EliA RNA Pol III is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to RNA polymerase III (RNA Pol III) in human serum as an aid in the diagnosis of systemic sclerosis (diffuse form) in conjunction with other laboratory and clinical findings. EliA RNA Pol III uses the EliA IgG method.

Device Description

EliA RNA Pol III is a semi-quantitative solid-phase fluoroenzymeimmunoassay, for the determination of autoantibodies against RNA polymerase III. The EliA RNA Pol III test System is fully integrated and automated system which comprises of assay-specific reagents, EliA method-specific reagents, and general reagents.

AI/ML Overview

Here's a breakdown of the acceptance criteria and study information for the EliA RNA Pol III device, based on the provided text:

Acceptance Criteria and Device Performance

Acceptance Criteria Category	Specific Criteria	Reported Device Performance	Comments
Analytical Precision (Phadia 250)	Within-Run %CV	1.7% to 3.8%	Meets precision requirements.
	Between-Run %CV	1.4% to 2.3%	Meets precision requirements.
	Between-Instrument %CV	0.7% to 3.0%	Meets precision requirements.
	Lot-to-Lot %CV	0.5% to 2.2%	Meets precision requirements.
	Total Imprecision %CV	2.9% to 5.3%	Meets precision requirements.
Within-Lab Imprecision (Phadia 250)	Within-Run %CV	2.0% to 2.2%	Meets precision requirements.
	Between-Run %CV	1.4% to 2.6%	Meets precision requirements.
	Between-Day %CV	0.9% to 1.7%	Meets precision requirements.
	Total Within-Lab Imprecision %CV	2.8% to 3.3%	Meets precision requirements.
Analytical Precision (Phadia 2500/5000 E-module)	Within-Run %CV	2.7% to 4.9%	Meets precision requirements.
	Between-Run %CV	0.8% to 2.3%	Meets precision requirements.
	Between-Instrument %CV	2.0% to 5.9%	Meets precision requirements.
	Total Imprecision %CV	4.7% to 6.7%	Meets precision requirements.
Linearity/Assay Reportable Range	R^2 for dilution ranges	1.00 (Phadia 250), 1.00 (Phadia 2500E)	Linearity demonstrated across the entire measuring range.
Hook Effect/Over the Range	Not applicable	Results above upper limit reported as ">192".	No hook effect observed.
Traceability	IgG calibrators traceable to IRP 67/86 of Human Serum Immunoglobulins A, G and M from WHO.	Achieved traceability through comparison to secondary standard or IRP.	Meets traceability requirements.
Stability (Shelf-life)	EliA RNA Pol III Wells stability	18 months at 2-8°C	Meets stability requirements.
Stability (On-board stability)	EliA RNA Pol III carriers stability	28 days at 2-8°C	Meets stability requirements.
Stability (Open Stability)	EliA RNA Pol III wells after opening	9 months at 2-8°C	Meets stability requirements.
Detection Limit (LoB, LoD, LoQ)	LoD/LoQ (target 0.7 EliA U/mL)	LoB: 0.0 U/mL (both instruments); LoD: 0.1-0.2 U/mL; LoQ: 0.3-0.4 U/mL	All results below and in support of the harmonized limits of 0.7 EliA U/mL.
Analytical Specificity (Interference)	Ratio of blank/spiked sample (target 0.94-1.04)	Ranged from 0.94 – 1.04	No interference observed from tested substances up to specified concentrations.
Reference Sera	CDC samples detected according to target values.	All 12 CDC samples detected according to target.	Meets reference sera performance.
Assay Cut-Off (Equivocal results considered negative)	Positive Percent Agreement vs. Predicate Device (QUANTA LITE)	79.2% (95% CI: 65.0 - 89.5)	Comparison to predicate device.
	Negative Percent Agreement vs. Predicate Device (QUANTA LITE)	100% (95% CI: 89.7 - 100)	Comparison to predicate device.
	Total Agreement vs. Predicate Device (QUANTA LITE)	87.8% (95% CI: 78.7 - 94.0)	Comparison to predicate device.
Assay Cut-Off (Equivocal results considered positive)	Positive Percent Agreement vs. Predicate Device (QUANTA LITE)	87.5% (95% CI: 74.8 - 95.3)	Comparison to predicate device.
	Negative Percent Agreement vs. Predicate Device (QUANTA LITE)	100% (95% CI: 89.7 - 100)	Comparison to predicate device.
	Total Agreement vs. Predicate Device (QUANTA LITE)	92.7% (95% CI: 84.8 - 97.3)	Comparison to predicate device.
Clinical Sensitivity (SSc diffuse, equivocal results evaluated as positive)	Sensitivity	25.0% (95% CI: 18.0% - 33.3%)	Specific to diagnosis of systemic sclerosis (diffuse form).
Clinical Specificity (SSc diffuse, equivocal results evaluated as positive)	Specificity	99.1% (95% CI: 97.8% - 99.8%)	Specific to diagnosis of systemic sclerosis (diffuse form).
Clinical Sensitivity (SSc diffuse, equivocal results evaluated as negative)	Sensitivity	22.7% (95% CI: 15.9% – 30.8%)	Specific to diagnosis of systemic sclerosis (diffuse form).
Clinical Specificity (SSc diffuse, equivocal results evaluated as negative)	Specificity	99.6% (95% CI: 98.5% - 99.9%)	Specific to diagnosis of systemic sclerosis (diffuse form).

Study Details

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

Test Set (Method Comparison with Predicate Device):
- Sample Size: 193 patient serum samples.
- Data Provenance: Not explicitly stated, but includes 126 samples with a diagnosis of SSc. The context suggests these are clinical samples.
Test Set (Clinical Sensitivity and Specificity):
- Sample Size: 596 clinically defined serum samples.
- Data Provenance: Clinically defined samples from patients with various diagnoses, including Systemic Sclerosis (diffuse and limited forms), and various control diseases/conditions (e.g., Celiac disease, Crohn's disease, SLE, RA, bacterial/viral infections). The text does not specify country of origin or if prospective/retrospective; however, "clinically defined serum samples" typically implies retrospective collection with known diagnoses.
Test Set (Assay Cut-Off):
- Sample Size: 70 apparently healthy blood donor samples and 17 target disease (Systemic Sclerosis, diffuse) samples.
- Data Provenance: "apparently healthy blood donor samples" and "target disease samples" (Systemic Sclerosis, diffuse). The text explicitly mentions "sera from Caucasian, African American, Hispanic and Asian population obtained from a blood bank" for the frequency distribution, which likely overlaps with the "apparently healthy blood donor samples" used for cut-off establishment.

3. Number of Experts and Qualifications for Ground Truth (Test Set):

Number of Experts: Not specified.
Qualifications of Experts: Not specified. The wording "clinically defined serum samples with a diagnosis" implies that the diagnoses used as ground truth were established clinically by medical professionals, but their specific roles, number, or years of experience are not detailed.

4. Adjudication Method (Test Set):

Adjudication Method: Not specified. The diagnostic labels for the clinical samples are presented as established fact without mention of an adjudication process.

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

Was an MRMC study done? No. This device is an in-vitro diagnostic (IVD) assay designed for automated measurement of antibodies, not for interpretation by human readers. Therefore, an MRMC comparative effectiveness study regarding human reader improvement with AI assistance is not applicable.

6. Standalone (Algorithm Only) Performance:

Was a standalone study done? Yes. The entire performance evaluation, including analytical performance, method comparison, and clinical sensitivity/specificity, evaluates the performance of the EliA RNA Pol III assay itself (the "algorithm only" in this context, as it's an automated system) without human interpretation as part of the primary measurement. The results are generated directly by the instrument.

7. Type of Ground Truth Used:

For Method Comparison: The ground truth was the results from the predicate device, QUANTA LITE RNA POL III ELISA.
For Clinical Sensitivity and Specificity: The ground truth was clinical diagnosis ("clinically defined serum samples with a diagnosis from patients with systemic sclerosis, diffuse," etc.).
For Assay Cut-Off: The ground truth was based on apparently healthy blood donors and clinically diagnosed Systemic Sclerosis, diffuse patients.

8. Sample Size for the Training Set:

Training Set (Assay Cut-Off establishment): This involved 70 apparently healthy blood donor samples and 17 target disease (Systemic Sclerosis, diffuse) samples. While not explicitly called a "training set," these samples were used to "define the cut-off," which is a form of model training/optimization for classification.
Other "training" data: The document mentions that "New batches of IgG calibrators are compared to a secondary standard (standardized with the IRP) or the IRP directly and adjusted accordingly to meet the correct concentration." This implies a continuous process of calibration and standardization, where previous data/standards (IRP) could be considered a form of "training" for the calibrators. However, a distinct, large-scale training set for an AI/algorithm in the conventional sense is not detailed as this is a chemical assay.

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

For Assay Cut-Off establishment: The ground truth for the 70 apparently healthy blood donors means they were confirmed healthy (presumably through standard screening). For the 17 Systemic Sclerosis (diffuse) samples, the ground truth was their clinical diagnosis.
For Calibrators: The ground truth for calibrators is established through their traceability to the International Reference Preparation (IRP) 67/86 of Human Serum Immunoglobulins A, G and M from WHO.

Summary

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September 13, 2021

Phadia AB % Sheryl Skinner Associate Director Quality and Regulatory Phadia US Inc. 4169 Commercial Avenue Portage, Michigan 49002

Re: K202541

Trade/Device Name: EliA RNA Pol III Regulation Number: 21 CFR 866.5100 Regulation Name: Antinuclear Antibody Immunological Test System Regulatory Class: Class II Product Code: NYO Dated: March 4, 2021 Received: March 5, 2021

Dear Sheryl Skinner:

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

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542 of the Act); 21 CFR 1000-1050.

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

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

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

Sincerely,

Ying Mao, Ph.D. Chief Division of Immunology and Hematology Devices OHT7: Office of In Vitro Diagnostics and Radiological Health Office of Product Evaluation and Quality Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K202541

Device Name EliA RNA Pol III

Indications for Use (Describe)

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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| > Prescription Use (Part 21 CFR 801 Subpart D)

Over-The-Counter Use (21 CFR 801 Subpart C)

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This 510(k) Summary is prepared in accordance with the requirements of 21 CFR Part 807.92.

Premarket Notification 510(k) No: K202541

Date of Summary Preparation: September 3, 2021

Manufacturer:	Phadia ABRapsgatan 7PP.O. Box 6460751 37 Uppsala, Sweden
Distributor:	Phadia US Inc.4169 Commercial AvenuePortage, MI 49002
Company Contact Person:	Sheryl SkinnerAssociate Director, Regulatory and QualityPhadia US Inc.4169 Commercial Avenue(269) 568-3603sheryl.skinner@thermofisher.com

Proprietary and Established Device Name:

EliA RNA Pol III

Regulatory Information:

Product Code:	NYO
Classification:	Class II
Regulation:	21 CFR 866.5100 – Antinuclear Antibody ImmunologicalTest System
Panel:	Immunology

Purpose of Submission:

New Device

Measurand:

lgG autoantibodies specific to RNA polymerase III (RNA Pol III) proteins

Type of Test:

Automated semi-quantitative solid phase fluoroenzymeimmunoassay

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Intended Use:

EliA RNA Pol III is intended for the in vitro semi-quantitative measurement of IgG antibodies directed to RNA polymerase III (RNA Pol III) in human serum as an aid in the diagnosis of systemic sclerosis (diffuse form) in coniunction with other laboratory and clinical findings. EliA RNA Pol III uses the EliA IgG method.

Indication(s) for Use:

Same as intended use

Special Conditions for Use:

Rx - For Prescription Use Only

Special Instrument Requirements:

For use on the Phadia 250 instrument and the Phadia 2500 and Phadia 5000 instrument series (E-modules).

Device Description:

EliA RNA Pol III is a semi-quantitative solid-phase fluoroenzymeimmunoassay, for the determination of autoantibodies against RNA polymerase III. The EliA RNA Pol III test System is fully integrated and automated system which comprises of assayspecific reagents, EliA method-specific reagents, and general reagents.

Assay-Specific Reagents include:

EliA RNA Pol III Wells: coated with human recombinant RNA polymerase III . proteins - 2 carriers (12 wells each), ready to use;
. EliA ANA 3 Positive Control 250 or 2500/5000: Human monoclonal antibodies in Tris buffer containing IqG antibodies to Ro52. Rib-P and RNA Pol III - 6 single use vials, 0.3 mL each, ready to use;
. EliA IgG/IgM/IgA Negative Control 250 or 2500/5000: Human blood preparation from healthy donors in PBS containing BSA, detergent and 0.095% sodium azide - 6 single-use vials, 0.3 mL each, ready to use;

EliA Method-Specific Reagents include:

. EliA Sample Diluent: PBS containing BSA, detergent and 0.095% sodium azide - 6 bottles, 48 mL each, ready to use; or 6 bottles, 400 mL each, ready to use;
. EliA IgG Conjugate 50 or 200: ß-Galactosidase labeled anti-lgG (mouse monoclonal antibodies) in PBS containing BSA and 0.06% sodium azide - 6 wedge shaped bottles, 5 mL each, ready to use; or 6 wedge shaped bottles, 19 mL each, ready to use;
. EliA IgG Calibrator Strips: Human IgG (0, 4, 10, 20, 100, 600 µg/L) in PBS containing BSA, detergent and 0.095% sodium azide - 5 strips, 6 single-use vials per strip, 0.3 mL each, ready to use:
. EliA IgG Curve Control Strips: Human IgG (20 ug/L) in PBS containing BSA, detergent and 0.095% sodium azide - 5 strips, 6 single-use vials per strip, 0.3 mL each, ready to use;
트 EliA IgG Calibrator Well: coated with mouse monoclonal antibodies – 4 carriers (16 wells each), ready to use;

Phadia AB, Rapsgatan 7P, P.O. Box 6460, 751 37 Uppsala, Sweden

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General Reagents include:

Development Solution: 0.01% 4-Methylumbelliferyl-β-D-galactoside, <0.0010% . preservative - 6 bottles (11 mL, 17 mL, or 112 mL each), sufficient for 6x >110, 6x >170. or 6x >1165 determinations:
.. Stop Solution: 4% Sodium Carbonate – 6 bottles (65 mL, 119 mL, or 2800 mL each), sufficient for 6x >292, 6x >560, or 6x >13100 determinations;
. Washing Solution Additive: detergent, preservative <0.13% - 6x 17.2 mL, 2x 86mL, or 4x 850 mL;
. Washing Solution Concentrate: phosphate buffer – 6x 80 mL, 2x 400 mL, or 1x 2800 mL.

Instrument System

EliA RNA Pol III is run on the Phadia 250 instrument and the E-module of the Phadia 2500 and Phadia 5000 instruments series. The instruments are automated platforms for EliA test procedures from sample and reagent handling to the processing of results.

Substantial Equivalence

QUANTA LITE RNA POL III ELISA, INOVA Diagnostics, Inc. (K070066)

	Similarities
Feature	Proposed DeviceEliA RNA Pol III	Predicate DeviceQUANTA LITE RNA POL III ELISA
Intended Use	EliA RNA Pol III is intended forthe in vitro semi-quantitativemeasurement of IgG antibodiesdirected to RNA polymerase III(RNA Pol III) in human serum asan aid in the diagnosis ofsystemic sclerosis (diffuse form)in conjunction with otherlaboratory and clinical findings.EliA RNA Pol III uses the EliAlgG method.	The QUANTA LITE RNA POL IIIELISA is a semiquantitativeenzyme-linked immunosorbentassay for the detection of IgGanti-RNA Polymerase IIIantibodies in patient sera. Thepresence of these antibodies,when considered in conjunctionwith other laboratory and clinicalfindings, is an aid in thediagnosis of systemic sclerosis(scleroderma) with increasedincidence of skin involvementand renal crisis.
Internal Controls	Positive and negative Controlprovided with EliA ANA 3Positive Control 250 and2500/5000 and EliA IgG/IgM/IgANegative Control 250 and2500/5000, resp.	Low Positive, High Positive andNegative Controlincluded in the kit
Assay technique	ELISA	Same

Comparison with Predicate Device:

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S C

Traditional 510(k) for EliA RNA Pol III

A.6 510k Summary_V6

Type of test	Semi-quantitative	Same
Reported Unit	EliA U/mL (arbitrary)	Units (arbitrary)

	Differences
Feature	Proposed Device	Predicate Device
	EliA RNA Pol III	QUANTA LITE RNA POL III ELISA
Antigen	Human recombinant RNApolymerase III protein	Purified RNA Pol III antigen
Instrumentation	EliA RNA Pol III uses the EliAIgG method on the instrumentsPhadia 250 and the E-modulesof the Phadia 2500 and Phadia5000 series.	ELISA-Reader needed
Sample Dilution	1:200	1:101
Reactiontemperature	37°C controlled	Room temperature, 20-26°C
Detectionantibody(conjugate)	IgG conjugate: anti-human IgGβ-Galactosidase (mousemonoclonal antibodies)	IgG conjugate: HRP IgGConjugate, (goat), anti-humanIgG
Signal	Fluorescence	Optical density
Calibration	6-point total IgG Calibration6 vials of human IgG atconcentrations of0 – 4 – 10 – 20 – 100 – 600µg/L	One-point calibration
Calibration curve	Option to store curve for up to28 days and run curve controlsin each assay for calibration	N/A
Interpretation ofresults	Negative < 7 EliA U/mLEquivocal 7-10 EliA U/mLPositive > 10 EliA U/mL	Negative < 20 UnitsWeak positive 20-39 UnitsModerate positive 40-80 UnitsStrong positive > 80 Units
Substrate	Development Solution 0.01 %4-Methylumbelliferyl-β-D-galactoside,<0.0010% preservative**Preservative: mixture of 5-chloro-2-methyl-2H-isothiazol-3-one [EC no. 247-500-7] and 2-methyl-2H-isothiazol-3-one [ECno. 220-239-6] (3:1)	TMB Chromogen

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Traditional 510(k) for EliA RNA Pol III

A.6 510k Summary V6

K202541.510kSummary.FINAL.docx

Standard/Guidance Document Referenced

CLSI EP25-A, Evaluation of Stability of In Vitro Diagnostic Reagents, . September 2009
. CLSI EP05-A3. Evaluation of Precision of Quantitative Measurement Procedures, September 2014
. CLSI EP6-A, Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach, April 2003
트 CLSI EP07-ED3, Interference Testing in Clinical Chemistry, September 2018
. CLSI EP37-ED1, Supplemental Tables for Interference Testing, September 2018
. CLSI EP09c-ED3, Measurement Procedure Comparison and Bias Estimation Using Patient Samples, June 2018
. CLSI EP17-A2, Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures, June 2012
CLSI EP28-A3c, Defining, Establishing, and Verifying Reference Intervals in . the Clinical Laboratory, October 2010

Test Principle

The EliA tests are fluorescence immunoassays for the detection and measurement of human antibodies based on EliA solid-phase components, which contain specific antigens for the antibodies to be measured.

The specific antigen for the antibodies to be detected is bound to the EliA solid phase component (EliA Well). The EliA wells are molded cups comparable to excised wells from a microtiter plate. They are made of polystyrene and are coated with the respective antigen. The wells are at the same time a holder of the coupled antigen for convenient automation and a reaction chamber with reaction/washing solution handling based on pipetting to add and aspiration to remove liquids. If present in the patient's specimen, antibodies to these proteins bind to their specific antigen. After washing away non-bound antibodies, enzyme-labeled antibodies against human IgG antibodies (EliA IgG Conjugate) are added to form an antibody-conjugate complex. After incubation, non-bound conjugate is washed away, and the bound complex is incubated with a Development Solution. After stopping the reaction, the fluorescence in the reaction mixture is measured. The assay directly measures the amount of antibody of interest bound to the antigen coating the EliA well, therefore the higher the value of fluorescent signal detected by the instrument, the higher the amount of antibody bound and detected in the sample tested. To evaluate test results, the response for patient samples is compared directly to the response for calibrators.

Performance Characteristics

1. Analytical performance:
- a) Precision/Reproducibility:

To determine the precision of the assay on the Phadia 250 instrument and the Phadia 2500 and Phadia 5000 instrument series, the variability was assessed on 5 samples.

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Three lots were used to determine the precision of the assay on Phadia 250 (totaling 252 replicate determinations per sample).

One lot was used to determine the precision of the assay on Phadia 2500E. which is a representative of the Phadia 2500 and Phadia 5000 instrument series.

EliA RNA Pol III on Phadia 250:

To determine the precision of the assay on the Phadia 250 instrument, the variability was assessed in a study with 21 runs by examining the samples in 252 replicates across 3 lots and 3 Phadia 250 instruments over 7 days. The data was calculated against the calibration curve from Day 1. The statistical evaluation was performed by Analysis of Variance. The results are given in the table below.

Mean EliA	Within-Run		Between-Run		Between-Instrument		Lot-to-Lot		Total Imprecision
U/mL	SD	%CV	SD	%CV	SD	%CV	SD	%CV	SD	%CV
3.1	0.1	2.3	0.1	2.2	0.1	3.0	0.1	1.5	0.2	4.6
7.5	0.1	1.7	0.1	1.4	0.2	2.3	0.1	0.7	0.3	3.3
9.1	0.2	2.0	0.2	1.9	0.1	1.5	0.0	0.5	0.3	3.2
19.2	0.4	2.1	0.3	1.7	0.1	0.7	0.2	0.9	0.6	2.9
171.9	6.6	3.8	4.0	2.3	3.4	2.0	3.8	2.2	9.2	5.3

Within-lab imprecision

To determine the within-lab precision of the assay, the variability was assessed in a study with 40 runs by examining the samples in 80 replicates on 1 instrument over 20 days. The data was calculated against the calibration curve from Day 1. The statistical evaluation was performed by Analysis of Variance. The results are given in the table below.

Mean(EliAU/mL)	Within-Run		Between-Run		Between-Day		TotalWithin-LabImprecision
	SD	%CV	SD	%CV	SD	%CV	SD	%CV
4.2	0.1	2.2	0.1	1.8	0.0	1.0	0.1	2.8
7.3	0.1	2.0	0.2	2.6	0.1	1.7	0.2	3.3
9.7	0.2	2.0	0.2	2.1	0.1	1.1	0.3	2.8
39.4	0.8	2.1	0.5	1.4	0.3	0.9	1.0	2.5

EliA RNA Pol III on the Phadia 2500 and Phadia 5000 instrument series: To determine the precision of the assay on the E-module of the Phadia 2500 and Phadia 5000 instrument series (E-module), the variability was assessed in a study with 21 runs by examining the samples in 84 replicates on 3 Phadia 2500E instruments over 7 days. The data was calculated against the calibration curve from Day 1.

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Image /page/9/Picture/0 description: The image contains the logo for Thermo Fisher Scientific. The words "Thermo Fisher" are in red, stacked on top of the word "SCIENTIFIC" which is in black. The logo is simple and clean, with a focus on the company name.

Traditional 510(k) for EliA RNA Pol III A.6 510k Summary V6

K202541.510kSummary.FINAL.docx

The statistical evaluation was performed by Analysis of Variance. The results are given in the table below.

MeanEliA	Within-Run		Between-Run		Between-Instrument		TotalImprecision
U/mL	SD	%CV	SD	%CV	SD	%CV	SD	%CV
3.0	0.1	4.7	0.1	2.0	0.1	2.3	0.2	5.6
7.5	0.2	2.7	0.2	2.0	0.3	3.3	0.4	4.7
9.1	0.4	4.0	0.1	1.1	0.3	3.8	0.5	5.6
27.0	0.9	3.2	0.2	0.8	1.6	5.9	1.8	6.7
170.4	8.3	4.9	4.0	2.3	3.4	2.0	9.8	5.8

b) Linearity/Assay Reportable Range:

3 serum samples were diluted in EliA Sample Diluent and tested on Phadia 250 and Phadia 2500E. The ratios of observed/expected values were calculated. The results are summarized below.

Phadia 250

Dilution Range(EliA U/mL)	Slope	Intercept	R2
0.8 – 31.7	0.99	-0.28	1.00
3.1 - 245.3	1.01	0.96	1.00
1.8 - 172.5	0.99	0.38	1.00

Phadia 2500E

Dilution Range(EliA U/mL)	Slope	Intercept	R2
0.7 – 28.3	1.02	0.19	1.00
3.0 - 270.3	1.01	-0.92	1.00
1.9 - 144.0	1.02	1.40	1.00

Linearity was shown for the entire measuring range.

Hook Effect/Over the Range Results:

Not applicable. Results above the upper limit of the measuring range are reported as ">192". No recommendations are made for dilution of samples outside measuring range in the Directions For Use.

c) Traceability, Stability, Expected Values (Controls, Calibrators, or Methods): Traceability:
The IgG calibrators are traceable (via unbroken chain of calibrations) to the International Reference Preparation (IRP) 67/86 of Human Serum Immunoqlobulins A. G and M from WHO. New batches of IgG calibrators are compared to a secondary standard (standardized with the IRP) or the IRP directly and adjusted accordingly to meet the correct concentration.

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The instrument measures specific IgG concentrations in uq/L. By using a conversion factor given by the lot-specific code of the EliA test well, the results are automatically converted to EliA U/mL.

Stability:

Data for open and closed real-time stability and on-board stability of EliA IgG reagents and general EliA reagents on Phadia 250 as well as on the Emodule of the Phadia 2500 and Phadia 5000 series were previously cleared with several other EliA tests, e.g. under K141375 (EliA M2 on Phadia 250). For the E-module of the Phadia 2500 and Phadia 5000 series, they were already cleared under K061165/A003 (EliA CCP).

Shelf-life:

The stability of EliA RNA Pol III Wells was evaluated with a real-time study. The results support stability of the test under the recommended storage of 2 - 8°C for up to 18 months.

On-board stability:

The on-board stability EliA RNA Pol III carriers (containing the antigen coated wells) was tested over 6 weeks using 3 positive and 2 negative samples only on the Phadia 250 instrument. As the storage conditions in the E-module of the Phadia 2500 and Phadia 5000 series are similar to the Phadia 250, the results can also be used for stability claims for these instruments. The on-board stability for the Phadia 250 was determined to be 28 days at 2-8°C.

Open Stability:

Stability after first opening of the foil bag containing the EliA RNA Pol III wells was initially tested with an accelerated and then verified in a real-time study. The real-time study confirmed the shelf-life of 9 months at 2-8°C after first opening which was assigned to EliA RNA Pol III wells.

d) Detection Limit:

Four blank and four low level samples were measured with two different reagent sets (two lots of antigen wells). The four blank samples were created from depleted IgG sera, each diluted with EliA Sample Diluent. The blank samples and the low-level samples were assayed in three runs using two different sets of EliA RNA Pol III Well lots over three different days on a Phadia 250 and Phadia 2500E each in 5-fold determination. For each instrument type, the total number of combined observations for blank and low-level samples is 120 (60 per reagent set, 15 per sample and

reagent set).

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The results are summarized in the table below:

Instrument	LoBEliA U/mL	LoDEliA U/mL	LoQEliA U/mL
Phadia 250	0.0	0.2	0.4
E-module of the Phadia 2500and Phadia 5000 series	0.0	0.1	0.3

The presented data have been acquired according to the CLSI Guideline EP17. The results are all below, and therefore in support of the current worldwide harmonized limits (LoD=LoQ, both set to 0.7 EliA U/mL).

e) Analytical specificity:
Endogenous and Exogenous Interference:

A study was run to investigate whether high concentrations of potentially interfering substances in serum, like bilirubin, hemoglobin, lipemic factor, rheumatoid factor, Ibuprofen, Losartan, Hydroxychloroquine, Azathioprine, Prednisone, Rituximab and Infliximab adversely affect the results of the new device.

Three serum samples (one negative sample, one sample with a concentration within the equivocal range, and one high positive sample) were prediluted in EliA Sample Diluent and spiked with the different interfering substances or blank solution. The samples were tested in triplicates. A calibration curve was run in duplicate. The runs were repeated twice. One batch of EliA antigen wells and one batch of system reagents were used throughout the studies.

The ratio of blank/spiked sample ranged from 0.94 – 1.04 for EliA RNA Pol III. No interference was observed up to the concentrations listed in the table below:

Potential InterferingCompound	Concentration inundiluted sample
Bilirubin F	40 mg/dL
Bilirubin C	40 mg/dL
Hemoglobin	1000 mg/dL
Lipemic factor	2000 mg/dL
Rheumatoid factor	500 IU/mL
Ibuprofen	21.9 mg/dL
Losartan	1.14 mg/dL
Hydroxychloroquine	0.23 mg/dL
Azathioprine	0.26 mg/dL
Prednisone	0.01 mg/dL
Rituximab	109 mg/dL
Infliximab	26.4 mg/dL

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Reference Sera:

Externally defined sera should be measured according to their target values as mentioned by the institution CDC. Using EliA RNA Pol III, all 12 CDC samples were detected according to their target.

Assay Cut-Off:

To define the cut-off, a study was performed using a cohort consisting of 70 apparently healthy blood donor samples and 17 target disease (Systemic Sclerosis, diffuse) samples. The samples were measured on a Phadia 250 instrument.

The cut-off was set as follows for EliA RNA Pol III:

<7 EliA U/mL	Negative
7-10 EliA U/mL	Equivocal
>10 EliA U/mL	Positive

In case of equivocal results, it is recommended to retest the patient after 8-12 weeks.

1. Comparison Studies:
- a) Method Comparison with Predicate Device:

A total of 193 patient serum samples, including 126 with the diagnosis of SSc, were used during this study. 82 patient samples were found within the measuring range, including 50 with the diagnosis of Systemic Sclerosis. The samples were analyzed with the EliA RNA Pol III and QUANTA LITE RNA POL III ELISA assay. The test was run in single determination and evaluated according to the Directions for Use. The results are summarized in the tables below:

EliA RNA Pol III: equivocal results considered negative

N = 82	QUANTA LITE RNAPOL III ELISApositive:≥ 20 Units	QUANTA LITERNA POL IIIELISA negative:< 20 Units	Total
EliA RNA Pol IIIpositive:> 10 EliA U/mL	38	0	38
EliA RNA Pol IIInegative:< 10 EliA U/mL	10	34	44
Total	48	34	82
	Calculation	Agreement (%)	95% CI

Confidential Page 10 (14)

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Traditional 510(k) for EliA RNA Pol III A.6 510k Summary_V6 K202

Total Agreement

2541.510kSummary.FINAL.docx
Positive PercentAgreement	$100% x 38 / 48$	79.2%
Negative PercentAgreement	$100% x 34 / 34$	100%

100% x (38 + 34) / 82

EliA RNA Pol III: equivocal results considered positive

N = 82	QUANTA LITERNA POL IIIELISA positive:≥ 20 Units	QUANTA LITERNA POL IIIELISA negative:< 20 Units	Total
EliA RNA Pol IIIpositive:>7 EliA U/mL	42	0	42
EliA RNA Pol IIInegative:< 7 EliA U/mL	6	34	40
Total	48	34	82
	Calculation	Agreement (%)	95% CI
Positive PercentAgreement	$100% x 42 / 48$	87.5%	74.8 - 95.3
Negative PercentAgreement	$100% x 34 / 34$	100%	89.7 - 100
Total Agreement	$100% x (42 + 34) / 82$	92.7%	84.8 - 97.3

b) Instrument Comparison:

Performance of EliA RNA Pol III was evaluated on the Phadia 250 and Phadia 2500E instrument using 64 positive, 13 equivocal and 23 negative serum samples.

The samples were analyzed in single determination on one Phadia 250 and one Phadia 2500E instrument each. The regression analysis results are summarized as follows:

	Intercept	Slope
Estimate	-0.67	1.05
95% Cl	-1.13 to -0.44	1.04 to 1.07

65.0 - 89.5

89.7 - 100 78.7 - 94.0

87.8%

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1. Clinical Studies:
- a) Clinical Sensitivity and Specificity:

596 clinically defined serum samples with a diagnosis from patients with systemic sclerosis, diffuse (n = 132), Systemic Sclerosis, limited (n = 91), Systemic Sclerosis/Raynaud Syndrome (n = 13), Celiac disease (n = 13), Crohn's disease (n = 12), CTD overlap Non-MCTD (n = 10), Dermatomyositis (n = 4), Polymyositis (n = 6), Graves' disease (n = 12), Primary antiphospholipid syndrome (n = 12), Primary biliary cirrhosis (n = 21), Sjögren's syndrome (n = 30), Systemic Lupus Erythematosus (n = 16), Type 1 Diabetes (n = 12), Ulcerative colitis (n = 11), cancer (n = 30), Mixed connective tissue disease (n = 10), Rheumatoid arthritis (n = 40), bacterial infections (n = 35), viral infections (n = 72), Hashimoto's disease (n = 10), Granulomatosis with Polyangiitis (n = 4) were used to determine sensitivity and specificity of the assay. The results are summarized in the tables below.

n=596	DiagnosticGroup	Disease Controls	Total
Positive test≥ 7 EliA U/mL	33	4	37
Negative test< 7 EliA U/mL	99	460	559
Total	132	464	596

EliA RNA Pol III - equivocal results evaluated as positive:

EliA RNA Pol III - equivocal results evaluated as negative:

Specificity (95% CI): 99.1% (97.8% - 99.8%)

n=596	DiagnosticGroup	DiseaseControls	Total
Positive test> 10 EliA U/mL	30	2	32
Negative test≤ 10 EliA U/mL	102	462	564
Total	132	464	596

Sensitivity (95% CI): 22.7% (15.9% – 30.8%) Specificity (95% CI): 99.6% (98.5% - 99.9%)

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Traditional 510(k) for EliA RNA Pol III A.6 510k Summary_V6

K202541.510kSummary.FINAL.docx

The table below shows the results for each clinical subgroup:

Diagnosis	totaln	positiven	positive%
Systemic Sclerosis, diffuse	132	30	23%
Target disease (Total)	132	30
Systemic Sclerosis, limited	91	1	1%
Systemic Sclerosis, RaynaudSyndrome	13	0	0%
Celiac disease	13	0	0%
Crohn's disease	12	0	0%
CTD overlap non-MCTD	10	0	0%
Dermatomyositis	4	0	0%
Polymyositis	6	0	0%
Grave's disease	12	0	0%
Primary antiphospholipid syndrome	12	0	0%
Primary biliary cirrhosis	21	0	0%
Sjögren's syndrome	30	0	0%
Systemic lupus erythematosus	16	0	0%
Type 1 Diabetes	12	0	0%
Ulcerative colitis	11	0	0%
Varied cancer	30	0	0%
Mixed connective tissue disease	10	0	0%
Rheumatoid arthritis	40	0	0%
Hashimoto's disease	10	0	0%
Granulomatosis with Polyangiitis	4	0	0%
Bacterial infections (Total)	35	0	0%
Viral infections (Total)	72	1	1%
Control disease	464	2
Total	596

b) Other Clinical Supportive Data: Not applicable.
1. Clinical Cut-Off: Same as assay cut-off.
1. Expected Values/Reference Range: Antibody prevalence in autoimmune patients varies widely depending on disease area. The proportion of sera from a normal population found positive for the antinuclear antibodies covered by the EliA RNA Pol III test is below 1%. Expected values may vary depending on the population tested.

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Traditional 510(k) for EliA RNA Pol III A.6 510k Summary_V6

K202541.510kSummary.FINAL.docx

The frequency distribution for antinuclear antibodies was investigated in a group of apparently healthy subjects equally distributed by age and gender, using sera from Caucasian, African American, Hispanic and Asian population obtained from a blood bank.

The results are given in the table below:

Test	n	MedianEliA U/mL	95th percentileEliA U/mL	99th percentileEliA U/mL
EliA RNA Pol III	568	<0.7	1.6	3.3

Proposed Labeling

The labeling is drafted in accordance with the requirements of 21 CFR Part 809.10.

Conclusion

All available data support that both immunoassays, the new device EliA RNA Pol III and its predicate device QUANTA LITE RNA POL III ELISA perform substantially equivalent.

Regulation Number and Section

§ 866.5100 Antinuclear antibody immunological test system.

(a)
Identification. An antinuclear antibody immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoimmune antibodies in serum, other body fluids, and tissues that react with cellular nuclear constituents (molecules present in the nucleus of a cell, such as ribonucleic acid, deoxyribonucleic acid, or nuclear proteins). The measurements aid in the diagnosis of systemic lupus erythematosus (a multisystem autoimmune disease in which antibodies attack the victim's own tissues), hepatitis (a liver disease), rheumatoid arthritis, Sjögren's syndrome (arthritis with inflammation of the eye, eyelid, and salivary glands), and systemic sclerosis (chronic hardening and shrinking of many body tissues).(b)
Classification. Class II (performance standards).