

ADVIA Centaur EPO Pediatric Ranges

As with all in vitro diagnostic assays, each laboratory should determine its own reference range(s) for the diagnostic evaluation of patient results. Consider these values as guidelines only.

2 Horn P, Pesce A. Reference Intervals – A User's Guide. Washington DC: AACC Press; 2005

{14}------------------------------------------------

L. Standardization:

The ADVIA Centaur EPO assay is traceable to the World Health Organization (WHO) 2ªª International Reference Preparation for Erythropoietin (Human, urinary derived); NIBSC code: 67/343. Assigned values for calibrators are traceable to this standard. The ADVIA Centaur EPO assay is also traceable to the 3rd World Health Organization (WHO) International Standard for Erythropoietin, recombinant, for bioassay; NIBSC code: 11/170

M. Conclusion:

Based on the performance characteristics the ADVIA Centaur EPO assay is substantially equivalent to the predicate device.

Regulation Number and Section

§ 864.7250 Erythropoietin assay.

(a)
Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This assay provides diagnostic information for the evaluation of erythrocytosis (increased total red cell mass) and anemia.(b)
Classification. Class II. The special control for this device is FDA's “Document for Special Controls for Erythropoietin Assay Premarket Notification (510(k)s).”