Rapidfret Oral Fluid Assay for Methamphetamine, Rapidfret Oral Fluid Methamphetamine Calibrators, Rapidfret Oral Fluid Methamphetamine Controls

K142293 · Biophor Diagnostics, Inc. · LAF · Sep 18, 2015 · Clinical Toxicology

Device Facts

• Record ID: K142293
• Device Name: Rapidfret Oral Fluid Assay for Methamphetamine, Rapidfret Oral Fluid Methamphetamine Calibrators, Rapidfret Oral Fluid Methamphetamine Controls
• Applicant: Biophor Diagnostics, Inc.
• Product Code: LAF · Clinical Toxicology
• Decision Date: Sep 18, 2015
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 862.3610
• Device Class: Class 2

Indications for Use

The RapidFRET Oral Fluid Assay for Methamphetamine is a homogeneous time-resolved fluorescence assay that is intended for prescription use in central laboratories only on the RapidFRET Integrated Workstation. The assay is used to perform a qualitative screen for methamphetamine at 50 ng/mL in neat oral fluid samples collected with the RapidEASE Oral Fluid Collector. This assay provides only a preliminary result. To obtain a confirmed analytical result, a more specific alternate chemical method such as GC/MS or LC/MS/MS is required. Professional judgment should be applied to any drug test result, particularly when using preliminary positive results. For In Vitro Diagnostic Use Only. The RapidFRET Oral Fluid Methamphetamine Calibrators and RapidFRET Oral Fluid Methamphetamine Controls are intended for use only with appropriate RapidFRET Oral Fluid Assay products and samples collected with the RapidEASE Oral Fluid Collector. The cutoff calibrator is used to determine the cutoff level and translate the assay measurement into a positive or negative result. The positive and negative controls are used to monitor laboratory systems, operators, precision, accuracy and assay conditions. For In Vitro Diagnostic Use Only.

Device Story

RapidFRET Oral Fluid Assay is a competitive homogeneous immunoassay for qualitative methamphetamine detection in oral fluid. Input: neat oral fluid collected via RapidEASE Oral Fluid Collector. Operation: drug in sample competes with donor-fluorophore-labeled drug for antibody-acceptor-fluorophore binding sites; binding brings fluorophores into proximity, enabling fluorescence resonance energy transfer (FRET). Output: FRET signal measured by RapidFRET Integrated Workstation; signal inversely proportional to drug concentration. Used in central laboratories by trained personnel. Results are preliminary; require confirmation via GC/MS or LC/MS/MS. Clinical decision-making relies on professional judgment; benefits include rapid screening for drug presence.

Clinical Evidence

Bench testing only. Precision evaluated over 20 days using spiked samples (0-200% of cutoff). Accuracy assessed by comparing 92 clinical samples against LC/MS/MS confirmation. Cross-reactivity and analytical specificity tested against structurally related compounds, common substances, and pH variations. Results demonstrate analytical sensitivity between 75% and 125% of cutoff and performance equivalent to the predicate.

Technological Characteristics

Competitive homogeneous time-resolved fluorescence immunoassay. Components: drug-specific antibody reagent and drug conjugate reagent. Platform: RapidFRET Integrated Workstation. Sample collection: RapidEASE Oral Fluid Collector (neat oral fluid, no diluent). Cutoff: 50 ng/mL. Controls/Calibrators: drug-specific materials for system monitoring.

Indications for Use

Indicated for qualitative screening of methamphetamine at 50 ng/mL in neat oral fluid samples. Intended for prescription use in central laboratories. Provides preliminary results requiring confirmation by GC/MS or LC/MS/MS.

Regulatory Classification

Identification

A methamphetamine test system is a device intended to measure methamphetamine, a central nervous system stimulating drug, in serum, plasma, and urine. Measurements obtained by this device are used in the diagnosis and treatment of methamphetamine use or overdose.

Special Controls

*Classification.* Class II (special controls). A methamphetamine 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

• Lin-Zhi International, Inc., LZI Oral Fluid Methamphetamine Enzyme Immunoassay (k131652)

Submission Summary (Full Text)

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

A. 510(k) Number:
K142293

B. Purpose for Submission:
New Device

C. Measurand:
Methamphetamine

D. Type of Test:
Qualitative Immunoassay

E. Applicant:
Biophor Diagnostics, Inc.

F. Proprietary and Established Names:
RapidFRET Oral Fluid Assay for Methamphetamine
RapidFRET Oral Fluid Methamphetamine Calibrators
RapidFRET Oral Fluid Methamphetamine Controls

G. Regulatory Information:

1. Regulation section:
21 CFR 862.3610, Methamphetamine test system
21 CFR 862.3200, Clinical toxicology calibrator
21 CFR 862.3280, Clinical toxicology control material

2. Classification:
Class II (test system, calibrator)
Class I, reserved (control material)

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3. Product code:
LAF, Gas Chromatography, Methamphetamine
DLJ, Calibrators, Drug Specific
LAS, Drug Specific Control Materials

4. Panel:
Toxicology (91)

H. Intended Use:

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

2. Indication(s) for use:
The RapidFRET Oral Fluid Assay for Methamphetamine is a homogeneous time-resolved fluorescence assay that is intended for prescription use in central laboratories only on the RapidFRET Integrated Workstation. The assay is used to perform a qualitative screen for methamphetamine at 50 ng/mL in neat oral fluid samples collected with the RapidEASE Oral Fluid Collector. This assay provides only a preliminary result. To obtain a confirmed analytical result, a more specific alternate chemical method such as GC/MS or LC/MS/MS is required. Professional judgment should be applied to any drug test result, particularly when using preliminary positive results. For In Vitro Diagnostic Use Only.

The RapidFRET Oral Fluid Methamphetamine Calibrators and RapidFRET Oral Fluid Methamphetamine Controls are intended for use only with appropriate RapidFRET Oral Fluid Assay products and samples collected with the RapidEASE Oral Fluid Collector. The cutoff calibrator is used to determine the cutoff level and translate the assay measurement into a positive or negative result. The positive and negative controls are used to monitor laboratory systems, operators, precision, accuracy and assay conditions. For In Vitro Diagnostic Use Only.

3. Special conditions for use statement(s):
For prescription use in Central Laboratories only.

4. Special instrument requirements:
For use with the RapidFRET Integrated Workstation.

I. Device Description:
The RapidFRET® Oral Fluid Assay for Methamphetamine is sold as a kit in two sizes. Each kit consists of 96 Well Microtiter Plates (round bottom plates), Methamphetamine Acceptor

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Reagent, Multi-Donor Reagent, Matrix Blank Reagent, the RapidEASE Oral Fluid Collector, Negative Calibrator (0 ng/mL), Cutoff Calibrator (50 ng/mL), Negative Control (25 ng/mL, 50% cutoff), and Positive Control (75 ng/mL, 150% cutoff).

The Calibrators and Controls Sets are required for running the assay and are purchased separately from the Assay Kit.

## J. Substantial Equivalence Information:

1. Predicate device name(s):

LZI Oral Fluid Methamphetamine Enzyme Immunoassay
LZI Oral Fluid Methamphetamine Calibrators
LZI Oral Fluid Methamphetamine Controls

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

k131652

3. Comparison with predicate:

| Similarities | | |
| --- | --- | --- |
| Item | Candidate Device
(Biophor RapidFRET Oral Fluid Assay
for Methamphetamine) | Predicate Device
LZI Oral Fluid Methamphetamine
Enzyme Immunoassay (K131652) |
| Intended Use | Qualitative determination of
methamphetamine in human oral fluid
in clinical setting. | Same |
| Neat Oral Fluid Cutoff Level | 50 ng/mL in neat oral fluid. | Same |
| Methodology | Competitive homogeneous
immunoassay. | Same |

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| Similarities | | |
| --- | --- | --- |
| Principle | Drugs in the oral fluid sample compete with the drug conjugate donor fluorophore for a fixed number of binding sites on the individual drug antibody acceptor reagents. When acceptor and donor fluorophores are brought into close proximity, through the binding event, fluorescent energy transfer is measured. The amount of drug in the specimen sample is inversely proportional to the assay signal as measured by time resolved fluorescence. | The assay is based on competition between drug in the sample and drug labeled with the donor fluorophore for a fixed number of binding sites on the antibody reagent. When acceptor and donor fluorophores are brought into close proximity through a binding event, energy transfer occurs. The fluorescence resonance energy transfer (FRET) signal is measured at the wavelength of the acceptor fluorophore following excitation of the donor and is inversely proportional to the amount of drug in the sample. |
| Differences | | |
| --- | --- | --- |
| Item | Candidate Device (Biophor RapidFRET Oral Fluid Assay for Methamphetamine) | Predicate Device LZI Oral Fluid Methamphetamine Enzyme Immunoassay (K131652) |
| Controls and Calibrator Levels | Calibrators are available at concentrations of 0 ng/mL and 50 ng/mL. Controls are available at concentrations of 25 ng/mL and 75 ng/mL. | Calibrators are available at concentrations of 0, 20, 50, 100, and 140 ng/mL. Controls are available at concentrations of 37.5 and 62.5 ng/mL. |
| Sample Collection | Neat oral fluid is collected with the RapidEASE Oral Fluid Collector via direct expectoration. No diluent is used and sample is stored in glass sample tube with inert screw cap. | Neat oral fluid is collected into a polypropylene collection tube via direct expectoration. No diluent is used and sample is stored in the collection tube. |
| Platform | RapidFRET Integrated Workstation available exclusively from Biophor Diagnostics, Inc. | Clinical chemistry analyzers |

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

None referenced.

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L. Test Principle:

The RapidFRET Oral Fluid Assay for Methamphetamine is an In Vitro Diagnostic competitive immunoassay used to detect methamphetamine in human oral fluid. This is a ready-to-use homogenous system that involves energy transfer between an acceptor fluorophore labeled to an antibody and a donor fluorophore labeled to drug. The assay is based on competition between drug in the sample and drug labeled with the donor fluorophore for a fixed number of binding sites on the antibody reagent. When acceptor and donor fluorophores are brought into close proximity through a binding event, energy transfer occurs. The fluorescence resonance energy transfer (FRET) signal is measured at the wavelength of the acceptor fluorophore and is inversely proportional to the amount of drug in the sample. A Cutoff Calibrator is used to translate the sample measurement into a positive or negative result. Controls are used to establish and monitor precision and accuracy.

M. Performance Characteristics (if/when applicable):

1. Analytical performance:

a. Precision/Reproducibility:

Three reagent lots of the RapidFRET Oral Fluid Assay for Methamphetamine were analyzed in 1-3 replicates per day on 1-4 plates per day for a minimum of 20 days. Negative oral fluid pools were spiked with methamphetamine derived from NIST weight traceable standards at 0%, 25%, 50%, 75%, 100%, 125%, 150%, 175% and 200% of the cutoff level corresponding to approximately 0, 12.5, 25, 37.5, 50, 62.5, 75, 87.5 and 100 ng/mL methamphetamine. Each spike level was processed through a RapidEASE collector and confirmed by LC/MS/MS.

Results for all lots are summarized below. All samples at concentrations higher than the cutoff reported positive results, and all samples at concentrations lower than the cutoff reported negative results.

| | 0% | 25% | 50% | 75% | 100% | 125% | 150% | 175% | 200% |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| POS | 0 | 0 | 0 | 0 | 47 | 264 | 264 | 264 | 264 |
| NEG | 264 | 264 | 264 | 264 | 217 | 0 | 0 | 0 | 0 |
| n | 264 | 264 | 264 | 264 | 264 | 264 | 264 | 264 | 264 |

b. Linearity/assay reportable range:

Not applicable.

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

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# Traceability

The cutoff calibrator and controls are prepared by spiking known concentrations of methamphetamine into synthetic oral fluid to obtain the cutoff level calibrator (50 ng/mL), and the positive (75 ng/mL) and negative (25 ng/mL) controls. The negative calibrator is drug free synthetic oral fluid. Calibrators and controls are prepared from *d*-methamphetamine (1 mg/mL) in methanol commercial primary standards from a vendor that uses NIST traceable weights and specific assays, such as HPLC and GC/MS, to confirm drug levels.

# Value Assignment – Calibrators and Controls

Calibrator and Control lots are value assigned during the manufacturing process in two stages. During the first stage following bottling and labeling, new lots are assayed against previously accepted, released and unexpired Calibrator and Control lot(s) using RapidFRET reagents. Results are qualitatively evaluated for performance relative to the previously accepted lots. If any of the new material does not pass this stage of evaluation it is not passed to the second stage.

During the second stage, each new manufactured lot of Calibrator or Control is quantitatively confirmed by a Mass Spectroscopy based method for target analyte concentration. Quantitative results within the sponsor’s acceptance criteria are released. Quantitative results that do not meet acceptance criteria are not released pending investigation and analysis.

# Calibrators and Controls Stability Studies

Real-time stability studies were conducted on multiple lots of RapidFRET Oral Fluid Calibrators and RapidFRET Oral Fluid Controls. The stability protocols for open and closed vial were reviewed and found acceptable. The open vial and closed vial study results support the open vial stability claim of 30 days and closed vial stability claim of 12 months when stored at 2 to 8 °C for the RapidFRET Oral Fluid Calibrators and RapidFRET Oral Fluid Controls.

# Sample Shipment - Stability Studies

A neat oral fluid pool was spiked with methamphetamine at 0%, 25%, 50%, 75%, 100%, 125%, 150%, 175%, and 200% of the cutoff, corresponding to 0, 12.5, 25.0, 37.5, 50.0, 62.5, 75.0, 87.5, and 100 ng/mL. Each spike was subsequently processed through a RapidEASE Oral Fluid Collection device to mimic actual collection process. Aliquots were stored and handled according to the collector insert.

One set of samples (9 concentrations) was shipped under high relative humidity conditions and a second set was shipped under low relative humidity conditions. Both sets were tested by RapidFRET and confirmed by LC/MS/MS prior to shipment. The procedure was to ship using an overnight commercial carrier from California to Maryland and back again to

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California and this was done twice over five days.

The "high humidity" set of samples was shipped under relative humidity conditions of 39-98% with a large majority of readings greater than 86.5% relative humidity. The temperatures for this set ranged from 10-25°C.

The "low humidity" set of samples was shipped under relative humidity conditions of 0-45% with a large majority of readings less than 15% relative humidity. The temperatures for this set ranged from 10.5-26.5°C.

Samples were quantitatively assayed by MS at study start and at various time points over the duration of the study. Methamphetamine recovery vs. day 1 at various time points over the course of the study ranged from 98.0 – 108.6% at high relative humidity and from 90.1 – 107.5% at low relative humidity conditions. The physical integrity of the RapidEASE sample tube was also evaluated following each shipment and no signs of degradation were noted.

## Sample Storage and Stability:

Conditions for oral fluid sample handling and storage were evaluated by preparing oral fluid samples with Methamphetamine at concentrations of 0, 12.5, 25.0, 37.5, 50.0, 62.5, 75.0, 87.5, and 100.0 ng/mL (0%, 25%, 50%, 75%, 100%, 125%, 150%, 175%, and 200% of the cutoff). Samples were processed through RapidEASE oral fluid collection devices and stored under various conditions including room temperature (21°C to 28°C) for 29 days, refrigerated (3°C to 7°C) for 86 days, and frozen (-10°C to -25°C) for 231 days. Samples were periodically sampled and analyzed by quantitative mass spectrometry. For each storage condition two sets of 9 levels each (18 total samples) were prepared and analyzed individually. Individual percent recoveries ranged from 89.6% to 109.6% after 29 days storage at room temperature, 88.8% to 110.4% after 86 days under refrigeration, and 88.8% to 107.2% after frozen storage for 231 days.

The sponsor states the following in their regarding sample storage: Samples may be stored at ambient temperatures for up to 7 days including shipping. Short term storage of samples is recommended at 2 – 8°C for up to 30 days. Samples should be frozen at -10°C to -25°C for long-term storage up to 7 months. All samples should be stored protected from light in collection vial with cap securely tightened.

## Sample Recovery Study

Recovery studies were conducted by aliquoting neat, human oral fluid pool into glass tubes and spiking with methamphetamine to achieve concentrations ranging from 0% of cutoff (0 ng/mL) to 200% of cutoff (100 ng/mL) in 5 replicates for each level in 25% (12.5 ng/mL) increments. Approximately half of the volume of each of these 'PRE-RapidEASE' samples was then removed and processed through a new RapidEASE Oral Fluid collector, mimicking as close as possible actual collection protocol, resulting in a 'POST-RapidEASE' sample. Both the PRE-RapidEASE and POST-RapidEASE for each spike level was

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confirmed for methamphetamine concentration by Mass Spectrometry. Results are summarized below:

| % Cutoff (ng/mL) | PRE-RapidEASE Mean Concentration (ng/mL) | POST-RapidEASE Mean Concentration (ng/mL) | Average Percent Recovery |
| --- | --- | --- | --- |
| 200 (100 ng/mL) | 104.2 | 102.5 | 98.3 |
| 175 (87.5 ng/mL) | 90.2 | 86.3 | 95.9 |
| 150 (75 ng/mL) | 78.2 | 80.0 | 102.3 |
| 125 (62.5 ng/mL) | 63.7 | 61.5 | 96.6 |
| 100 (50.0 ng/mL | 53.4 | 52.5 | 98.3 |
| 75 (37.5 ng/mL) | 37.8 | 37.4 | 99.1 |
| 50 (25.0 ng/mL) | 25.6 | 25.6 | 100.0 |
| 25 (12.5 ng/mL) | 12.7 | 12.2 | 96.4 |
| 0 (0.0 ng/mL) | Not detected | Not detected | Not applicable |

d. Detection limit:

Not applicable.

e. Analytical specificity:

An analytical specificity study of the assay to evaluate the interference from non-structurally and structurally related compounds was performed. The study design and results are described below.

Non-Structurally Related Compounds

Potential interference from structurally unrelated drugs and metabolites were evaluated by spiking these compounds at high concentrations into pooled negative oral fluid and in pooled oral fluid spiked with methamphetamine at $\pm 50\%$ of the cutoff (25 ng/mL and 75 ng/mL). No interference was observed with the following structurally unrelated compounds when tested up to a concentration of $30,000~\mathrm{ng/mL}$. 11-Hydroxy-$\Delta$-9-THC, $\Delta$-8-THC and O-Desmethylvenlafaxine were tested up to a concentration of $3000~\mathrm{ng/mL}$; LSD was tested up to a concentration of $1,500~\mathrm{ng/mL}$). Results are summarized below:

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| Cotinine | Dexbrompheniramine | Nicotine |
| --- | --- | --- |
| (-) Epinephrine | Dextromethorphan | Nitrazepam |
| (+) Brompheniramine | D-Glucose | N-Methylephedrine |
| (+) Chlorpheniramine | Diacetylmorphine (Heroin) | Norcocaine |
| (+) Naproxen | Diazepam | Nordiazepam |
| (+/-) Chlorpheniramine | Dihydrocodeine | Norketamine |
| (+/-) Epinephrine | Diphenhydramine | Normorphine |
| Isoprenaline | Diphenylhydantoin | Norpropoxyphene |
| (+/-) Methadone | Dopamine | Nortriptyline |
| (+/-) Pseudoephedrine | Doxepin | O-Desmethylvenlafaxine |
| (R, 2R) Pseudoephedrine | Doxylamine | Oxalic acid |
| 11-Hydroxy-Δ-9-THC (tetrahydrocannabinol) | d-Propoxyphene | Oxazepam |
| 4-Aminophenylsulfone | Ecgonine | Oxycodone |
| 4-Dimethylaminoantipyrine | Ecgonine methyl ester | Oxymorphone |
| 4-Hydroxy-PCP (phencyclidine) | EDDP (ethylidene-dimethyl - diphenylpyrrolidine) | Pantoprazole |
| 6-Monoacetylmorphine | Erythromycin | PCM (PCP Analog) |
| Acetaminophen | Ethylmorphine | Penicillin G |
| Acetylsalicylic acid | Fenoprofen | Pentazocine |
| Alprazolam | Fentanyl | Pentobarbital |
| Amitriptyline | Flunitrazepam | Perphenazine |
| Amobarbital | Fluoxetine | Phencyclidine |
| Ampicillin | Flurazepam | Phendimetrazine |
| Aprobarbital | Furosemide | Pheniramine |
| Ascorbic acid | Gentisic Acid | Phenobarbital |
| Aspartame | Glipizide | Phenothiazine |
| Atropine | Guaiacol glycerol | Phentermine |
| Benzocaine | Hydrocodone | Phenylpropanolamine |
| Benzoylecgonine | Hydromorphone | Prazepam |
| Bromazepam | Ibuprofen | Primidone |
| Buprenorphine | Imipramine | Promethazine |
| Butabarbital | Isoxsuprine | Protriptyline |
| Butalbital | Ketamine | Quetiapine |
| Caffeine | Levorphanol | Quinidine |
| Cannabidiol | Lidocaine | Rifampin |
| Cannabinol | Loperamide | Secobarbital |
| Carbamazepine | Lorazepam | Sulindac |
| Chlordiazepoxide | l-Phenylalanine | Theophylline |
| Chlorothiazide | LSD (Lysergic acid diethylamide) | Tramadol |
| Chlorpromazine | Maprotiline | Triazolam |

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| Clobazam | Medazepam | Trifluoperazine |
| --- | --- | --- |
| Clomipramine | Meperidine | Trimipramine |
| Clonazepam | Methadol | Tyramine |
| Clorazepate | Methaqualone | Venlafaxine |
| Cocaethylene | Methylphenidate | Δ-8-THC |
| Cocaine | Morphine | Δ-9-THC |
| Codeine | Morphine-3bDG | Δ-9-THC acid |
| Creatine | Nalorphine | Hydroxy-bupropion |
| Cyclizine | Naloxone | Dihydrobupropion |
| Cyclobenzaprine | Naltrexone | |
| Desipramine | Niacinamide | |

## Structurally Related Compounds

Potential interference from structurally related drugs and metabolites was evaluated by spiking these compounds at the concentrations below into neat oral fluid that did not contain methamphetamine. Compounds causing a positive result were titrated to determine the cutoff equivalence level. The cross-reactivity of structurally related compounds is summarized in the table below:

| Compound | Concentration yielding a result equivalent to a sample at the methamphetamine cutoff concentration (ng/mL) | Percent Cross-reactivity |
| --- | --- | --- |
| (-)-Ephedrine | 5,100 | 1.0 |
| Benzodioxolylbutanamine (BDB) | 16,000 | 0.3 |
| Phenethylamine | 5,700 | 0.9 |
| Chloroquine | 2,300 | 2.2 |
| d-Amphetamine | 3,500 | 1.4 |
| Fenfluramine | 290 | 17 |
| l-Methamphetamine | 300 | 17 |
| l-Phenylephrine | 9,400 | 0.5 |
| 4-methylethcathinone (4-MEC) | 4545 | 1.1 |
| 3,4-methylenedioxy-N-methylcathinone (Methylone) | 3333 | 1.5 |
| MBDB (Butylone) | 28 | 179 |
| 3,4-Methylenedioxyamphetamine (MDA) | 12,700 | 0.4 |
| 3,4-Methylenedioxyethylamphetamine (MDEA) | 1,100 | 4.5 |
| D,L 3,4-Methylenedioxymethamphetamine (MDMA) | 126 | 40 |
| Mephentermine | 1,500 | 3.3 |

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| (para-Methoxyamphetamine) PMA | 9,100 | 0.5 |
| --- | --- | --- |
| para-Methoxy-N-methylamphetamine (PMMA) | 87 | 57 |
| Procaine | 24,000 | 0.2 |
| Ranitidine | 8,300 | 0.6 |
| Trimethobenzamide | 730 | 6.8 |
| d-Ephedrine | 30,000 | <2 |
| l-Amphetamine | 30,000 | <2 |
| Procainamide | 30,000 | <2 |

# Potential Interferents and Common Substances

An interference study was performed to evaluate potential interference from endogenous substances that may be present in the oral fluid samples. Aliquots of a neat oral fluid pool were prepared and spiked with the potential interferent and with methamphetamine at concentrations of $0\mathrm{ng / mL}$ and approximately 25 and $75\mathrm{ng / mL}$ $(\pm 50\%)$ of the cutoff). Samples were then processed through a RapidEASE Oral Fluid Collector per Instructions and screened using the RapidFRET Oral Fluid Assay for Methamphetamine. All zero concentration samples and at $-50\%$ of cutoff gave negative results, and all samples at $+50\%$ of cutoff gave positive results. The potential interferents and concentrations tested are listed below:

| Potential Interferent | Concentration Tested |
| --- | --- |
| Human Serum Albumin (HSA) | 1.0 mg/mL |
| Alcohol (Ethanol) | 1% v/v |
| Baking Soda | 6% w/v |
| Whole Blood | 0.4% v/v |
| Hemoglobin | 0.5 mg/mL |
| Hydrogen Peroxide, OTC (3%) | 6% v/v |
| Sodium Chloride | 18 ng/mL |
| pH 5, 6, 7, 8, 9 | N/A |
| Cholesterol | 45 ng/mL |
| Denture Adhesive | 0.6% w/v |
| Ascorbic Acid | 1 mg/mL |
| Bilirubin | 150 ug/mL |
| IgA | 0.1 mg/mL |
| IgG | 0.5 mg/mL |
| IgM | 0.1 mg/mL |

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To evaluate potential interference from additional food and dental products, volunteers used the indicated product according to common practice or product instructions, then provided an oral fluid sample using the RapidEASE Oral Fluid Collector. Each oral fluid sample was then spiked with methamphetamine to approximately 25 and 75 ng/mL (±50% of the cutoff) and analyzed. All samples at -50% of cutoff gave negative results, and all samples at +50% of cutoff gave positive results. Results of the study are summarized below:

| Product | Amount Used by Volunteer |
| --- | --- |
| Antiseptic Mouthwash | 1 oz. |
| Cough Syrup | 1 teaspoon |
| Cranberry Juice | 6 oz. |
| Orange Juice | 8 oz. |
| Tooth Paste | 1 gram |
| Chewing Tobacco | 1 gram |
| Cigarettes | 1 cigarette |
| Chewing Gum | 1 piece |
| Hard Candy | 1 piece |
| Teeth Whitening Strips | 2 strips |
| Cola | 12 oz. |
| Water | 6 oz. |
| Antacid | 2 x 500 mg tablets |
| Coffee | 8 oz. |
| Tea | 8 oz. |

f. Assay cut-off:

Characterization of how the device performs analytically around the claimed cutoff concentration of 50 ng/mL methamphetamine is described in the precision section, M.1.a. above.

2. Comparison studies:

a. Method comparison with predicate device:

Neat oral fluid was collected with the RapidEASE Oral Fluid Collector from volunteers. A total of 92 samples were randomized and blinded to the instrument operator and assayed using the RapidFRET Oral Fluid Assay for Methamphetamine and LC/MS/MS reference method. Samples are categorized in the Method Comparison table according to the sum of d and l-methamphetamine concentration as measured by LC/MS/MS. The screening method is calibrated to d-methamphetamine. The results from the method comparison study are summarized in the table below:

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| Candidate Device Results | LC/MS/MS concentration (ng/mL) | | | |
| --- | --- | --- | --- | --- |
| | Less than half the cutoff by LC/MS/MS analysis | Near cutoff (between 50% below the cutoff and the cutoff concentration) | Near cutoff positive (between the cutoff and 50% above the cutoff concentration) | High Positive (greater than 50% above the cutoff concentration) |
| Positive | 8* | 2** | 5 | 39 |
| Negative | 33 | 3 | 0 | 2† |

* Discordant sample resolution

| Sample number | Methamphetamine (ng/mL) | MDMA (3,4-methylenedioxy-methamphetamine) concentration (ng/mL) |
| --- | --- | --- |
| 2439 | 0 | 211 |
| 2158 | 0 | 241 |
| 2496 | 0 | 250 |
| 2491 | 36.8 | 439 |
| 4313 | 40.4 | 1880 |
| 2316 | 0 | 1940 |
| 2442 | 0 | 2020 |
| 2477 | 0 | 4310 |

The screening method cross-reacts with MDMA at approximately 39.7%, resulting in positive screening results even with methamphetamine levels below the cutoff concentration of 50 ng/mL.

** Discordant sample resolution

| Sample number | Methamphetamine (ng/mL) | MDMA (3,4-methylenedioxy-methamphetamine) concentration (ng/mL) | 4-methylethcathinone (4-MEC) concentration (ng/mL) | 3,4-methylenedioxy-N-methylcathinone (Methylone) concentration (ng/mL) |
| --- | --- | --- | --- | --- |
| 2486 | 0 | 10 | 7240 | 47,000 |
| 2830 | 0 | 13.6 | 0 | 8920 |

The screening method cross-reacts with MDMA at approximately 39.7%, with 4-MEC at approximately 1.1%, and with Methylone at approximately 1.5%. These cross-reactivities can result in positive screening results even with methamphetamine levels below the cutoff concentration of 50 ng/mL.

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† Discordant sample resolution

| Sample number | d-methamphetamine (ng/mL) | l-methamphetamine (ng/mL) |
| --- | --- | --- |
| 2393 | 0 | 150 |
| 2382 | 28.4 | 114 |

The screening method is calibrated to d-methamphetamine. The screening method cross-reacts with l-methamphetamine at approximately $17\%$ . These samples contained mostly l-methamphetamine and therefore produced false negative screening results.

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:

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.