Predicate Devices

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Reference Devices

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AI/ML (Artificial Intelligence / Machine Learning)

No
The document describes a standard immunoassay and associated reagents and controls. There is no mention of AI or ML in the intended use, device description, or performance studies. The analysis of patient data for monitoring disease progression is based on changes in quantitative immunoassay values, not AI/ML algorithms.

Therapeutic

No
The device is an immunoassay for in vitro quantitative determination of a biomarker, intended as an aid in monitoring disease progression. It is a diagnostic tool, not a therapeutic device used for treatment or prevention of disease.

Diagnostic

Yes

The "Intended Use / Indications for Use" section states that the assay is to be used "as an aid in monitoring disease progression during the course of disease and treatment in lung cancer patients," which is a diagnostic purpose.

SaMD (Software as a Medical Device)

No

The device description clearly outlines physical components including reagents, calibrators, and control materials, which are integral to the device's function as an immunoassay. It also mentions use on "Elecsys and cobas e immunoassay analyzers," which are hardware devices.

IVD (In Vitro Diagnostic)

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

Here's why:

Intended Use: The "Intended Use / Indications for Use" section explicitly states "Immunoassay for the in vitro quantitative determination of fragments of cytokeratin 19 in human serum and plasma...". The term "in vitro" is a key indicator of an IVD.
Sample Type: The assay is performed on human serum and plasma, which are biological samples taken from the body but tested outside of it.
Purpose: The assay is used as an "aid in monitoring disease progression during the course of disease and treatment in lung cancer patients." This is a diagnostic purpose, providing information about a patient's health status.
Device Description: The description details the components of the system (immunoassay, calibrator, control) which are typical for an IVD system used for laboratory testing.
Performance Studies: The document describes performance studies conducted on human samples to evaluate the assay's effectiveness in a clinical context.

All of these factors align with the definition of an In Vitro Diagnostic device.

PCCP (Predetermined Change Control Plan) Authorized

N/A

Overview

Intended Use / Indications for Use

Elecsys CYFRE 21-1:

Immunoassay for the in vitro quantitative determination of fragments of cytokeratin 19 in human serum and plasma (Li-Heparin. K2-EDTA and K3-EDTA). The assay is to be used as an aid in monitoring disease progression during the course of disease and treatment in lung cancer patients. Serial testing for patient CYFRA 21-1 assay values should be used in conjunction with other clinical methods used for monitoring lung cancer.

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

Elecsys CYFRA 21-1 CalSet:

CYFRA 21-1 is used for calibrating the quantitative Elecsys CYFRA 21-1 assay on the Elecsys and cobas e immunoassay analyzers.

Elecsys PreciControl Tumor Marker:

PreciControl Tumor Marker is used for quality control of Elecsys immunoassays on Elecsys and cobas e immunoassay analyzers.

Product codes (comma separated list FDA assigned to the subject device)

OVK, JIT, JJY

Device Description

(1) The Elecsys CYFRA 21-1 Immunoassay is a two-step sandwich immunoassay with streptavidin microparticles, a biotinylated monoclonal cytokeratin 19-specific antibody, and a monoclonal cytokeratin 19-specific antibody labeled with a ruthenium complex and electrochemiluminescence detection. Results are determined using a calibration curve that is generated specifically on each instrument by a 2 point calibration master curve (5-pointcalibration) provided with the reagent bar code.

(2) The Elecsys CYFRA 21-1 CalSet is a lyophilized product consisting of Cytokeratin from cell culture of the cell line MCF-7 in two concentrations ranges (approximately 0 ng/mL and 50 ng/mL) in a cytokeratin free human serum matrix with preservative. During manufacture, the analyte is spiked ito the matrix at the desired concentration levels.

(3) The Elecsys PreciControl Tumor Marker (CYFRA 21-1) is a lyophilized control serum in two concentration ranges (approximately 3.29 ng/mL and 27.2 ng/mL).

Note: The reagent, calibrator, and the quality control materials are all packaged separately.

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

Not Found

Intended User / Care Setting

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

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

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Summary of Performance Studies (study type, sample size, AUC, MRMC, standalone performance, key results)

Precision (Human Sera):
Internal precision was evaluated with a seven-member panel (five pooled patient samples and two controls: Precicontrol TM Level 1 and 2). Protocol: single determination in four separate aliquots (two runs/day) for 21 operating days on cobas e 411.
External precision: two replicates of each control (PC TM 1 and PC TM 2) and six human serum samples per run, two runs per day for 20 days at three external sites. Samples run in randomized order. Repeatability and Intermediate precision calculated per CLSI EP5-A3.

Limit of Blank (LoB):
Determined according to CLSI EP17-A2 as the 95th percentile of blank-sample measurements. Values for five analyte-free serum samples determined with one reagent lot on one cobas e 411 analyzer over three days for six runs total. Samples measured in two-fold determination for each run (60 measuring points total). LoB: 0.0946 ng/mL.

Limit of Detection (LoD):
Determined as the smallest amount of analyte detectable with 95% probability, according to CLSI EP17-A2. Values for five low-level human serum samples determined with one reagent lot on one cobas e 411 analyzer over three days for six runs total. Samples measured in two-fold determination for each run (60 measuring points total). LoD: 0.1813 ng/mL.

Limit of Quantitation (LoQ):
Determined according to CLSI EP17-A2. Values for 8 spiked human serum samples, diluted to concentrations between LoB and 2x LoQ, determined with one reagent lot on one cobas e411 analyzer across five runs over five days. Each run calibrated separately using a two-point calibration. Samples measured in single determination (200 measuring points total). LoQ: 0.286 ng/mL with allowable error of 30%.

Linearity:
Assessed on cobas e 411 immunoassay analyzer. Six dilution series prepared from six different spiked human samples (three serum, three plasma), each with at least 11 dilutions. Each sample measured 3-fold within one run. Data determined in accordance with CLSI EP6-A. Linearity check performed with first order (linear) regression and then with higher order models (quadratic and cubic).

Analytical Specificity:
Determined using human serum samples spiked with potential cross-reacting compounds. Spiked and unspiked reference samples measured in duplicate or triplicate on one cobas e411 analyzer. Specificity determined using three spiked human samples prepared into 11 dilution series, tested with one reagent lot. Samples spiked with potential cross-reactant compounds at three different concentrations each.

Endogenous Interferences:
Effect of six endogenous interfering substances (Hemoglobin, Biotin, Intralipid, Bilirubin, and Rheumatoid Factor) on CYFRA 21-1 quantitation tested on one cobas e 411 analyzer using spiked serum pools. Three human serum samples (low, mid, high CYFRA 21-1) tested for each interferent.

HAMA Effect:
Effect of human anti-mouse antibodies (HAMA) on Elecsys CYFRA 21-1 assay assessed on cobas e411 analyzer. Suitable HAMA serum spiked with CYFRA 21-1 analyte at two concentrations (3.21 ng/mL and 96.3 ng/mL). Control (human serum) samples spiked similarly. Series of 11 dilutions of HAMA sample and control sample prepared and measured in duplicate. Recovery calculated by comparison to reference (no HAMA) sample. Mean result was 98.51% recovery.

High-Dose Hook Effect:
Assessed on cobas e 411 analyzer. Human serum pools spiked with analyte up to 2,470ng/mL and subsequently diluted. Expected values calculated from spiked concentration and dilution factor. Hook concentration reported corresponds to analyte concentration generating a signal ≥10% above upper measuring range limit. No high-dose hook effect at analyte concentrations up to 2427 ng/mL.

Exogenous Interferences - Drugs:
Effect of 27 pharmaceutical compounds on analyte quantitation determined by comparing values from samples spiked with drugs versus reference aliquots, tested on cobas e 411 analyzer. Spiked aliquots tested in three-fold determination.

Serum/Plasma Comparison:
Effect of anticoagulants on analyte quantitation determined by comparing values from samples drawn into SST, Serum, Li-Heparin Plasma, K2-, K3-EDTA and Plasma primary tubes and PST. At least 44 serum/plasma pairs per sample type tested on one cobas e 411 analyzer. Passing/Bablok regression analysis used.

Reagent Stability:

Study 1. Reagent Stability (On Board): Tested on one cobas e 411 analyzer. Fresh reagent Rack Pack placed, calibrated. Reference values determined. On day 36, 50, and 64, same samples determined with same reagent kit (on-board), using calibration of day 0 and day 64 respectively. Four spiked human serum samples tested in duplicate.
Study 2. Reagent Stability (After Opening): Determined on cobas e 411 analyzer by comparing stability for three kits of same lot. All opened Day 0. One kit placed on analyzer, calibrated, reference values determined. Two stored at 2-8°C. After 50 and 92 days, one stored kit placed, calibrated, and original test samples measured. Five human serum (HS) samples tested in duplicate.
Study 3. Reagent Stability (Real-Time Shelf Life): Reagent stored at 2-8°C. Tested at T=0 and specified intervals over shelf life (+1 month). Average on-test recovery calculated as percent recovery compared to reference value. Two controls tested in duplicate.

Sample Stability:

Study 1. Sample Stability (Freeze/Thaw Cycles): Four samples of human serum and plasma types collected, stored at -80°C. Tested after one freeze/thaw cycle with Elecsys CYFRA 21-1 assay. Measurements in three-fold determination on one cobas e 411 analyzer. Recovery calculated as percent of reference value or deviation in ng/mL.
Study 2. Sample Stability (at -15 to -25°C): Four samples of human serum and plasma types collected, stored at -80°C. Tested after storage at -15 to -25°C for 6 and 12 weeks. Measurements in three-fold determination on one cobas e 411 analyzer. Recovery calculated as percent of reference value or deviation in ng/mL.
Study 3. Sample Stability (at 2-8°C): Four samples of human serum and plasma types collected, stored at -80°C. Tested after storage for up to 14 days at 2-8°C. Measurements in three-fold determination on one cobas e 411 analyzer. Recovery calculated as percent of reference value or deviation in ng/mL.
Study 4. Sample Stability (at 15-25°C): Four samples of human serum and plasma types collected, stored at -80°C. Tested after storage for up to 5 days at 15-25°C. Measurements in three-fold determination on one cobas e 411 analyzer. Recovery calculated as percent of reference value or deviation in ng/mL.

Calibration Stability:

Study 1. Calibration (Lot) Stability: Determined by comparing calibration for three kits of same lot. Day 0, first kit opened, calibrated, samples measured. Same samples measured with fresh opened kits of same lot, using same calibration, after 36 and 64 days on one cobas e 411. Five human serum samples (≥ 1 ng/mL spiked with CYFRA 21-1) measured in duplicate with one reagent lot in one run/day.
Study 2. Calibration (On Board) Stability: Tested on one cobas e 411 immunoassay analyzer. One reagent kit opened, samples measured Day 0. Same samples retested after 8 days with new opened reagent bottle (on-board condition) using Day 0 calibration. Recovery calculated based on initial values. Four human serum samples (> 1 ng/mL spiked with CYFRA 21-1) measured in duplicate with one reagent lot on a cobas e 411 in one run/day.

Calibrator Stability:

Calibrator Value Assignment: Values assigned by comparing to master calibrators traceable to WHO, NIST, or in-house standards. Elecsys CYFRA 21-1 CalSet assayed against reference preparations; target values and ranges assigned. Standardized against in-house reference system. Values assigned using cobas e 411 analyzer. Six independent series performed. Samples tested in duplicate. Sample recovery (%) calculated as median of each sample relative to target value.
Study 1. Calibrator Stability (On-Board): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept on-board at 20-25°C for 6 hours. On-test recovery calculated as signal (counts) of reference value.
Study 2. Stress Stability (Post Reconstitution at 35°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at 35°C for 3 weeks. On-test recovery calculated as signal (counts) of reference value.
Study 3. Post Reconstitution Storage Stability (2-8°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at 2-8°C for 8 days. On-test recovery calculated as signal (counts) of reference value.
Study 4. Post Reconstitution Stability (-15 to -25°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at -25°C to -15°C for 9 weeks. On-test recovery calculated as signal (counts) of reference value.
Study 5. Real-time Shelf Life Stability: CalSet material stored at 2-8°C. Tested at T=0 and specified intervals over shelf life (up to 30 months) in duplicate. Testing performed using PreciControl Tumor Marker 1 and 2 (stored at -20℃). Average on-test recovery calculated as percent recovery compared to reference value.

PreciControl Tumor Marker:

PreciControl Value Assignment: Values assigned using cobas e 411 Immunoassay Analyzer. Six independent series performed. Samples tested in duplicate. Sample recovery (%) calculated as median of each sample relative to target value.
Study 1. Calibrator Stability (On-Board): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept on-board at 20-25°C for 6 hours.
Study 2. Open-Vial Stability (Post Reconstitution at 20-25°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at 20-25°C for 25 hours.
Study 3. Post Reconstitution Storage Stability (2-8°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at 2-8°C for 15 days.
Study 4. Post Reconstitution Stability (-15 to -25°C): Reference and on-test materials tested in duplicate with one reagent lot in one run/day on one cobas e 411 analyzer. On-test material opened and kept at -25°C to -15°C for 5 weeks.
Study 5. Real-time Shelf Life Stability: PreciControl material stored at 2-8°C. Tested at T=0 and specified intervals over shelf life (up to 39 months) in duplicate. Testing performed using PreciControl Tumor Marker 1 and 2 (stored at -20℃). Average on-test recovery calculated as percent recovery compared to reference value.

Clinical performance data:
Study to assess effectiveness of Elecsys CYFRA 21-1 assay in monitoring disease status in lung cancer patients. Sample size: 83 patients with serial serum samples. Total 398 samples measured (86 baseline, 315 monitoring values). Subjects had ≥ 3 blood draws over time. Positive change in CYFRA 21-1 defined as an increase of at least 50% greater than previous value.
Key results: 44.1% (26/59) of patient samples with a positive change correlated with disease progression. 91.0% (233/256) of patient serial samples with no significant change in CYFRA 21-1 value correlated with no progression.

Key Metrics (Sensitivity, Specificity, PPV, NPV, etc.)

Percent (%) change in Elecsys CYFRA 21-1	Sensitivity (%)	Specificity (%)	NPV	PPV
30	49.2	87.1	88.1	46.8
40	44.1	89.8	87.5	50.0
50	44.1	91.0	87.6	53.1
60	39.0	91.4	86.7	51.1
70	35.6	93.4	86.3	55.3

Predicate Device(s): If the device was cleared using the 510(k) pathway, identify the Predicate Device(s) K/DEN number used to claim substantial equivalence and list them here in a comma separated list exactly as they appear in the text. List the primary predicate first in the list.

Fujirebio CYFRA 21-1 EIA kit, Elecsys HE4 CalSet, Elecsys PreciControl Tumor Marker

Reference Device(s): Identify the Reference Device(s) K/DEN number and list them here in a comma separated list exactly as they appear in the text.

Not Found

Predetermined Change Control Plan (PCCP) - All Relevant Information for the subject device only (e.g. presence / absence, what scope was granted / cleared under the PCCP, any restrictions, etc).

Not Found

Regulation Number and Section

§ 866.6010 Tumor-associated antigen immunological test system.

(a)
Identification. A tumor-associated antigen immunological test system is a device that consists of reagents used to qualitatively or quantitatively measure, by immunochemical techniques, tumor-associated antigens in serum, plasma, urine, or other body fluids. This device is intended as an aid in monitoring patients for disease progress or response to therapy or for the detection of recurrent or residual disease.(b)
Classification. Class II (special controls). Tumor markers must comply with the following special controls: (1) A guidance document entitled “Guidance Document for the Submission of Tumor Associated Antigen Premarket Notifications (510(k)s) to FDA,” and (2) voluntary assay performance standards issued by the National Committee on Clinical Laboratory Standards.

Summary

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Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002

December 14, 2016

Roche Diagnostics Mr. Adam Clark Regulatory Affairs Consultant 9115 Hague Road Indianapolis, IN 46250

Re: K160915

Trade/Device Name: Elecsys CYFRA 21-1 Elecsys CYFRA 21-1 CalSet Elecsys PreciControl Tumor Marker Regulation Number: 21 CFR 866.6010 Regulation Name: Tumor-Associated Antigen Immunological Test System Regulatory Class: II Product Code: OVK, JIT Dated: November 7, 2016 Received: November 8, 2016

Dear Mr. Clark:

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. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

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

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR 803); good manufacturing practice requirements

1

as set forth in the quality systems (OS) regulation (21 CFR Part 820); and if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Industry and Consumer Education at its toll-free number (800) 638 2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm. 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

http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

You may obtain other general information on your responsibilities under the Act from the Division of Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.

Sincerely yours,

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FOR

Leonthena R. Carrington, MS, MBA, MT(ASCP) Director Division of Immunology and Hematology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K160915

Device Name Elecsys CYFRA 21-1 Elecsys CYFRA 21-1 CalSet Elecsys PreciControl Tumor Marker

Indications for Use (Describe)

Elecsys CYFRE 21-1:

Immunoassay for the in vitro quantitative determination of tragments of cytokeratin 19 in human serum and plasma (Li-Heparin. K2-EDTA and K3-EDTA). The assay is to be used as an aid in monitoring disease progression during the course of disease and treatment in lung cancer patients. Serial testing for patient CYFRA 21-1 assay values should be used in conjunction with other clinical methods used for monitoring lung cancer.

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

Elecsys CYFRA 21-1 CalSet:

CYFRA 21-1 is used for calibrating the quantitative Elecsys CYFRA 21-1 assay on the Elecsys and cobas e immunoassay analyzers.

Elecsys PreciControl Tumor Marker:

PreciControl Tumor Marker is used for quality control of Elecsys immunoassays on Elecsys and 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)

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510(k) Summary

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

Submitter Name	Roche Diagnostics
Address	9115 Hague Road
Indianapolis, IN 46250
Contact	Adam Clark
Phone317)521-4371
Email: adam.clark@roche.com
Date Prepared	November 7, 2016
Proprietary Name	1. Elecsys CYFRA 21-1

Elecsys CYFRA 21-1 CalSet
Elecsys PreciControl Tumor Marker |
| Common Name | 1. CYFRA 21-1
CYFRA 21-1 CalSet
Tumor Marker Control |
| Classification Name | 1. Tumor-Associated antigen immunological test system
Secondary, calibrator
Multi-analyte controls, all kinds (assayed and unassayed) |
| Product Codes | 1. OVK; 866.6010
JIT; 862.1150
JJY; 862.1660 |
| Predicate Devices | 1. Fujirebio CYFRA 21-1 EIA kit
Elecsys HE4 CalSet
Elecsys PreciControl Tumor Marker |
| Establishment Registration | Roche Diagnostics GmbH in Mannheim, Germany: 9610126
Roche Diagnostics GmbH in Penzberg, Germany: 9610529
Roche Diagnostics in the United States: 1823260 |

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1. DEVICE DESCRIPTION

(1) The Elecsys CYFRA 21-1 Immunoassay is a two-step sandwich immunoassay with streptavidin microparticles, a biotinylated monoclonal cytokeratin 19-specific antibody, and a monoclonal cytokeratin 19-specific antibody labeled with a ruthenium complex and electrochemiluminescence detection. Results are determined using a calibration curve that is generated specifically on each instrument by a 2 point calibration master curve (5-pointcalibration) provided with the reagent bar code.

(2) The Elecsys CYFRA 21-1 CalSet is a lyophilized product consisting of Cytokeratin from cell culture of the cell line MCF-7 in two concentrations ranges (approximately 0 ng/mL and 50 ng/mL) in a cytokeratin free human serum matrix with preservative. During manufacture, the analyte is spiked ito the matrix at the desired concentration levels.

(3) The Elecsys PreciControl Tumor Marker (CYFRA 21-1) is a lyophilized control serum in two concentration ranges (approximately 3.29 ng/mL and 27.2 ng/mL).

Note: The reagent, calibrator, and the quality control materials are all packaged separately.

INDICATIONS FOR USE 2.

(1) Immunoassay for the in vitro quantitative determination of fragments of cytokeratin 19 in human serum and plasma (Li-Heparin, K2-EDTA and K3-EDTA). The assay is to be used as an aid in monitoring disease progression during the course of disease and treatment in lung cancer patients. Serial testing for patient CYFRA 21-1 assay values should be used in conjunction with other clinical methods used for monitoring lung cancer.

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

(2) CYFRA 21-1 CalSet is used for calibrating the quantitative Elecsys CYFRA 21-1 assay on the Elecsys and cobas e immunoassay analyzers.

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(3) PreciControl Tumor Marker is used for quality control of Elecsys immunoassays on Elecsys and cobas e immunoassay analyzers.

TECHNOLOGICAL CHARACTERISTICS 3.

Comparison of Assays, Similarities and Differences
Table 1

Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831)	Candidate Device:
Elecsys CYFRA 21-1
	General Assay Features
Intended Use/
Indications
for Use	The CYFRA 21-1 EIA kit is intended
for the quantitative determination of
soluble cytokeratin 19 fragments in
human serum. The assay is to be used as
an aid in monitoring disease progression
during the course of disease and
treatment in lung cancer patients. Serial
testing for patient CYFRA 21-1 assay
values should be used in conjunction
with other clinical methods used for
monitoring lung cancer.	Immunoassay for the in vitro
quantitative determination of fragments
of cytokeratin 19 in human serum and
plasma (Li-Heparin, K2-EDTA and K3-
EDTA). The assay is to be used as an
aid in monitoring disease progression
during the course of disease and
treatment in lung cancer patients. Serial
testing for patient CYFRA 21-1 assay
values should be used in conjunction
with other clinical methods used for
monitoring lung cancer.
The electrochemiluminescence
immunoassay “ECLIA” is intended for
use on Elecsys and cobas e
immunoassay analyzers.
Assay
Protocol	Quantitative enzyme immunometric
assay	Quantitative sandwich
immunoassay
Detection
Protocol	Spectrophotometry	Electrochemiluminescence
Applications	Manual assay	18 minute application
Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831)	Candidate Device:
Elecsys CYFRA 21-1
	General Assay Features
Instrument
Platform	Microtiter plate	cobas e 411
Sample
Volume	50 μL	20 μL
Sample
Type	Human serum	Human serum and plasma (Li-
Heparin, K2-EDTA and K3-
EDTA)
Reagents	The Fuijirebio CYFRA 21-1 EIA is a
solid phase, non-competitive
immunoassay based on two monoclonal
antibodies (derived from mice) directed
against two separate antigenic
determinants of soluble fragments of
cytokeratin 19. Calibrators, controls and
patient samples are incubated together
with biotinylated Anti-CYFRA 21-1
MAb and horseradish peroxidase (HRP)
labeled Anti-CYFRA 21-1 MAb in
streptavidin coated microstrips. After
washing, buffered
Substrate/Chromogen reagent
(hydrogen peroxide and 3, 3', 5, 5'
tetramethylbenzidine) is added to each
well and the enzyme reaction is allowed
to proceed. During the enzyme reaction
a blue color will develop if antigen is
present. The color intensity is
determined in a microplate
spectrophotometer at 620 nm.	The Elecsys CYFRA 21-1 assay is
a sandwhich immunoassay
biotinylated monoclonal
cytokeratin 19-specific antibody
and a monoclonal cytokeratin 19-
specific antibody labeled with a
ruthenium complex [Tris(2,2'- bipyridyl)ruthenium(II)-complex
(Ru(bpy))]
Calibrator	CYFRA 21-1 A, B, C, D, E and F	Elecsys CYFRA 21-1 CalSet
Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831) and Elecsys HE4
PreciControl (K112624)	Candidate Device:
Elecsys CYFRA 21-1
	General Assay Features
Calibration
Interval	Calibration curves are determined
from the 6 calibrators and are run
in duplicate with each assay run.	Calibration must be performed once per
reagent lot using fresh reagent (i.e. not
more than 24 hours since the reagent kit
was registered on the analyzer.)
Renewed Calibration is recommended as
follows:
After 8 weeks when using the same
reagent lotAfter 7 days when using the same
reagent kit on the analyzerAs required: e.g. quality control
findings outside the specified limits
Controls	Elecsys HE4 PreciControl (K112624)	Elecsys PreciControl Tumor Marker
Traceability/
Standardization	In K100831, the sponsor states:
There is no recognized reference
standard for CYFRA 21-1. The
concentration of the CYFRA 21-1
EIA primary calibrators has been
assigned by using Roche CK19
antigen. The methods used for
value assignment of the CYFRA
21 primary and secondary
calibrators were provided.	Elecsys CYFRA 21-1 has been
standardized against the Enzymun-Test
CYFRA 21-1 method.
Reagent Stability	Microplate – 2-8°C until expiry
date stated on the plate
Biotin Anti-CYFRA 21-1 – 2-8°C
until expiry date stated on the vial
Tracer, HRP Anti-CYFRA 21-1 –
2-8°C until expiry date stated on
the vial
TMB HRP-Substrate – 2-8°C until
expiry date stated on the vial	Unopened at 2-8°C – up to stated
expiration date.
After opening at 2-8°C – 12 weeks.
On the analyzer – 8 weeks
Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831)	Candidate Device:
Elecsys CYFRA 21-1
	Labeled Performance Characteristics
Measuring
Range	0.5 - 50 ng/mL	0.5 – 100 ng/mL
Precision	On the Fujirebio Test System
Within-run
n=160
3.70% CV @ 2.69 ng/mL
5.10% CV @ 2.91 ng/mL
4.30% CV @ 7.28 ng/mL
4.10% CV @ 7.63 ng/mL
2.40% CV @ 17.4 ng/mL
2.60% CV @ 18.6 ng/mL
2.60% CV @ 33.4 ng/mL
3.80% CV @ 35.7 ng/mL
Total
6.10% CV @ 2.69 ng/mL
8.40% CV @ 2.91 ng/mL
6.90% CV @ 7.28 ng/mL
7.30% CV @ 7.63 ng/mL
5.30% CV @ 17.4 ng/mL
4.90% CV @ 18.6 ng/mL
4.90% CV @ 33.4 ng/mL
5.60% CV @ 35.7 ng/mL	cobas e 411
Within-run (will be labeled repeatability)
n=84
HS1: 4.8% CV @ 0.53 ng/mL
HS2: 2.1% CV @ 1.31 ng/mL
HS3: 2.1% CV @ 1.37 ng/mL
HS4: 1.9% CV @ 2.80 ng/mL
HS5: 1.3% CV @ 18.98 ng/mL
HS6: 1.3% CV @ 79.37 ng/mL
PC 1: 1.2% CV @ 2.66 ng/mL
PC 2: 0.7% CV @ 21.73 ng/mL
Total (will be labeled Intermediate)
HS1: 9.6% CV @ 0.53 ng/mL
HS2: 4.0% CV @ 1.31 ng/mL
HS3: 4.6% CV @ 1.37 ng/mL
HS4: 2.6% CV @ 2.80 ng/mL
HS5: 2.2% CV @ 18.98 ng/mL
HS6: 2.4% CV @ 79.37 ng/mL
PC 1: 4.3% CV @ 2.66 ng/mL
PC 2: 0.7% CV @ 21.73 ng/mL

6

7

Table 1, continued

8

9

Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831)	Candidate Device:
Elecsys CYFRA 21-1
Labeled Performance Characteristics
LoB	Not determined	0.0946 ng/mL
LoD	0.12 ng/mL	0.1813 ng/mL
LoQ	0.21 ng/mL with the highest
allowable imprecision of 17.78%	0.286 ng/mL with a total allowable error
of 30%
Lower
Detection
Limit	0.5 ng/mL	0.5 ng/mL
Hook
Effect	No high-dose hook effect in samples
up to 1100 ng/mL	There is no high-dose hook effect at
analyte concentrations up to 2427 ng/mL
Limitations	The assay is inaffected by:
Triglycerides ≤ 30 mg/mL Bilirubin1 ≤ 0.2 mg/mL Hemoglobin ≤ 5.0 mg/mL Total Protein ≤ 120 mg/mL The CYFRA 21-1 EIA assay was
evaluated using specimens with
HAMA and Rheumatoid Factor (FR)
to further assess the assay specificity.
Six specimens positive for HAMA
and five specimens positive for RF
were evaluated for % recovery with
CYFRA 21-1 antigen spiked into
each specimen at approximately 5
and 25 ng/mL. Mean recovery
results are summarized in the
following table:
Clinical
Condition Number
of
specimens Mean %
Recovery HAMA 6 98 RF 5 101 1A mixture of conjugated (10%) and
unconjugated (90%) bilirubin.			The assay is unaffected by:
Lipemia (Intralipid®) ≤ 1500 mg/dL Biotin ≤ 50 ng/mL Bilirubin ≤ 66 mg/dL Hemoglobin ≤ 1500 mg/dL Rheumatoid Factor ≤ 1500 IU/mL Human Serum Albumin ≤ 7 g/dL The CYFRA 21-1 assay was evaluated
using specimens with HAMA to further
assess the assay specificity. A specified
HAMA serum was spiked with analyte to
two different analyte concentrations (3.21
ng/mL and 96.3 ng/mL). As a control, a
serum without interferent (related basic
serum) was spiked analogously. The
samples were measured in duplicates.
The Mean result was 98.51% recovery.

10

Assay Comparison
Feature	Predicate Device:
Fujirebio CYFRA 21-1 EIA
(K100831)	Candidate Device:
Elecsys CYFRA 21-1
Labeled Performance Characteristics
External
Method
Comparison
(Fujirebio vs.
Elecsys)	Slope
Unweighted 0.911
Deming (95% CI: 0.86, 0.96)	Intercept Pearson r
-1.4 0.955
(95% CI: -2.86, 0.06)
Clinical
Sensitivity and
Specificity	The following table shows the
resulting sensitivities and
specificities of the CYFRA 21-1
EIA at various percent changes in
CYFRA 21-1 EIA concentrations,
together with the positive predictive
values (PPV) and negative
predictive values (NPV) for the
population tested (85 sequential
pairs from patients with disease
progression and 229 sequential pairs
from patients with no progression).	The following table shows the resulting
sensitivities and specificities of the
Elecsys CYFRA 21-1 assay compared
to the disease status at various changes
in concentration over time. Sensitivity is
represented as a concordance of the
Elecsys CYFRA 21-1 to progression of
disease. Specificity is represented as a
concordance of the Elecsy CYFRA 21-1
to no progression of disease.
	Percent
Change in
CYFRA 21-1	Sensitivity Specificity
(%) (%)
	30	52.9 84.3
	40	48.2 85.6
	50	45.9 87.3
	60	44.7 88.2
	70	43.5 89.5
	Percent
Change in
CYFRA 21-1	Sensitivity Specificity
(%) (%)
	30	49.2 87.1
	40	44.1 89.8
	50	44.1 91.0
	60	39.0 91.4
	70	35.6 93.4

11

CalSet Comparison
Feature	Predicate Device:
Elecsys HE4 CalSet (K112624)	Candidate Device:
Elecsys CYFRA 21-1 CalSet
Intended Use	Elecsys HE4 CalSet is used for
calibrating the quantitative Elecsys
HE4 assay on the Elecsys and cobas
e immunoassay analyzers.	CYFRA 21-1 CalSet is used for
calibrating the quantitative Elecsys
CYFRA 21-1 assay on the Elecsys and
cobas e immunoassay analyzers.
Levels	Two	Two
Matrix	Human Serum	Human Serum
Format	Lyophilized	Lyophilized
Stability	Unopened:
• Store at 2-8°C up to the
stated expiration date.
After reconstitution:
• At 2-8°C: 7 days
• At -20°C: 8 weeks (freeze
only once).
• On Elecsys 2010/cobas e
411 at 20-25°C: Up to 5
hours.
• On MODULAR
ANALYTICS E170/cobas e
601: Use only once.	Unopened:
• Store at 2-8°C up to the stated
expiration date.
After reconstitution:
• At 2-8°C: 7 days
• At -15--25°C: 8 weeks (freeze
only once).
• On cobas e 411 at 20-25°C: Up
to 5 hours.
Handling	Dissolve the contents of one bottle
carefully by adding exactly 1.0mL
of distilled water and allow to stand
closed for 15 minutes to reconstitute.
Mix carefully, avoiding the
formation of foam. Transfer the
reconstituted calibrator into the
empty labeled snap-cap bottles
supplied.	Dissolve the contents of one bottle
carefully by adding exactly 1.0mL of
distilled water and allow to stand closed
for 15 minutes to reconstitute. Mix
carefully, avoiding the formation of
foam. Transfer the reconstituted
calibrator into the empty labeled snap-
cap bottles supplied.

12

Table 1, continued

| Feature | Predicate Device:
PreciControl Tumor Marker
(K050387) | Candidate Device:
PreciControl Tumor Marker
Modified to Include CYFRA 21-1 |
|--------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Intended Use | PreciControl Tumor Marker is used
for quality control of Elecsys
immunoassays on Elecsys
immunoassay analyzers. | PreciControl Tumor Marker is used for
quality control of Elecsys
immunoassays on Elecsys and cobas e
immunoassay analyzers. |
| Analyte | AFP: approx.. 8 & 100 IU/mL
CEA: approx.. 5 & 50 mL
CA 15-3 II: approx. 20 & 100 U/mL
CA 125 II: approx. 35 & 100 U/mL
Ferritin: approx.. 25 & 200 ng/mL
fPSA: approx.. 1 & 10 ng/mL
tPSA: approx.. 4 & 40 ng/mL
CA 19-9: approx.. 20 & 100 U/mL | AFP: approx. 8 & 100 IU/mL
CEA: approx. 5 & 50 mL
CA 15-3 II: approx. 20 & 100 U/mL
CA 125 II: approx. 35 & 100 U/mL
Ferritin: approx. 25 & 200 ng/mL
fPSA: approx. 1 & 10 ng/mL
tPSA: approx. 4 & 40 ng/mL
CA 19-9: approx. 20 & 100 U/mL
CYFRA 21-1: approx. 3 & 27 ng/mL |
| Levels | Two | Same |
| Matrix | Human Serum | Same |
| Format | Lyophilized | Same |
| Handling | Dissolve the contents of one bottle
carefully by adding exactly 3.0mL
of distilled water and allow to stand
closed for 30 minutes to reconstitute.
Mix carefully, avoiding the
formation of foam. Transfer the
reconstituted calibrator into the
empty labeled snap-cap bottles
supplied. | Same |
| Stability | On the analyzer: up to 5 hours
20-25°C: 24 hours
2-8°C: 2 weeks
-20°C: 1 month (freeze only once) | Same |

NON-CLINICAL PERFORMANCE EVALUATION 4.

Non-clinical performance evaluations for the Elecsys CYFRA 21-1 assay executed with the study briefly summarized.

13

Precision (Human Sera) 4.1.

Internal precision of the Elecsys CYFRA 21-1 assay was evaluated with a seven-member panel consisting of five pooled patient samples and two controls (Precicontrol TM Level 1 and 2) were measured. The protocol consisted of testing the samples in single determination in four separate aliquots (divided in two runs per day) for 21 operating days. The measurements were performed on the cobas e 411 with one reagent lot, performing rackpack calibration according to instruction for use.

External precision consisted of testing two replicates of each control (PC TM 1 and PC TM 2) and six human serum samples per run, two runs per day for 20 days at three external sites. The samples were run in randomized order on the analyzer.

Repeatability and Intermediate precision were calculated according to CLSI EP5-A3.

4.2. Limit of Blank (LoB)

For the analytical sensitivity studies, the Limit of Blank was determined according to CLSI EP17-A2. The LoB was determined as the 95th percentile of blank-sample measurements. The distribution of values for five analyte-free serum samples was determined with one reagent lot on one cobas e 411 anaylzer over three days for six runs total.

The samples were measured in two-fold determination for each run. A total of 60 measuring points were collected.

4.3. Limit of Detection (LoD)

For the analytical sensitivity studies, the Limit of Detection was determined according is the smallest amount of analyte that can be detected with 95% probability. The distribution of values for five low-level human serum samples was determined with one reagent lot on one cobas e 411 anaylzer over three days for six runs total.

The samples were measured in two-fold determination for each run. A total of 60 measuring points were collected.

14

4.4. Limit of Quantitation (LoQ)

LoO of the CYFRA 21-1 assay has been determined according to CLSI EP17-A2.

The distribution of values for 8 spiked human serum samples each diluted to concentrations which covered the range between LoB and 2x LoQ has been determined with one reagent lot on one cobas e411 analyzer with five runs distributed over five days. Each run was calibrated separately using a two-point calibration. Samples were measured in single determination in each run. In summary, a total of 200 measuring points were collected.

4.5. Linearity

The linearity of the CYFRA 21-1 assay was assessed on the cobas e 411 immunoassay analyzer; six dilution series were prepared from six different spiked human samples. Three dilution series were performed with serum samples and the other three with plasma samples. Each dilution series included at least 11 dilutions. Each sample was measured 3-fold within one run and the measured concentrations were plotted against the expected sample concentration.

The linearity data was determined in accordance with CLSI EP6-A. In a first step, a linearity check was performed with a first order (linear) regression and then with higher order models (quadratic and cubic).

Analytical Specificity 4.6.

The specificity of the Elecsys CYFRA 21-1assay was determined using human serum samples spiked with potential cross-reacting compounds. The spiked and unspiked reference samples were measured in duplicate or triplicate on one cobas e411 analyzer. Specificity was determined using three spiked human samples to prepare dilution series of 11 dilutions that were tested with one reagent lot. Samples were spiked with potential cross-reactant compounds at three different concentrations each. Cross-reactivity results will be reported in the method sheet.

4.7. Endogenous Interferences

The effect on quantitation of CYFRA 21-1 in the presence of six endogenous interfering substances (Hemoglobin, Biotin, Intralipid, Bilirubin', and Rheumatoid Factor) was tested using one cobas e 411 analyzer. Spiked serum pools were used for testing.

For each potential interferent, three human serum samples (containing low, mid, and high concentrations of CYFRA 21-1) were tested. 1 A mixture of conjugated (10%) and unconjugated (90%) bilirubin.

4.8. HAMA Effect

The effect of the presence of human anti-mouse antibodies (HAMA) on the Elecsys CYFRA 21-1assay was assessed on the cobas e411 analyzer.

15

A suitable HAMA serum was spiked with CYFRA 21-1 analyte to two different analyte concetrations (3.21 ng/mL and 96.3 ng/mL. In parallel, a control (human serum) sample was spiked with the same levels of analyte. For each analyte concentration, a series of 11 dilutions of the HAMA sample and control sample were each prepared and measured in duplicate. The recovery for each sample was calculated by comparison to the reference (no HAMA) sample.

4.9. High-Dose Hook Effect

The high-dose hook effect of the Elecsys CYFRA 21-1assay was assessed on the cobas e 411 analyzer. Human serum pools were spiked with analyte up to a concentration of 2,470ng/mL. These samples were subsequently diluted with negative human serum.

The expected values for the diluted samples were calculated using the known spiked CYFRA 21-1 analyte concentration of the undiluted sample and the appropriate dilution factor.

The hook concentration reported corresponds to the analyte concentration that generates a signal ≥10% above the upper limit of the measuring range

4.10. Exogenous Interferences - Drugs

The effect on quantitation of analyte in the presence of drugs was determined by comparing values obtained from samples spiked with 27 pharmaceutical compounds into two human serum samples at differing analyte concentrations and tested on the cobas e 411 analyzer.

The CYFRA 21-1 concentration of the spiked aliquots was tested in three-fold determination and compared to the CYFRA 21-1 concentration for the reference aliquot (also tested in three-fold determination).

4.11. Serum/Plasma Comparison

The effect on quantitation of analyte in the presence of anticoagulants with the Elecsys CYFRA 21-1 assay was determined by comparing values obtained from samples drawn into Serum Separator Tubes (SST). Serum, Li-Heparin Plasma, K2-, K3-EDTA and Plasma primary tubes and Plasma Separation Tubes (PST). At least 44 serum/plasma pairs per sample type were tested on one cobas e 411 analyzer. For Serum / Plasma Separation Tube comparison, measurements were performed on five samples in duplicate on the cobas e 601 assay. Potential effects were assessed by Passing/Bablok regression analysis.

4.12. Reagent Stability

To test reagent stability, three studies were executed.

4.12.1. Study 1. Reagent Stability (On Board)

Reagent on-board stability for the Elecsys CYFRA 21-1 assay was tested on one cobas e 411 analyzer.

16

A fresh reagent Rack Pack was placed on the analyzer and calibrated. Reference values for the samples tested were determined. On day 36, 50 and day 64 the same samples were determined with the same reagent kit (kept on board the instrument) using the calibration curve of day 0 and day 64 respectively.

Samples tested in duplicate include four spiked human serum samples.

4.12.2. Study 2. Reagent Stability (After Opening)

Reagent stability after first opening for the Elecsys CYFRA 21-1 assay was determined on a cobas e 411 immunoassay analyzer by comparing the reagent stability for three kits of the same lot. All reagent kits were opened on day 0. One kit was placed on the analyzer and calibrated and reference values for the samples tested were determined.

The other two kits were stored at 2 to 8°C. After 50 and 92 days, one of the stored kits was placed on the analyzer and calibrated, and the original test samples were measured.

Samples tested in duplicate include five human serum (HS) samples.

4.12.3. Study 3. Reagent Stability (Real-Time Shelf Life)

In the real-time, shelf-life stability study, the Elecsys CYFRA 21-1assay reagent was stored at 2-8°C. The stored reagent was tested at time point 0 (at manufacture), then again at specified intervals over the shelf life of the device (up to the planned shelf life plus one month). The average on-test recovery was calculated as percent recovery compared to the reference value.

Samples tested in duplicate included two controls.

4.13. Sample Stability

To test reagent stability, four studies were executed.

4.13.1. Study 1. Sample Stability (Freeze/Thaw Cycles)

Four samples of each human serum and plasma types were collected and stored at -80°C. The samples were tested after one freeze/thaw cycle with the Elecsys CYFRA 21-1 assay.

Measurement were performed in three-fold determination on one cobas e 411 analyzer, and recovery was calculated as percent of the reference value or as a deviation in ng/mL.

4.13.2. Study 2. Sample Stability (at -15 to -25°C)

Four samples of each human serum and plasma types were collected and stored at -80°C. The samples were tested after storage at -15 to -25°C for 6 and 12 weeks with the Elecsys CYFRA 21-1 assay.

Measurement were performed in three-fold determination on one cobas e 411 analyzer, and recovery was calculated as percent of the reference value or as a deviation in ng/mL.

17

4.13.3. Study 3. Sample Stability (at 2-8°C)

Four samples of each human serum and plasma types were collected and stored at -80°C. The samples were tested after storage for up to14 days at 2-8°C with the Elecsys CYFRA 21-1assay.

Measurement were performed in three-fold determination on one cobas e 411 analyzer, and recovery was calculated as percent of the reference value or as a deviation in ne/mL.

4.13.1. Study 4. Sample Stability (at 15-25°C)

Four samples of each human serum and plasma types were collected and stored at -80°C. The samples were tested after storage for up to 5 days at 15-25℃ with the Elecsys CYFRA 21-1 assay.

Measurement were performed in three-fold determination on one cobas e 411 analyzer, and recovery was calculated as percent of the reference value or as a deviation in ng/mL.

4.14. Calibration Stability

To test calibration stability, two studies were executed.

4.14.1. Study 1. Calibration (Lot) Stability

The stability of lot calibration was determined by comparing the calibration for three kits of the same lot. On day 0, the first reagent kit was opened and calibrated, and samples were measured. The same samples were also measured with fresh opened kits of the same lot, using the same calibration established by the first kit, after 36 and 64 days on one cobas e 411.

Five human serum samples (samples ≥ 1 ng/mL spiked with CYFRA 21-1) were measured in duplicate with one reagent lot in one run per day.

4.14.2. Study 2. Calibration (On Board) Stability

On-board calibration stability for the CYFRA 21-1 assay was tested on one cobas e 411 immunoassay analyzer. One reagent kit was opened and samples were measured on day 0. The same samples were then retested after 8 days with a new opened reagent bottle kept at 20 ± 3℃ (on-board condition) using the calibration from day 0. Recovery was calculated based on the initial values.

Four human serum samples > 1 ng/mL spiked with CYFRA 21-1) were measured in duplicate with one reagent lot on a cobas e 411 in one run per day..

4.15. Calibrator Stability

The CYFRA 21-1 CalSet was evaluated for value assignment and stability: five studies were executed.

18

4.15.1. Calibrator Value Assignment

Roche Diagnostics maintains a set of master calibrators for each assay that has values assigned that are traceable to various method "gold standards". These standards can either be WHO, NIST, or in-house developed standards.

The Elecsys CYFRA 21-1 CalSet is assayed and compared to these reference preparations, and target values and ranges are assigned.

In the case of this submission, the CYFRA 21-1 assay has been standardized against an in-house reference system.

Values are assigned using the cobas e 411 analyzer. Six independent series are performed. All samples are tested in duplicate. The sample recovery (%) is calculated as the median of each sample as a reference to the target value.

4.15.2. Study 1. Calibrator Stability (On-Board)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept on-board at 20 to 25°C for 6 hours. On-test recovery was calculated as the signal (counts) of the reference value.

4.15.3. Study 2. Stress Stability (Post Reconstitution at 35°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at 35°C for 3 weeks.

On-test recovery was calculated as the signal (counts) of the reference value.

4.15.4. Study 3. Post Reconstitution Storage Stability(2-8°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at 2 to 8°C for 8 days.

On-test recovery was calculated as the signal (counts) of the reference value.

4.15.5. Study 4. Post Reconstitution Stability(-15 to -25°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at -25°C to -15°C for 9 weeks.

On-test recovery was calculated as the signal (counts) of the reference value.

4.15.6. Study 5. Real-time Shelf Life Stability

In the real-time stability study, the CYFRA 21-1 CalSet material was stored at 2 to 8°C. The stored CalSet reagents were tested at time point T=0 and at specified intervals over the shelf life of the device up to 30 months in duplicate. Testing was performed using PreciControl Turnor Marker 1 and 2 (stored at -20℃).

19

The average on-test recovery was calculated as percent recovery compared to the reference value (Assigned value for PreciControl Tumor Marker 1 and 2).

4.16. PreciControl Tumor Marker

PreciControl Tumor Marker was evaluated for value assignment and stability; five studies were executed.

4.16.1. PreciControl Value Assignment

Values are assigned using the cobas e 411 Immunoassay Analyzer. Six independent series are performed. All samples are tested in duplicate. The sample recovery (%) is calculated as the median of each sample as a reference to the target value.

4.16.2. Study 1. Calibrator Stability (On-Board)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept on-board at 20 to 25°C for 6 hours.

4.16.3. Study 2. Open-Vial Stability (Post Reconstitution at 20-25°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at 20 to 25°C for 25 hours.

4.16.4. Study 3. Post Reconstitution Storage Stability(2-8°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at 2 to 8°C for 15 days.

4.16.5. Study 4. Post Reconstitution Stability(-15 to -25°C)

Reference and on-test materials were tested in duplicate with one reagent lot in one run per day on one cobas e 411 analyzer. The on-test material was opened and kept at -25°C to -15°C for 5 weeks.

4.16.6. Study 5. Real-time Shelf Life Stability

In the real-time stability study, the PreciControl material was stored at 2 to 8°C. The stored PreciControl reagents were tested at time point T=0 and at specified intervals over the shelf life of the device up to 39 months in duplicate. Testing was performed using PreciControl Turnor Marker 1 and 2 (stored at -20℃).

The average on-test recovery was calculated as percent recovery compared to the reference value (Assigned value for PreciControl Tumor Marker 1 and 2).

20

5. CLINICAL PERFORMANCE EVALUATION

Expected values

Normal CYFRA 21-1 values are expected to be ≤ 2.37 ng/mL.

The following table shows the results of three separate cohorts. The first study represents the distribution of expected results from 240 apparently healthy men and women equally divided into smokers and nonsmokers. The second study represents the distribution of expected results from 195 benign disease conditions other than cancers, and the third study represents the distribution of expected results from different cancers, including lung cancer.

| | No. of
subjects | 0.3- 2.37
ng/mL | 2.38- 5.0
ng/mL | 5.01- 20.0
ng/mL | 20.01- 100
ng/mL | > 100
ng/mL |
|---------------------------------------------------------------------|--------------------|--------------------|--------------------|---------------------|---------------------|----------------|
| Apparently
healthy | 240 | | | | | |
| All normals | 240 | 228 | 12 | 0 | 0 | 0 |
| Nonsmokers | 120 | 111 | 9 | 0 | 0 | 0 |
| Smokers | 120 | 117 | 3 | 0 | 0 | 0 |
| Normal females | 125 | 119 | 6 | 0 | 0 | 0 |
| Nonsmokers | 63 | 59 | 4 | 0 | 0 | 0 |
| Smokers | 62 | 60 | 2 | 0 | 0 | 0 |
| Normal males | 115 | 109 | 6 | 0 | 0 | 0 |
| Nonsmokers | 57 | 52 | 5 | 0 | 0 | 0 |
| Smokers | 58 | 57 | 1 | 0 | 0 | 0 |
| | No. of
subjects | 0.3-2.37
ng/mL | 2.38- 5.0
ng/mL | 5.01- 20.0
ng/mL | 20.01- 100
ng/mL | > 100
ng/mL |
| Investigational Elecsys CYFRA 21-1 distribution of values by cohort | | | | | | |
| Benign conditions | 195 | | | | | |
| Benign lung
disease | 75 | 70 | 5 | 0 | 0 | 0 |
| CHFb) | 40 | 29 | 11 | 0 | 0 | 0 |
| Benign kidney
disease | 40 | 8 | 24 | 8 | 0 | 0 |
| Benign liver
disease | 40 | 35 | 4 | 1 | 0 | 0 |
| Cancer | 440 | | | | | |
| Lung cancer | 120 | 53 | 33 | 27 | 5 | 2 |
| Bladder cancer | 40 | 13 | 9 | 12 | 5 | 1 |
| Breast cancer | 40 | 32 | 5 | 3 | 0 | 0 |
| Cervical cancer | 40 | 28 | 11 | 1 | 0 | 0 |
| ESCCc) | 40 | 21 | 12 | 6 | 1 | 0 |
| GI tract cancer | 40 | 23 | 10 | 6 | 1 | 0 |
| Head and neck
cancer | 40 | 29 | 11 | 0 | 0 | 0 |
| Prostate cancer | 40 | 37 | 1 | 2 | 0 | 0 |
| Ovarian cancer | 40 | 25 | 8 | 5 | 2 | 0 |

21

b) CHF = Congestive heart failure

22

c) ESCC = Esophageal squamous cell carcinoma

Each laboratory should investigate the transferability of the expected values to its own patient population and if necessary determine its own reference ranges.

Specific performance data

Representative performance data on the analyzers are given below. Results obtained in individual laboratories may differ.

Clinical performance data

The effectiveness of the Elecsys CYFRA 21-1 assay as an aid in monitoring of disease status in lung cancer patients was determined by assessing changes in CYFRA 21-1 levels in serial serum samples from 83 patients compared to changes in disease status. A total of 398 samples were measured, including 86 baseline values and 315 monitoring values. Subjects had ≥ 3 blood draws over time.

A positive change in CYFRA 21-1 was defined as an increase in the value that was at least 50 % greater than the previous value of the test. This level of change takes into account the analytical variability of the assay. 44.1 % (26/59) of the patient samples with a positive change correlated with the disease progression while 91.0 % (233/256) of the patient serial samples with no significant change in CYFRA 21-1 value correlated with no progression. The following table presents the data in a 2 x 2 format.

23

Investigational Elecsys CYFRA 21-1 elevation vs. disease progression
		Disease progression
		No progression	Progression	Total
Investigational Elecsys
CYFRA 21-1 elevation	Not elevated	233	33	267
	Elevated	23	26	48
	Total	256	59	315

No set cutoff exists for CYFRA 21-1. The clinical performance of other percent changes in serial samples are presented below. Clinicians may choose to use these other values to enhance the sensitivity or specificity of the assay, depending on their needs.

Cutoff values and corresponding performance measurements
Percent (%)
change in
Elecsys CYFRA
21-1	Sensitivity (%)	Specificity (%)	NPVa)	PPVb)
30	49.2	87.1	88.1	46.8
40	44.1	89.8	87.5	50.0
50	44.1	91.0	87.6	53.1
60	39.0	91.4	86.7	51.1
70	35.6	93.4	86.3	55.3

a) NPV = negative predictive value

b) PPV = positive predictive value

CONCLUSIONS 6.

The information provided in this 510(k) Premarket Notification will support a determination of substantial equivalence for the Elecsys CYFRA 21-1 test system. The data supports a safe, effective device which performs as well as or better than the predicate device.