ENZYME IMMUNOASSAY FOR THE DETECTION OF SALIVARY CORTISOL

K051733 · Drg Intl., Inc. · NHG · Dec 7, 2005 · Clinical Chemistry

Device Facts

• Record ID: K051733
• Device Name: ENZYME IMMUNOASSAY FOR THE DETECTION OF SALIVARY CORTISOL
• Applicant: Drg Intl., Inc.
• Product Code: NHG · Clinical Chemistry
• Decision Date: Dec 7, 2005
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.1205
• Device Class: Class 2

Intended Use

The DRG Salivary Cortisol ELISA Test is an enzyme immunoassay for the quantitative in vitro diagnostic measurement of active free cortisol (hydrocortisone and hydroxycorticosterone) in saliva. Measurements of cortisol are used in the diagnosis and treatment of disorders of the adrenal gland.

Device Story

The DRG Salivary Cortisol ELISA is an enzyme immunoassay used in clinical laboratories to measure free cortisol in human saliva. The device utilizes a competitive binding principle: cortisol in the patient sample competes with horseradish peroxidase-conjugated cortisol for binding sites on mouse polyclonal cortisol-antiserum coated onto microplate wells. After incubation and washing, a TMB substrate is added; the resulting optical density is inversely proportional to the cortisol concentration in the sample. Healthcare providers use these quantitative results to assist in diagnosing and managing adrenal gland disorders. The test provides a diagnostic tool for clinicians to assess adrenal function via non-invasive salivary sampling.

Clinical Evidence

No clinical trials were performed. Evidence consists of analytical bench testing and method comparison studies. Method comparison against a commercial LIA method (n=41, range 3.10–13.01 ng/mL) showed correlation r=0.9876. Comparison against a commercial EIA method (n=30, range <0.5–11.86 ng/mL) showed correlation r=0.95742. Precision studies (intra-assay CV 5.55–6.96%; inter-assay CV 5.82–7.47%) and linearity studies (1.04–77 ng/mL) support analytical performance. Reference range established using 109 healthy adult samples (0.12–1.47 µg/dL).

Technological Characteristics

Enzyme-linked immunosorbent assay (ELISA) using competitive binding principle. Microplate wells coated with mouse polyclonal cortisol-antiserum. Detection via horseradish peroxidase conjugate and TMB substrate. Optical density measurement at 450 nm. Usable range: 0.537–77 ng/mL. Manual or automated plate reader required.

Indications for Use

Indicated for the quantitative measurement of active free cortisol in saliva for the diagnosis and treatment of adrenal gland disorders in adult patients.

Regulatory Classification

Identification

A cortisol (hydrocortisone and hydroxycorticosterone) test system is a device intended to measure the cortisol hormones secreted by the adrenal gland in plasma and urine. Measurements of cortisol are used in the diagnosis and treatment of disorders of the adrenal gland.

Submission Summary (Full Text)

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K051733

# DEC 7 2005

# SUMMARY OF SAFETY AND EFFECTIVENESS FOR DRG SALIVARY CORTISOL ELISA

| Manufacturer: | DRG International, Inc.<br>1167 U.S. Highway 22<br>Mountainside, NJ 07092 |
|----------------------|----------------------------------------------------------------------------------------------------------------|
| Contact Information: | Lehnus & Associates<br>Gary Lehnus<br>150 Cherry Lane Rd.<br>East Stroudsburg, PA 18301<br>Tel: [phone] |

#### Device Name / Classification:

The device trade name is the DRG SLV Cortisol ELISA having FDA assigned name: Cortisol test system, 21 CFR, 862.1205, categorized as Class II medical devices for the Clinical Chemistry and Clinical Toxicology Panel, as Product Code NHG.

# Test Principle

The DRG Salivary Cortisol ELISA KIT is based on the competition principle and the microplate separation. An unknown amount of Cortisol present in the sample and a fixed amount of Cortisol coniugated with horseradish peroxidase compete for the binding sites of mouse polyclonal Cortisol-antiserum coated onto the wells. After one hour incubation the microplate is washed to stop the competition reaction. After addition of the TMB substrate solution the concentration of Cortisol is inversely proportional to the optical density measured.

# Device Intended Use:

An enzyme immunoassay for the quantitative in vitro diagnostic measurement of active free cortisol (hydrocortisone and hydroxycorticosterone) in saliva. Measurements of cortisol are used in the diagnosis and treatment of disorders of the adrenal gland.

# Device Performance

# Normal Range Study

In order to determine the normal range of SLV cortisol, 109 saliva samples from adult male and female apparently healthy subjects, ages 20 to 80 years, were collected in the morning and analyzed using the DRG SLV Cortisol ELISA kit. The following range was calculated from this study.

0.12 - 1.47 µg/dL or 1.2 - 14.7 ng/mL (AM collection) Adults:

# Method comparison

Studies were performed to compare the DRG SLV Cortisol test to commercially available tests. One study evaluated saliva samples from 114 subjects ages 40 to 70 years. The samples were run in duplicate on the DRG test and another commercially available LIA method to determine the concentration of Cortisol in the samples. A correlation of 0.872 was obtained versus this method.

A second study was performed using saliva samples from seventy-two (72) saliva samples collected from 40 - 70 year old men and run in duplicate on DRG

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and another commercially available EIA test. A correlation of 0.936 was observed compared to another EIA method.

Another study was performed comparing 28 saliva samples to a reference LC-MS method. A correlation of r = 0.89056 with a formula of y = 1.0144x + 1.7762 was obtained to this method.

To further demonstrate substantial equivalence of the DRG SLV test, additional expanded comparison studies were requested. One expanded study evaluated saliva samples from 40 subjects ages 25 - 65 years. The samples were run in duplicate on the DRG test and another commercially available LIA method to determine the concentration of Cortisol in the samples. An overall correlation of 0.9795 and a regression formula of y = 0.9588x - 0.0338 was obtained versus this method.

A second expanded study was performed using 40 saliva samples collected from men and women ages 25 - 65 years and run in duplicate on DRG and another commercially available EJA test. A correlation of 0.9920 with a regression formula of y = 1.0722x + 0.1482 was observed compared to another EIA method.

The DRG Cortisol ELISA test demonstrated equivalent performance to commercially available ELISA and LIA methods for detection of Cortisol in saliva.

# Sensitivity

The lowest detectable level of Cortisol that can be distinguished from the Zero Standard is 0.537 ng/mL or 0.0537 ug/dl at the 95 % confidence limit.

#### Specificity

The following materials have been evaluated for cross reactivity. The percentage indicates cross reactivity at 50% displacement compared to Cortisol.

| Steroid | % Cross<br>reaction |
|-----------------------------------|---------------------|
| Cortisol | 100% |
| Corticosterone | 29.00% |
| Cortisone | 3.00% |
| 11-Deoxycortisol | < 1,00% |
| 17-OH Progesterrone | < 0,50% |
| Prednisone | <0,10% |
| Progesterone | < 0,10% |
| Dexamethazone | < 0,10% |
| Desoxycorticosterone | < 0,10% |
| Dehydroepiandrosterone<br>sulfate | < 0,10% |
| Estradiol | < 0,10% |
| Estriol | < 0,10% |
| Estrone | < 0,10% |
| Testosterone | < 0,10% |

#### Reproducibility

#### Intra-Assay

The intra-assay variation was determined by replicate measurements of 4 saliva samples using DRG ELISA kit. The within assay variability is shown below:

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# Inter-Assay

The inter-assay (between-run) variation was determined by quadruplicate measurements of commercial control samples in three different days runs.

| Mean | 24.29 ng/mL | 40.85 ng/mL |
|--------|-------------|-------------|
| SD | 1.81 ng/mL | 2.38 ng/mL |
| CV (%) | 7.47 | 5.82 |
| n= | 12 | 12 |

# Inter-Lot

The Inter-Lot (between-lot) variation was determined by duplicate measurements of five saliva samples in three different kit lots. The between run variability is shown below.

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# Recovery

Recovery of the DRG ELISA was determined by adding increasing amounts of the analyte to three different saliva samples containing different amounts of endogenous analyte. Each sample (non-spiked and spiked) was assayed and analyte concentrations of the samples were calculated from the standard curve. The percentage recoveries were determined by comparing expected and measured values of the samples

| Sample | Endogenous<br>cortisol<br>ng/ml | Added<br>cortisol<br>ng/ml | Measured OD<br>mean of duplicate<br>(450 nm) | Measured Conc.<br>SLV cortisol<br>ng/ml | Expected<br>conc<br>ng/ml | Recovery<br>(%) |
|--------|---------------------------------|----------------------------|----------------------------------------------|-----------------------------------------|---------------------------|-----------------|
| 1 | 0.90 | 0.00 | 1.284 | 0.90 | | |
| | | 40.00 | 0.175 | 38.74 | 40.90 | 94.7 |
| | | 20.00 | 0.262 | 22.45 | 20.90 | 107.4 |
| | | 10.00 | 0.421 | 11.50 | 10.90 | 105.5 |
| | | 5.00 | 0.608 | 6.42 | 5.90 | 108.8 |
| 2 | 8.37 | 0.00 | 0.518 | 8.37 | | |
| | | 40.00 | 0.160 | 43.57 | 48.37 | 90.1 |
| | | 20.00 | 0.225 | 27.59 | 28.37 | 97.3 |
| | | 10.00 | 0.321 | 17.00 | 18.37 | 92.5 |
| | | 5.00 | 0.367 | 14.07 | 13.37 | 105.2 |
| 3 | 14.60 | 0.00 | 0.357 | 14.61 | | |
| | | 40.00 | 0.144 | 50.31 | 54.61 | 92.1 |
| | | 20.00 | 0.187 | 35.55 | 34.60 | 102.7 |
| | | 10.00 | 0.246 | 24.52 | 24.60 | 99.7 |
| | | 5.00 | 0.279 | 20.60 | 19.60 | 105.1 |

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# Linearity

Three samples (saliva) containing different amounts of analyte were serially diluted to 1:64 with zero standard and assayed with the DRG ELISA. The percentage recovery was calculated by comparing the expected and measured values for SLV cortisol. An assay linearity of 0.537 - 77 ng/mL has been identified as the usable range. Samples above this range must be diluted and re-run.

| | | Sample 1 | Sample 2 | Sample 3 |
|--------------------|-------|----------|----------|----------|
| Concentr. | ng/ml | 33.13 | 80.00 | 23.23 |
| Average % Recovery | | 107.0 | 99.1 | 97.5 |
| Range of | from | 101.1 | 97.8 | 92.4 |
| % Recovery | to | 114.0 | 99.6 | 104.4 |

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Image /page/4/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo consists of a stylized eagle with three stripes forming its wing and tail feathers. The text "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA" is arranged in a circular fashion around the eagle.

Public Health Service

Food and Drug Administration 2098 Gaither Road Rockville MD 20850

DEC 7 2005

DRG International, Inc. c/o Mr. Gary Lehnus Lehnus & Associates 150 Cherry Lane Rd. East Stroudsburg, PA 18301

Re: k051733

Trade/Device Name: DRG Salivary Cortisol ELISA test Regulation Number: 21 CFR 864.1205 Regulation Name: Cortisol (hydrocortisone and hydroxycorticosterone) test system Regulatory Class: Class II Product Code: NHG Dated: November 16, 2005 Received: November 21, 2005

Dear Mr. Lehnus:

We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to such additional controls. Existing major regulations affecting your device can be found in Title 21, Code of Federal Regulations (CFR), Parts 800 to 895. In addition, FDA may publish further announcements concerning your device in the Federal Register.

Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); and good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820).

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This letter will allow you to begin marketing your device as described in your Section 510(k) premarket notification. The FDA finding of substantial equivalence of your device to a legally promated predicate device results in a classification for your device and thus, permits your device to proceed to the market.

If you desire specific information about the application of labeling requirements to your device, or questions on the promotion and advertising of your device, please contact the Office of In Vitro Diagnostic Device Evaluation and Safety at [phone]. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21CFR Part 807.97). You may obtain other general information on your responsibilities under the Act from the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number [phone] or [phone] or at its Internet address http://www.fda.gov/cdrh/industry/support/index.html.

Sincerely yours,

Alberto Guts

Alberto Gutierrez, Ph.D. Director Division of Chemistry and Toxicology Office of In Vitro Diagnostic Device Evaluation and Safety Center for Devices and Radiological Health

Enclosure

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# Indications for Use

510(k) Number (if known):

Device Name: _________________________________________________________________________________________________________________________________________________________________

Indications For Use:

An enzyme immunoassay for the quantitative in vitro diagnostic measurement of active free All Che Immanoused Trenvroxycorticosterone) in saliva. Measurements of cortisol are used in the diagnosis and treatment of disorders of the adrenal gland.

Prescription Use X (Part 21 CFR 801 Subpart D) AND/OR

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

(Please DO NOT WRITE BELOW THIS LINE - CONTINUE ON ANOTHER PAGE IF NEEDED)

Concurrence of CDRH, Office of Device Evaluation (OIVD)

Division Sign-Off

**Division Sign-Off**

Office of In Vitro Diagnostic Ice Evaluation and Safety

510(k) K051733

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