THERMO SCIENTIFIC QMS TACROLIMUS ASSAY AND CALIBRATORS

K123343 · Microgenics Corp. · MLM · Jul 11, 2013 · Clinical Chemistry

Device Facts

• Record ID: K123343
• Device Name: THERMO SCIENTIFIC QMS TACROLIMUS ASSAY AND CALIBRATORS
• Applicant: Microgenics Corp.
• Product Code: MLM · Clinical Chemistry
• Decision Date: Jul 11, 2013
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1678
• Device Class: Class 2

Indications for Use

The QMS® Tacrolimus Immunoassay is intended for the quantitative determination of tacrolimus in human whole blood on automated clinical chemistry analyzers. The results obtained are used as an aid in the management of kidney, liver, and heart transplant patients receiving tacrolimus therapy. This in vitro diagnostic device is intended for clinical laboratory use only. The QMS® Tacrolimus Calibrator set is intended for use in calibration of the QMS® Tacrolimus Immunoassay.

Device Story

The QMS® Tacrolimus Immunoassay is an in vitro diagnostic test for human whole blood samples. It utilizes a homogeneous particle-enhanced turbidimetric inhibition immunoassay principle. The assay involves a manual pretreatment step where whole blood is extracted using a zinc sulfate reagent. The processed sample is then analyzed on automated clinical chemistry analyzers. The device uses competition between tacrolimus in the sample and drug-coated microparticles for antibody binding sites; the rate of absorbance change is measured photometrically. The presence of tacrolimus inhibits the agglutination reaction, resulting in a concentration-dependent decrease in the rate of absorbance change. Results are used by clinicians to monitor and manage tacrolimus therapy in transplant patients. The device provides quantitative results that assist in adjusting patient medication dosages to maintain therapeutic levels.

Clinical Evidence

Bench testing only. Precision evaluated per CLSI EP5-A2 (n=40 runs). Linearity/recovery validated across 1-30 ng/mL range. Limit of Quantitation (LoQ) established at 0.9 ng/mL. Specificity tested against metabolites and co-administered drugs. Method comparison performed against LC-MS/MS and predicate using 383 patient samples (kidney, liver, heart), showing Deming slopes 1.09-1.13 and correlation R > 0.94.

Technological Characteristics

Homogeneous particle-enhanced turbidimetric inhibition immunoassay. Reagents: anti-tacrolimus monoclonal antibody (rabbit), tacrolimus-coated microparticles, and zinc sulfate extraction reagent. Form factor: liquid, ready-to-use. Analyzed via photometric absorbance change on automated clinical chemistry analyzers. Storage: 2-8°C for reagents, -20°C for calibrators.

Indications for Use

Indicated for quantitative determination of tacrolimus in human whole blood to aid in management of kidney, liver, and heart transplant patients receiving tacrolimus therapy. For clinical laboratory use only.

Regulatory Classification

Identification

A tacrolimus test system is a device intended to quantitatively determine tacrolimus concentrations as an aid in the management of transplant patients receiving therapy with this drug. This generic type of device includes immunoassays and chromatographic assays for tacrolimus.

Special Controls

*Classification.* Class II (special controls). The special control is “Class II Special Controls Guidance Document: Cyclosporine and Tacrolimus Assays; Guidance for Industry and FDA.” See § 862.1(d) for the availability of this guidance document.

Predicate Devices

• Abbott ARCHITECT® Tacrolimus Assay (k070820)

Submission Summary (Full Text)

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510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION
DECISION SUMMARY
ASSAY ONLY TEMPLATE

A. 510(k) Number:
K123343

B. Purpose for Submission:
New device

C. Measurand:
Tacrolimus

D. Type of Test:
Quantitative immunoassay

E. Applicant:
Microgenics Corporation

F. Proprietary and Established Names:
Thermo Scientific QMS® Tacrolimus Immunoassay
Thermo Scientific QMS® Tacrolimus Calibrators

G. Regulatory Information:
1. Regulation section:
21 CFR §862.1678, Tacrolimus Test System
21 CFR §862.1150, Calibrator
2. Classification: Class II
3. Product code: MLM, JIT
4. Panel: Toxicology (91)

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H. Intended Use:

1. Intended use(s):
See indication(s) for use below

2. Indication(s) for use:
The QMS® Tacrolimus Immunoassay is intended for the quantitative determination of tacrolimus in human whole blood on automated clinical chemistry analyzers. The results obtained are used as an aid in the management of kidney, liver, and heart transplant patients receiving tacrolimus therapy. This in vitro diagnostic device is intended for clinical laboratory use only.

The QMS® Tacrolimus Calibrator set is intended for use in calibration of the QMS® Tacrolimus Assay.

3. Special conditions for use statement(s):
For prescription use only.

4. Special instrument requirements:
Beckman AU 680 Analyzer.

I. Device Description:

The QMS® Tacrolimus Assay consists of the following separately packaged reagents in ready-to-use liquid form:

Extraction reagent
R1: < 1.0% anti-tacrolimus monoclonal antibody (rabbit), < 0.09% sodium azide.
R2: < 0.3% tacrolimus-coated microparticles, < 0.09% sodium azide.

Calibrators are prepared in a human whole blood matrix. The calibrator kit contains six liquid ready-to use levels of tacrolimus, with target concentrations of 0.0, 2.0, 5.0, 10.0, 20.0, and 30.0 ng/mL. The package insert contains the following caution:
"This product contains human sourced and/or potentially infectious components. Components sourced from human blood have been tested and found to be nonreactive for HBsAg, anti-HIV 1/2, and anti-HCV. No known test method can offer complete assurance that products derived from human sources or inactivated microorganisms will not transmit infection. Therefore, it is recommended that all human sourced materials be considered potentially infectious and handled with appropriate biosafety practices."

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J. Substantial Equivalence Information:

1. Predicate device name(s):
ARCHITECT® Tacrolimus Assay
ARCHITECT® Tacrolimus Calibrators

2. Predicate 510(k) number(s):
K070820

3. Comparison with predicate:

| Feature | Proposed Device | Predicate Device (k070820) |
| --- | --- | --- |
| Proprietary Name | Thermo Scientific QMS® Tacrolimus Assay
Thermo Scientific QMS® Tacrolimus Calibrator Set | ARCHITECT® Tacrolimus Assay
ARCHITECT® Tacrolimus Calibrators |
| Intended Use | The Thermo Scientific QMS® Tacrolimus Assay is intended for the quantitative determination of tacrolimus in human whole blood on automated clinical chemistry analyzers. The results obtained are used as an aid in the management of kidney, liver, and heart transplant patients receiving tacrolimus therapy. This in vitro diagnostic device is intended for clinical laboratory use only.
The Thermo Scientific QMS® Tacrolimus Calibrator set is intended for use in calibration of the Thermo Scientific QMS® Tacrolimus Assay. | Same except that assay indication is for kidney and liver transplant patients; does not include heart transplant patients. |
| Test Principle | The assay is based on competition between drug in the sample and drug coated onto a microparticle for antibody binding sites of the tacrolimus antibody reagent. The tacrolimus-coated microparticle reagent is agglutinated in the presence of the anti-tacrolimus antibody reagent and in the absence of | Tacrolimus present in the sample binds to the anti-tacrolimus coated microparticles. After a delay, tacrolimus acridinium-labeled conjugate is added to the reaction mixture. The tacrolimus on the acridinium-labeled conjugate competes for the available binding |

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| | any competing drug in the sample. The rate of absorbance change is measured photometrically. When a sample containing tacrolimus is added, the agglutination reaction is partially inhibited, slowing down the rate of absorbance change. A concentration-dependent agglutination inhibition curve can be obtained with the maximum rate of agglutination at the lowest tacrolimus concentration and the lowest agglutination rate at the highest tacrolimus concentration. | sites on the microparticles. Following incubation, the microparticles are washed, and pre-trigger and trigger solutions are added to the reaction mixture. The resulting chemiluminescent reaction is measured as relative light units (RLUs). An indirect relationship exists between the amount of tacrolimus in the sample and the RLUs detected by the ARCHITECT® System optics. |
| --- | --- | --- |
| Sample Matrix | Human Whole Blood | Same |
| Sample Preparation | Manual Pretreatment | Same |
| Reagent | Liquid Ready-to-Use (Antibody reagent, Tacrolimus coated microparticle reagent, and Extraction reagent) | Liquid Ready-to-Use (Antibody coated microparticle reagent, Tacrolimus conjugate reagent, and Assay diluent) |
| Calibrator | Liquid Ready-to-Use, six levels (0.0, 2.0, 5.0, 10.0, 20.0, and 30.0 ng/mL) | Liquid Ready-to-Use, six levels (0.0, 3.0, 6.0, 12.0, 20.0, and 30.0 ng/mL) |
| Assay Range | 1.0 to 30.0 ng/mL | 2.0 to 30.0 ng/mL |

# K. Standard/Guidance Document Referenced (if applicable):

CLSI EP5-A2, Evaluation of Precision Performance of Quantitative Measurement Methods.

CLSI EP9-A2, Method Comparison and Bias Estimation using Patient Samples.

# L. Test Principle:

The Thermo Scientific QMS Tacrolimus Assay is a homogeneous particle-enhanced turbidimetric immunoassay based on competition between drug in the sample and drug coated onto a microparticle for antibody binding sites of the tacrolimus antibody reagent. The tacrolimus-coated microparticle reagent is agglutinated in the presence of the anti-tacrolimus antibody reagent and in the absence of any competing drug in the sample. The rate of absorbance change is measured photometrically at $700\mathrm{nm}$ . When a sample containing tacrolimus is added, the agglutination reaction is partially inhibited, slowing down the rate of absorbance change. A concentration-dependent agglutination inhibition curve can be obtained

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with the maximum rate of agglutination at the lowest tacrolimus concentration and the lowest agglutination rate at the highest tacrolimus concentration.

## M. Performance Characteristics (if/when applicable):

Performance was validated on the Beckman AU 680 Analyzer.

### 1. Analytical performance:

#### a. Precision/Reproducibility:

Precision was determined at various concentration levels across the assay range. Three whole blood pools were comprised of samples obtained from patients taking tacrolimus and three negative whole blood pools spiked with tacrolimus were tested for precision. Following CLSI protocol EP5-A2, each tacrolimus sample was assayed in replicates of two, twice a day for 20 days for a total of 40 runs on two reagent lots. Each run per day was separated by at least two hours. Evaluations were performed at the manufacturer's site and at external sites. Results were similar across lots. Representative results from manufacturer site testing are shown below:

| Samples | Mean (ng/mL) | Within-Run SD (ng/mL) | Within-Run CV (%) | Total-Run SD (ng/mL) | Total-Run CV (%) |
| --- | --- | --- | --- | --- | --- |
| Spiked Sample 1 | 3.1 | 0.2 | 4.9 | 0.2 | 7.2 |
| Spiked Sample 2 | 10.1 | 0.2 | 2.0 | 0.4 | 4.3 |
| Spiked Sample 3 | 21.4 | 0.5 | 2.5 | 1.2 | 5.8 |
| Patient Pooled Sample 1 | 3.2 | 0.1 | 4.5 | 0.2 | 7.1 |
| Patient Pooled Sample 2 | 10.4 | 0.3 | 2.6 | 0.5 | 5.0 |
| Patient Pooled Sample 3 | 25.0 | 0.5 | 2.0 | 1.6 | 6.3 |

Results from a representative external site are shown below. Similar precision was observed for both external sites.

| Samples | Mean (ng/mL) | Within-Run | Within-Run | Total-Run SD (ng/mL) | Total-Run CV (%) |
| --- | --- | --- | --- | --- | --- |
| Sample 1 | 4.21 | 0.2 | 3.5 | 0.3 | 7.3 |
| Sample 2 | 8.3 | 0.2 | 2.3 | 0.2 | 2.7 |
| Sample 3 | 16.2 | 0.3 | 1.7 | 0.5 | 2.9 |
| Sample 4 | 22.4 | 0.7 | 3.1 | 0.9 | 3.8 |

#### b. Linearity

Linearity and Recovery evaluations:

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Results of the linearity and recovery studies are consistent with the Sponsor's claimed range of 1-30 ng/mL.

High tacrolimus samples (including spiked whole blood and pooled patient samples) were serially mixed with tacrolimus-free samples to attain 13 concentrations ranging from 0.8 to 30 ng/mL tacrolimus. The high sample expected concentration was based on a chromatographic method; the remainder were based on percent dilution of the high sample. Replicate samples (n=4) were measured using the QMS method and the means were compared to expected values. Linear regression results are as follows:

| | Lot 1 spiked | Lot 1 pooled patient sample | Lot 2 spiked | Lot 2 pooled patient sample |
| --- | --- | --- | --- | --- |
| Linear regression | Y=1.03x+0.05 | Y=1.02x+0.06 | Y=1.02x+0.06 | Y=1.00x+0.05 |
| Correlation coefficient R | 0.99 | 0.99 | 0.99 | 0.99 |
| Range of % recoveries (for each level separately) | 92%-103% | 94%-101% | 94%-108% | 96%-103% |

A representative table of recoveries across the range is shown below for one of the lots using spiked samples.

| | QMS measurement (ng/mL) | | | | |
| --- | --- | --- | --- | --- | --- |
| % dilution of high Sample | Rep 1 | Rep 2 | Rep 3 | Rep 4 | Average percent recovery |
| 100.0 | 30.3 | 28.7 | 30.1 | 29.4 | 100.0 |
| 90.0 | 25.6 | 26.6 | 26.8 | 26.4 | 98.8 |
| 80.0 | 23.2 | 22.9 | 23.1 | 23.8 | 98.1 |
| 70.0 | 19.5 | 19.5 | 19.8 | 19.5 | 94.4 |
| 60.0 | 17.2 | 17.3 | 17.8 | 17.5 | 98.2 |
| 50.0 | 14.5 | 14.7 | 14.7 | 14.8 | 99.1 |
| 40.0 | 11.2 | 11.0 | 11.4 | 11.1 | 94.3 |
| 30.0 | 8.4 | 8.5 | 8.7 | 8.5 | 95.9 |
| 20.0 | 5.7 | 5.7 | 5.9 | 5.7 | 97.0 |
| 10.0 | 3.0 | 3.0 | 3.3 | 3.2 | 105.5 |
| 5.0 | 1.5 | 1.5 | 2.0 | 1.4 | 108.0 |
| 3.3 | 0.9 | 1.1 | 0.6 | 1.3 | 99.7 |
| 2.8 | 0.7 | 1.0 | 0.6 | 0.8 | 96.4 |

A separate recovery evaluation of the Thermo Scientific QMS assay was also performed using USP materials. A stock solution containing a high

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concentration of known concentrations of USP tacrolimus were spiked into negative whole blood samples to yield final expected concentrations of 3, 10, 15, 22, and 25 ng/mL of tacrolimus. Replicates of each concentration level were separately extracted and measured with the QMS assay. Mean values for each concentration level were compared with the expected values based on USP materials (and verified using chromatographic methods). Two reagent lots were evaluated. Results were similar for both lots. Representative results are tabulated below:

| Description | Sample 1 | Sample 2 | Sample 3 | Sample 4 | Sample 5 |
| --- | --- | --- | --- | --- | --- |
| Mean Measured Value (ng/mL) | 2.7 | 10.6 | 17.8 | 21.8 | 27.4 |
| SD (ng/mL) | 0.3 | 0.2 | 0.5 | 0.4 | 0.8 |
| CV (%) | 9.3 | 1.9 | 2.8 | 1.9 | 3.1 |
| Expected Value by LCMS (ng/mL) | 2.7 | 9.8 | 18.0 | 19.8 | 27.0 |
| Average % Recovery | 100.6% | 107.9% | 98.7% | 110.1% | 101.3% |

## Manual dilution:

Testing was performed to validate the manual dilution protocol recommended by the manufacturer in the package insert. The following types of samples were prepared for testing: (i) spiked tacrolimus in negative whole blood at a concentration of 45 ng/mL, (ii) patient sample with additional tacrolimus added to 40 ng/mL, and iii) tacrolimus patient samples with concentration close to but within the upper limit of the reportable range. The samples were diluted 1:1 with Calibrator A and measured by the Thermo Scientific QMS Tacrolimus Immunoassay. Percent recoveries (based on measured/expected concentration) were determined for each sample (tested with two reagent lots). Results are shown below:

| Initial concentration | Lot 1 % recovery after dilution | Lot 2 % recovery after dilution |
| --- | --- | --- |
| 46.8 | 99 | 97 |
| 40.4 | 95 | 96 |
| 25.1 | 105 | 104 |
| 26.8 | 105 | 104 |
| 26.6 | 103 | 101 |

## c. Traceability, Stability, Expected values (controls, calibrators, or methods):

Traceability and Value Assignment:

Thermo Scientific QMS® Tacrolimus Calibrator set was prepared by gravimetric addition of USP reference standard tacrolimus into human whole

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blood matrix at 0, 2, 5, 10, 20, and 30 ng/mL target concentrations to form Cal A to Cal F. Calibrators are traceable to USP material. Value assignment is based on USP material and verified by chromatographic methods. The QMS® Tacrolimus Calibrators are traceable to USP reference standard, through value assignment. The reported estimated uncertainty in assigned values range from 0.21 ng/mL for Cal B to 0.65 ng/mL for Cal F.

## Stability:

Real-time testing for opened and closed Thermo Scientific QMS® Calibrators was performed. Protocols and acceptance criteria were reviewed and deemed acceptable. The Sponsor's claims of stability up to 15 months at -20°C and for 42 days at 2-8°C when stored tightly capped are supported.

## d. Detection limit:

To determine the Limit of Quantitation (LOQ), low concentration samples were prepared gravimetrically using USP tacrolimus gravimetrically spiked into negative whole blood at low concentrations. Each sample was tested once per run, twice a day for 30 days for a total of n=60. All replicate measurements were from separately extracted samples. LoQ was defined as the point where the upper 95% confidence interval of %CV was < 20 and mean recovery was within +/-10% (relative to material assigned based on USP standards). Based on these criteria, a LoQ of 0.9 ng/mL was determined.

To verify precision and accuracy (by recovery) at the claimed LoQ, samples were further tested by preparing spiked samples to 0.9 ng/mL and evaluating precision and recovery on two instruments with two reagent lots. Each sample was tested in 4 replicates per day for a total of 5 days. Summary results are shown below:

| | Lot 1 | Lot 2 |
| --- | --- | --- |
| Expected conc (ng/mL) | 0.90 | 0.90 |
| Mean (ng/mL) | 0.92 | 0.92 |
| % recovery | 102.0 | 101.9 |
| SD (ng/mL) | 0.14 | 0.13 |
| %CV | 14.7 | 13.8 |
| N | 40 | 40 |

## e. Analytical specificity:

Specificity studies were conducted for (i) major metabolites of tacrolimus, (ii) medications routinely co-administered with tacrolimus, and (iii) other over-the-counter drugs. Concentrations tested and results are tabulated below.

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| Tacrolimus Metabolites | Metabolite Conc (ng/mL) | Expected Tacrolimus Conc(ng/mL) | | Measured Tacrolimus Conc (ng/mL) | | Recovery (%) | | Cross Reactivity (%) | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | Lot 1 | Lot 2 | Lot 1 | Lot 2 | Lot 1 | Lot 2 | Lot 1 | Lot 2 |
| M-I (13-O-demethyl) | 20.0 | 5.8 | 5.8 | 7.6 | 7.6 | 138.2 | 131.6 | 10.5 | 9.2 |
| | 20.0 | 13.2 | 13.3 | 15.3 | 14.8 | 115.7 | 111.6 | 10.3 | 7.7 |
| M-II (31-O-demethyl) | 20.0 | 5.6 | 5.7 | 5.5 | 5.9 | 97.0 | 102.3 | -0.8 | 0.7 |
| | 20.0 | 13.0 | 13.2 | 13.3 | 13.1 | 102.1 | 99.2 | 1.3 | -0.5 |
| M-III (15-O-demethyl) | 20.0 | 5.4 | 5.3 | 6.8 | 6.0 | 125.3 | 114.6 | 6.8 | 3.8 |
| | 20.0 | 12.0 | 12.4 | 13.7 | 13.0 | 113.9 | 104.3 | 8.3 | 2.7 |
| M-IV (12-hydroxy) | 20.0 | 4.9 | 5.0 | 6.2 | 6.1 | 126.5 | 122.8 | 6.5 | 5.7 |
| | 20.0 | 11.9 | 12.0 | 13.8 | 14.1 | 115.7 | 117.5 | 9.3 | 10.5 |
| M-IV (12-hydroxy) | 3.5 | 14.6 | 14.6 | 18.8 | 18.7 | 127.2 | 128.1 | 113.4 | 117.1 |
| | 3.3 | 21.2 | 21.2 | 25.5 | 27.0 | 120.3 | 127.2 | 130.3 | 174.8 |
| Tacrolimus Metabolites | Metabolite Conc (ng/mL) | Expected Tacrolimus Conc (ng/mL) | Measured Tacrolimus Conc (ng/mL) | Recovery (%) | Cross Reactivity (%) |
| --- | --- | --- | --- | --- | --- |
| M-VII (13,15-O-didemethyl) | 20.0 | 5.4 | 7.3 | 135.2 | 9.3 |
| | 20.0 | 13.4 | 14.7 | 109.7 | 6.7 |
| M-VII (13,15-O-didemethyl) + M-VI (13,31-o-didemethyl) | 20.0 | 5.4 | 5.8 | 107.4 | 2.2 |
| | 20.0 | 13.4 | 13.8 | 103.0 | 2.0 |
| Co-administrated Immunosuppressant | Co-administered Concentration (ng/mL) | Expected Tacrolimus Concentration (ng/mL) | Measured Tacrolimus Concentration (ng/mL) | Percent Recovery (%) |
| --- | --- | --- | --- | --- |
| Cyclosporine / Cyclosporin A | 10,000 | 5.2 | 5.4 | 103.8 |
| | | 12.1 | 12.7 | 105.0 |
| Everolimus | 100 | 5.1 | 5.0 | 99.3 |
| | | 11.9 | 11.4 | 95.8 |
| Mycophenolic acid | 10,000 | 5.4 | 5.7 | 104.9 |
| | | 12.0 | 12.1 | 100.8 |

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| Sirolimus (Rapamycin) | 300 | 5.3 | 6.7 | 107.9 |
| --- | --- | --- | --- | --- |
| | | 12.8 | 13.4 | 104.4 |
| Hydrocortisol | 100,000 | 5.4 | 5.4 | 100.6 |
| | | 12.0 | 12.4 | 103.0 |
| Prednisolone | 100,000 | 5.2 | 5.3 | 101.9 |
| | | 12.1 | 12.9 | 106.9 |
| Prednisone | 100,000 | 5.2 | 5.3 | 101.3 |
| | | 12.1 | 12.3 | 101.9 |

In addition, other commonly co-administered and OTC drugs were tested tested at 5 and 12 ng/mL tacrolimus. No significant interference was observed. A complete list of the compounds and results are tabulated in the package insert.

Endogenous substances were tested at 5 and 12 ng/mL tacrolimus. No significant interference was observed. Concentrations tested and results are shown below.

| Potential Interferent | High concentration tested | Recovery (%) for each Concentration of Tacrolimus: | |
| --- | --- | --- | --- |
| | | 5 ng/mL | 12 ng/mL |
| Albumin | 12 g/dL | 98.5 | 90.2 |
| Bilirubin | 60 mg/dL | 97.5 | 96.7 |
| Cholesterol | 500 mg/dL | 93.6 | 94.4 |
| Creatinine | 5 mg/dL | 98.8 | 101.8 |
| Triglyceride | 1500 mg/dL | 96.6 | 93.8 |
| Uric acid | 20 mg/dL | 103.6 | 99.0 |
| IgG | 12 g/dL | 94.9 | 96.1 |
| Rheumatoid factor | 500 IU/mL | 104.9 | 106.7 |
| HAMA | 400 ng/mL | 102.0 | 106.2 |
| hematocrit | 63.9% (low value 12%) | 104.0 | 105.6 |

f. Assay cut-off:

Not applicable – this is a quantitative assay.

2. Comparison studies:

a. Method comparison with predicate device:

Method comparison studies were conducted according to CLSI EP-9. Adult trough (pre-dose) patient samples from heart, liver, and kidney allograft recipients from multiple clinical sites were measured by the Thermo-Scientific

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QMS® Tacrolimus assay and by an LC-MS/MS (as well as by the Abbott ARCHITECT® Tacrolimus assay). The majority of samples were from patients $>9$ months post-transplant. Patients within the population tested received drug regimens of tacrolimus either alone or with co-administered immuosuppressants such as mycophenolic acid, or corticosteroids. Samples, were tested on the Beckman AU680® clinical analyzer with the QMS assay at both Thermo Fisher Scientific and two external evaluation sites. Two QMS reagent lots were evaluated. Results of regression analyses are shown below. These results are based on analyses of single measurements for each sample by the QMS assay.

| Comparative method | Transplant type | N | Slope (Deming) | Intercept (Deming) | Correlation (R) |
| --- | --- | --- | --- | --- | --- |
| Lot 1 QMS v. Site 1 LC-MS/MS | Combined | 383 | 1.111 | 0.53 | 0.972 |
| | Kidney | 136 | 1.095 | 0.91 | 0.978 |
| | Liver | 133 | 1.131 | 0.23 | 0.964 |
| | Heart | 114 | 1.114 | 0.40 | 0.972 |
| Lot 1 QMS v. Site 2 LC-MS/MS | Combined | 232 | 1.130 | 0.71 | 0.967 |
| | Kidney | 104 | 1.135 | 0.84 | 0.975 |
| | Liver | 66 | 1.120 | 0.51 | 0.969 |
| | Heart | 62 | 1.131 | 0.71 | 0.949 |

Similar results were obtained with both lots.

Additional patient sample testing was performed with the QMS assay at external sites and results were compared to results obtained at the manufacturer's site. Results are shown below:

| method | N | Slope (Deming) | Intercept (Deming) | Correlation (R) |
| --- | --- | --- | --- | --- |
| QMS External site 1 v. QMS manufacturer | 157 | 1.055 (1.034 to 1.077) | -0.18 (-0.37 to 0.01) | 0.992 |
| QMS External site 2 v. QMS manufacturer | 158 | 1.034 (-0.42 to -0.12) | -0.27 (-0.42 to -0.12) | 0.995 |

# b. Matrix comparison:

Not applicable. The assay is intended for use with EDTA whole blood only.

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3. Clinical studies:
a. Clinical Sensitivity:
Not applicable
b. Clinical specificity:
Not applicable.
c. Other clinical supportive data (when a. and b. are not applicable):
Not applicable.

4. Clinical cut-off:
Not applicable; this is a quantitative assay.

5. Expected values/Reference range:
The following is stated in the package insert:
The optimal therapeutic range for tacrolimus in whole blood has not been established with this assay. The therapeutic ranges for tacrolimus may vary depending on clinical factors and on the methodology used. Given the heterogeneity of the patient's clinical state, clinicians should establish a desired therapeutic management range based on their own experience as well as each patient's clinical requirements. Changes to treatment regimen should not be based solely on tacrolimus values. Differences in sensitivity to immunosuppressive and nephrotoxic effects of tacrolimus, co-administration of other immunosuppressants, type of transplant, time post-transplant and a number of other factors contribute to different requirements for optimal blood levels of tacrolimus. Optimal ranges may vary depending on the test used, and therefore should be established for each commercial test. Values obtained with different assay methods cannot be used interchangeably due to differences in methodology and cross reactivity, nor should correction factors be applied. Consistent use of one assay for individual patients is recommended.

N. Proposed Labeling:
The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

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

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