IMMUNALYSIS AMPHETAMINE ELISA FOR ORAL FLUIDS

K051579 · Immunalysis Corporation · DKZ · Oct 21, 2005 · Clinical Toxicology

Device Facts

• Record ID: K051579
• Device Name: IMMUNALYSIS AMPHETAMINE ELISA FOR ORAL FLUIDS
• Applicant: Immunalysis Corporation
• Product Code: DKZ · Clinical Toxicology
• Decision Date: Oct 21, 2005
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.3100
• Device Class: Class 2

Indications for Use

The Immunalysis Amphetamine ELISA test system utilizes an Enzyme Linked Immunoassay (ELISA) for the qualitative detection of amphetamine in oral fluid samples collected with the Quantisal™ oral fluid collection device using a cutoff of 50 ng/mL of d-amphetamine. This in vitro diagnostic device is intended for clinical laboratory use only. The Immunalysis amphetamine ELISA Kit for Oral Fluids provides only a preliminary analytical test result. A more specific alternate chemical method must be used in order to obtain a confirmed analytical result. Gas chromatography/ mass spectrometry (GS-MS) is the preferred confirmatory method (1). Clinical and Professional judgment should be applied to any drug of abuse test result, particularly when preliminary positive results are obtained.

Device Story

Immunalysis Amphetamine ELISA is an in-vitro diagnostic test system for qualitative detection of amphetamine in oral fluid. Samples are collected using the Quantisal oral fluid collection device. The assay utilizes Enzyme Linked Immunoassay (ELISA) technology to detect amphetamine at a 50 ng/mL cutoff. The device is intended for use in clinical laboratories by trained personnel. The test provides preliminary analytical results; positive findings require confirmation via a more specific alternate chemical method, preferably Gas Chromatography/Mass Spectrometry (GC-MS). Results are intended to assist healthcare providers in clinical decision-making regarding drug abuse, requiring professional judgment for interpretation.

Clinical Evidence

No clinical data provided. Bench testing only.

Technological Characteristics

Enzyme Linked Immunoassay (ELISA) test system. Qualitative detection of amphetamine in oral fluid. Cutoff concentration: 50 ng/mL d-Amphetamine. Requires Quantisal oral fluid collection device. In-vitro diagnostic use.

Indications for Use

Indicated for qualitative detection of amphetamine in oral fluid samples for clinical laboratory use. Intended for use as a preliminary analytical test; requires confirmation by alternate chemical method (e.g., GC-MS).

Regulatory Classification

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).

Submission Summary (Full Text)

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

A. 510(k) Number:
k051579

B. Purpose for Submission:
New device

C. Measurand:
Amphetamine

D. Type of Test:
Qualitative immunoassay using oral fluid

E. Applicant:
Immunalysis Corporation

F. Proprietary and Established Names:
Immunalysis Amphetamine ELISA for Oral Fluids

G. Regulatory Information:
1. Regulation section:
21 CFR §862.3100, Amphetamine Test System
2. Classification:
Class II
3. Product code:
DKZ
4. Panel:
Toxicology (91)

H. Intended Use:
1. Intended use(s):
See indications for use below.
2. Indication(s) for use:
The Immunalysis Amphetamine ELISA test system utilizes an Enzyme Linked Immunoassay (ELISA) for the qualitative detection of amphetamine in oral fluid samples collected with the Quantisal™ oral fluid collection device using a cutoff of 50 ng/mL of d-amphetamine. This in vitro diagnostic device is intended for clinical laboratory use only.

The Immunalysis amphetamine ELISA Kit for Oral Fluids provides only a preliminary analytical test result. A more specific alternate chemical method must be used in order to obtain a confirmed analytical result. Gas chromatography/ mass spectrometry (GS-MS) is the preferred confirmatory method (1). Clinical and Professional judgment should be applied to any drug of abuse test result, particularly when preliminary positive results are obtained.

3. Special conditions for use statement(s):
See above.

4. Special instrument requirements:

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This device must be read on a spectrophotometer that reads wavelengths of 450 nm and 620 nm.

I. Device Description:
The ELISA assay contains a micro-plate coated with monoclonal mouse antibody to amphetamine (with a spacer chain) and the following reagents: amphetamine conjugate, negative and positive controls, a cut-off calibrator, enzyme-labeled amphetamine, and stop reagent. The device also consists of a saliva collection device. An oral fluid specimen is collected by placing the collection device, a cellulose pad affixed to a propylene stem under the tongue until approximately one milliliter saliva has saturated the pad. A blue indicator on the stem indicates when enough sample has been collected. The collector is transferred to a provided polypropylene tube containing preservative buffer (3 ml) and closed, ready for transport or storage.

J. Substantial Equivalence Information:
1. Predicate device name(s):
DRI Amphetamines EIA Assay
2. Predicate 510(k) number(s):
K934891
3. Comparison with predicate:

| Similarities | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Analyte | Amphetamine | Methamphetamine and Amphetamine |
| Methodology | Immunoassay (EIA) | Immunoassay (ELISA) |
| Differences | | |
| --- | --- | --- |
| Item | Device | Predicate |
| Test Matrix | Oral Fluid | Urine |
| Cutoff | 50 ng/mL | 1000 ng/mL |

K. Standard/Guidance Document Referenced (if applicable):
None referenced.

L. Test Principle:
Enzyme-labeled drug and drug present in the sample compete for limited antibody binding sites. Binding of the enzyme-labeled drug inhibits its reaction with the substrate, thereby influencing the rate of absorbance change measured by the instrument. The rate of absorbance change is proportional to the concentration of drug in the sample. Concentrations of controls and unknowns are calculated from the standard curve. Results are read at 450 and 620 nm.

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M. Performance Characteristics (if/when applicable):

1. Analytical performance:

a. Precision/Reproducibility:

Precision was tested by spiking negative oral fluid with 0, 25, 50, 75, and 100 ng/mL of d-methamphetamine; this corresponded to 0, 50%, 100%, 150%, and 200% of the cutoff. One milliliter of the spiked fluids were pipetted onto the collection pad of the oral fluid collector then processed as per instructions.

Intra-assay precision was estimated with sixteen replicates of each concentration analyzed in one run. Results are shown below:

| amphetamine (ng/mL) | Mean OD | Std Dev | CV |
| --- | --- | --- | --- |
| 0 | 1.79 | 0.138 | 7.71 % |
| 25 (50% c/o) | 0.726 | 0.052 | 7.29 % |
| 50 (100% c/o) | 0.463 | 0.025 | 5.45 % |
| 75 (150% c/o) | 0.304 | 0.015 | 5.18 % |
| 100 (200% c/o) | 0.255 | 0.019 | 7.49 % |

Inter-assay precision was assessed by eight replicates of each concentration run in 10 different assay runs (2 per day over 5 days). Results are expressed as B/B0% where B = absorbance of sample and B0 = absorbance of the zero calibrator.

| | amphetamine Concentration (ng/mL) | | | |
| --- | --- | --- | --- | --- |
| | 25 (50% c/o) | 50 (100% c/o) | 75 (150% c/o) | 100 (200% c/o) |
| Mean | 34.1 | 22.2 | 14.2 | 12.4 |
| Std.Dev | 2.4 | 1.6 | 1.1 | 0.90 |
| % CV | 7.1 | 7.0 | 8.0 | 7.2 |

Reproducibility of the oral fluid collection device was assessed by collecting oral fluid from 50 subjects with a pre-weighed collector and tube as per the package instructions. After the volume indicator turned blue, the collector and tube were weighed and the net weight of the saliva was determined and converted to volume (mLs). Results are shown below:

| Avg. Vol. (mL) | Std. Dev. | C.V. | Mean + 3 SD (mL) | Mean - 3 SD (mL) |
| --- | --- | --- | --- | --- |
| 0.993 | 0.029 | 2.88% | 1.079 | 0.907 |

b. Linearity/assay reportable range:

Not applicable. This assay is intended for qualitative use.

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c. Traceability, Stability, Expected values (controls, calibrators, or methods): All calibrator and control stock solutions are prepared from commercially available solutions. Amphetamine concentration is confirmed by GC/MS analysis. The error concentration allowed is +/-15% of the nominal concentration. Drug concentrations at 50%, 100% and 200% of the cutoff concentration are stored at 2-8 degrees C for stability testing. Results of testing up to 4 months indicate stability (within +/-15%). Further studies are ongoing.

Stability of specimens within the collection device was determined by spiking a pool of negative saliva with drug at a concentration near the cutoff. Samples were stored at 4°C or at room temperature (22-25°C). The specimens kept at room temperature were assayed in duplicate by GC-MS after 7 days, 14 days and 30 days; samples kept at 4°C were assayed were assayed in duplicate by GC-MS after 14 days and 30 days. Results are shown below:

| Day | amphetamine spike (ng/mL) | % initial value | amphetamine spike (ng/mL) | % initial value |
| --- | --- | --- | --- | --- |
| 0 | 40.03 | 100 | 81.90 | 100 |
| 7 | 41.30 | 103.2 | 80.34 | 98.1 |
| 14 | 41.57 | 103.8 | 72.70 | 88.77 |
| 21 | 39.04 | 97.53 | 76.12 | 92.94 |
| 30 | 38.40 | 95.93 | 69.24 | 84.54 |
| 0 | 40.03 | 100 | 81.90 | 100 |
| 14 | 40.00 | 99.9 | 69.40 | 84.7 |
| 30 | 36.71 | 91.7 | 74.08 | 90.5 |

A shipping study showed that recovery of drug in samples after 4 days in transit at temperatures ranging from 69-93°F, was similar to recovery of observed for samples held at 2-8°C.

d. Detection limit:

See the Precision/Reproducibility section above for performance around the stated cutoff concentration.

e. Analytical specificity:

Cross-reactivity of structurally similar compounds was determined by spiking concentrations of potentially cross-reacting drugs into synthetic oral fluid. Results are shown below:

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| Test compound | Conc. Tested (ng/mL) | Cross-Reactivity |
| --- | --- | --- |
| d-amphetamine | 25-75 | 100% |
| l-amphetamine | 250 | 9.7% |
| d-methamphetamine | 1000 | <0.1% |
| dl-methamphetamine | 1000 | ND |
| l-methamphetamine | 5000 | <0.02% |
| dl-MDMA | 500 | ND |
| dl-MDA | 200 | 179% |
| dl-MDEA | 1000 | ND |
| (+) pseudoephedrine | 1000 | 3.7% |
| (-) pseudoephedrine | 500 | <1% |
| phenolpropanolamine | 5000 | <0.2 |
| (+) ephedrine | 10000 | ND |
| (-) ephedrine | 10000 | 0.42% |
| fenfluramine | 1000 | ND |
| diphenhydramine | 10000 | ND |

Structurally unrelated compounds were spiked into synthetic oral fluid at a concentration of 10000 ng/mL each; none of the compounds tested affected assay results. The list of compounds tested is in the package insert.

Commonly ingested substances were tested for interference. Results are shown below:

| Compound | Mean Abs | B/B0% | POS/NEG |
| --- | --- | --- | --- |
| Synthetic Oral Fluid | 2.799 | 100 | |
| 25 ng/mL amphetamine | 1.438 | 51.38 | NEG |
| 50 ng/mL amphetamine | 0.735 | 26.26 | Cutoff |
| 100 ng/mL amphetamine | 0.499 | 17.81 | POS |
| | | | |
| | 25 ng/mL amp spike | | 75 ng/mL amp spike |
| | Mean Abs | B/B0% | POS/NEG |
| Distilled water | 1.409 | 50.32 | NEG |
| Sugar water solution | 1.268 | 45.3 | NEG |
| Toothpaste slurry | 1.146 | 41.0 | NEG |
| Cranberry juice | 1.164 | 41.6 | NEG |
| Baking Soda sol'n | 1.207 | 43.1 | NEG |
| Orange juice | 0.86 | 30.8 | NEG |
| Cola | 1.015 | 36.3 | NEG |
| Cough syrup* | 1.411 | 50.4 | NEG |
| Mouthwash | 1.217 | 43.6 | NEG |

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f. Assay cut-off:

Performance around the assay cut-off of 50 ng/mL is demonstrated in the intra-assay precision section above.

The Substance Abuse and Mental Health Services Administration (SAMHSA) currently recommend 50 ng/mL as a cutoff level for amphetamine oral fluid tests.

2. Comparison studies:

a. Method comparison with predicate device:

Oral fluid and urine samples were collected at the same time from admitted amphetamine users in a clinical setting. Urine samples were tested by the predicate assay using a cutoff of 1000 ng/mL. Oral fluid samples were tested in duplicate using a screening cutoff of 50 ng/mL. All samples were tested by GC/MS at an independent facility. Twenty two samples contained amphetamine (according to GCMS) either below, or near the cutoff concentration.

Comparison of Immunalysis Amphetamine Oral Fluid Assay and the Predicate Urine Assay

| | Predicate
Urine
Assay | | |
| --- | --- | --- | --- |
| | | Pos | Neg |
| Amphetamine
Oral Fluid
Assay | Pos | 39 | 2 |
| | Neg | 33 | 111 |

Comparison of Immunalysis Amphetamine Oral Fluid Assay and GC/MS

| | GC/MS | | |
| --- | --- | --- | --- |
| | | Pos | Neg |
| Amphetamine
Oral Fluid Assay | Pos | 38 | 3 |
| | Neg | 0 | 144 |

Positive agreement: 92.7 %
Negative agreement: 100 %
Overall agreement: 98.3 %

b. Matrix comparison:

Not applicable; this device is intended for use with oral fluid only.

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):

4. Clinical cut-off: Not applicable.
5. Expected values/Reference range: Not applicable.

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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