← Product Code [DKZ](/submissions/TX/subpart-d%E2%80%94clinical-toxicology-test-systems/DKZ) · K033743

# COZART EIA AMPHETAMINES ORAL FLUID MICROPLATE KIT (K033743)

_Cozart Bioscience , Ltd. · DKZ · Jun 3, 2004 · Clinical Toxicology · SESE_

**Canonical URL:** https://fda.innolitics.com/submissions/TX/subpart-d%E2%80%94clinical-toxicology-test-systems/DKZ/K033743

## Device Facts

- **Applicant:** Cozart Bioscience , Ltd.
- **Product Code:** [DKZ](/submissions/TX/subpart-d%E2%80%94clinical-toxicology-test-systems/DKZ.md)
- **Decision Date:** Jun 3, 2004
- **Decision:** SESE
- **Submission Type:** Traditional
- **Regulation:** 21 CFR 862.3100
- **Device Class:** Class 2
- **Review Panel:** Clinical Toxicology

## Indications for Use

The Cozart® EIA Amphetamines Oral Fluid Microplate Kit is intended for use in clinical and forensic laboratories when used in conjunction with the Cozart® RapiScan Oral Fluid collection system. Using this collection system it provides qualitative screening results for amphetamines in human oral fluid at a cutoff concentration of 15ng/ml. This is equal to 45ng/ml in neat oral fluid as the collection system involves a 1:3 dilution of the sample. This assay is for professional use only and provides only a preliminary analytical test result. Clinical consideration and professional judgement must be applied to any drug of abuse test result, particularly in evaluating a preliminary positive result. In order to obtain a more confirmed analytical result a more specific alternative chemical method is needed. Gas Chromatography/Mass Spectrometry (GCMS) is the preferred confirmatory method.

## Device Story

The Cozart® EIA Amphetamines Oral Fluid Microplate Kit is a laboratory-based competitive ELISA for detecting amphetamines in human oral fluid. Input consists of oral fluid samples collected via the Cozart® RapiScan system (1:3 dilution). The assay utilizes a microtitre plate coated with antibody and a Horse Radish Peroxidase enzyme conjugate. The reaction is measured spectrophotometrically at 450nm. The device is operated by laboratory professionals in clinical or forensic settings. Results are qualitative; positive findings require confirmation by GC/MS. The device aids in identifying potential amphetamine presence, supporting clinical or forensic decision-making.

## Clinical Evidence

Bench testing only. Method comparison study of 163 oral fluid samples compared to GC/MS confirmation. Results: 67 samples screened positive (62 confirmed positive, 5 false positives); 96 samples screened negative (94 confirmed negative, 2 false negatives). Overall agreement with GC/MS was 96%. Precision studies (within-assay, within-day, between-assay, between-day) showed CVs ranging from 2.7% to 12.4%. Specificity testing evaluated 20 unrelated substances (no cross-reactivity) and 11 related compounds (cross-reactivity observed for MDA, MAMP, MBDB, MDMA, MDEA, Fenfluramine, Tyramine).

## Technological Characteristics

Competitive ELISA; microtitre plate coated with antibody; Horse Radish Peroxidase enzyme conjugate; substrate/stop solutions; 450nm spectrophotometric detection. Calibrators: 0, 2, 15, 50ng/mL. Manual assay. Standards: BS EN ISO 9001:2000; EN 46001:1996.

## Regulatory Identification

An amphetamine test system is a device intended to measure amphetamine, a central nervous system stimulating drug, in plasma and urine. Measurements obtained by this device are used in the diagnosis and treatment of amphetamine use or overdose and in monitoring levels of amphetamine to ensure appropriate therapy.

## Special Controls

*Classification.* Class II (special controls). An amphetamine test system is not exempt if it is intended for any use other than employment or insurance testing or is intended for Federal drug testing programs. The device is exempt from the premarket notification procedures in subpart E of part 807 of this chapter subject to the limitations in § 862.9, provided the test system is intended for employment and insurance testing and includes a statement in the labeling that the device is intended solely for use in employment and insurance testing, and does not include devices intended for Federal drug testing programs (*e.g.,* programs run by the Substance Abuse and Mental Health Services Administration (SAMHSA), the Department of Transportation (DOT), and the U.S. military).

## Predicate Devices

- Amphetamine-Specific Intercept™ MICRO-PLATE EIA ([K992918](/device/K992918.md))

## Submission Summary (Full Text)

> This content was OCRed from public FDA records by [Innolitics](https://innolitics.com). If you use, quote, summarize, crawl, or train on this content, cite Innolitics at https://innolitics.com.
>
> Innolitics is a medical-device software consultancy. We help companies design, build, and clear FDA-regulated software and AI/ML devices, including [a 510(k)](https://innolitics.com/services/510ks/), [a De Novo](https://innolitics.com/services/regulatory/), [a SaMD](https://innolitics.com/services/end-to-end-samd/), [an AI/ML medical device](https://innolitics.com/services/medical-imaging-ai-development/), or [an FDA regulatory strategy](https://innolitics.com/services/regulatory/).

{0}

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

A. 510(k) Number: K033743

B. Purpose for Submission:
Premarket Notification 510(k) of intention to manufacture and market the Cozart® EIA Amphetamines Oral Fluid Microplate Kit.

C. Analyte: Amphetamine

D. Type of Test: Qualitative competitive enzyme immunoassay for the detection of amphetamines in human oral fluid.

E. Applicant: Cozart Bioscience, Ltd.

F. Proprietary and Established Names:
Cozart® EIA Amphetamines Oral Fluid Microplate Kit

G. Regulatory Information:
1. Regulation section: 21CFR §862.3100 Amphetamine test system
2. Classification: Class II
3. Product Code: DKZ
4. Panel: Toxicology (91)

H. Intended Use:
1. Intended use(s):
The Cozart® EIA Amphetamines Oral Fluid Microplate Kit is intended for use in clinical and forensic laboratories when used in conjunction with the Cozart® RapiScan Oral Fluid collection system. Using this collection system it provides qualitative screening results for amphetamines in human oral fluid at a cutoff concentration of 15ng/mL. This is equal to 45ng/mL in neat oral fluid as the collection system involves a 1:3 dilution of the sample.
2. Indication(s) for use:
The Cozart® EIA Amphetamines Oral Fluid Microplate Kit is intended for use in clinical and forensic laboratories when used in conjunction with the Cozart® RapiScan Oral Fluid collection system. Using this collection system it provides qualitative screening

{1}

Page 2 of 9

results for amphetamines in human oral fluid at a cutoff concentration of 15ng/mL. This is equal to 45ng/mL in neat oral fluid as the collection system involves a 1:3 dilution of the sample.

3. Special condition for use statement(s):

This assay is for professional use only and provides only a preliminary analytical result. Clinical consideration and professional judgement must be applied to any drug of abuse test result, particularly in evaluating a preliminary positive result. In order to obtain a more confirmed analytical result a more specific alternative chemical method is needed. Gas Chromatography/ Mass Spectrophotometry (GC/MS) is the preferred confirmatory method.

4. Special instrument Requirements:

Automated microtitre plate reader with a 450nm filter. Automated microtitre plate washing machine, manual microtiter plate washer or 350μL eight-channel pipette for dispensing diluted wash buffer.

I. Device Description:

The Cozart® EIA Amphetamine Oral Fluid Microplate Kit is supplied with the following reagents –

- a microtitre plate coated with antibody
- enzyme conjugate reagent
- wash buffer
- substrate solution
- stop solution
- four calibrators (0, 2, 15, and 50 ng/mL Amphetamine in synthetic oral fluid matrix)

Materials required but not provided –

- positive and negative controls
- automated microtitre plate reader with a 450nm filter
- precision pipettes with disposable tips.
- automated microtitre plate washing machine
- manual microtitre plate washer or 350μL eight-channel pipette for dispensing diluted wash buffer
- a timer for timing 30 minute intervals
- a clean measuring cylinder for dilution of wash buffer concentrate
- distilled or deionized water

J. Substantial Equivalence Information:

1. Predicate device name(s):

Amphetamine-Specific Intercept™ MICRO-PLATE EIA

{2}

Page 3 of 9

2. Predicate K number(s): K992918

3. Comparison with predicate:

|  Parameter | Cozart® EIA Amphetamine Oral Fluid Microplate Kit | Amphetamine-Specific Intercept™ MICRO-PLATE EIA  |
| --- | --- | --- |
|  Intended Use | Qualitative test for amphetamines in human oral fluid with a 45ng/mL cutoff. Recommended confirmation of positive results by GC/MS. | Qualitative test for amphetamines in human oral fluid with a 100ng/mL cutoff. Recommended confirmation of positive results by GC/MS.  |
|  Target Population | Clinical and forensic samples. | Clinical samples  |
|  Design | Competitive ELISA | Competitive ELISA  |
|  Enzyme | Horse Radish Peroxidase | Horse Radish Peroxidase  |
|  Results | Read spectrophotometrically at 450nm. | Read spectrophotometrically at 450nm.  |
|  Calibrators | 0, 2, 15, 50ng/mL | 0, 100ng/mL  |
|  Matrix | Human Oral Fluid | Human Oral Fluid  |
|  Controls | None supplied but Cozart recommends using external controls. | 50, 200 ng/mL  |
|  Method Comparison | 163 samples were tested, 67 screened positive for amphetamines, of which 62 were confirmed positive by GC/MS. 96 samples screened negative for amphetamines and 94 were confirmed negative by GC/MS. 96% Agreement as compared to GC/MS. | 89% Agreement as compared to GC/MS.  |
|  Precision | CV (%) of 2.7 – 12.4% | CV (%) of 3.5 – 7.9%  |
|  Sensitivity | 1.2 ng/mL | Unknown  |
|  Specificity | 20 potential interferents tested – none cross-reacted. | 47 potential interferents tested – none cross-reacted.  |

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

Cut-Off validation Study was conducted with cutoff concentration of $\pm 50\%$ of the value. Interference screening to identify exogenous and endogenous compounds was done in accordance with the NCCLS EP7-A.

{3}

Page 4 of 9

L. Test Principle: The Cozart® EIA Amphetamine Oral Fluid Microplate Kit is a competitive enzyme immunoassay for the detection of amphetamines in human oral fluid. The wells of the microtitre strips are coated with anti-Amphetamine antibody. During the first incubation, the horseradish peroxidase (HRP) labelled amphetamine derivative competes with the free amphetamine in the patients sample for the anti-Amphetamine antibody binding sites on the microtitre strips. The wells are washed to remove any excess enzyme material prior to addition of the TMB substrate solution. Addition of the stop solution terminates the reaction and absorbances are read spectrophotometrically at 450nm.

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

### 1. Analytical performance:

#### a. Precision/Reproducibility:

Four calibrators (0, 2, 15, and 50ng/mL) and three samples (0, 7.7 and 22.5ng/mL) were tested in duplicate every day for twenty days. The samples were a negative, a negative sample fortified with amphetamine at 50% below the cutoff and a negative sample fortified with amphetamine at 50% above the cutoff. The within assay, within day, between assay, between day and total precision for the absorbance readings is displayed below.

## Inter-Assay Precision

|  Sample | Between Assay  |   |   |
| --- | --- | --- | --- |
|   |  Mean | SD | CV%  |
|  Cal 1 | 2.287 | 0.140 | 6.11  |
|  Cal 2 | 1.406 | 0.099 | 7.02  |
|  Cal 3 | 0.848 | 0.069 | 8.11  |
|  Cal 4 | 0.575 | 0.063 | 10.91  |
|  Sample 1 | 2.252 | 0.159 | 7.05  |
|  Sample 2 | 1.090 | 0.104 | 9.55  |
|  Sample 3 | 0.771 | 0.075 | 9.73  |

## Inter-Assay Precision

|  Sample | Between Day  |   |   |
| --- | --- | --- | --- |
|   |  Mean | SD | CV%  |
|  Cal 1 | 2.287 | 0.132 | 5.79  |
|  Cal 2 | 1.406 | 0.092 | 6.54  |
|  Cal 3 | 0.848 | 0.066 | 7.84  |
|  Cal 4 | 0.575 | 0.061 | 10.59  |
|  Sample 1 | 2.252 | 0.153 | 6.78  |
|  Sample 2 | 1.090 | 0.094 | 8.66  |
|  Sample 3 | 0.771 | 0.073 | 9.41  |

{4}

Page 5 of 9

# Intra-Assay Precision

|  Sample | Within Assay  |   |   |
| --- | --- | --- | --- |
|   |  Mean | SD | CV%  |
|  Cal 1 | 2.320 | 0.06 | 2.74  |
|  Cal 2 | 1.427 | 0.041 | 2.96  |
|  Cal 3 | 0.861 | 0.029 | 3.35  |
|  Cal 4 | 0.586 | 0.036 | 7.11  |
|  Low | 2.286 | 0.063 | 2.85  |
|  Medium | 1.109 | 0.042 | 3.62  |
|  High | 0.784 | 0.03 | 3.94  |

# Intra-Assay Precision

|  Sample | Within Day  |   |   |
| --- | --- | --- | --- |
|   |  Mean | SD | CV%  |
|  Cal 1 | 2.350 | 0.076 | 3.38  |
|  Cal 2 | 1.446 | 0.057 | 4.14  |
|  Cal 3 | 0.872 | 0.033 | 3.98  |
|  Cal 4 | 0.593 | 0.036 | 6.97  |
|  Low | 2.315 | 0.077 | 3.47  |
|  Medium | 1.122 | 0.065 | 5.81  |
|  High | 0.794 | 0.036 | 4.75  |

# Total Precision

|  Sample | Within Assay  |   |   |
| --- | --- | --- | --- |
|   |  Mean | SD | CV%  |
|  Cal 1 | 2.287 | 0.154 | 6.73  |
|  Cal 2 | 1.406 | 0.110 | 7.82  |
|  Cal 3 | 0.848 | 0.075 | 8.84  |
|  Cal 4 | 0.575 | 0.071 | 12.35  |
|  Low | 2.252 | 0.173 | 7.68  |
|  Medium | 1.090 | 0.118 | 10.83  |
|  High | 0.771 | 0.081 | 10.54  |

b. Linearity/assay reportable range:
Not applicable. This test is for qualitative determinations.

c. Traceability (controls, calibrators, or method):
Each of the four cocaine calibrators supplied with the Cozart EIA Amphetamines Oral Fluid Microplate Kit were confirmed by GC/MS, the results are displayed below.

{5}

Page 6 of 9

|  Calibrator Value (ng/mL) | GC/MS Cocaine Concentration (ng/mL)  |
| --- | --- |
|  0 | 0  |
|  2 | 1.67  |
|  15 | 14.67  |
|  50 | 50.33  |

The sponsor will make available to the user a calibrator (control) protocol through Cozart Technical Services. Information for technical assistance is provided in the package insert.

d. Detection limit:

The sensitivity was calculated by testing the 0 calibrator twenty times in a single assay. A calibration curve was plotted with the four calibrators. The mean absorbance of the 20 zeros minus two standard deviations was calculated. The sensitivity was calculated by reading this absorbance value off the calibration curve. The sensitivity of the Cozart EIA Amphetamines Oral Fluid Microplate EIA is 1.2ng/mL.

e. Analytical specificity:

Twenty potentially interfering unrelated substances were tested for cross reactivity in the Cozart Amphetamine Oral Fluid Kit and none were found to cross react. Eleven related compounds were tested and seven showed a level of cross reactivity. The substances were prepared in a negative oral fluid sample prior to testing in the assay. A calibration curve was plotted using the four calibrators to estimate the concentration of each sample.

{6}

Page 7 of 9

|  Compound | ng/mL Tested | Apparent Amphetamine ng/mL | % Cross Reactivity  |
| --- | --- | --- | --- |
|  MDA | 5 | 10.8 | 216  |
|   | 10 | 18.8 | 188  |
|  MAMP | 1,000 | 9.1 | 0.91  |
|   | 5,000 | 86.6 | 1.73  |
|  MBDB | 5,000 | 5.2 | 0.10  |
|   | 10,000 | 19.9 | 0.20  |
|  MDMA | 1,000 | 10.0 | 1.00  |
|   | 5,000 | 90.0 | 1.80  |
|  MDEA | 5,000 | 7.7 | 0.15  |
|   | 10,000 | 22.3 | 0.22  |
|  Fenfluramine | 100,000 | 6.2 | 0.01  |
|  Tyramine | 1,000 | 3.1 | 0.31  |
|   | 5,000 | 42.9 | 0.86  |
|  (+) Ephedrine | 10,000 | 0.9 | 0.009  |
|   | 100,000 | 26.4 | 0.026  |
|  (-) Ephedrine | 100,000 | 2.8 | 0.003  |
|  (+) Pseudo Ephedrine | 100,000 | 10.3 | 0.010  |
|  (-) Pseudo Ephedrine | 100,000 | <2 | <0.002  |
|  |   |   |   |
|  Morphine | 100,000 | <2 | <0.002  |
|  Temazepam | 100,000 | <2 | <0.002  |
|  Cocaine | 100,000 | <2 | <0.002  |
|  Chloroquine | 100,000 | <2 | <0.002  |
|  Diazepam | 100,000 | <2 | <0.002  |
|  Amitriptyline HCL | 100,000 | <2 | <0.002  |
|  Dextromethorphan | 100,000 | <2 | <0.002  |
|  Ranitidine | 100,000 | 6.6 | 0.007  |
|  Ascorbic acid | 100,000 | <2 | <0.002  |
|  DL-Propranolol | 100,000 | <2 | <0.002  |
|  Caffeine | 100,000 | <2 | <0.002  |
|  Acetylsalicylic acid (Aspirin) | 100,000 | <2 | <0.002  |
|  Acetaminophen (Paracetamol) | 100,000 | <2 | <0.002  |
|  Quinalbarbitone | 100,000 | <2 | <0.002  |
|  Methylphenidate | 100,000 | <2 | <0.002  |
|  Buprenorphine | 10,000 | <2 | <0.002  |
|  Phenobarbital | 100,000 | <2 | <0.002  |
|  Nicotine | 100,000 | <2 | <0.002  |
|  LSD | 100,000 | <2 | <0.002  |
|  Cotinine | 100,000 | <2 | <0.002  |

{7}

# f. Assay cut-off:

Testing samples at the cutoff concentration,  $50\%$  above and  $50\%$  below was carried out to validate the cutoff concentration. The  $50\mathrm{ng / mL}$  calibrator was diluted with a negative oral fluid sample to 7.5, 15, and  $22.5\mathrm{ng / mL}$ . Ten replicates of each concentration were tested along with the four calibrators. The mean absorbance of the ten replicates of the  $15\mathrm{ng / mL}$  calibrator was used as the cutoff. The absorbances obtained for the  $7.5\mathrm{ng / mL}$  calibrator were all higher than the  $15\mathrm{ng / mL}$  cutoff calibrator. Similarly the absorbances obtained for the  $22.5\mathrm{ng / mL}$  calibrator were all lower than the  $15\mathrm{ng / mL}$  cutoff calibrator.

|  Sample Concentration | 7.5ng/mL  |
| --- | --- |
|  Mean Absorbance | 0.857  |
|  SD | 0.04  |
|  CV (%) | 4.51  |
|  Sample Concentration | 15ng/mL  |
| --- | --- |
|  Mean Absorbance | 0.723  |
|  SD | 0.03  |
|  CV (%) | 4.74  |
|  Sample Concentration | 22.5ng/mL  |
| --- | --- |
|  Mean Absorbance | 0.654  |
|  SD | 0.02  |
|  CV (%) | 3.29  |

# 2. Comparison studies:

# a. Method comparison with predicate device:

163 samples were tested through the Cozart® EIA Amphetamine Oral Fluid Microplate Kit. 67 samples screened positive for amphetamines, these consisted of 62 samples from drug users attending drug dependency units. And 5 samples spiked at  $60\mathrm{ng / mL}$  to give samples around the cutoff. 62 of the samples that screened positive for amphetamines were then confirmed positive y GC/MS. 3 of the unconfirmed positive samples were confirmed positive for MDMA at  $&gt;180\mathrm{ng / mL}$  and  $149\mathrm{ng / mL}$  but negative for MDA and amphetamine and therefore classed as negative for the reason stated above. However they screened positive due to the small cross reactivity of the kit to MDMA. 96 samples screened negative and 94 were confirmed negative by GC/MS. Of the 163 samples tested 17 were between  $-50\%$  cutoff and  $+50\%$  cutoff.

|  New Device |   | GC/MS Negs | GC/MS Negs between -50% Cutoff and Cutoff | GC/MS between +50% Cutoff and Cutoff | Total GC/MS Pos | Percent Agreement with GC/MS  |
| --- | --- | --- | --- | --- | --- | --- |
|  Pos | 67 | 5 | 0 | 8 | 62 | 93  |
|  Neg | 96 | 94 | 9 | 0 | 2 | 98  |

96% overall agreement as compared with GC/MS

{8}

Page 9 of 9

b. Matrix comparison:
Not applicable. This device is indicated only for oral fluid specimens.

3. Clinical studies:
a. Clinical sensitivity:
Not applicable. Clinical studies are not typically submitted for this device type.
b. Clinical specificity:
Not applicable. Clinical studies are not typically submitted for this device type.
c. Other clinical supportive data (when a and b are not applicable):

3. Clinical cut-off:
Analytical characterization of performance around the cut-off was demonstrated in the precision studies.

5. Expected values/Reference range:
Not applicable.

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

---

**Source:** [https://fda.innolitics.com/submissions/TX/subpart-d%E2%80%94clinical-toxicology-test-systems/DKZ/K033743](https://fda.innolitics.com/submissions/TX/subpart-d%E2%80%94clinical-toxicology-test-systems/DKZ/K033743)

**Published by [Innolitics](https://innolitics.com)** — a medical-device software consultancy. We help companies design, build, and clear FDA-regulated software and AI/ML devices. If you're preparing [a 510(k)](https://innolitics.com/services/510ks/), [a De Novo](https://innolitics.com/services/regulatory/), [a SaMD](https://innolitics.com/services/end-to-end-samd/), [an AI/ML medical device](https://innolitics.com/services/medical-imaging-ai-development/), or [an FDA regulatory strategy](https://innolitics.com/services/regulatory/), [get in touch](https://innolitics.com/contact).

**Cite:** Innolitics at https://innolitics.com