VAP-NT CHOLESTEROL TEST

K062026 · Atherotech, Inc. · CHH · Mar 6, 2007 · Clinical Chemistry

Device Facts

• Record ID: K062026
• Device Name: VAP-NT CHOLESTEROL TEST
• Applicant: Atherotech, Inc.
• Product Code: CHH · Clinical Chemistry
• Decision Date: Mar 6, 2007
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1175
• Device Class: Class 1

Indications for Use

The VAP-NT cholesterol test is a device intended to measure total cholesterol, and its component lipoprotein fractions from VLDL to HDL, in fasting serum. Cholesterol measurements are used in the diagnosis and treatment of disorders involving excess cholesterol in the blood and lipid and lipoprotein metabolism disorders.

Device Story

VAP-NT Cholesterol Test is an in vitro diagnostic device used to measure total cholesterol and lipoprotein fractions (VLDL to HDL) in fasting serum samples. The device is intended for use in clinical laboratory settings to assist physicians in diagnosing and managing lipid metabolism disorders and conditions related to excess blood cholesterol. The test provides quantitative measurements of cholesterol components, which healthcare providers use to assess patient lipid profiles and guide clinical decision-making regarding treatment and monitoring of lipid-related disorders.

Clinical Evidence

No clinical data provided; device is a standard cholesterol test system.

Technological Characteristics

Ultracentrifugation-based density gradient separation; enzymatic colorimetric detection (cholesterol esterase/oxidase). Requires ultracentrifuge, spectrophotometer, and fluid handling fixture. Reagent: Roche CH (k952127). Connectivity: Bar-coded sample identification. Software: Mathematical deconvolution of absorbance curves into lipoprotein subcurves. Sterilization: Not specified.

Indications for Use

Indicated for the measurement of total cholesterol and lipoprotein fractions (VLDL to HDL) in fasting serum for the diagnosis and treatment of disorders involving excess blood cholesterol and lipid/lipoprotein metabolism disorders.

Regulatory Classification

Identification

A cholesterol (total) test system is a device intended to measure cholesterol in plasma and serum. Cholesterol measurements are used in the diagnosis and treatment of disorders involving excess cholesterol in the blood and lipid and lipoprotein metabolism disorders.

Submission Summary (Full Text)

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# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY AND INSTRUMENT COMBINATION TEMPLATE

A. 510(k) Number:
k062026

B. Purpose for Submission:
Notification of intent to manufacture and market a new device

C. Measurand:
Cholesterol, Low Density Lipoprotein Cholesterol (LDL-C), Very Low Density Lipoprotein Cholesterol (VLDL-C) and High Density Lipoprotein Cholesterol (HDL-C)

D. Type of Test:
Cholesterol, LDL-C, VLDL-C, and HDL-C is determined enzymatically using cholesterol esterase and cholesterol oxidase.

E. Applicant:
Atherotech, Inc

F. Proprietary and Established Names:
Proprietary – VAP-NT Cholesterol Test
Established – Cholesterol, LDL, VLDL, HDL

G. Regulatory Information:
1. Regulation section:
21 CFR 862.1175 – Cholesterol
21 CFR 862.1475 LDL-C, VLDL-C and HDL-C
2. Classification:
Class I, meets the limitation to the exemption (21 CFR 862.9 (c) (4))
3. Product code:
CHH, MRR, LBS

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4. Panel:
75, Chemistry

H. Intended Use:

1. Intended use(s):
See Indications for Use below

2. Indication(s) for use:
The VAP-NT Cholesterol test is a device intended to measure total cholesterol and its component lipoprotein fractions from VLDL, LDL and HDL, in fasting serum. Cholesterol measurements are used in the diagnosis and treatment of disorders involving excess cholesterol in the blood and lipid and lipid metabolism disorders.

3. Special conditions for use statement(s):
For in vitro diagnostic use only
For professional use only

4. Special instrument requirements:
Ultracentrifuge, Spectrophotometer

I. Device Description:

The VAP-NT Cholesterol Test is an in vitro diagnostic medical device used in conjunction with laboratory equipment. The VAP-NT Cholesterol Test is intended to measure total cholesterol, and its component lipoprotein fractions VLDL, LDL and HDL, in fasting serum.

Test Methodology

The VAP-NT Test method consists of a sample preparation phase in which the serum is mixed with potassium bromide (KBr) to adjust the density and layered beneath a saline solution. The sample is spun using an ultracentrifuge and vertical rotor to obtain a density-gradient.

The centrifuge tube is placed into a fluid handling fixture which drains the tube from the bottom and uses piston pumps to control the flow rates. The centrifuged specimen is drained at a defined rate and mixed with a commercially available reagent (Roche CH). The sample is mixed with reagent in a continuous flow by drawing each fluid alternately through a "y" shaped connector. The sample/reagent is heated as it flows through the temperature bath. The reacted specimen is delivered in a continuous flow process to the spectrometer flow cells for absorbance measurements.

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A cholesterol profile (absorbance curve) is obtained by plotting the individual absorbance measurements on the Y-coordinate and their relative position in the density gradient on the X-coordinate. The relative position is calculated from the sample drain time.

The VAP-NT software algorithm mathematically deconvolutes the absorbance curve into major lipoprotein curves. The area under each curve is directly proportional to the cholesterol value (i.e. mg/dl). Total Cholesterol is determined as a sum of the area under all subcurves.

The VAP-NT algorithm is based on the premise that the curve shape for individual lipoprotein class and subclass is generally consistent. These fixed subcurve peak parameters are analogous to the use of fixed density ranges for the separation of each lipoprotein class in the $\beta$ quantification method and electrophoretic methods.

# J. Substantial Equivalence Information:

1. Predicate device name(s):

Quantimetrix Lipoprint, Roche Diagnostics Reagent

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

Quantimetrix Lipoprint k010337 & k013662

Roche Diagnostics Reagent k952127

3. Comparison with predicate:

Substantial Equivalency Comparison- Similarities and Differences

| Parameter | VAP-NT Cholesterol Test | Lipoprint | Roche Cholesterol Reagent |
| --- | --- | --- | --- |
| 510k Number | pending | k010337 and k013662 | k952127 |
| Device Name | VAP-NT Cholesterol Test | LipoPrint LDL and HDL | Cholesterol Assay |
| Intended Use | The VAP cholesterol test is a device intended to measure total cholesterol, and its component lipoprotein fractions ,VLDL, LDL and HDL, in fasting serum. Cholesterol measurements are used in the diagnosis and treatment of disorders involving excess cholesterol in the blood and lipid and lipoprotein metabolism disorders | Intended to measure lipoprotein cholesterol (for lipoprotein fractions and subfractions from VLDL to HDL) in fasting serum or plasma with a Total Cholesterol concentration of ≥ 100 mg/dl. Lipoprotein cholesterol measurements are use as an aid in evaluating lipid metabolism disorders when used in conjunction with other lipid tests, patient risk | Enzymatic in vitro test for the direct quantitative determination of cholesterol in human serum and plasma on automated clinical chemistry analyzers |

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| Parameter | VAP-NT Cholesterol Test | Lipoprint | Roche Cholesterol Reagent |
| --- | --- | --- | --- |
| | | assessment and clinical evaluation | |
| Measurand | Total Cholesterol LDL Cholesterol, HDL Cholesterol, VLDL Cholesterol | LDL Cholesterol, HDL Cholesterol, VLDL Cholesterol | Total Cholesterol |
| Methodology | Multi-step procedure that combines separation by ultracentrifugation, colorimetric enzyme based determination of cholesterol and spectrophotometric characterization of the reaction. | Electrophoretic separation | Colorimetric- enzyme based |
| Testing Environment | Clinical Laboratory | Clinical Laboratory | Clinical Laboratory |
| Reporting Units | mg/dl | mg/dl | mg/dl |
| Sample Type | Serum | Serum or plasma | Serum or plasma |
| Controls/Calibration | Labeling recommends Calibration/ Control materials and procedures | Labeling recommends Calibration/ Control materials and procedures | Labeling recommends Calibration/ Control materials and procedures |
| Reporting Values | Numeric Values Peak chart | Numeric Values Peak chart | Analyzer dependant |
| Specimen dilution | Requires mixture (dilution) of sample with potassium bromide and saline. | Requires mixture (dilution) of sample with loading gel. | Allows for dilution and dilution factor to obtain results |
| Parameter | VAP-NT Cholesterol Test | Lipoprint | Roche Cholesterol Reagent |
| Sample Type | Serum | Serum or plasma | Serum or plasma |
| Sample Size | 1-2 mL | < 1mL | Instrumentation dependant |
| Reagent Principal | Cholesterol is determined enzymatically using cholesterol esterase and cholesterol oxidase. The color intensity generated is directly proportional to the concentration of cholesterol and can be determined photometrically. | N/A | Cholesterol is determined enzymatically using cholesterol esterase and cholesterol oxidase. The color intensity generated is directly proportional to the concentration of cholesterol and can be determined photometrically. |
| Temperature of Reaction | 37C | Ambient | 37C |
| Assay Range | 32-785 mg/dl Total Cholesterol, | 13-695 mg/dl LDL 5-260 mg/dl HDL | 3-800 mg/dl |

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| Parameter | VAP-NT Cholesterol Test | Lipoprint | Roche Cholesterol Reagent |
| --- | --- | --- | --- |
| | 10-200 mg/dl HDL
60-250 mg/dl LDL
7-61 mg/dl for VLDL | 11-140 mg/dl VLDL | |
| Expected Values | According to the recommendations of NCEP Adult Treatment Panel III | According to the recommendations of NCEP Adult Treatment Panel III | According to the recommendations of NCEP Adult Treatment Panel III |
| Risk to Patient | Provides diagnostic information, which may indirectly affect the patient such that incorrect or delayed information could result in non-serious injury to the patient | Provides diagnostic information, which may indirectly affect the patient such that incorrect or delayed information could result in non-serious injury to the patient | Provides diagnostic information, which may indirectly affect the patient such that incorrect or delayed information could result in non-serious injury to the patient |
| Labeling | Targeted to the professional market | Targeted to the professional market | Targeted to the professional market |

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

Guidance for 510k’s on Cholesterol Tests for Clinical Laboratory, Physicians’ Office laboratory and Home Use

This device has not been tested by the Cholesterol Reference Method Laboratory Network

## L. Test Principle:

Cholesterol is determined enzymatically using cholesterol esterase and cholesterol oxidase. The color intensity generated is directly proportional to cholesterol concentration and can be determined photometrically using spectrophotometers. The VAP-NT software interprets the photometric reaction.

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

### 1. Analytical performance:

#### a. Precision/Reproducibility:

Quality Control Material was the test sample material. A total of three concentrations, near medical decision points, (low, intermediate, high) were used in the precision evaluation of the VAP-NT Cholesterol Test. The table below illustrates the approximate analyte values for each of the pools.

#### Data Collection

Cholesterol measurements were obtained for all samples using the VAP-NT Cholesterol Test procedure. All data collection took place after device-familiarization runs.

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Data was obtained over a 20 day period, using a single set of analytical equipment. Two (2) batches were run per day, and an additional 2 duplicate batches per day, for a total of 80 batches. Data for 2 replicate samples at each pool level were obtained for each batch run. One sample in a single run was excluded due to a centrifuge error. Each replicate specimen yielded analytical results for TC, HDL, LDL and VLDL, for a total of 160 data points per analyte, per level, with the exception of the high level analyte which yielded 159 data points.

The following within-run and between -run results were obtained from the analysis of the 20-day data collection period for the three levels of control material.

| Total Cholesterol | | | Within-run | | Between Run | | Between Day | | Total | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | n | Mean | SD | CV | SD | CV | SD | CV | SD | CV |
| High | 160 | 333.1 | 3.59 | 1.1% | 2.24 | 0.7% | 1.7 | 0.5% | 4.56 | 1.4% |
| Mid | 159 | 227 | 3.96 | 1.7% | 0 | 0% | 0.54 | 0.2% | 4 | 1.8% |
| Mid | 160 | 197 | 2.86 | 1.5% | 1.43 | 0.7% | 0 | 0% | 3.2 | 1.6% |
| Mid | 160 | 162 | 2.66 | 1.6% | 2.04 | 1.3% | 1.64 | 1.0% | 3.74 | 2.3% |
| Low | 160 | 98.9 | 1.96 | 2.0% | 0 | 0% | 1.06 | 1.1% | 2.22 | 2.2% |
| HDL | | | Within-run | | Between Run | | Between Day | | Total | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | n | Mean | SD | CV | SD | CV | SD | CV | SD | CV |
| High | 159 | 90.7 | 1.5 | 1.6% | 0 | 0% | 0.18 | 0.2% | 1.51 | 1.7% |
| Mid | 160 | 60.05 | 0.98 | 1.6% | 0.43 | 0.7% | 0.61 | 1.0% | 1.23 | 2% |
| Mid | 160 | 52.7 | 0.88 | 1.7% | 0 | 0% | 0 | 0% | 0.88 | 1.7% |
| Mid | 160 | 43.7 | 0.65 | 1.5% | 0.54 | 1.2% | 0.65 | 1.5% | 1.06 | 2.4% |
| Low | 160 | 34.4 | 0.6 | 1.8% | 0 | 0% | 0.36 | 1.0% | 0.7 | 2.0% |
| LDL | | | Within-run | | Between Run | | Between Day | | Total | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | n | Mean | SD | CV | SD | CV | SD | CV | SD | CV |
| High | 160 | 213 | 2.98 | 1.4% | 1.3 | 0.6% | 1.79 | 0.8% | 3.71 | 1.7% |
| Mid | 159 | 137.5 | 4.03 | 2.9% | 0 | 0% | 1.55 | 1.1% | 4.32 | 3.1% |
| Mid | 160 | 117 | 2.65 | 2.3% | 1.02 | 0.9% | 0 | 0% | 2.84 | 2.4% |
| Mid | 160 | 93.9 | 2.47 | 2.6% | 1.68 | 1.8% | 0 | 0% | 2.99 | 3.2% |
| Low | 160 | 65.6 | 2.46 | 3.7% | 0 | 0% | 0 | 0% | 2.46 | 3.7% |
| VLDL | | | Within-run | | Between Run | | Between Day | | Total | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | n | Mean | SD | CV | SD | CV | SD | CV | SD | CV |
| High | 159 | 52.45 | 2.22 | 4.2% | 0 | 0% | .46 | 0.9% | 2.27 | 4.3% |
| Mid | 160 | 29.64 | 2.59 | 8.7% | 0 | 0% | 1.17 | 4.0% | 2.84 | 9.6% |
| Mid | 160 | 27.16 | 1.75 | 6.4% | 0 | 0% | 0.91 | 3.3% | 1.97 | 7.2% |
| Mid | 160 | 24.8 | 1.73 | 7.0% | 1.09 | 4.4% | 0% | 0% | 2.04 | 8.2% |
| Low | 160 | 9.36 | 1.75 | 18.7% | 0 | 0% | 0.16 | 1.7% | 1.76 | 18/8% |

## b. Linearity/assay reportable range:

A total of 15 serum pools were created using a high analyte starting serum pool. The starting pool (sample 4) was either serially diluted or serially concentrated to achieve a wide range of each measured analyte. The table below illustrates the theoretical pool ranges for TC.

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Based on the above dilution ratios, the table below illustrates the estimated subclass values for the serial dilutions

| Sample | Relative Concentration | Est. HDL | Est. LDL | Est. VLDL |
| --- | --- | --- | --- | --- |
| 1 | 0.1 | 6 | 16 | 4 |
| 2 | 0.4 | 24 | 64 | 16 |
| 3 | 0.7 | 42 | 112 | 28 |
| 4 | 1.0 | 60 | 160 | 40 |
| 5 | 1.3 | 78 | 208 | 52 |
| 6 | 1.6 | 96 | 256 | 64 |
| 7 | 1.9 | 114 | 304 | 76 |
| 8 | 2.2 | 132 | 352 | 88 |
| 9 | 2.5 | 150 | 400 | 100 |
| 10 | 2.8 | 168 | 448 | 112 |
| 11 | 3.1 | 186 | 496 | 124 |
| 12 | 3.4 | 204 | 544 | 136 |
| 13 | 3.7 | 222 | 592 | 148 |
| 14 | 4.0 | 240 | 640 | 160 |
| 15 | 4.3 | 258 | 688 | 172 |

## Data Collection

Cholesterol measurement was obtained for all samples using the VAP-NT Cholesterol Test procedure. Data for 8 replicate samples at each pool level were obtained. A total of 12 batches were run. Two runs were rejected due to improper dilution techniques. Data collection took place within a single day, using a single set of analytical equipment. Three ultracentrifuges were used. The replicate samples were sequenced randomly for each run. Three individual samples (one each from Pool 2, 10 and 14) were eliminated for analysis due to a centrifuge error (tube collapsed). Each replicate specimen yielded analytical results for TC, HDL, LDL and VLDL, for a total of 117 data points per

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analyte.

## Data Analysis

The data was plotted and examined for excessive differences (errors) and potential outliers. The plot was created of the results with the observed results (Y) on the vertical axis and the theoretical sample concentration on the (X) horizontal axis.

$$
\begin{array}{l}
\text{Cholesterol } y = 0.9758x + 7.397 r^2 = 0.9997 \text{ for the range of } 32 - 785 \text{ mg/dL} \\
\text{HDL-C } y = 0.9645x + 3.4159 r^2 = 0.9998 \text{ for the range of } 10 - 200 \text{ mg/dL} \\
\text{LDL-C } y = 0.9867x - 5.2267 r^2 = 0.9992 \text{ for the range of } 60 - 250 \text{ mg/dL} \\
\text{VLDL-C } y = 1.1325x + 2.89 r^2 = 0.9986 \text{ for the range of } 7 - 61 \text{ mg/dL}
\end{array}
$$

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

The VAP-NT Cholesterol Test uses a commercially available cholesterol reagent from Roche Diagnostics. This reagent was reviewed and cleared for commercial distribution by the FDA under k952127.

## d. Detection limit:

A limit of detection study was performed based upon CLSI EP-17 – Protocols for determination of Limits of Detection and Limits for Quantitation. The Limit of blank (LOB) was assessed on 60 saline/KBr Samples. In addition 60 serum samples were used to establish the Limit of detection (LOD) per CLSI EP-17. The following results were obtained.

| Analyte | LOB | LOD |
| --- | --- | --- |
| TC | 6.1 | 29.5 |
| HDL | 2.6 | 9.4 |
| LDL | 2.6 | 14.1 |
| VLDL | 1.4 | 6.0 |

## e. Analytical specificity:

### Interferences

The VAP-NT Cholesterol Test uses a commercially available cholesterol reagent from Roche Diagnostics. This reagent was reviewed and cleared for commercial distribution by the FDA under 510(k) K952127.

As a part of the Roche reagent development, interference studies/analysis were performed and yielded the following information:

**Icterus:** No significant interference from conjugated bilirubin up to an I index of 25. No significant interference from unconjugated bilirubin up to an I index of 10.

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Hemolysis: No significant interference from hemolysis up to an H index of 700.

Lipemia: No significant interference from lipemia up to an L index of 1250.

As the VAP-NT test uses a reagent with known limitations, the interfering substances and conditions for the Roche reagent have been incorporated into the VAP-NT Cholesterol Test labeling.

In accordance with the NCEP ATP III guidelines, the VAP-NT cholesterol test procedure will recommend patient samples be obtained from fasting donors.

f. Assay cut-off:

Not applicable

2. Comparison studies:

a. Method comparison with predicate device:

The VAP-NT was compared to standard laboratory methods for the measurement of total cholesterol and cholesterol subclasses. For Total Cholesterol, the VAP-NT was compared to the Abell-Kendall method and Beta Quantification, and for the subclasses the VAP-NT was compared to Beta Quantification. The results of the study yielded to following method comparison data for the VAP-NT.

| | Total Cholesterol (BQ) | | HDL | | LDL | | VLDL | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | VAP | Ref | VAP | Ref. | VAP | Ref. | VAP | Ref. |
| n= | 492 | 492 | 416 | 416 | 416 | 416 | 336 | 336 |
| Ranges | 89-404 | | 24-90 | | 53-188 | | 0-60 | |
| Slope | 1.0373 | | 0.9645 | | 1.0037 | | 0.951965 | |
| Intercept | -4.3966 | | 2.8310 | | 0.6887 | | 2.52943 | |
| Coefficient of Determination (r²) | 0.9917 | | 0.9766 | | 0.9870 | | 0.884145 | |
| Average Bias (mg/dl) | 6.09 | | 2.46 | | 3.58 | | See table below | |
| Average Bias % | 2.49% | | 5.41% | | 3.33% | | See table below | |

VLDL Bias

| Group | N | Range (mg/dL) | Average Bias mg/dL | Bias % | Bias SD |
| --- | --- | --- | --- | --- | --- |
| Low | 112 | 9-17 | 2.69 | 19.07 | 1.9 |
| Mid | 112 | 18-29 | 3.35 | 13.69 | 2.7 |
| High | 112 | 30-60 | 3.90 | 10.63 | 3.6 |

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b. Matrix comparison:

Not Applicable.

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

5. Expected values/Reference range:

**Expected Values**

Guidelines for reference ranges have been suggested by the Panel of the National Institutes of Health's Cholesterol Consensus Development Conference and adopted by the National Cholesterol Education Program. The suggested guidelines of the Panel are as follows:

| Total Cholesterol Value | Risk levels Classification |
| --- | --- |
| < 200 mg/dL (5.17 mMol/L) | Desirable blood cholesterol |
| 200-239 mg/dL (5.17 - 6.18 mMol/L) | Borderline-high blood cholesterol |
| ≥ 240 mg/dL (6.20 mMol/L) | High blood cholesterol |
| HDL Cholesterol | Classification |
| <1.0 mmol/L 40.00 mg/dL | Low (undesirable, High risk) |
| ≥60.00 mg/dl | High (desirable, Low risk) |
| LDL Level | Classification |
| <100 mg/dL | Optimal |
| 100 - 129 mg/dL | Near optimal/above optimal |

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130 - 159 mg/dL
Borderline High
160 - 189 mg/dL
High
≥ 190 mg/dL
Very High

| VLDL Level | Classification |
| --- | --- |
| ≤30 mg/dL | Normal |
| >30 mg/dL | Not Normal |

N. Instrument Name:
VAP-NT Cholesterol

O. System Descriptions:
1. Modes of Operation:
Random channel, batch mode analyzer

2. Software:
FDA has reviewed applicant’s Hazard Analysis and software development processes for this line of product types:
Yes ☐ X ☐ or No ☐

3. Specimen Identification:
Bar Coded sample Identification

4. Specimen Sampling and Handling:
The VAP-NT Test method consists of a sample preparation phase in which serum is mixed with Potassium Bromide solution to adjust density and layered beneath a saline solution, the sample is then spun using an ultra-centrifuge and vertical rotor to obtain a density gradient.

5. Calibration:
The VAP-NT system is calibrated so that the total area under the VAP profile yields the measured total cholesterol (in mg/dL). The Calibrator material for use with the VAP-NT Cholesterol Test is bulk, filtered serum obtained from an outside research laboratory.

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6. Quality Control:

The Control material for use with the VAP-NT Cholesterol Test is bulk, filtered serum obtained from an outside research laboratory.

Atherotech recommends that a full rotor (batch) of control samples be run and reviewed at the beginning of each shift/work period to verify analysis is within specification.

P. Other Supportive Instrument Performance Characteristics Data Not Covered In The "Performance Characteristics" Section above:

Not Applicable

Q. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

R. Conclusion:

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