Elecsys Anti-HAV II

K190428 · Roche Diagnostics · LOL · Aug 13, 2019 · Microbiology

Device Facts

• Record ID: K190428
• Device Name: Elecsys Anti-HAV II
• Applicant: Roche Diagnostics
• Product Code: LOL · Microbiology
• Decision Date: Aug 13, 2019
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.3310
• Device Class: Class 2
• Attributes: Pediatric

Intended Use

Immunoassay for the in vitro qualitative detection of total antibodies (IgG and IgM) to hepatitis A virus (HAV) in human pediatric (ages 2 through 21 years) and adult serum and plasma (Li-Heparin, potassium EDTA, Na-Citrate, Na-Heparin). The assay, in conjunction with other serological and clinical information, is indicated as an aid in the clinical laboratory diagnosis of acute or past hepatitis A virus infection in persons with signs or symptoms of hepatitis and in persons at increased risk for hepatitis A infection, or as an aid to identify HAV susceptible individuals and to determine the presence of an antibody response to HAV in vaccine recipients. The electrochemiluminescence immunoassay "ECLIA" is intended for use on the cobas e immunoassay analyzers. Assay performance characteristics have not been established for immunocompromised or immunosuppressed patients. This assay has not been FDA cleared or approved for the screening of blood or plasma donors.

Device Story

Elecsys Anti-HAV II is an in vitro diagnostic immunoassay for qualitative detection of total anti-HAV antibodies (IgG/IgM) in human serum/plasma. Operated on cobas e 601 analyzers by laboratory personnel. Uses competition principle: sample antibodies compete with biotinylated monoclonal anti-HAV antibodies for binding to HAV antigen; ruthenium-labeled monoclonal anti-HAV antibodies bind to remaining HAV antigen. Electrochemiluminescence (ECLIA) signal inversely proportional to antibody concentration in sample. Software automatically calculates results by comparing signal to cutoff value from calibration. Used in clinical labs to aid diagnosis of acute/past HAV infection, identify susceptibility, or confirm vaccine response. Benefits include rapid (18-minute) automated testing, supporting clinical decision-making for hepatitis management.

Clinical Evidence

Method comparison study of 961 subjects (prospective, retrospective, remnant) across three sites using cobas e 601. Compared to predicate Elecsys Anti-HAV. Overall PPA 99.8% (501/502) and NPA 95.2% (437/459). Cohorts included routine, hospitalized, increased risk, symptomatic, acute HAV, and pediatric. Seroconversion panel testing (n=4) showed equivalent performance to predicate. Vaccination study (n=49) showed 100% agreement with predicate.

Technological Characteristics

Electrochemiluminescence immunoassay (ECLIA) on cobas e 601 analyzer. Reagents: streptavidin-coated microparticles, HAV antigen, biotinylated monoclonal anti-HAV antibody, ruthenium-labeled monoclonal anti-HAV antibody. Sample volume: 20 µL. Calibration: 2-point, 28-day stability. Connectivity: cobas e 601 platform. Software: automated signal processing and cutoff comparison.

Indications for Use

Indicated for pediatric (ages 2-21) and adult patients requiring diagnosis of acute or past HAV infection, identification of HAV susceptibility, or assessment of vaccine-induced antibody response. Not for use in immunocompromised/immunosuppressed patients or blood/plasma donor screening.

Regulatory Classification

Identification

HAV serological assays are devices that consist of antigens and antisera for the detection of hepatitis A virus-specific IgM, IgG, or total antibodies (IgM and IgG), in human serum or plasma. These devices are used for testing specimens from individuals who have signs and symptoms consistent with acute hepatitis to determine if an individual has been previously infected with HAV, or as an aid to identify HAV-susceptible individuals. The detection of these antibodies aids in the clinical laboratory diagnosis of an acute or past infection by HAV in conjunction with other clinical laboratory findings. These devices are not intended for screening blood or solid or soft tissue donors.

Special Controls

*Classification.* Class II (special controls). The special control is “Guidance for Industry and FDA Staff: Class II Special Controls Guidance Document: Hepatitis A Virus Serological Assays.” See § 866.1(e) for the availability of this guidance document.

Predicate Devices

• Elecsys Anti-HAV (K100903)

Submission Summary (Full Text)

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Image /page/0/Picture/0 description: The image shows 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 square are the words "U.S. FOOD & DRUG" in blue, with the word "ADMINISTRATION" below it.

August 13, 2019

Roche Diagnostics Jamie Ferguson Regulatory Affairs Principal 9115 Hague Road Indianapolis, Indiana 46250

Re: K190428

Trade/Device Name: Elecsys Anti-HAV II Regulation Number: 21 CFR 866.3310 Regulation Name: Hepatitis A Virus (HAV) Serological Assays Regulatory Class: Class II Product Code: LOL, QCH Dated: February 20, 2019 Received: February 22, 2019

Dear Jamie Ferguson:

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

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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 ([e-mail]) or phone [phone] or 301-796-7100).

Sincerely,

for

Maria Garcia, Ph.D. Chief Division of Microbiology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

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### Indications for Use

510(k) Number (if known) K190428

Device Name Elecsys Anti-HAV II

### Indications for Use (Describe)

Immunoassay for the in vitro qualitative detection of total antibodies (IgG and IgM) to hepatitis A virus (HAV) in human pediatric (ages 2 through 21 years) and adult serum and plasma (Li-heparin, potassium EDTA, Na-hebarin). The assay, in conjunction with other serological and clinical information, is indicated as an aid in the clinical laboratory diagnosis of acute or past hepatitis A virus infection in persons with signs or symptoms of hepatitis and in persons at increased risk for hepatitis A infection, or as an aid to identify HAV susceptible individuals and to determine the presence of an antibody response to HAV in vaccine recipients.

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

Assay performance characteristics have not been established for immunocompromised or immunosuppressed patients. This assay has not been FDA cleared or approved for the screening of blood or plasma donors.

| Type of Use (Select one or both, as applicable) | |
|------------------------------------------------------------------------------------------------|---------------------------------------------|
| <span style="text-decoration: overline;">X</span> Prescription Use (Part 21 CFR 801 Subpart D) | Over-The-Counter Use (21 CFR 801 Subpart C) |

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# Elecsys Anti-HAV II 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.

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

The purpose of this Traditional 510(k) Premarket Notification is to obtain FDA review and clearance for the Elecsys Anti-HAV II.

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| Submitter Name | Roche Diagnostics |
|--------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Address | 9115 Hague Road<br>P.O. Box 50416<br>Indianapolis, IN 46250-0457 |
| Contact | Jamie Ferguson<br>Phone: [phone]<br>Fax: [phone]<br>Email: [e-mail] |
| | Tammy Dean<br>Phone: [phone]<br>Fax: [phone]<br>Email: [e-mail] |
| Date Prepared | February 20, 2019 |
| Proprietary Name | Elecsys Anti-HAV II<br>PreciControl Anti-HAV II |
| Common Name | Anti-HAV II<br>PreciControl Anti-HAV II |
| Classification Name | 1. Hepatitis A virus (HAV) Serological Assays<br>2. Assayed Quality Control Material for Clinical Microbiology Assays |
| Product Codes,<br>Regulation Numbers | 1. LOL, 21 CFR §866.3310<br>2. QCH, 21 CFR §866.3920, Quality Control |
| Predicate Devices | Elecsys Anti-HAV (K100903) |
| Establishment Registration | For the Elecsys Anti-HAV II, the establishment registration number for Roche<br>Diagnostics GmbH in Mannheim, Germany is 9610126, and for Penzberg,<br>Germany, 9610529. The establishment registration number for Roche<br>Diagnostics in the United States is 1823260. |

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#### DEVICE DESCRIPTION 1.

Elecsys Anti-HAV II is a second generation assay by Roche Diagnostics for the in vitro qualitative detection of total antibodies (IgG and IgM) to the hepatitis A virus (HAV) in human pediatric (ages 2 through 21 years) and adult serum and plasma. It is intended for use on the cobas e 601 immunoassay analyzer. The cobas e family of analyzers employs the electrochemiluminescence "ECLIA" technology. The assay is an 18-minute assay utilizing a competition principle.

Results are determined automatically by the software by comparing the electrochemiluminescence signal obtained from the reaction product of the sample with the signal of the cutoff value previously obtained by calibration.

The reagent rackpack working solutions include:

- M: Streptavidin-coated microparticles .
- R1: HAV Ag (cell culture) .
- R2: Biotinylated monoclonal anti-HAV antibody, monoclonal Anti-HAV antibody . labeled with ruthenium complex
- AHAV 2 Cal1: Negative Calibrator 1 (human serum) .
- AHAV 2 Cal2: Positive Calibrator 2 (anti-HAV (human), approximately 60 IU/L in . human serum)

PreciControl Anti-HAV II is a ready-for-use control serum based on human serum both in the negative and positive concentration range. The controls are used for monitoring the performance of the Elecsys Anti-HAV II immunoassay. PreciControl Anti-HAV II is sold separately from the Elecsys Anti-HAV II immunoassay reagent.

#### 2. INTENDED USE

Immunoassay for the in vitro qualitative detection of total antibodies (IgG and IgM) to hepatitis A virus (HAV) in human pediatric (ages 2 through 21 years) and adult serum and plasma (Li-Heparin, potassium EDTA, Na-Citrate, Na-Heparin). The assay, in conjunction with other

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serological and clinical information, is indicated as an aid in the clinical laboratory diagnosis of acute or past hepatitis A virus infection in persons with signs or symptoms of hepatitis and in persons at increased risk for hepatitis A infection, or as an aid to identify HAV susceptible individuals and to determine the presence of an antibody response to HAV in vaccine recipients.

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

Assay performance characteristics have not been established for immunocompromised or immunosuppressed patients. This assay has not been FDA cleared or approved for the screening of blood or plasma donors.

#### 3. TECHNOLOGICAL CHARACTERISTICS

Elecsys Anti-HAV II utilizes electrochemiluminescence "ECLIA" technology for the qualitative detection of total antibodies (IgG and IgM) to the hepatitis A virus (HAV) in human pediatric (ages 2 through 21 years) and adult serum and plasma on the cobas e 601 immunoassay analyzer.

The following tables compare the Elecsys Anti-HAV II with its predicate device, Elecsys Anti-HAV (K100903).

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#### Assay Comparison Table 1:

| Feature | Elecsys Anti-HAV (K100903) | Elecsys Anti-HAV II |
|-------------------------------------|---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|-------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|
| Intended Use/Indications for<br>Use | Immunoassay for the in vitro qualitative<br>detection of total antibodies (IgM and<br>IgG) to hepatitis A virus in human serum<br>and plasma (K2-EDTA). The assay is<br>intended for use as an aid in the<br>laboratory diagnosis of past or<br>acute/recent hepatitis A infection. Assay<br>results, in conjunction with other<br>laboratory results and clinical<br>information, may be used to provide<br>presumptive evidence of infection with<br>hepatitis A virus in persons with signs or<br>symptoms of hepatitis and in persons at<br>risk for hepatitis A infection, or used as an<br>aid to determine the presence of antibody<br>response to HAV in vaccine recipients.<br>The electrochemiluminescence<br>immunoassay "ECLIA" is intended for<br>use on Elecsys and cobas e immunoassay<br>analyzers.<br>This assay is not intended for screening<br>blood or solid or soft tissue donors. Assay<br>performance characteristics have not been<br>established for immunocompromised or<br>immunosuppressed patients. The users are<br>responsible for establishing their own<br>assay performance characteristics in these<br>populations. | Immunoassay for the in vitro qualitative<br>detection of total antibodies (IgG and<br>IgM) to hepatitis A virus (HAV) in<br>human pediatric (ages 2 through 21<br>years) and adult serum and plasma (Li-<br>Heparin, potassium EDTA, Na-Citrate,<br>Na-Heparin). The assay, in conjunction<br>with other serological and clinical<br>information, is indicated as an aid in the<br>clinical laboratory diagnosis of acute or<br>past hepatitis A virus infection in<br>persons with signs or symptoms of<br>hepatitis and in persons at increased risk<br>for hepatitis A infection, or as an aid to<br>identify HAV susceptible individuals<br>and to determine the presence of an<br>antibody response to HAV in vaccine<br>recipients.<br>The electrochemiluminescence<br>immunoassay "ECLIA" is intended for<br>use on cobas e immunoassay analyzers.<br>Assay performance characteristics have<br>not been established for<br>immunocompromised or<br>immunosuppressed patients. This assay<br>has not been FDA cleared or approved<br>for the screening of blood or plasma<br>donors. |
| Assay Method | Competition principle binding protein | Same |
| Detection Method | Electrochemiluminescence | Same |
| Applications/Test Time | 18 minutes | Same |
| Instrument Platform | Elecsys 2010, cobas e 411, cobas e 601,<br>cobas e 602, and MODULAR<br>ANALYTICS E170 | cobas e 601 |
| Sample volume | 50 µL | 20 µL |
| Sample Type/Matrix | Human serum, plasma | Same |
| Sample Anticoagulants | K2-EDTA | Li-Heparin, potassium EDTA, Na-<br>Citrate, Na-Heparin |
| Calibrator | Anti-HAV Cal1 and Cal2 (packed in kit) | Anti-HAV II Cal1 and Cal2 (packed in<br>kit) |
| Calibration Method | 2-point calibration | Same |
| Feature | Elecsys Anti-HAV (K100903) | Elecsys Anti-HAV II |
| Calibration Interval | Calibration must be performed once per<br>reagent lot using fresh reagent (i.e. not<br>more than 24 hours since the reagent kit<br>was registered on the analyzer). Renewed<br>calibration is recommended as follows:<br>• after 1 month (28 days) when using<br>the same reagent lot<br>• after 7 days (when using the same<br>reagent kit on the analyzer)<br>• as required: e.g. quality control<br>findings outside the defined limits | Same |
| Controls | PreciControl Anti-HAV | PreciControl Anti-HAV II |
| Traceability/Standardization | Second International Standard for Anti<br>Hepatitis A, Immunoglobulin, Human,<br>NIBSC code: 97/646 | Same |
| Reagent Stability | Store at 2-8°C.<br>Do not freeze.<br>Store the Elecsys reagent kit upright in<br>order to ensure complete availability of<br>the microparticles during automatic<br>mixing prior to use.<br>Unopened at 2-8°C, up to stated<br>expiration date.<br>After opening at 2-8°C, 8 weeks.<br>On the analyzers at 20-25°C, 7 days or 4<br>weeks when stored alternative in the<br>refrigerator and on the analyzer, with the<br>total time onboard on the analyzer not<br>exceeding 40 hours. | Store at 2-8°C.<br>Do not freeze.<br>Store the Elecsys reagent kit upright in<br>order to ensure complete availability of<br>the microparticles during automatic<br>mixing prior to use.<br>Unopened at 2-8°C, up to stated<br>expiration date.<br>After opening at 2-8°C, 8 weeks.<br>On the analyzers at 20-25°C, 8 weeks. |
| Feature | Elecsys Anti-HAV (K100903) | Elecsys Anti-HAV II |
| Limitations | Bilirubin < 855 µmol/L or < 50 mg/dL<br>Hemolysis < 0.623 mmol/L or < 1.0 g/dL<br>Lipemia < 1500 mg/dL<br>Biotin < 205 nmol/L or < 50 ng/mL<br>Criterion: Recovery within +/- 20% of<br>initial value<br>Samples should not be taken from patients<br>receiving therapy with high biotin doses<br>(i.e. > 5 mg/day) until at least 8 hours<br>following the last biotin administration.<br>In vitro tests were performed on 18<br>commonly used pharmaceuticals<br>(Acetylcystein, Ampicillin, Ascorbic acid,<br>Ca Dobesilate, Cyclosporine, Cefoxitin,<br>Heparin, Intralipid, Levodopa,<br>Methyldopa, Metronidazole,<br>Phenylbutazone, Tetracycline,<br>Acetylsalicylic Acid, Rifampicin,<br>Acetaminophen, Ibuprofen and<br>Theophylline). No interference with the<br>assay was found.<br>In rare cases, interference due to<br>extremely high titers of antibodies to<br>analyte specific antibodies, streptavidin or<br>ruthenium can occur. These effects are<br>minimized by suitable test design. | Bilirubin < 1129 µmol/L or ≤ 66 mg/dL<br>Hemoglobin ≤ 0.621 mmol/L or ≤ 1000<br>mg/dL<br>Intralipid ≤ 2000 mg/dL<br>Rheumatoid Factors ≤ 1400 IU/mL<br>IgG ≤ 7.0 g/dL<br>IgA ≤ 1.6 g/dL<br>IgM ≤ 1.0 g/dL<br>Serum Albumin ≤ 7 g/dL<br>Criterion: > 1.0 COI +/- 20% recovery,<br>≤ 1.0 COI +/- 0.20 recovery<br>Negative specimens with biotin<br>concentrations up to 100 ng/mL<br>demonstrated ≤ 11 % negative bias in<br>COI values. Biotin concentrations<br>greater than 100 ng/mL lead to higher<br>negative bias and in consequence can<br>lead to false positive Elecsys Anti-HAV<br>II results. Some studies have shown that<br>serum concentrations of biotin can reach<br>up to 355 ng/mL within the first hour<br>after ingestion for subjects consuming<br>supplements of 20 mg biotin per day and<br>up to 1160 ng/mL in plasma for subjects<br>consuming a single dose of 300 mg<br>biotin.<br>In vitro tests were performed on 18<br>commonly used pharmaceuticals<br>(Acetylcystein, Ampicillin-Na, Ascorbic<br>acid, Ca Dobesilate, Cyclosporine,<br>Cefoxitin, Doxycycline, Heparin,<br>Levodopa, Methyldopa +1.5,<br>Metronidazole, Phenylbutazone,<br>Tetracycline, Acetylsalicylic Acid,<br>Rifampicin, Acetaminophen, Ibuprofen<br>and Theophylline). No interference with<br>the assay was found.<br>In rare cases, interference due to<br>extremely high titers of antibodies to<br>analyte specific antibodies, streptavidin<br>or ruthenium can occur. These effects are<br>minimized by suitable test design. |

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#### NON-CLINICAL PERFORMANCE EVALUATION 4.

Non-clinical performance evaluation for Elecsys Anti-HAV II is briefly summarized below.

#### Precision 4.1.

#### 4.1.1. Repeatability and Within-Laboratory Precision

Precision measurements were conducted to evaluate repeatability (within-run precision) and within-laboratory precision (intermediate precision) in a study based on the protocol of CLSI EP05-A3. Precision was evaluated on a single cobas e 601 immunoassay analyzer. One Elecsys Anti-HAV II reagent lot was evaluated. The protocol consisted of testing 2 aliquots each of two levels of control and 5 human sera per run, 2 runs per day for 21 days. Calibration was performed on day 1 and on day 17. Serum samples were human serum sample pools.

| | | Repeatability | | Within-Laboratory<br>(Intermediate) Precision | | |
|--------------------|------------|---------------|--------|-----------------------------------------------|--------|----|
| Sample | Mean (COI) | SDa)<br>(COI) | CV (%) | SD<br>(COI) | CV (%) | n |
| Human serum 1 | 1.42 | 0.016 | 1.1 | 0.028 | 2.0 | 84 |
| Human serum 2 | 1.15 | 0.012 | 1.1 | 0.022 | 1.9 | 84 |
| Human serum 3 | 0.955 | 0.009 | 0.9 | 0.022 | 2.3 | 84 |
| Human serum 4 | 0.665 | 0.009 | 1.3 | 0.020 | 2.9 | 84 |
| Human serum 5 | 0.006 | 0.0002 | 3.0 | 0.0002 | 3.3 | 84 |
| PCb) Anti-HAV II 1 | 1.30 | 0.015 | 1.1 | 0.024 | 1.8 | 84 |
| PC Anti-HAV II 2 | 0.339 | 0.005 | 1.5 | 0.009 | 2.7 | 84 |

Table 2: Repeatability and Within-Laboratory Precision Results

SD = Standard Deviation a)

PC = PreciControl b)

#### 4.2. Analytical Sensitivity

#### 4.2.1. Determination of Cut-off Sensitivity

The cut-off of the Elecsys Anti-HAV II assay was established with internal studies, and the validation of the cut-off was performed by external clinical studies. In order to determine the cut-off sensitivity, the 2nd International Standard for Anti-Hepatitis A, Immunoglobulin, Human,

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NIBSC code: 97/646 was serially diluted to 10 samples and tested fourfold with three different reagent and calibrator lots. The cut-off of COI = 1.0 corresponds to ≤ 25.4 IU/L.

| Dilution-<br>factor/<br>Sample | Target value<br>[IU/L] | Measuring cell 1 [COI] | | Measuring cell 2 [COI] | | [COI]<br>Mean |
|--------------------------------|------------------------|------------------------|-------|------------------------|-------|---------------|
| | | Run 1 | Run 2 | Run 1 | Run 2 | |
| Ref-Std_01 | 0 | 1.31 | 1.28 | 1.31 | 1.30 | 1.30 |
| Ref-Std_02 | 10.2 | 1.16 | 1.17 | 1.17 | 1.16 | 1.17 |
| Ref-Std_03 | 20.3 | 1.05 | 1.03 | 1.06 | 1.04 | 1.05 |
| Ref-Std_04 | 30.5 | 0.921 | 0.923 | 0.923 | 0.946 | 0.928 |
| Ref-Std_05 | 59.9 | 0.628 | 0.626 | 0.637 | 0.646 | 0.634 |
| Ref-Std_06 | 99.7 | 0.365 | 0.361 | 0.365 | 0.369 | 0.365 |
| Ref-Std_07 | 200 | 0.098 | 0.098 | 0.104 | 0.102 | 0.100 |
| Ref-Std_08 | 300 | 0.035 | 0.034 | 0.036 | 0.035 | 0.035 |
| Ref-Std_09 | 400 | 0.017 | 0.017 | 0.017 | 0.017 | 0.017 |
| Ref-Std_10 | 500 | 0.011 | 0.011 | 0.019 | 0.011 | 0.013 |
| Cut-off sensitivity | | 24.6 IU/L | | | | |

### Table 3: Lot MP01

### Table 4: Lot PoQ

| Dilution-<br>factor/<br>Sample | Target value<br>[IU/L] | Measuring cell 1 [COI] | | Measuring cell 2 [COI] | | [COI]<br>Mean |
|--------------------------------|------------------------|------------------------|-------|------------------------|-------|---------------|
| | | Run 1 | Run 2 | Run 1 | Run 2 | |
| Ref-Std_01 | 0 | 1.28 | 1.28 | 1.27 | 1.28 | 1.28 |
| Ref-Std_02 | 10.2 | 1.14 | 1.15 | 1.17 | 1.17 | 1.16 |
| Ref-Std_03 | 20.3 | 1.05 | 1.06 | 1.04 | 1.04 | 1.05 |
| Ref-Std_04 | 30.5 | 0.941 | 0.944 | 0.934 | 0.942 | 0.940 |
| Ref-Std_05 | 59.9 | 0.663 | 0.663 | 0.674 | 0.665 | 0.667 |
| Ref-Std_06 | 99.7 | 0.389 | 0.399 | 0.399 | 0.401 | 0.397 |
| Ref-Std_07 | 200 | 0.113 | 0.112 | 0.115 | 0.115 | 0.114 |
| Ref-Std_08 | 300 | 0.039 | 0.038 | 0.040 | 0.040 | 0.040 |
| Ref-Std_09 | 400 | 0.018 | 0.018 | 0.018 | 0.019 | 0.018 |
| Ref-Std_10 | 500 | 0.011 | 0.011 | 0.011 | 0.011 | 0.011 |
| Cut-off sensitivity | | | | 25.4 IU/L | | |

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| Dilution-factor/<br>Sample | Target value<br>[IU/L] | Measuring cell 1 [COI] | | Measuring cell 2 [COI] | | [COI]<br>Mean | |
|----------------------------|------------------------|------------------------|-------|------------------------|-------|---------------|--|
| | | Run 1 | Run 2 | Run 1 | Run 2 | | |
| Ref-Std_01 | 0 | 1.25 | 1.25 | 1.26 | 1.27 | 1.26 | |
| Ref-Std_02 | 10.2 | 1.11 | 1.10 | 1.13 | 1.15 | 1.12 | |
| Ref-Std_03 | 20.3 | 0.976 | 0.975 | 0.982 | 0.984 | 0.980 | |
| Ref-Std_04 | 30.5 | 0.867 | 0.859 | 0.873 | 0.872 | 0.868 | |
| Ref-Std_05 | 59.9 | 0.551 | 0.548 | 0.563 | 0.559 | 0.555 | |
| Ref-Std_06 | 99.7 | 0.269 | 0.274 | 0.276 | 0.276 | 0.274 | |
| Ref-Std_07 | 200 | 0.048 | 0.048 | 0.049 | 0.048 | 0.048 | |
| Ref-Std_08 | 300 | 0.016 | 0.016 | 0.016 | 0.016 | 0.016 | |
| Ref-Std_09 | 400 | 0.010 | 0.010 | 0.010 | 0.010 | 0.010 | |
| Ref-Std_10 | 500 | 0.009 | 0.009 | 0.009 | 0.009 | 0.009 | |
| Cut-off sensitivity | | 20.1 IU/L | | | | | |

### Table 5: Lot P02

#### 4.3. Human Anti-Mouse Antibodies (HAMA)

The effect on detection of analyte in the presence of HAMA using the Elecsys Anti-HAV II was determined on the cobas e 601 immunoassay analyzer using 11 human serum samples. Only samples that are double positive for HAMA and anti-HAV antibodies were used, since HAMA interference would lead to false negative results in the competitive assay format. No HAMA interference was found within the predefined acceptance criteria.

#### Endogenous Interferences 4.4.

#### Hemoglobin/Bilirubin/Lipemia 4.4.1.

The purpose of this study was to evaluate endogenous substances for potential interference with the parameters measured on the Elecsys Anti-HAV II on the cobas e 601 immunoassay analyzer. For each interfering substance, four human serum samples were tested in accordance with CLSI EP07-A2.

#### 4.4.2. Rheumatoid Factor Interference

The recovery of analyte values in the presence of endogenous interfering substances was determined with the Elecsys Anti-HAV II on the cobas e 601 immunoassay analyzer.

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One aliquot of each serum sample was spiked with the interfering substance, and another aliquot with the same volume of the solvent (buffer matrix) of the interfering substance (dilution pool). The interfering pool was then diluted into the dilution pool in 10% increments.

The recovery for each sample was calculated by comparison to the reference sample.

#### Serum Albumin, IgG, IgA, and IgM Interference 4.4.3.

One aliquot of each serum sample was spiked with the interfering substance (lyophilisate). In this case, another aliquot without any additives (since the interfering substance is a lyophilisate) was used as dilution pool. The interfering pool was then diluted into the dilution pool in 14.3% increments.

The deviation or recovery for each sample was calculated by comparison to the reference sample.

### Conclusion:

All results met the pre-defined acceptance criteria, demonstrating no interference from endogenous substances up to the levels shown in the table below.

| Interferent | Interfering substance<br>measured up to | No interference<br>seen up to |
|-----------------------|-----------------------------------------|-------------------------------|
| Intralipid® (Lipemia) | 2000 mg/dL | 2000 mg/dL |
| Bilirubin | 66 mg/dL | 66 mg/dL |
| Hemoglobin | 1000 mg/dL | 1000 mg/dL |
| Rheumatoid Factor | 2000 IU/mL | 1400 IU/mL |
| human Serum albumin | 7.00 g/dL | 7.00 g/dL |
| human IgG | 7.00 g/dL | 7.00 g/dL |
| human IgM | 1.00 g/dL | 1.00 g/dL |
| human IgA | 1.60 g/dL | 1.60 g/dL |

Table 6: Summary of Results - Endogenous Interfering Substances

#### 4.5. Biotin

The purpose of this study was to evaluate biotin for potential interference with the parameters measured on the Elecsys Anti-HAV II on the cobas e 601 immunoassay analyzer. Five human serum samples were tested in accordance with CLSI EP07-A2. Negative specimens with biotin

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concentrations up to 100 ng/mL demonstrated ≤11% negative bias in COI values. Biotin concentrations greater than 100 ng/mL lead to higher negative bias and in consequence can lead to false positive Elecsys Anti-HAV II results. Refer to Limitations for additional information on biotin interference.

#### 4.6. Analytical Specificity/Cross-Reactivity

The effect on detection of analyte in the presence of potential cross-reacting antibodies using the Elecsys Anti-HAV II was determined on the cobas e 601 immunoassay analyzer with native human serum and plasma sample pools.

Anti-HAV negative samples containing potential cross-reacting antibodies to other infectious diseases were measured with Elecsys Anti-HAV and Elecsys Anti-HAV II. In total, 10 samples with acute Hepatitis B infection, 10 samples with acute Hepatitis C infection, 10 samples with HIV infection, 10 samples with EBV infection, 10 samples with Anti-CMV antibodies, 10 samples with Anti-HSV antibodies, 10 samples with Toxoplasma Gondii infection, 10 samples with Treponema Pallidum infection, 10 samples with Anti-Mumps/Rubeola antibodies, 10 samples with Anti-Rubella antibodies, 10 samples with Anti-Parvovirus B19 antibodies and 10 samples with ANA Autoimmune antibodies were tested.

No cross-reactivity with other infectious agents was found.

#### Exogenous Interferences – Drugs 4.7.

The recovery of analyte values in the presence of drugs was determined by comparing values obtained from samples spiked with 18 commonly used pharmaceutical compounds (Acetylcystein, Ampicillin-Na, Ascorbic acid, Ca-Dobesilate, Cyclosporine, Cefoxitin, Doxycycline, Heparin, Levodopa, Methyldopa +1.5, Metronidazole, Phenylbutazone, Tetracycline, Acetylsalicylic Acid, Rifampicin, Acetaminophen, Ibuprofen and Theophylline) with the reference sample (unspiked). Four human serum samples (native human serum pools) were used and tested on the cobas e 601 immunoassay analyzer according to CLSI EP07-A2. The drug concentrations tested correspond at least to the five times maximum daily doses. The four serum samples were divided into aliquots and spiked with the potential interferents. The reference sample without interferent was spiked with the respective amount of solvent only.

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The Elecsys Anti-HAV II concentration (as COI) of the spiked aliquots was determined in threefold determination and compared to the Elecsys Anti-HAV II result determined for the reference aliquot (also in three-fold determination) on one coba…