Elecsys Folate III

K233060 · Roche Diagnostics · CGN · Jun 17, 2024 · Clinical Chemistry

Device Facts

• Record ID: K233060
• Device Name: Elecsys Folate III
• Applicant: Roche Diagnostics
• Product Code: CGN · Clinical Chemistry
• Decision Date: Jun 17, 2024
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1295
• Device Class: Class 2

Indications for Use

Binding assay for the in vitro quantitative determination of folate in erythrocytes (red blood cells, RBC). Folate measurements are used in the diagnosis and treatment of anemia. The electrochemiluminescence immunoassay, "ECLIA" is intended for use on cobas e immunoassay analyzers.

Device Story

Elecsys Folate III is an in vitro diagnostic binding assay for quantitative folate measurement in erythrocytes. Input: whole blood samples treated with anticoagulants (heparin or EDTA). Process: samples are mixed with ascorbic acid, incubated (~90 min, 20-25 °C) to lyse erythrocytes and stabilize intracellular folate; resulting hemolysate is analyzed via competitive electrochemiluminescence immunoassay (ECLIA) on cobas e immunoassay analyzers. Output: folate concentration determined via instrument-specific calibration curve. Used in clinical laboratories by trained personnel. Results assist clinicians in diagnosing and managing anemia. Benefits include standardized, automated quantification of folate levels to support clinical decision-making.

Clinical Evidence

Bench testing only. Precision evaluated per CLSI EP05-A3 (21-day and 5-day multi-site/multi-lot studies). Linearity assessed per CLSI EP06-Ed2 (103-656 ng/mL range). Interference testing (endogenous/exogenous) performed per CLSI EP07-Ed3. Method comparison (N=119) against predicate showed Passing-Bablok slope 1.04, intercept 14.6, r=0.991. LoB/LoD/LoQ determined per CLSI EP17-A2.

Technological Characteristics

Electrochemiluminescence immunoassay (ECLIA). Reagents: pretreatment reagents, streptavidin-coated microparticles, ruthenium-labeled folate binding protein, biotin-labeled folate. Analyzed on cobas e 801. Measuring interval: 120-620 ng/mL. Biotin tolerance: 1200 ng/mL. Calibration: 2-point calibration with master curve via cobas link.

Indications for Use

Indicated for the in vitro quantitative determination of folate in erythrocytes (red blood cells, RBC) to aid in the diagnosis and treatment of anemia in patients.

Regulatory Classification

Identification

A folic acid test system is a device intended to measure the vitamin folic acid in plasma and serum. Folic acid measurements are used in the diagnosis and treatment of megaloblastic anemia, which is characterized by the presence of megaloblasts (an abnormal red blood cell series) in the bone marrow.

Predicate Devices

• Roche Diagnostics Elecsys Folate RBC (K103716)

Submission Summary (Full Text)

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FDA U.S. FOOD & DRUG ADMINISTRATION

# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

ASSAY ONLY

## I Background Information:

A 510(k) Number

K233060

B Applicant

Roche Diagnostics

C Proprietary and Established Names

Elecsys Folate III

D Regulatory Information

| Product Code(s) | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| CGN | Class II | 21 CFR 862.1295 - Folic Acid Test System | CH - Clinical Chemistry |

## II Submission/Device Overview:

A Purpose for Submission:

Modified device

B Measurand:

Folate

C Type of Test:

Quantitative; electrochemiluminescence immunoassay

Food and Drug Administration

10903 New Hampshire Avenue

Silver Spring, MD 20993-0002

www.fda.gov

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## III Intended Use/Indications for Use:

### A Intended Use(s):
See Indications for Use below.

### B Indication(s) for Use:
Binding assay for the in vitro quantitative determination of folate in erythrocytes (red blood cells, RBC). Folate measurements are used in the diagnosis and treatment of anemia. The electrochemiluminescence immunoassay, "ECLIA" is intended for use on cobas e immunoassay analyzers.

### C Special Conditions for Use Statement(s):
Rx - For Prescription Use Only

For in vitro diagnostic use

### D Special Instrument Requirements:
Cobas e 801 analyzer

## IV Device/System Characteristics:

### A Device Description:
The Elecsys Folate III (RBC Application) is a modified version of the predicate assay to increase tolerance to biotin interference. The Elecsys Folate III (RBC Application) reagent pack includes the following reagents:

- PT1 Pretreatment reagent 1
- PT2 Pretreatment reagent 2
- M Streptavidin-coated microparticles
- R1 Folate-binding protein~Ru (bpy)
- R2 Folate~biotin

Other materials required to run the assay, but not included in the kit include: CalSet Folate, PreciControl Varia and Folate RBC Hemolyzing Reagent.

### B Principle of Operation:
Elecsys Folate III (RBC Application) is a competition principle assay with a total duration of 27 minutes. Whole blood treated with anticoagulants is mixed with ascorbic acid solution and incubated at 20-25 °C for lysis of the erythrocytes to take place, with liberation and stabilization of the intracellular folate. Fifteen (15 µL) of resulting hemolysate sample is incubated with the folate pretreatment reagents 1 and 2. Bound folate is released from endogenous folate binding proteins. The pretreated sample is then incubated with the ruthenium-labeled folate binding protein forming a folate complex, the amount of which is dependent upon the analyte

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concentration in the sample. After addition of streptavidin-coated microparticles and folate labeled with biotin, the unbound sites of the ruthenium labeled folate binding protein become occupied, with formation of a ruthenium labeled folate binding protein-folate biotin complex. The entire complex becomes bound to the solid phase via interaction of biotin and streptavidin. The reaction mixture is aspirated into the measuring cell where the microparticles are magnetically captured onto the surface of the electrode. Unbound substances are then removed with ProCell II M. Application of a voltage to the electrode then induces chemiluminescent emission which is measured by a photomultiplier. Results are determined via a calibration curve which is instrument-specifically generated by 2-point calibration and a master curve provided via the cobas link.

V Substantial Equivalence Information:

A Predicate Device Name(s): Elecsys Folate RBC

B Predicate 510(k) Number(s): K103716

C Comparison with Predicate(s):

| Device & Predicate Device(s): | K233060 | K103716 |
| --- | --- | --- |
| Device Trade Name | Elecsys Folate III | Elecsys Folate RBC |
| General Device Characteristic Similarities | | |
| Intended Use/Indications For Use | Quantitative determination of folate. Measurements are used in the diagnosis and treatment of anemia. | Same |
| Assay Technology | Electro-chemiluminescence immunoassay | Same |
| Analytical Measuring Interval | 120 - 620 ng/mL | Same |
| Sample Matrix | Hemolysate | Same |
| General Device Characteristic Differences | | |
| Biotin tolerance | ≤ 1200 ng/mL | ≤ 21 ng/mL |

VI Standards/Guidance Documents Referenced:

CLSI EP05-A3: Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline - Third Edition

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CLSI EP06 2nd Edition: Evaluation of the Linearity of Quantitative Measurement Procedures - 2nd Edition

CLSI EP07-Ed3: Interference Testing in Clinical Chemistry - 3rd Edition

CLSI EP09c: Method Procedure Comparison and Bias Estimation Using Patient Samples; Approved Guideline – Third Edition

CLSI EP17-A2: Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures; Approved Guideline - Second Edition

## VII Performance Characteristics (if/when applicable):

### A Analytical Performance:

#### 1. Precision/Reproducibility:

Precision studies were conducted for both 21 days and 5 days with the Elecsys Folate III (RBC Application) to evaluate repeatability (within-run precision) and intermediate precision (within-laboratory precision) according to the CLSI guideline EP05-A3.

#### Repeatability and Intermediate Precision (21 Days)

Precision was evaluated on one cobas e 801 analyzer with one reagent lot. Five human hemolysate samples prepared from whole blood samples collected in K3-EDTA tubes and each of the three levels of control were tested in two replicates per run, two runs per day for 21 days (n=84 measurements per sample). Repeatability and intermediate imprecision were calculated in accordance with CLSI EP05-A3. Results are summarized in the table below.

Summary of 21-Day Precision Results

| Sample | Mean (ng/mL) | Repeatability (within-run) | | Intermediate Precision(within-lab) | |
| --- | --- | --- | --- | --- | --- |
| | | SD (ng/mL) | CV (%) | SD (ng/mL) | CV (%) |
| Sample 1 | 152 | 5.73 | 3.8 | 6.17 | 4.1 |
| Sample 2 | 206 | 6.14 | 3.0 | 7.17 | 3.5 |
| Sample 3 | 252 | 6.70 | 2.7 | 8.28 | 3.3 |
| Sample 4 | 363 | 8.01 | 2.2 | 10.0 | 2.8 |
| Sample 5 | 605 | 10.7 | 1.8 | 14.6 | 2.4 |
| Control 1 | 222 | 6.88 | 3.1 | 8.14 | 3.7 |
| Control 2 | 391 | 11.1 | 2.8 | 11.5 | 2.9 |
| Control 3 | 500 | 10.8 | 2.2 | 13.1 | 2.6 |

#### Lot-to-Lot Reproducibility (5 days)

A multi-site-multi-lot reproducibility study was performed at three sites using three reagent lots (1 site used all three lots and the other 2 sites used 1 lot each). Seven human hemolysate

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samples prepared from whole blood samples collected in K3-EDTA tubes and three control levels were tested. Samples were tested in five replicates per day for five days on one cobas e 801 analyzer per site, per lot for a total N=125 measurements per sample. Repeatability and intermediate imprecision were calculated in accordance with CLSI EP05-A3 and results are summarized below.

Lot-to-Lot Reproducibility - All Lots Combined

| | | | Reproducibility | |
| --- | --- | --- | --- | --- |
| Sample | N | Mean | SD (ng/mL) | CV (%) |
| Hemolysate 1 | 125 | 128 | 10.5 | 8.20 |
| Hemolysate 2 | 125 | 149 | 10.6 | 7.12 |
| Hemolysate 3 | 125 | 201 | 10.2 | 5.08 |
| Hemolysate 4 | 125 | 247 | 10.7 | 4.35 |
| Hemolysate 5 | 125 | 356 | 14.1 | 3.95 |
| Hemolysate 6 | 125 | 601 | 20.4 | 3.41 |
| Hemolysate 7 | 125 | 605 | 28.3 | 4.67 |
| Control 1 | 125 | 212 | 10.7 | 5.06 |
| Control 2 | 125 | 382 | 15.0 | 3.93 |
| Control 3 | 125 | 490 | 17.5 | 3.56 |

Between-Lot and Between-Day Summary

| | | | Between-site | | Between-lot | | Between-day | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Sample | N | Mean | SD | CV (%) | SD | CV (%) | SD | CV (%) |
| Hemolysate 1 | 125 | 128 | 6.69 | 5.23 | 4.36 | 3.41 | 5.13 | 4.02 |
| Hemolysate 2 | 125 | 149 | 6.71 | 4.52 | 3.43 | 2.31 | 5.44 | 3.66 |
| Hemolysate 3 | 125 | 201 | 2.61 | 1.30 | 3.57 | 1.77 | 6.13 | 3.05 |
| Hemolysate 4 | 125 | 247 | 3.62 | 1.47 | 2.80 | 1.13 | 6.32 | 2.56 |
| Hemolysate 5 | 125 | 356 | 8.93 | 2.51 | 1.34 | 0.38 | 4.99 | 1.40 |
| Hemolysate 6 | 125 | 601 | 8.06 | 1.34 | 15.1 | 2.52 | 8.45 | 1.41 |
| Hemolysate 7 | 125 | 605 | 16.8 | 2.77 | 19.5 | 3.23 | 7.37 | 1.22 |
| Control 1 | 125 | 212 | 5.41 | 2.55 | 0 | 0 | 5.19 | 2.44 |
| Control 2 | 125 | 382 | 7.82 | 2.05 | 2.87 | 0.75 | 8.33 | 2.18 |
| Control 3 | 125 | 490 | 12.2 | 2.48 | 2.14 | 0.44 | 7.53 | 1.54 |

# 2. Linearity:

Linearity of the Elecsys Folate III (RBC Application) was assessed using human hemolysate samples prepared from whole blood samples collected in K3-EDTA tubes on the cobas e 801 analyzer. The studies were performed in accordance with CLSI EP06-Ed2 using three series comprised of 9-10 levels of samples ranging from $\sim 103$ to $656\mathrm{ng / mL}$ . Samples were tested using one lot of reagent, run in 4 replicates within a single run. The linearity data was analyzed using a weighted linear regression model according to CLSI EP06-Ed2. The deviation from linearity did not exceed $5\%$ within the claimed measuring interval.

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The results support the Elecsys Folate III (RBC Application)'s measuring range of 120 - 620 ng/mL.

## Dilution Study

The sponsor provided information to support that samples with folate concentrations above the measuring range can be diluted with Elecsys Folate RBC Hemolyzing Reagent. For the Elecsys Folate III (RBC Application), the recommended manual dilution is 1:2. The concentration of the diluted sample must be ≥265 ng/mL.

## 3. Analytical Specificity/Interference:

### Interference - Endogenous substances

An interference study was performed using eight endogenous substances (biotin, lipemia, bilirubin, rheumatic factor, albumin, human IgG, human IgM, and human IgA) for potential interference with the Elecsys Folate III (RBC Application) on the cobas e 801 analyzer. For each potentially interfering substance, three human hemolysate samples prepared from whole blood samples collected in K3-EDTA tubes containing low, mid, and high concentrations of 142 ng/mL, 359 ng/mL, 488 ng/mL were tested with 5 replicates per sample. Recovery (absolute deviation or % recovery) was calculated for each sample compared to the reference (control sample with no potential interferent added). Results are summarized below.

| Interferent | Highest concentration tested with no significant interference observed |
| --- | --- |
| Biotin | 1200 ng/mL |
| Lipemia (Intralipid) | 1500 mg/dL |
| Bilirubin | 29 mg/dL |
| Rheumatoid Factor | 1000 IU/mL |
| Serum Albumin | 7.0 g/dL |
| Human IgG | 1.6 g/dL |
| Human IgM | 1 g/dL |
| Human IgA | 0.4 g/dL |

### Interference - Exogenous substances

An interference study was conducted to evaluate 17 drugs and human erythropoietin for potential interference with the Elecsys Folate III (RBC Application). Two human hemolysate samples prepared from whole blood samples collected in K3-EDTA tubes with folate concentrations of ~150 ng/mL and ~430 ng/mL were used for the testing. Recovery was calculated as percent recovery compared to the reference sample (control sample with no potential interferent added). Results are summarized below.

| Drug | Highest concentration tested with no significant interference observed |
| --- | --- |
| Acetyl cysteine | 150 mg/L |
| Acetylsalicylic Acid | 30 mg/L |

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| Ampicillin-Na | 75 mg/L |
| --- | --- |
| Ascorbic acid | 52.5 mg/L |
| Cefoxitin | 750 mg/L |
| Doxycycline | 18 mg/L |
| Heparin | 3300 IU/L |
| Levodopa | 7.5 mg/L |
| Methyldopa | 22.5 mg/L |
| Metronidazole | 123 mg/L |
| Rifampicin | 48 mg/L |
| Acetaminophen | 156 mg/L |
| Cyclosporine | 1.8 mg/L |
| Ibuprofen | 219 mg/L |
| Theophylline | 60 mg/L |
| Phenylbutazone | 321 mg/L |
| Itraconazole | 30 mg/L |
| Erythropoietin | 2000 u/mL |

## Cross-reactivity

A cross-reactivity study was performed in which potential cross-reactants were added at defined concentrations to one hemolysate sample prepared from a whole blood sample collected in K3-EDTA tubes. Samples were measured on the cobas e 801 analyzer and cross-reactivity was calculated and summarized in the table below.

| Cross reactant | Highest Con. Tested (ng/mL) | Highest % Cross-reactivity |
| --- | --- | --- |
| Amethopterin | 750 | 1.7 |
| Aminopterin | 750 | 2.0 |
| Folinic acid | 750 | 2.6 |

4. Assay Reportable Range:

See section VII.A.2 Linearity.

5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):

Traceability: The Elecsys Folate III (RBC Application) is traceable to a commercially available assay.

Stability

The Elecsys Folate III (RBC Application) reagent kit can be stored on-board the analyzers for up to 112 days (16 weeks). A new calibration of the kit kept on-board is recommended every 28 days.

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6. Detection Limit:

Determination of the Limit of Blank (LoB), Limit of Detection (LoD) and Limit of Quantification (LoQ) for the Elecsys Folate III (RBC Application) was determined in accordance with CLSI EP17-A2 guideline. For the LoQ study, the highest CV% reported across all lots in the study was 10.4%. Results are summarized below.

| Matrix | LoB (ng/mL) | LoD (ng/mL) | LoQ (ng/mL) |
| --- | --- | --- | --- |
| Hemolysate | 45 | 70 | 120 |

7. Assay Cut-Off:

Not applicable.

B Comparison Studies:

1. Method Comparison with Predicate Device:

A method comparison study was performed comparing the performance of the Elecsys Folate III (RBC Application) (candidate device, Y) versus the predicate device, Elecsys Folate RBC (non-biotin remediated, X) on one cobas e 801 analyzer. The study was conducted in accordance with CLSI Guideline EP09c-A3.

A total of 119 hemolysate samples prepared from native whole blood samples collected in K3-EDTA tubes (including 99 native samples, 10 spiked samples and 10 diluted samples) that span the measuring range were tested with one replicate per sample. Samples were measured with one lot of reagents. Results are summarized in the tables below (Passing-Bablok).

| Sample N=119 | Passing/Bablok |
| --- | --- |
| Sample concentration range 132 - 618 ng/mL | |
| Slope | 1.04 |
| Intercept | 14.6 |
| Linear Correlation coefficient r | 0.991 |

2. Matrix Comparison:

The effect of anticoagulants on the measurement of the analyte with the Elecsys Folate III (RBC Application) was assessed by comparing values obtained from native whole blood samples drawn into Na-heparin and K3-EDTA tubes. At least 40 sample pairs were tested in singleton with one reagent lot on one cobas e 801 analyzer. Passing/Bablok regression analysis was performed. Results are summarized below.

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| K3-EDTA vs. Na-Heparin | Passing/Bablok |
| --- | --- |
| N | 53 |
| Range (ng/mL) | 135-601 ng/mL |
| Slope (95% CL) | 1.07 |
| Correlation coefficient r | 0.992 |

C Clinical Studies:

1. Clinical Sensitivity:
Not applicable

2. Clinical Specificity:
Not applicable

3. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):
Not applicable

D Clinical Cut-Off:
Not applicable

E Expected Values/Reference Range:
Previously established in K082340.

VIII Proposed Labeling:
The labeling supports the finding of substantial equivalence for this device.

IX Conclusion:
The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

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