ACE ALKALINE PHOSPHATASE REAGENT, AMYLASE REAGENT, ALT REAGENT, AST REAGENT

K131351 · Alfa Wassermann Diagnostic Technologies, LLC · CJE · Aug 15, 2013 · Clinical Chemistry

Device Facts

• Record ID: K131351
• Device Name: ACE ALKALINE PHOSPHATASE REAGENT, AMYLASE REAGENT, ALT REAGENT, AST REAGENT
• Applicant: Alfa Wassermann Diagnostic Technologies, LLC
• Product Code: CJE · Clinical Chemistry
• Decision Date: Aug 15, 2013
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1050
• Device Class: Class 2

Indications for Use

The ACE Alkaline Phosphatase Reagent is intended for the quantitative determination of alkaline phosphatase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Measurements of alkaline phosphatase are used in the diagnosis and treatment of liver, bone, parathyroid, and intestinal diseases. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only. The ACE Amylase Reagent is intended for the quantitative determination of α-amylase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Amylase measurements are used primarily for the diagnosis and treatment of pancreatitis (inflammation of the pancreas). This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only. The ACE ALT Reagent is intended for the quantitative determination of alanine aminotransferase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Alanine aminotransferase measurements are used in the diagnosis and treatment of certain liver diseases (e.g., viral hepatitis and cirrhosis) and heart diseases. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only. The ACE AST Reagent is intended for the quantitative determination of aspartate aminotransferase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Measurements of aspartate aminotransferase are used in the diagnosis and treatment of certain types of liver and heart disease. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only.

Device Story

Reagents for quantitative photometric/colorimetric analysis of ALP, Amylase, ALT, and AST; used on ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Input: serum or lithium heparin plasma samples. Operation: automated photometric measurement of reaction rates (e.g., NADH oxidation for ALT/AST; p-nitrophenol formation for ALP). Output: enzyme activity levels (U/L). Used in clinical/physician office labs by trained personnel. Results assist clinicians in diagnosing liver, bone, parathyroid, intestinal, pancreatic, and heart conditions.

Clinical Evidence

Bench testing only. Precision studies (n=20-24) and matrix comparison studies (serum vs. plasma) performed across three platforms (ACE, ACE Alera, ACE Axcel). Matrix comparison showed high correlation (r > 0.99) and slopes near 1.0. Linearity, LoB, LoD, and LoQ established for ACE Alera system.

Technological Characteristics

Photometric/colorimetric assay reagents. ALP: p-nitrophenyl phosphate substrate. Amylase: 2-chloro-p-nitrophenyl-α-D-maltotrioside. ALT/AST: NADH-based kinetic assays. Automated analysis on ACE/ACE Alera/ACE Axcel systems. No calibration required. Traceable to IFCC or ERM-AD454/457 standards.

Indications for Use

Indicated for quantitative determination of ALP, amylase, ALT, and AST activity in serum and lithium heparin plasma for diagnosis/treatment of liver, bone, parathyroid, intestinal, pancreatic, and heart diseases. Intended for clinical and physician office laboratories.

Regulatory Classification

Identification

An alkaline phosphatase or isoenzymes test system is a device intended to measure alkaline phosphatase or its isoenzymes (a group of enzymes with similar biological activity) in serum or plasma. Measurements of alkaline phosphatase or its isoenzymes are used in the diagnosis and treatment of liver, bone, parathyroid, and intestinal diseases.

Predicate Devices

• ACE Alkaline Phosphatase Reagent (K931786)
• ACE Amylase Reagent (K931786)
• ACE ALT Reagent (K930104)
• ACE AST Reagent (K930104)

Submission Summary (Full Text)

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

A. 510(k) Number:
k131351

B. Purpose for Submission:
New reagents (Alkaline Phosphatase, Amylase, Alanine amino transferase, Aspartate amino transferase) added onto ACE Alera instrument (k123018)

Addition of lithium heparin plasma samples to already cleared reagents on the ACE (k930104) and ACE Axcel (k113389) instruments.

C. Measurand:
Alkaline Phosphatase, Amylase, Alanine Amino Transferase (ALT/SGPT), Aspartate Amino Transferase (AST/SGOT)

D. Type of Test:
Quantitative, photometric/colorimetric methods

E. Applicant:
Alfa Wassermann Diagnostic Technologies, LLC

F. Proprietary and Established Names:
ACE Alkaline Phosphatase Reagent
ACE Amylase Reagent
ACE ALT Reagent
ACE AST Reagent

G. Regulatory Information:
| Product Code | Classification | Regulation | Panel |
| --- | --- | --- | --- |
| CJE | Class II | 21 CFR 862.1050
Alkaline Phosphatase or Isoenzymes Test System | Clinical Chemistry (75) |
| CIJ | Class II | 21 CFR 862.1, Amylase Test System | Clinical Chemistry (75) |

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| CKA | Class I, meets limitations of exemption per 862.9(c)(9) | 21 CFR 862.1030 Alanine amino transferase (ALT/SGPT) Test System | Clinical Chemistry (75) |
| --- | --- | --- | --- |
| CIT | Class II | 21 CFR 862.1100, Aspartate amino transferase (AST/SGOT) Test System | Clinical Chemistry (75) |

H. Intended Use:

1. Intended use(s):

See Indication(s) for use.

2. Indication(s) for use:

The ACE Alkaline Phosphatase Reagent is intended for the quantitative determination of alkaline phosphatase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Measurements of alkaline phosphatase are used in the diagnosis and treatment of liver, bone, parathyroid, and intestinal diseases. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only.

The ACE Amylase Reagent is intended for the quantitative determination of $\alpha$ -amylase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Amylase measurements are used primarily for the diagnosis and treatment of pancreatitis (inflammation of the pancreas). This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only.

The ACE ALT Reagent is intended for the quantitative determination of alanine aminotransferase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Alanine aminotransferase measurements are used in the diagnosis and treatment of certain liver diseases (e.g., viral hepatitis and cirrhosis) and heart diseases. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only.

The ACE AST Reagent is intended for the quantitative determination of aspartate aminotransferase activity in serum and lithium heparin plasma using the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. Measurements of aspartate aminotransferase are used in the diagnosis and treatment of certain types of liver and heart disease. This test is intended for use in clinical laboratories and physician office laboratories. For in vitro diagnostic use only.

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3. Special conditions for use statement(s):

For in vitro diagnostic use only

For prescription use and point-of-care settings

4. Special instrument requirements:

For use on the ACE, ACE Axcel and ACE Alera Clinical Chemistry Systems.

I. Device Description:

The ACE Alkaline Phosphatase Reagent is composed of two reagent bottles (Buffer and Substrate Reagent). The reagents contain AMP Buffer (pH 10.45), magnesium acetate, p-nitrophenyl phosphate.

The ACE Amylase Reagent is composed of a single reagent bottle. The reagents contain 2-chloro-p-nitrophenyl-α-D-maltotrioside, sodium chloride, calcium acetate, potassium thiocyanate and MES buffer (pH 6.0).

The ACE ALT Reagent consists of two reagent bottles (Substrate and Coenzyme). The reagents contain L-alanine, α-ketoglutarate, nicotinamide adenine dinucleotide, reduced (NADH), lactate dehydrogenase and Tris buffer.

The ACE AST Reagent consists of two reagent bottles (Substrate and Coenzyme). The reagents contain L-aspartate, α-ketoglutarate, nicotinamide adenine dinucleotide, reduced (NADH), malate dehydrogenase, lactate dehydrogenase and Tris buffer.

J. Substantial Equivalence Information:

1. Predicate device name(s):

ACE Alkaline Phosphatase Reagent
ACE Amylase Reagent
ACE ALT Reagent
ACE AST Reagent

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

K931786 (ACE Alkaline Phosphatase Reagent and ACE Amylase Reagent)
K930104 (ACE ALT Reagent and ACE AST Reagent)

3. Comparison with predicate:

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1. ACE Alkaline Phosphatase Reagent:

| ALP | Candidate Device | Predicate Device K931786 |
| --- | --- | --- |
| Intended Use/Indications for Use | The ACE Alkaline Phosphatase Reagent is intended for the quantitative determination of alkaline phosphatase activity. | Same |
| Platforms | ACE, ACE *Alera*^{®}, and ACE Axcel Clinical Chemistry | ACE Clinical Chemistry System |
| Method | Photometric | Same |
| Calibration Stability | Not a calibrated test | Same |
| On-Board Stability | SA2002: 20 Days
RX2002: 7 Days | Same |
| Sample Type | Serum and lithium heparin plasma | Serum |
| Sample Volume | 4 μL | Same |
| Reaction Volume | 169 μL | Same |
| Expected Values | 44 - 147 U/L | Same |
| Measuring Range | 9 - 1400 U/L | Same |

2. ACE Amylase Reagent:

| AMYLASE | Candidate Device | Predicate Device K931786 |
| --- | --- | --- |
| Intended Use/Indications for Use | The ACE Amylase Reagent is intended for the quantitative determination of α-amylase activity. | Same |
| Platforms | ACE, ACE *Alera*^{®}, and ACE Axcel Clinical Chemistry Systems | ACE Clinical Chemistry System |
| Method | Photometric | Same |
| Calibration Stability | Not a calibrated test | Same |
| On-Board Stability | 30 Days | Same |
| Sample Type | Serum and lithium heparin plasma | Serum |
| Sample Volume | 3 μL | Same |
| Reaction Volume | 168 μL | Same |
| Expected Values | 20 - 104 U/L | Same |
| Measuring Range | 9 - 1900 U/L | Same |

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3. ACE ALT Reagent:

| ALT | Candidate Device | Predicate Device K930104 |
| --- | --- | --- |
| Intended Use/Indications for Use | The ACE ALT Reagent is intended for the quantitative determination of alanine aminotransferase activity | Same |
| Platforms | ACE, ACE *Alera*^{®}, and ACE Axcel Clinical Chemistry Systems | ACE Clinical Chemistry System |
| Method | Photometric | Same |
| Calibration Stability | Not a calibrated test | Same |
| On-Board Stability | 30 Days | Same |
| Sample Type | Serum and lithium heparin plasma | Serum |
| Sample Volume | 13 μL | Same |
| Reaction Volume | 185 μL | Same |
| Expected Values | 5 - 30 U/L | Same |
| Measuring Range | 4 - 480 U/L | Same |

4. ACE AST Reagent:

| AST | Candidate Device | Predicate Device K930104 |
| --- | --- | --- |
| Intended Use/Indications for Use | The ACE AST Reagent is intended for the quantitative determination of aspartate aminotransferase activity. | Same |
| Platforms | ACE, ACE *Alera*^{®}, and ACE Axcel Clinical Chemistry Systems | ACE Clinical Chemistry System |
| Method | Photometric | Same |
| Calibration Stability | Not a calibrated test | Same |
| On-Board Stability | 30 Days | Same |
| Sample Type | Serum and lithium heparin plasma | Serum |
| Sample Volume | 13 μL | Same |
| Reaction Volume | 185 μL | Same |
| Expected Values | 7 - 31 U/L | Same |
| Measuring Range | 4 - 450 U/L | Same |

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

Evaluation of Precision Performance of Quantitative Measurement Methods (CLSI EP5-A2)

Method Comparison and Bias Estimation Using Patient Samples (CLSI EP9-A2IR)

Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures (CLSI EP17-A2)

Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach (CLSI EP6-A)

Interference Testing in Clinical Chemistry (CLSI EP7-A2)

L. Test Principle:

In the ACE Alkaline Phosphatase Reagent assay, alkaline phosphatase catalyzes the hydrolysis of colorless p-nitrophenyl phosphate to p-nitrophenol and inorganic phosphate. In an alkaline solution (pH 10.5), p-nitrophenol is in the phenoxide form and has a strong absorbance at 408 nm. The rate of increase in absorbance, monitored bichromatically at 408 nm/486 nm, is directly proportional to the alkaline phosphatase activity in the sample.

In the ACE Amylase Reagent assay, α-amylase hydrolyzes the 2-chloro-p-nitrophenyl-α-D-maltotrioside substrate to release 2-chloro-p-nitrophenol and form 2-chloropnitrophenyl-α-D-maltoside, maltotriose and glucose. The rate of increase in absorbance, monitored bichromatically at 408 nm/647 nm, is directly proportional to the α-amylase activity in the sample.

In the ACE ALT Reagent assay, alanine Aminotransferase converts the L-alanine and α-ketoglutarate substrates in the reagent to L-glutamate and pyruvate. Lactate dehydrogenase (LDH) catalyzes the oxidation of the reduced cofactor to the cofactor. The rate of conversion of the reduced cofactor to the cofactor can be determined by monitoring the decrease in absorbance bichromatically at 340 nm/647 nm. This rate of conversion from the reduced cofactor to the cofactor is a function of the activity of ALT in the sample.

In the ACE AST Reagent assay, aspartate aminotransferase converts the L-aspartate and α-ketoglutarate in the reagent to oxaloacetate and L-glutamate. The oxaloacetate undergoes reduction with simultaneous oxidation of NADH to NAD⁺ in the malate dehydrogenase catalyzed indicator reaction. NADH absorbs strongly at 340 nm, whereas NAD⁺ does not. Therefore, the rate of conversion of NADH to NAD⁺ can be determined by monitoring the decrease in absorbance bichromatically at 340 nm/647 nm. This rate of conversion from NADH to NAD⁺ is a function of the activity of AST in the sample. Lactate dehydrogenase is added to prevent interference from endogenous pyruvate, which is normally present in blood.

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M. Performance Characteristics (if/when applicable):

1. Analytical performance:

a. Precision/Reproducibility:

Precision studies were conducted in house and at three Physician Office Laboratories (POL) with trained operators typically found in these settings, following CLSI guidance document EP5-A2. Three samples each of low, mid and high analyte serum were run on the ACE Alera Clinical Chemistry System in duplicate, for at least 20 days at 2 runs per day (N=80). Results are summarized below:

Serum POL precision study:

| ALP | | ACE Alera | | | | |
| --- | --- | --- | --- | --- | --- | --- |
| Lab | Sample | Mean (U/L) | Within-Run | | Total | |
| | | | SD | %CV | SD | %CV |
| In-House | 1 | 60 | 1.1 | 1.8% | 1.3 | 2.1% |
| POL 1 | 1 | 56 | 0.8 | 1.4% | 1.7 | 3.0% |
| POL 2 | 1 | 59 | 1.2 | 2.0% | 2.1 | 3.5% |
| POL 3 | 1 | 56 | 1.4 | 2.5% | 3.0 | 5.4% |
| In-House | 2 | 653 | 4.5 | 0.7% | 7.0 | 1.1% |
| POL 1 | 2 | 626 | 7.4 | 1.2% | 16.7 | 2.7% |
| POL 2 | 2 | 659 | 4.2 | 0.6% | 18.0 | 2.7% |
| POL 3 | 2 | 640 | 5.9 | 0.9% | 21.9 | 3.4% |
| In-House | 3 | 1192 | 9.4 | 0.8% | 13.4 | 1.1% |
| POL 1 | 3 | 1135 | 19.0 | 1.7% | 25.0 | 2.2% |
| POL 2 | 3 | 1209 | 9.6 | 0.8% | 29.3 | 2.4% |
| POL 3 | 3 | 1165 | 6.6 | 0.6% | 37.2 | 3.2% |
| AMY | | ACE Alera | | | | |
| --- | --- | --- | --- | --- | --- | --- |
| Lab | Sample | Mean (U/L) | Within-Run | | Total | |
| | | | SD | %CV | SD | %CV |
| In-House | 1 | 39 | 0.8 | 2.1% | 1.4 | 3.5% |
| POL 1 | 1 | 38 | 0.9 | 2.4% | 1.7 | 4.4% |
| POL 2 | 1 | 40 | 1.3 | 3.2% | 1.3 | 3.2% |
| POL 3 | 1 | 39 | 1.0 | 2.5% | 1.1 | 2.8% |
| In-House | 2 | 747 | 4.4 | 0.6% | 7.3 | 1.0% |
| POL 1 | 2 | 723 | 4.7 | 0.6% | 7.0 | 1.0% |
| POL 2 | 2 | 770 | 4.3 | 0.6% | 6.1 | 0.8% |
| POL 3 | 2 | 747 | 5.8 | 0.8% | 7.4 | 1.0% |
| In-House | 3 | 1437 | 11.6 | 0.8% | 12.8 | 0.9% |
| POL 1 | 3 | 1388 | 19.5 | 1.4% | 21.6 | 1.6% |
| POL 2 | 3 | 1500 | 10.3 | 0.7% | 11.7 | 0.8% |
| POL 3 | 3 | 1435 | 8.4 | 0.6% | 14.4 | 1.0% |

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| ALT | | ACE Alera | | | | |
| --- | --- | --- | --- | --- | --- | --- |
| Lab | Sample | Mean (U/L) | Within-Run | | Total | |
| | | | SD | %CV | SD | %CV |
| In-House | 1 | 32 | 1.1 | 3.5% | 1.2 | 3.9% |
| POL 1 | 1 | 28 | 1.4 | 5.1% | 2.4 | 8.4% |
| POL 2 | 1 | 26 | 0.9 | 3.6% | 2.1 | 8.2% |
| POL 3 | 1 | 29 | 2.1 | 7.5% | 2.4 | 8.4% |
| In-House | 2 | 190 | 4.0 | 2.1% | 4.1 | 2.1% |
| POL 1 | 2 | 193 | 1.7 | 0.9% | 2.0 | 1.0% |
| POL 2 | 2 | 194 | 2.2 | 1.1% | 2.5 | 1.3% |
| POL 3 | 2 | 195 | 3.5 | 1.8% | 5.0 | 2.6% |
| In-House | 3 | 307 | 4.0 | 1.3% | 4.1 | 1.3% |
| POL 1 | 3 | 309 | 3.1 | 1.0% | 3.6 | 1.2% |
| POL 2 | 3 | 314 | 2.3 | 0.7% | 2.6 | 0.8% |
| POL 3 | 3 | 310 | 8.5 | 2.8% | 9.1 | 3.0% |
| AST | | ACE Alera | | | | |
| --- | --- | --- | --- | --- | --- | --- |
| Lab | Sample | Mean (U/L) | Within-Run | | Total | |
| | | | SD | %CV | SD | %CV |
| In-House | 1 | 28 | 1.2 | 4.5% | 1.7 | 6.0% |
| POL 1 | 1 | 26 | 1.0 | 3.8% | 1.3 | 5.1% |
| POL 2 | 1 | 27 | 2.0 | 7.5% | 2.4 | 8.9% |
| POL 3 | 1 | 29 | 2.9 | 9.9% | 2.9 | 9.9% |
| In-House | 2 | 222 | 4.8 | 2.2% | 7.1 | 3.2% |
| POL 1 | 2 | 220 | 2.0 | 0.9% | 3.2 | 1.5% |
| POL 2 | 2 | 233 | 3.8 | 1.6% | 5.0 | 2.1% |
| POL 3 | 2 | 229 | 5.3 | 2.3% | 7.2 | 3.2% |
| In-House | 3 | 406 | 2.9 | 0.7% | 6.8 | 1.7% |
| POL 1 | 3 | 416 | 7.8 | 1.9% | 9.2 | 2.2% |
| POL 2 | 3 | 428 | 5.1 | 1.2% | 5.6 | 1.3% |
| POL 3 | 3 | 417 | 8.2 | 2.0% | 12.1 | 2.9% |

Additional precision studies were performed in-house using 3 concentrations of lithium heparin plasma samples over a period of 3 or more days $(n = 20)$ . All samples were tested twice a day in duplicate on the ACE, ACE Axcel, and ACE Alera Clinical Chemistry Systems. The within-run and total precision results are summarized in the tables below.

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Plasma in-house precision study:

| Precision (SD, %CV) | | | | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| ALP U/L | ACE Mean | Within-Run | Total | Alera Mean | Within-Run | Total | Axcel Mean | Within-Run | Total |
| Plasma Low | 76 | 1.9, 2.5% | 2.8, 3.7% | 75 | 0.8, 1.1% | 3.4, 4.6% | 74 | 1.1, 1.5% | 3.7, 5.1% |
| Plasma Mid | 614 | 5.8, 0.9% | 24.4, 4.0% | 609 | 5.1, 0.8% | 20.2, 3.3% | 613 | 3.4, 0.6% | 20.5, 3.3% |
| Plasma High | 1163 | 6.8, 0.6% | 33.5, 2.9% | 1149 | 5.9, 0.5% | 32.9, 2.9% | 1155 | 7.6, 0.7% | 35.7, 3.1% |
| Precision (SD, %CV) | | | | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| AMY U/L | ACE Mean | Within-Run | Total | Alera Mean | Within-Run | Total | Axcel Mean | Within-Run | Total |
| Plasma Low | 41 | 0.7, 1.8% | 1.3, 3.2% | 42 | 0.5, 1.3% | 1.5, 3.5% | 41 | 1.0, 2.5% | 2.5, 6.0% |
| Plasma Mid | 806 | 8.1, 1.0% | 12.5, 1.5% | 801 | 4.4, 0.5% | 13.1, 1.6% | 805 | 5.3, 0.7% | 12.2, 1.5% |
| Plasma High | 1604 | 15.4, 1.0% | 29.3, 1.8% | 1596 | 18.8, 2.1% | 33.8, 2.1% | 1604 | 21.6, 1.3% | 25.0, 1.6% |
| Precision (SD, %CV) | | | | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| ALT U/L | ACE Mean | Within-Run | Total | Alera Mean | Within-Run | Total | Axcel Mean | Within-Run | Total |
| Plasma Low | 32 | 0.9, 2.7% | 1.3, 4.0% | 32 | 0.8 2.6% | 1.5, 4.7% | 34 | 1.0, 2.9% | 1.4, 4.1% |
| Plasma Mid | 112 | 1.1, 1.0% | 1.5, 1.4% | 112 | 0.9, 0.8% | 1.1, 1.0% | 113 | 1.0, 0.9% | 1.5, 1.4% |
| Plasma High | 219 | 1.1, 0.5% | 1.6, 0.7% | 219 | 2.1, 0.9% | 3.0, 1.4% | 222 | 1.7, 0.7% | 2.7, 1.2% |
| Precision (SD, %CV) | | | | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| AST U/L | ACE Mean | Within-Run | Total | Alera Mean | Within-Run | Total | Axcel Mean | Within-Run | Total |
| Plasma Low | 26 | 0.8, 3.2% | 1.4, 5.4% | 26 | 0.6, 2.5% | 1.0, 3.7% | 26 | 0.9, 3.5% | 1.0, 4.0% |
| Plasma Mid | 157 | 1.6, 1.0% | 2.3, 1.4% | 157 | 1.5, 0.9% | 1.7, 1.1% | 158 | 1.4, 0.9% | 2.0, 1.3% |
| Plasma High | 304 | 3.5, 1.2% | 4.5, 1.5% | 303 | 2.6 0.9% | 3.9, 1.3% | 305 | 3.2, 1.1% | 4.3, 1.4% |

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# b. Linearity/assay reportable range:

Serum samples were spiked with the appropriate analyte and used to determine linearity on the ACE Alera Clinical Chemistry System. The assigned values of the highest and lowest sample were set to their mean values. The assigned values of the other levels were calculated by multiplying the mean value by the dilution proportions used. All samples were run in triplicate. The linear regression correlation between the expected values and the measured values is summarized below:

| ACE Reagents | Low level tested | Upper level tested | Linear Regression Equation | R2 |
| --- | --- | --- | --- | --- |
| ALP | 4 U/L | 1401 U/L | y = 0.998x - 0.5 | 0.9993 |
| Amylase | 4 U/L | 2012 U/L | y = 1.013x + 0.2 | 0.9974 |
| ALT | 3.1 U/L | 503.8U/L | y = 1.007x - 0.17 | 0.9992 |
| AST | 3 U/L | 491.3 U/L | y = 1.013x + 0.24 | 0.9992 |

The linearity data provided by the sponsor support the following reportable range claims:

| Analyte tested | Assay range |
| --- | --- |
| ALP (U/L) | 9 - 1400 |
| Amylase (U/L) | 9 - 1900 |
| ALT (U/L) | 4 - 480 |
| AST (U/L) | 4 - 450 |

# Automatic Dilution Study:

Sponsor performed additional auto-dilution studies to confirm the auto-dilution function (1:2 for ALP and 1:4 for Amylase, ALT and AST) on the ACE Alera analyzer for plasma and serum samples by comparing the auto-diluted results to the manual dilution results. ALT, AST, ALP and Amylase samples recovered within $\pm 10\%$ bias.

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

The ALP assay and the Amylase assay are traceable to an IFCC traceable method. The ALT assay is traceable to ERM-AD454 and the AST assay is traceable to ERM-AD457.

Calibration is not needed for these four assays.

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# d. Detection limit:

The Limit of blank (LoB), limit of detection (LoD), and limit of quantification (LoQ) were determined according to CLSI EP17-A with the ACE Alera Clinical Chemistry System. For the LoB and LoD studies, five low samples and five blanks $(n = 60$ reps, 20 reps per day) were tested over three days on two ACE Alera Clinical Chemistry Systems. For the LoQ studies, five samples $(n = 40$ reps, 8 reps per run) were tested in five separate runs over five days. LoQ values are based on inter-assay precision of $\leq 20\%$ CV. The results are as follows:

| ACE Alera | ALP | Amylase | ALT | AST |
| --- | --- | --- | --- | --- |
| LoB (U/L) | 2.8 | 0.2 | 1.6 | 2.2 |
| LoD (U/L) | 4.8 | 0.9 | 3.1 | 3.3 |
| LoQ (U/L) | 4.8 | 5.6 | 4.1 | 3.3 |

Sponsor claims the following measuring ranges for the assays:

| Analyte | Assay range |
| --- | --- |
| ALP (U/L) | 9 - 1400 |
| Amylase (U/L) | 9 - 1900 |
| ALT (U/L) | 4 - 480 |
| AST (U/L) | 4 - 450 |

# e. Analytical specificity:

Interference studies were performed to determine the effects from potential interferents. The various concentration of interferent was spiked into serum pools containing analytes at normal and abnormal concentrations. All samples were tested in triplicate. Six interferent levels and the control samples were tested for each interferent $(n = 7)$ . For lipemia interference studies, Intralipid was used as the added interferent with ALP and Amylase, while avian triglycerides concentrate was used with ALT and AST. Bias greater than $+ / - 10\%$ defines significant interference by the sponsor. The results of the highest concentration tested without significant interference are as follows:

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| Interferents on ACE Alera | No Significant Interference at or below: | | | |
| --- | --- | --- | --- | --- |
| | ALP | Amylase | ALT | AST |
| Icterus | 70.6 mg/dL | 30.0 mg/dL | 50 mg/dL | 50 mg/dL |
| Hemolysis | 62.5 mg/dL | 62.5 mg/dL | 500 mg/dL | 62.5 mg/dL |
| Lipemia | 1000 mg/dL | 1000 mg/dL | 419 mg/dL | 439 mg/dL |
| Ascorbic Acid | 6 mg/dL | 6 mg/dL | 6 mg/dL | 6 mg/dL |

Limitations are included in the package insert to reflect these results. Since hemolysis affects ALP, Amylase, and AST results, the sponsor has the following limitations in the labeling:

"Specimens showing any indication of hemolysis should not be analyzed."

For a comprehensive list of drugs and other substances which can affect assay in serum please refer to the literature given by Young, et. Al.

Young, D.S., et.al., Effects of drugs on Clinical Laboratory Tests, $5^{\text{th}}$ edition, AACC press, Washington D.C. (2000).

f. Assay cut-off:

Not applicable

# 2. Comparison studies:

a. Method comparison with predicate device:

Method comparison studies were completed at 3 POC sites following CLSI document EP9-A2. Samples were run on the ACE Clinical Chemistry System at Alfa Wassermann and the results were compared against those gathered on ACE Alera Clinical Chemistry Systems at 3 Physician Office Labs. For each test method, at least 40 determinations were made in singlicate for serum samples drawn from the same individuals on each platform. To test across the assay reportable ranges, 5-6 samples were spiked per analyte. No samples were diluted with saline.

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The following chart summarizes the POL method comparison studies:

| ALP | ACE (in-house) vs. ACE Alera (POL) | | |
| --- | --- | --- | --- |
| | POL 1 | POL 2 | POL 3 |
| N | 50 | 50 | 50 |
| Range (mg/dL) | 58 to 1199 | 58 to 1199 | 58 to 1199 |
| Slope | 0.997 | 1.029 | 1.010 |
| Intercept | -4.6 | -4.1 | -6.6 |
| Correlation Coefficient | 0.9992 | 0.9991 | 0.9986 |
| AMY | ACE (in-house) vs. ACE Alera (POL) | | |
| --- | --- | --- | --- |
| | POL 1 | POL 2 | POL 3 |
| N | 51 | 51 | 51 |
| Range (mmol/L) | 28 to 1732 | 28 to 1732 | 28 to 1732 |
| Slope | 0.960 | 1.010 | 0.990 |
| Intercept | 3.0 | 5.8 | 3.7 |
| Correlation Coefficient | 0.9991 | 0.9995 | 0.9995 |
| ALT | ACE (in-house) vs. ACE Alera (POL) | | |
| --- | --- | --- | --- |
| | POL 1 | POL 2 | POL 3 |
| N | 50 | 50 | 50 |
| Range (mmol/L) | 6 to 442 | 6 to 442 | 6 to 442 |
| Slope | 1.019 | 1.012 | 0.970 |
| Intercept | -0.5 | -3.5 | 2.4 |
| Correlation Coefficient | 0.9986 | 0.9985 | 0.9977 |
| AST | ACE (in-house) vs. ACE Alera (POL) | | |
| --- | --- | --- | --- |
| | POL 1 | POL 2 | POL 3 |
| N | 50 | 50 | 50 |
| Range (mmol/L) | 6 to 413 | 6 to 413 | 6 to 413 |
| Slope | 1.028 | 1.040 | 1.004 |
| Intercept | 1.4 | 0.5 | 1.8 |
| Correlation Coefficient | 0.9995 | 0.9992 | 0.9995 |

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# b. Matrix comparison:

A matrix comparison study was performed using paired serum/lithium heparin plasma samples, with levels of each analyte that cover the measuring ranges of the assays, in singlicate on the ACE, ACE Alera, and ACE Axcel Clinical Chemistry Systems. To cover the assay range, 4-5 spiked samples were tested per analyte and 6 diluted samples were tested for ALP. Linear regression analyses (Deming) were performed using serum (x) and lithium heparin plasma (y), with the following results:

Matrix Comparison Studies

| Reagent | Results | | |
| --- | --- | --- | --- |
| | ACE | ACE Alera | ACE Axcel |
| ALP (U/L) | Pairs: 108 | Pairs: 108 | Pairs: 62 |
| | Range: 9-1274 | Range: 9-1202 | Range: 11-1222 |
| | Slope: 0.998 | Slope: 0.9952 | Slope: 0.9982 |
| | Intercept: -8.3 | Intercept: -6.4 | Intercept: -6.5 |
| | Correlation: 0.9980 | Correlation: 0.9952 | Correlation: 0.9982 |
| Amylase (U/L) | Pairs: 104 | Pairs: 101 | Pairs: 52 |
| | Range: 11-1766 | Range: 11-1703 | Range: 10-1890 |
| | Slope: 0.977 | Slope: 0.979 | Slope: 0.994 |
| | Intercept: 1.7 | Intercept: 0.9 | Intercept: -1.76 |
| | Correlation: 0.9995 | Correlation: 0.9994 | Correlation: 0.9996 |
| ALT (U/L) | Pairs: 54 | Pairs: 52 | Pairs:56 |
| | Range: 4-460 | Range: 5-463 | Range: 6-469 |
| | Slope: 1.003 | Slope: 1.000 | Slope: 0.985 |
| | Intercept: -3.6 | Intercept: -3.6 | Intercept: -3.35 |
| | Correlation: 0.9994 | Correlation: 0.9986 | Correlation: 0.9993 |
| AST (U/L) | Pairs: 53 | Pairs: 52 | Pairs: 52 |
| | Range: 4-404 | Range: 4-396 | Range: 4-408 |
| | Slope: 0.981 | Slope: 0.999 | Slope: 1.001 |
| | Intercept: 0.7 | Intercept: -0.6 | Intercept: 0.22 |
| | Correlation: 0.9992 | Correlation: 0.9989 | Correlation: 0.9989 |

# 3. Clinical studies:

a. Clinical Sensitivity:

Not applicable

b. Clinical specificity:

Not applicable

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c. Other clinical supportive data (when a. and b. are not applicable):

Not applicable

4. Clinical cut-off:

Not applicable

5. Expected values/Reference range:

Reference values for serum and plasma are provided in the labeling according to literature as follows:

ALP¹: 44 - 147 U/L
Amylase²: 20 - 104 U/L
ALT²: 5 - 30 U/L
ASP²: 7 - 31 U/L

1. Dugdale, D., Zieve, D. (05/30/2011). ALP- blood test: MedlinePlus Medical Encyclopedia.
2. Wu, A.H.B. (Ed.), Tietz Clinical Guide to Laboratory Tests, 4th Edition, Saunders Elsevier, S. Louis, Mo (2006).

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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