First Sign Drug of Abuse MDMA Tests are immunoassay tests. The test can detect MDMA in human urine. The cut-off value is 500 ng/mL.. The tests are available in a Cup format and a Dip Card format.

The tests provide only preliminary test results. If you want to get a confirmed result, you must use a more specific chemical method. Gas Chromatography/Mass Spectrometry is the preferred confirmatory method. Clinical consideration and professional judgment should be used when you get any drug of abuse test result. It should be used particularly when the preliminary result is positive.

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

First Sign Drug of Abuse EDDP Tests are immunoassay tests. The test can detect EDDP in human urine. The cut-off value is 300 ng/mL. The tests are available in a Cup format and a Dip Card format.

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

First Sign Drug of Abuse Nortriptyline Tests are immunoassay tests. The test can detect Nortriptyline in human urine. The cut-off value is 1,000 ng/mL. The tests are available in a Cup format and a Dip Card format.

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

Device Description

First Sign™ Drug of Abuse MDMA Test, First Sign™ Drug of Abuse EDDP Test and First Sign™ Drug of Abuse Nortriptyline Test are immunochromatographic assays. Each assay test is a lateral flow system for the qualitative detection of MDMA, or EDDP or Nortriptyline in human urine. The products are single-use in vitro diagnostic devices, which come in the formats of Dip Cards or Cups. Each test kit contains a Test Device (in one of the two formats), a package insert and a urine cup for sample collection. Each test device is sealed with a desiccant in an aluminum pouch.

AI/ML Overview

The provided document describes the performance characteristics of three drug of abuse tests: First Sign® Drug of Abuse MDMA Test, First Sign® Drug of Abuse EDDP Test, and First Sign® Drug of Abuse Nortriptyline Test. These are qualitative immunoassay tests designed to detect specific drugs in human urine.

Here's an analysis of the acceptance criteria and study details:

1. Table of Acceptance Criteria and Reported Device Performance

The document does not explicitly state formal acceptance criteria thresholds in terms of specific percentages for accuracy, sensitivity, or specificity. However, the performance is reported through various studies, and the implied acceptance criteria are that the device performs reliably at and around the defined cut-off values, and that lay users can operate the device effectively.

Below is a table summarizing the reported device performance, categorized by drug:

First Sign® Drug of Abuse MDMA Test

Performance Metric	Acceptance Criteria (Implied)	Reported Device Performance (Dip Card / Cup Format)
Precision (Agreement at cut-off)	Consistent results at concentrations below and above cut-off; minimal false negatives/positives at cut-off.	Laboratory Operators: At cut-off (500 ng/mL), 47-48 positive / 2-3 negative for all 3 lots (out of 50 tests). At +/-25%, +/-50%, +/-75%, +/-100% cut-off, results were consistently 50-/0+ or 50+/0-.
Cut-off Verification	All positive at and above +25% cut-off; all negative at and below -25% cut-off.	Verified: All positive at and above +25% cut-off (598 ng/mL); all negative at and below -25% cut-off (358 ng/mL).
Comparison Studies (Expert)	High agreement with GC/MS results.	Dip Card: Viewer A: 1 discordant negative (GC/MS 561 ng/mL). Viewer B: 1 discordant positive (GC/MS 468 ng/mL). Viewer C: 1 discordant positive (GC/MS 474 ng/mL) and 1 discordant negative (GC/MS 544 ng/mL). Cup Format: Viewer A: 1 discordant positive (GC/MS 477 ng/mL). Viewer B: 1 discordant negative (GC/MS 517 ng/mL). Viewer C: 1 discordant positive (GC/MS 470 ng/mL).
Lay-User Study (Accuracy)	High percentage of correct results, especially at concentrations away from cut-off.	Dip Card: 100% correct from -100% to -25% cut-off and +50% to +75% cut-off. 95% correct at +25% cut-off (19/20 Positive). Cup Format: 100% correct from -100% to -50% cut-off and +25% to +75% cut-off. 95% correct at -25% cut-off (1 discordant positive).
Lay-User Study (Ease of Use)	Instructions easily followed.	All lay users indicated instructions were easily followed. Flesch-Kincaid Grade Level of 7.
Interference	No interference from common substances at 100 µg/mL.	Extensive list of 90+ compounds showed no interference.
Specificity (Cross-reactivity)	Defined cross-reactivity for related compounds, no detection for unrelated compounds.	MDA (6.3%), MDEA (50%), Ephedrine (1.3%). d-methamphetamine, d-amphetamine, l-amphetamine, l-methamphetamine: Not Detected at 100000 ng/mL.
Urine Specific Gravity/pH Effects	No effect on results across range (pH 4-9, SG 1.000-1.035).	Results were all positive for samples at and above +25% Cut-Off and all negative for samples at and below -25% Cut-Off within the tested ranges, indicating no significant effect.

First Sign® Drug of Abuse EDDP Test

Performance Metric	Acceptance Criteria (Implied)	Reported Device Performance (Dip Card / Cup Format)
Precision (Agreement at cut-off)	Consistent results at concentrations below and above cut-off; minimal false negatives/positives at cut-off.	Laboratory Operators: At cut-off (300 ng/mL), 47-48 positive / 2-3 negative for all 3 lots (out of 50 tests). At +/-25%, +/-50%, +/-75%, +/-100% cut-off, results were consistently 50-/0+ or 50+/0-.
Cut-off Verification	All positive at and above +25% cut-off; all negative at and below -25% cut-off.	Verified: All positive at and above +25% cut-off (410 ng/mL); all negative at and below -25% cut-off (235 ng/mL).
Comparison Studies (Expert)	High agreement with GC/MS results.	Dip Card: Viewer A: 1 discordant negative (GC/MS 340 ng/mL) and 1 discordant positive (GC/MS 276 ng/mL). Viewer B: 1 discordant positive (GC/MS 269 ng/mL). Viewer C: 1 discordant negative (GC/MS 344 ng/mL) and 1 discordant positive (GC/MS 260 ng/mL). Cup Format: Viewer A: 1 discordant negative (GC/MS 344 ng/mL) and 1 discordant positive (GC/MS 269 ng/mL). Viewer B: 1 discordant negative (GC/MS 340 ng/mL). Viewer C: 1 discordant negative (GC/MS 342 ng/mL) and 1 discordant positive (GC/MS 266 ng/mL).
Lay-User Study (Accuracy)	High percentage of correct results, especially at concentrations away from cut-off.	Dip Card: 100% correct from -100% to -50% cut-off and +25% to +75% cut-off. 90% correct at -25% cut-off (2 discordant positives). Cup Format: 100% correct from -100% to -50% cut-off and +25% to +75% cut-off. 95% correct at -25% cut-off (1 discordant positive).
Lay-User Study (Ease of Use)	Instructions easily followed.	All lay users indicated instructions were easily followed. Flesch-Kincaid Grade Level of 7.
Interference	No interference from common substances at 100 µg/mL.	Extensive list of 90+ compounds showed no interference.
Specificity (Cross-reactivity)	Defined cross-reactivity for related compounds, no detection for unrelated compounds.	EDDP (100%). EMDP, Disopyramide, Methadone, LAAM, Alpha Methadol, Doxylamine: Not Detected at 100000 ng/mL.
Urine Specific Gravity/pH Effects	No effect on results across range (pH 4-9, SG 1.000-1.035).	Results were all positive for samples at and above +25% Cut-Off and all negative for samples at and below -25% Cut-Off within the tested ranges, indicating no significant effect.

First Sign® Drug of Abuse Nortriptyline Test

Performance Metric	Acceptance Criteria (Implied)	Reported Device Performance (Dip Card / Cup Format)
Precision (Agreement at cut-off)	Consistent results at concentrations below and above cut-off; minimal false negatives/positives at cut-off.	Laboratory Operators: At cut-off (1000 ng/mL), 47-48 positive / 2-3 negative for all 3 lots (out of 50 tests). At +/-25%, +/-50%, +/-75%, +/-100% cut-off, results were consistently 50-/0+ or 50+/0-.
Cut-off Verification	All positive at and above +25% cut-off; all negative at and below -25% cut-off.	Verified: All positive at and above +25% cut-off (1180 ng/mL); all negative at and below -25% cut-off (720 ng/mL).
Comparison Studies (Expert)	High agreement with GC/MS results.	Dip Card: Viewer A: 1 discordant positive (GC/MS 863 ng/mL). Viewer B: 1 discordant negative (GC/MS 1069 ng/mL) and 1 discordant positive (GC/MS 851 ng/mL). Viewer C: 1 discordant negative (GC/MS 1125 ng/mL) and 1 discordant positive (GC/MS 879 ng/mL). Cup Format: Viewer A: 1 discordant positive (GC/MS 851 ng/mL) and 1 discordant negative (GC/MS 1084 ng/mL). Viewer B: 1 discordant positive (GC/MS 870 ng/mL). Viewer C: 1 discordant negative (GC/MS 1135 ng/mL).
Lay-User Study (Accuracy)	High percentage of correct results, especially at concentrations away from cut-off.	Dip Card: 100% correct from -100% to -50% cut-off and +25% to +75% cut-off. 95% correct at -25% cut-off (1 discordant positive). Cup Format: 100% correct from -100% to -50% cut-off, and +50% to +75% cut-off. 95% correct at -25% cut-off and +25% cut-off.
Lay-User Study (Ease of Use)	Instructions easily followed.	All lay users indicated instructions were easily followed. Flesch-Kincaid Grade Level of 7.
Interference	No interference from common substances at 100 µg/mL.	Extensive list of 90+ compounds showed no interference.
Specificity (Cross-reactivity)	Defined cross-reactivity for related compounds, no detection for unrelated compounds.	Nortriptyline (100%). Other TCAs and related compounds show varying cross-reactivity (e.g., Amitriptyline 67%, Desipramine 100%, Imipramine 167%). Maprotiline, Promethazine, Norclomipramine: Not Detected at 100000 ng/mL.
Urine Specific Gravity/pH Effects	No effect on results across range (pH 4-9, SG 1.000-1.035).	Results were all positive for samples at and above +25% Cut-Off and all negative for samples at and below -25% Cut-Off within the tested ranges, indicating no significant effect.

2. Sample Size Used for the Test Set and Data Provenance

Precision Studies:
- For each drug (MDMA, EDDP, Nortriptyline) and each format (Cup, Dip Card): 9 concentration levels (from -100% cut-off to +100% cut-off).
- At each concentration level: 50 tests (2 runs/day for 25 days).
- Total for precision per drug/format: 9 concentrations * 50 tests/concentration = 450 tests.
- Total for precision across all 3 drugs and 2 formats: 3 * 2 * 450 = 2700 tests.
- Data Provenance: "Prepared by spiking drug in negative urine samples." This suggests laboratory-controlled samples, likely from a US lab given the FDA submission. Retrospective, as samples were prepared and then tested.
Cut-off Verification Studies:
- For each drug and format: 4 concentration levels ( -50% cut-off, cut-off, +25% cut-off, +50% cut-off).
- Total of 150 samples "equally distributed" meaning approximately 37-38 samples per concentration level. These were tested using three different lots.
- Data Provenance: Laboratory-controlled samples, likely from a US lab. Retrospective.
Interference and Specificity Studies:
- The document implies that these were conducted with spiked samples in drug-free urine, testing against three lots of each device for all formats. No specific sample numbers are given for individual interference compounds or cross-reactants, but the method suggests a systematic laboratory study.
- Data Provenance: Laboratory-controlled samples, likely from a US lab. Retrospective.
Effect of Urine Specific Gravity and Urine pH:
- Urine samples with varying specific gravity and pH were spiked with target drugs at 25% below and 25% above cut-off levels. Tested using three lots of each device for all formats. No specific sample number provided, but implies a comprehensive set to cover the specified ranges.
- Data Provenance: Laboratory-controlled samples, likely from a US lab. Retrospective.
Comparison Studies (Clinical Samples):
- For each drug and each format: 80 unaltered clinical samples (40 negative, 40 positive).
- Total for comparison studies across all 3 drugs and 2 formats: 3 * 2 * 80 = 480 clinical samples.
- Data Provenance: "Unaltered clinical samples" implies real human urine samples from an unspecified source (likely US-based for FDA submission). The study is retrospective, as samples were collected and then tested.
Lay-User Study:
- For each drug: 280 lay persons.
- Each participant tested one blind-labeled urine sample. There were 7 concentration levels tested (-100%, -75%, -50%, -25%, +25%, +50%, +75% of cut-off).
- For each concentration and format (Dip Card / Cup): 20 samples.
- Total samples given to lay users per drug: 7 concentrations * 20 samples/concentration * 2 formats = 280 samples. This matches the 280 lay persons.
- Data Provenance: Laboratory-prepared urine samples (spiked drug into drug-free pooled urine). This is a prospective simulation of real-world use with prepared samples. The study was performed at "three intended user sites". Given the FDA submission, these are likely US sites.

3. Number of Experts and Qualifications for Ground Truth

Precision, Cut-off, Interference, Specificity, Urine SG/pH Studies:
- The ground truth (drug concentration) for all these studies was established by GC/MS (Gas Chromatography/Mass Spectrometry). This is a highly accurate and widely accepted reference method for drug quantification in toxicology. The document does not specify a separate "expert" for interpreting GC/MS results, as the instrument provides quantitative values which are then used to prepare samples.
Comparison Studies (Clinical Samples):
- The ground truth for the 80 clinical samples per drug/format was established by GC/MS results.
- The device results were interpreted by three different laboratory assistants. Their qualifications are not explicitly stated beyond "laboratory assistants," but it can be inferred they are trained professionals in a laboratory setting.
Lay-User Study:
- The ground truth for the prepared urine samples was established by GC/MS to confirm the spiked drug concentrations.
- No "experts" were used to establish ground truth for how the lay users interpreted the test, as the study's purpose was to evaluate if lay users could correctly interpret the device results against the known (GC/MS confirmed) concentration.

4. Adjudication Method for the Test Set

Precision Studies, Cut-off Verification, Interference, Specificity, Urine SG/pH Studies: The ground truth used was quantitative GC/MS results, which serves as a definitive reference standard. There's no indication of an adjudication process among multiple readers for these lab-controlled tests; the output of the device (positive/negative line) was compared directly to the GC/MS confirmed concentration.
Comparison Studies (Clinical Samples): The document mentions "three different laboratory assistants" running the tests and comparing them to GC/MS results. It does not describe an adjudication method (like 2+1 or 3+1) among these three assistants where they would resolve discrepancies in their readings. Instead, individual discrepancies are noted in the "Discordant Results" tables for each viewer. This suggests independent assessment rather than a consensus-seeking or adjudicated process for the readers.
Lay-User Study: Each lay user interpreted their own test result individually against the known GC/MS confirmed concentration. No adjudication among lay users was described.

5. Multi-Reader Multi-Case (MRMC) Comparative Effectiveness Study

No, a typical MRMC comparative effectiveness study was not explicitly conducted as described for AI vs. human readers.
The comparison studies did involve multiple readers (three laboratory assistants) reviewing clinical cases, which provides some multi-reader data. However, this was a standalone performance evaluation of the device being read by humans, not a comparison of human performance with vs. without AI assistance. The device itself is an immunoassay, not an AI algorithm providing assistance in interpreting complex images or data.
Therefore, an "effect size of how much human readers improve with AI vs without AI assistance" is not applicable here.

6. Standalone (Algorithm Only) Performance

Yes, in essence, the "Analytical Performance" section (Precision, Cut-off, Interference, Specificity) represents the standalone performance of the devices. These studies evaluate the intrinsic performance of the immunoassay devices under controlled laboratory conditions, without human interpretation variability, by comparing the device's qualitative output (presence/absence of lines) against known, quantitatively confirmed drug concentrations.
The "Comparison Studies" with laboratory assistants and "Lay-user studies" then evaluate the human-in-the-loop performance.

7. Type of Ground Truth Used

All studies (Analytical, Comparison, Lay-User) consistently used Gas Chromatography/Mass Spectrometry (GC/MS) results as the gold standard ground truth.
GC/MS is a laboratory-based, highly accurate, and quantitative method for identifying and measuring specific substances in a sample, making it a robust ground truth for drug testing.

8. Sample Size for the Training Set

Not applicable. These are immunoassay devices (lateral flow tests), not machine learning or AI algorithms that require a "training set." The performance characteristics are derived from intrinsic chemical and physical properties, evaluated through these detailed analytical and clinical studies.

9. How the Ground Truth for the Training Set Was Established

Not applicable, as no training set was used.

Summary

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Image /page/0/Picture/1 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo consists of a circular seal with the text "DEPARTMENT OF HEALTH & HUMAN SERVICES - USA" around the perimeter. Inside the circle is a stylized image of three human profiles facing to the right, with flowing lines representing hair or movement.

Auqust 17,2016

Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-0002

W.H.P.M., INC. C/O JOE SHIA LSI CONSULTING 504 EAST DIAMOND AVE., SUITE I GAITHERSBURG MD 20877

Re: K160793

Trade/Device Name: First Sign Drug of Abuse MDMA Cup Test: First Sign Drug of Abuse MDMA Dip Card Test; First Sign Drug of Abuse EDDP Cup Test; First Sign Drug of Abuse EDDP Dip Card Test; First Sign Drug of Abuse Nortriptyline Cup Test; First Sign Drug of Abuse Nortriptyline Dip Card Test Regulation Number: 21 CFR 862.3620 Regulation Name: Methadone test system Regulatory Class: II Product Code: DJR, LAF, LFG Dated: June 29, 2016 Received: July 7, 2016

Dear Joe Shia:

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. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading.

If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. 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); medical device reporting of medical device-related adverse events) (21 CFR 803); good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820); and if applicable, the

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electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.

If you desire specific advice for your device on our labeling regulations (21 CFR Parts 801 and 809), please contact the Division of Industry and Consumer Education at its toll-free number (800) 638 2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/Resourcesfor You/Industry/default.htm. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to

http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.

You may obtain other general information on your responsibilities under the Act from the Division of Industry and Consumer Education at its toll-free number (800) 638-2041 or (301) 796-7100 or at its Internet address

http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.

Sincerely yours,

Katherine Serrano -S

For: Courtney H. Lias, Ph.D.

Director Division of Chemistry and Toxicology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health

Enclosure

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Indications for Use

510(k) Number (if known) K160793

Device Name

First Sign Drug of Abuse MDMA Cup Test; First Sign Drug of Abuse MDMA Dip Card Test First Sign Drug of Abuse EDDP Cup Test; First Sign Drug of Abuse EDDP Dip Card Test First Sign Drug of Abuse Nortriptyline Cup Test; First Sign Drug of Abuse Nortriptyline Dip Card Test

Indications for Use (Describe)

First Sign Drug of Abuse MDMA Cup Test

First Sign Drug of Abuse MDMA Dip Card Test

First Sign Drug of Abuse MDMA Tests are immunoassay tests. The test can detect MDMA in human urine. The cut-off value is 500 ng/mL.. The tests are available in a Cup format and a Dip Card format.

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

First Sign Drug of Abuse EDDP Cup Test First Sign Drug of Abuse EDDP Dip Card Test First Sign Drug of Abuse EDDP Tests are immunoassay tests. The test can detect EDDP in human urine. The cut-off value is 300 ng/mL. The tests are available in a Cup format and a Dip Card format.

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

First Sign Drug of Abuse Nortriptyline Cup Test

First Sign Drug of Abuse Nortriptyline Dip Card Test

For in vitro diagnostic use only. The tests are intended for over-the-counter and for prescription use.

Type of Use (Select one or both, as applicable)

X Prescription Use (Part 21 CFR 801 Subpart D)

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

CONTINUE ON A SEPARATE PAGE IF NEEDED.

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1. Date: March 15, 2016
1. Submitter W.H.P.M., Inc. 5358 Irwindale Ave. Irwindale, CA 91706
1. Contact person: John Wan W.H.P.M., Inc. 5358 Irwindale Ave. Irwindale, CA 91706 Telephone: 626-443-8480 Fax: 626-443-8065 Email: johnwan@whpm.com
1. Device Name: First Sign® Drug of Abuse MDMA Cup Test First Sign® Drug of Abuse MDMA Dip Card Test First Sign® Drug of Abuse EDDP Cup Test First Sign® Drug of Abuse EDDP Dip Card Test First Sign® Drug of Abuse Nortriptyline Cup Test First Sign® Drug of Abuse Nortriptyline Dip Card Test
MDMA Urine Test Common Name: EDDP Urine Test Nortriptyline Urine Test

Product Code	Class	CFR #	Panel
LAF	Class II	21 CFR, 862.3610 Methamphetamine Test System	Toxicology
DJR	Class II	21 CFR, 862.3620 Methadone Test System	Toxicology
LFG	Class II	21 CFR, 862.3910 Tricyclic Antidepressant DrugsTest System	Toxicology

1. Predicate Devices:
  K142800, Co-Innovation Rapid Single/Multi Drug Test K140748, Co-Innovation One Step Single/Multi Drug Test
1. Intended Use First Sign® Drug of Abuse MDMA Cup Test First Sign® Drug of Abuse MDMA Dip Card Test

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First Sign® Drug of Abuse MDMA Tests are immunoassay tests. The test can detect MDMA in human urine. The cut-off value is 500 ng/mL. The tests are available in a Cup format and a Dip Card format.

First Sign® Drug of Abuse EDDP Cup Test First Sign® Drug of Abuse EDDP Dip Card Test First Sign® Drug of Abuse EDDP Tests are immunoassay tests. The test can detect EDDP in human urine. The cut-off value is 300 ng/mL. The tests are available in a Cup format and a Dip Card format.

First Sign® Drug of Abuse Nortriptyline Cup Test

First Sign® Drug of Abuse Nortriptyline Dip Card Test

First Sign® Drug of Abuse Nortriptyline Tests are immunoassay tests. The test can detect Nortriptyline in human urine. The cut-off value is 1,000 ng/mL. The tests are available in a Cup format and a Dip Card format.

7. Device Description

8. Substantial Equivalence Information

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A summary comparison of features of the candidate device and the predicate device is provided in Table 1, Table 2 & Table 3.

Table 1: Features Comparison of First Sign® Drug of Abuse MDMA Test and the Predicate
Device

Item	Candidate DeviceFirst Sign® Drug of Abuse MDMA Test	Predicate Device(K142800)Co-InnovationMulti Drug Test
Indication(s)for Use	For the qualitative determination ofMDMA in human urine.	Same
Calibrator	MDMA	Same
Methodology	Competitive binding, lateral flowimmunochromatographic assays based onthe principle of antigen antibodyimmunochemistry.	Same
Specimen Type	Human urine	Same
Cut-Off Values	500 ng/mL	Same
IntendedPopulation	For over-the-counter and prescriptionuses.	Same
Configurations	Cup, Dip Card	Same

Table 2: Features Comparison of First Sign® Drug of Abuse EDDP Test and the Predicate
Device
Item	Candidate DeviceFirst Sign® Drug of Abuse EDDP Test	Predicate Device(K140748)Co-InnovationMulti Drug Test
Indication(s)for Use	For the qualitative determination ofEDDP in human urine.	Same
Calibrator	EDDP	Same
Methodology	Competitive binding, lateral flowimmunochromatographic assays based onthe principle of antigen antibodyimmunochemistry.	Same
Specimen Type	Human urine	Same
Cut-Off Values	300 ng/mL	Same
IntendedPopulation	For over-the-counter and prescriptionuses.	Same
Configurations	Cup, Dip Card	Same

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Table 3: Features Comparison of First Sign® Drug of Abuse Nortriptyline Test and the Predicate Device

Item	Candidate DeviceFirst Sign® Drug of Abuse NortriptylineTest	Predicate Device(K140748)Co-InnovationMulti Drug Test
Indication(s)for Use	For the qualitative determination ofNortriptyline in human urine.	Same
Calibrator	Nortriptyline	Same
Methodology	Competitive binding, lateral flowimmunochromatographic assays based onthe principle of antigen antibodyimmunochemistry.	Same
Specimen Type	Human Urine	Same
Cut-Off Values	1000 ng/mL	Same
IntendedPopulation	For over-the-counter and prescriptionuses.	Same
Configurations	Cup, Dip Card	Same

9. Test Principle

Each assay test is a lateral flow chromatographic immunoassay. During testing, a urine specimen migrates upward by capillary action. If target drugs are present in the urine specimen below its cut-off concentration, it will not saturate the binding sites of its specific antibody (monoclonal mouse antibody) coated on the particles. The antibody-coated particles will then be captured by immobilized drugconjugate and a visible colored line will show up in the test line region. The will not form in the test line region if the target drug level exceeds its cut-off concentration because it will saturate all the binding sites of the antibody coated on the particles. A band should form in the control region of the devices regardless of the presence of drug or metabolite in the sample.

10. Performance Characteristics

First Sign® Drug of Abuse MDMA Test

Analytical Performance

a. Precision

Precision studies were carried out for samples with concentrations of -100% cut-off, -50% cut-off, -25% cut-off, at the cut-off, +25% cut-off, +75% cut-off, +75% cut-off and +100% cut-off. These samples were prepared by spiking drug in negative urine samples. Each drug concentration was confirmed by GC/MS. All sample aliquots were blind-labeled and randomized

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by the person who prepared samples and did not take part in the sample testing. For each concentration, tests were performed two runs per day for 25 days by three different operators for each format of devices. Different set of operators tested each format. The results obtained are summarized in the following tables:

	Result	-100%Cut-off	-75%Cut-off	-50%Cut-off	-25%Cut-off	Cut-off	+25%Cut-off	+50%Cut-off	+75%Cut-off	+100%Cut-off
Drug	Lot 1	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-
Drug	Lot 2	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-
Drug	Lot 3	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-

50-/0+

3-/47+

2-/48+

50+/0-

MDMA Dip Card Format

b. Linearity

Not applicable.

Lot 4

Lot 5

Lot 6

50-/0+

c. Stability

The devices are stable at 39-86ºF (4-30ºC) for 24 months based on the accelerated stability study at 50°C. Control materials are not provided with the device. The labeling provides information on how to obtain control materials.

d. Cut-off

A total of 150 samples equally distributed at concentrations of -50% cut-off; cutoff; +25% cut-off; +50% cut-off were tested using three different lots of each device by three different operators. Results were all positive at and above +25% cut-off and all negative at and below -25% cut-off for MDMA. The following cut-off value for the test devices have been verified.

Test	Calibrator	Cut-off (ng/mL)
First Sign® Drug of Abuse MDMA Test	MDMA	500

e. Interference

Potential interfering substances found in human urine of physiological conditions were added to drug-free urine and to urine containing target drugs at 25% below and 25% above cut-off levels. These urine samples were tested using three lots of each device for all formats.

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Compounds that showed no interference at a concentration of 100µg/mL are summarized in the following tables. There were no differences observed for different formats.

4-Acetamidophenol	(L) - Epinephrine	Pentobarbital
Acetophenetidin	Erythromycin	Perphenazine
N-Acetylprocainamide	β-Estradiol	Phencyclidine
Acetylsalicylic acid	Estrone-3-sulfate	Phenelzine
Aminopyrine	Ethyl-p-aminobenzoate	Phenobarbital
Amitryptyline	Fenoprofen	Phentermine
Amobarbital	Furosemide	Trans-2-phenylcyclopropylamine hydrochloride
Amoxicillin	Gentisic acid	L-Phenylephrine
Ampicillin	Hemoglobin	β-Phenylethylamine
L-Ascorbic acid	Hydralazine	Phenylpropanolamine
Apomorphine	Hydrochlorothiazide	Prednisolone
Aspartame	Hydrocodone	Prednisone
Atropine	Hydrocortisone	Procaine
Benzilic acid	O-Hydroxyhippuric acid	Promazine
Benzoic acid	3-Hydroxytyramine	Promethazine
Benzoylecgonine	Ibuprofen	DL-Propranolol
Bilirubin	Imipramine	D-Propoxyphene
(±) - Brompheniramine	Iproniazid	D-Pseudoephedrine
Buspiron	(±) - Isoproterenol	Quinacrine
Caffeine	Isoxsuprine	Quinidine
Cannabidiol	Ketamine	Quinine
Cannabinol	Ketoprofen	Ranitidine
Chloralhydrate	Labetalol	Salicylic acid
Chloramphenicol	Levorphanol	Secobarbital
Chlordiazepoxide	Loperamide	Serotonin (5- Hydroxytyramine)
Chlorothiazide	Maprotiline	Sulfamethazine
(±) - Chlorpheniramine	Meperidine	Sulindac
Chlorpromazine	Meprobamate	Sustiva
Chloroquine	Methadone	Temazepam
Cholesterol	Morphine-3-β-Dglucuronide	Tetracycline
Clomipramine	Morphine sulfate	Tetrahydrocortisone 3-(β-Dglucuronide)
Clonidine	Nalidixic acid	Tetrahydrozoline
Cocaethylene	Naloxone	Thebaine
Cocaine hydrochloride	Naltrexone	Theophynine
Codeine	Naproxen	Thiamine
Cortisone	Niacinamide	Thioridazine
(-) Cotinine	Nifedipine	Tolbutamide
Creatinine	Nimesulidate	Trazodone
Deoxycorticosterone	Norcodein	Triamterene
Dextromethorphan	Norethindrone	DL-Tyrosine
Diclofenac	D-Norpropoxyphene	Trifluoperazine
Diazepam	Noscapine	Trimethoprim
Diflunisal	D,L-Octopamine	Trimipramine
Digoxin	Oxalic acid	Tryptamine
Dicylomine	Oxazepam	D L-Tryptophan
Diphenhydramine	Oxolinic acid	Tyramine
5,5 - Diphenylhydantoin	Oxycodone	Uric acid
Doxylamine	Oxymetazoline	Verapamil
Ecgonine hydrochloride	Papaverine	Zomepirac
Ecgonine methylester	Penicillin-G
1R,2S Ephedrine	Pentazocinehydrochloride

{10}------------------------------------------------

f. Specificity

To test the specificity, drug metabolites and other components that are likely to interfere in urine samples were tested using three lots of each device for all formats. The obtained lowest detectable concentration was used to calculate the cross-reactivity. There were no differences observed for different formats.

Drug	Concentration (ng/ml)	% Cross-Reactivity
Methylenedioxymethamphetamine (MDMA)	500	100%
3,4-Methylenedioxyamphetamine (MDA)	8000	6.3%
3,4-Methylenedioxyethylamphetamine (MDEA)	1000	50%
Ephedrine	40000	1.3%
d-methamphetamine	Negative at 100000	Not Detected
d-amphetamine	Negative at 100000	Not Detected
l-amphetamine	Negative at 100000	Not Detected
l-methamphetamine	Negative at 100000	Not Detected

g. Effect of Urine Specific Gravity and Urine pH

To investigate the effect of urine specific gravity and urine pH, urine samples with a range of 1.000 to 1.035 specific gravity or urine samples with a range of pH 4 to 9 were spiked with target drugs at 25% below and 25% above cut-off levels. These samples were tested using three lots of each

{11}------------------------------------------------

device for all formats. Results were all positive for samples at and above +25% Cut-Off and all negative for samples at and below -25% Cut-Off. There were no differences observed for different formats.

h. Comparison Studies
The method comparison studies for the First Sign® Drug of Abuse MDMA Test was performed in-house with three different laboratory assistants for each format of the device. Operators ran 80 (40 negative and 40 positive) unaltered clinical samples were blind labeled and compared to GC/MS results. The results are presented in the tables below:

Dip Cardformat		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% andcut-off)	NearCutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan +50%)
Viewer A	Positive	0	0	0	13	26
	Negative	10	10	20	1	0
Viewer B	Positive	0	0	1	14	26
	Negative	10	10	19	0	0
Viewer C	Positive	0	0	1	13	26
	Negative	10	10	19	1	0

Discordant Results

Viewer	Sample Number	GC/MS (ng/mL) Result	Dipcard Format Viewer Results
Viewer A	2014111947	561	Negative
Viewer B	2014102405	468	Positive
Viewer C	2014111901	474	Positive
Viewer C	2014102302	544	Negative

Cupformat		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% andcut-off)	NearCutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan +50%)
Viewer A	Positive	0	0	1	14	26
	Negative	10	10	19	0	0
Viewer B	Positive	0	0	0	13	26
	Negative	10	10	20	1	0

{12}------------------------------------------------

Viewerし	Positive			C60
	Negative	1 Cﻟﺘﻌﻠﻴﻘﺎﺕ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘﺤﺪﺓ ﺍﻟﻤﺘ	1 C

Discordant Results

Viewer	Sample Number	GC/MS (ng/mL)Result	Cup FormatViewer Results
Viewer A	2014102306	477	Positive
Viewer B	2014111950	517	Negative
Viewer C	2014102315	470	Positive

i. Lay-user study

A lay user study was performed at three intended user sites with 280 lay persons testing the MDMA devices. They had diverse educational and professional backgrounds and ranged in age from 21 to > 50 years. Urine samples were prepared at the following concentrations; negative, +/-75%, +/-50%, +/-25% of the cutoff by spiking drugs into drug free-pooled urine specimens. The concentrations of the samples were confirmed by GC/MS. Each sample was aliquoted into individual containers and blind-labeled. Each participant was provided with the package insert, 1 blind labeled sample and a device. The results are summarized below.

% of Cutoff	Number ofsamples	MDMA Concentration byGC/MS(ng/mL)	Lay person resultsNo. ofPositive	Lay person resultsNo. ofNegative	Thepercentage ofcorrect results(%)
-100% Cutoff	20	0	0	20	100%
-75% Cutoff	20	115	0	20	100%
-50% Cutoff	20	237	0	20	100%
-25% Cutoff	20	358	0	20	100%
+25% Cutoff	20	598	19	1	95%
+50% Cutoff	20	755	20	0	100%
+75% Cutoff	20	912	20	0	100%

Comparison between GC/MS and Lay Person Results (MDMA DipCard)

Comparison between GC/MS and Lay Person Results (MDMA Cup)

	Number	MDMA Concentration by	Lay person results		The
% of Cutoff	ofsamples	GC/MS(ng/mL)	No. ofPositive	No. ofNegative	percentage ofcorrect results(%)
-100% Cutoff	20	0	0	20	100%
-75% Cutoff	20	115	0	20	100%
-50% Cutoff	20	237	0	20	100%
-25% Cutoff	20	358	1	19	95%
+25% Cutoff	20	598	20	0	100%
+50% Cutoff	20	755	20	0	100%
+75% Cutoff	20	912	20	0	100%

{13}------------------------------------------------

Lay-users were also given surveys on the ease of understanding the package insert instructions. All lay users indicated that the device instructions can be easily followed. A Flesch-Kincaid reading analysis was performed on each package insert and the scores revealed a reading Grade Level of 7.

j. Clinical Studies
Not applicable.

First Sign® Drug of Abuse EDDP Test

Analytical Performance

a. Precision
Precision studies were carried out for samples with concentrations of -100% cut-off, -50% cut-off, -25% cut-off, at the cut-off, +25% cut-off, +75% cut-off, +75% cut-off and +100% cut-off. These samples were prepared by spiking drug in negative urine samples. Each drug concentration was confirmed by GC/MS. All sample aliquots were blind-labeled and randomized by the person who prepared samples and did not take part in the sample testing. For each concentration, tests were performed two runs per day for 25 days by three different operators for each format of devices. Different set of operators tested each format. The results obtained are summarized in the following tables:

Result	-100%Cut-off	-75%Cut-off	-50%Cut-off	-25%Cut-off	Cut-off	+25%Cut-off	+50%Cut-off	+75%Cut-off	+100%Cut-off
Drug
Lot 1	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 2	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 3	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-

EDDP Dip Card Format

b. Linearity
Not applicable.
c. Stability

{14}------------------------------------------------

d. Cut-off
A total of 150 samples equally distributed at concentrations of -50% cut-off; cutoff; +25% cut-off; +50% cut-off were tested using three different lots of each device by three different operators. Results were all positive at and above +25% cut-off and all negative at and below -25% cut-off for EDDP. The following cut-off value for the test devices have been verified.

Test	Calibrator	Cut-off (ng/mL)
First Sign® Drug of Abuse EDDPTest	EDDP	300

e. Interference
Potential interfering substances found in human urine of physiological or pathological conditions were added to drug-free urine and to urine containing target drugs at 25% below and 25% above cut-off levels. These urine samples were tested using three lots of each device for all formats.

Compounds that showed no interference at a concentration of 100µg/mL are summarized in the following tables. There were no differences observed for different formats.

Acetaminophen	Ecgonine hydrochloride	O-Hydroxyhippuric acid
Acetophenetidin	Ecgonine methylester	Oxalic acid
Acetylsalicylic acid	(IR,2S)(-)Ephedrine	Oxazepam
Amobarbital	Erythromycin	Oxolinic acid
Aminopyrine	β-Estradiol	Oxycodone
Amitryptyline	Estrone-3-sulfate	Oxymetazoline
Amoxicillin	Ethyl-p-aminobenzoate	Papaverine
DL-Amphetamine sulfate	Fenoprofen	Penicillin-G
Ampicillin	Furosemide	Pentazocine
Apomorphine	Gentisic acid	Pentobarbital
Ascorbic acid	Hemoglobin	Perphenazine
Aspartame	Hydralazine	Phencyclidine
Atropine	Hydrochlorothiazide	Phenelzine
Benzilic acid	Hydrocodone	Phenobarbital
Benzoic acid	Hydrocortisone	Phentermine
Benzoylecgonine	p-Hydroxyamphetamine	β-Phenylethylamine
Bilirubin	p-Hydroxymethamphetamine	Phenylpropanolamine
Brompheniramine	3-Hydroxytyramine	Prednisolone

{15}------------------------------------------------

Caffeine	Ibuprofen	Prednisone
Cannabidiol	Imipramine	Procaine
Cannabinol	(-) Isoproterenol	Promazine
Chloralhydrate	Isoxsuprine	Promethazine
Chloramphenicol	Ketamine	Quinidine
Chlorothiazide	Ketoprofen	Quinine
(±) - Chlorpheniramine	Labetalol	Ranitidine
Chlorpromazine	Levorphanol	Salicylic acid
Chloroquine	Loperamide	Secobarbital
Cholesterol	L-Phenylephrine	Serotonin
Clomipramine	Maprotiline	Sulfamethazine
Clonidine	Meperidine	Sulindac
Cocaine hydrochloride	Meprobamate	Temazepam
Codeine	Methamphetamine	Tetracycline
(-) Cotinine	Methoxyphenamine	Tetrahydrocortisone 3- (β-D-glucuronide)
Cortisone	(±) - 3,4-Methylenedioxy-amphetamine hydrochloride	Tetrahydrozoline
Creatinine	(±)-3,4-Methylenedioxy-methamphetamine hydrochloride	Thebaine
Deoxycorticosterone	Morphine Sulfate	Thiamine
Dextromethorphan	Morphine-3-β-D glucuronide	Thioridazine
Diazepam	N-Acetylprocainamide	Triamterene
Diclofenac	Nalidixic acid	Trifluoperazine
Diflunisal	Naloxone	Trimethoprim
Digoxin	Naltrexone	Trimipramine
Diphenhydramine	Naproxen	Tryptamine
D-Norpropoxyphene	Niacinamide	DL-Tryptophan
D-Propoxyphene	Nifedipine	Tyramine
D,L-Tyrosine	Norcodein	Uric acid
DL-Octopamine	Norethindrone	Verapamil
DL-Propranolol	Noscapine	Zomepirac

f. Specificity

Drug	Concentration (ng/ml)	% Cross-Reactivity
------	-----------------------	--------------------

{16}------------------------------------------------

EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine)	300	100%
EMDP (2-Ethyl-5-methyl-3,3-diphenylpyrroline)	Negative at 100000	Not Detected
Disopyramide	Negative at 100000	Not Detected
Methadone	Negative at 100000	Not Detected
LAAM (Levo-alpha-acetylmethadol)	Negative at 100000	Not Detected
Alpha Methadol	Negative at 100000	Not Detected
Doxylamine	Negative at 100000	Not Detected

g. Effect of Urine Specific Gravity and Urine pH
To investigate the effect of urine specific gravity and urine pH, urine samples with a range of 1.000 to 1.035 specific gravity or urine samples with a range of pH 4 to 9 were spiked with target drugs at 25% below and 25% above cut-off levels. These samples were tested using three lots of each device for all formats. Results were all positive for samples at and above +25% Cut-Off and all negative for samples at and below -25% Cut-Off. There were no differences observed for different formats.
h. Comparison Studies
The method comparison studies for the First Sign® Drug of Abuse EDDP Test was performed in-house with three different laboratory assistants for each format of the device. Operators ran 80 (40 negative and 40 positive) unaltered clinical samples were blind labeled and compared to GC/MS results. The results are presented in the tables below:

Dip Cardformat		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% andcut-off)	NearCutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan +50%)
Viewer A	Positive	0	0	1	13	26
	Negative	10	10	19	1	0
Viewer B	Positive	0	0	1	14	26
	Negative	10	10	19	0	0
Viewer C	Positive	0	0	1	13	26
	Negative	10	10	19	1	0

{17}------------------------------------------------

Viewer	Sample Number	GC/MS (ng/mL)Result	DipCard FormatViewer Results
Viewer A	94911951	340	Negative
Viewer A	94910121	276	Positive
Viewer B	94911312	269	Positive
Viewer C	94911296	344	Negative
Viewer C	94911562	260	Positive

Cupformat		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% andcut-off)	NearCutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan +50%)
Viewer A	Positive	0	0	1	13	26
	Negative	10	10	19	1	0
Viewer B	Positive	0	0	0	13	26
	Negative	10	10	20	1	0
Viewer C	Positive	0	0	1	13	26
	Negative	10	10	19	1	0

Discordant Results

Viewer	Sample Number	GC/MS (ng/mL) Result	Cup Format Viewer Results
Viewer A	94911296	344	Negative
Viewer A	94910742	269	Positive
Viewer B	94911951	340	Negative
Viewer C	94911928	342	Negative
Viewer C	94910755	266	Positive

i. Lay-user study
A lay user study was performed at three intended user sites with 280 lay persons testing the EDDP devices. They had diverse educational and professional backgrounds and ranged in age from 21 to > 50 years. Urine samples were prepared at the following concentrations; negative, +/-75%, +/-50%, +/-25% of the cutoff by spiking drugs into drug free-pooled urine specimens. The concentrations of the samples were confirmed by GC/MS. Each sample was aliquoted into individual containers and blind-labeled. Each participant was provided with the package insert, 1 blind labeled sample and a device. The results are summarized below.

Comparison between GC/MS and Lay Person Results (EDDP DipCard)

{18}------------------------------------------------

% of Cutoff	Number of samples	EDDP Concentration by GC/MS (ng/mL)	Lay person results		The percentage of correct results (%)
-100% Cutoff	20	0	No. of Positive	No. of Negative	100%
-75% Cutoff	20	81	0	20	100%
-50% Cutoff	20	157	0	20	100%
-25% Cutoff	20	235	2	18	90%
+25% Cutoff	20	410	20	0	100%
+50% Cutoff	20	485	20	0	100%
+75% Cutoff	20	566	20	0	100%

Comparison between GC/MS and Lay Person Results (EDDP Cup)

% of Cutoff	Number of samples	EDDP Concentration by GC/MS (ng/mL)	Lay person results		The percentage of correct results (%)
			No. of Positive	No. of Negative
-100% Cutoff	20	0	0	20	100%
-75% Cutoff	20	81	0	20	100%
-50% Cutoff	20	157	0	20	100%
-25% Cutoff	20	235	1	19	95%
+25% Cutoff	20	410	20	0	100%
+50% Cutoff	20	485	20	0	100%
+75% Cutoff	20	566	20	0	100%

j. Clinical Studies
Not applicable.

First Sign® Drug of Abuse Nortriptyline Test

Analytical Performance

a. Precision
Precision studies were carried out for samples with concentrations of -100% cut-off, -50% cut-off, -25% cut-off, at the cut-off, +25% cut-off, +75% cut-off, +75% cut-off and +100% cut-off. These samples were prepared by spiking drug in negative urine samples. Each drug concentration was confirmed by GC/MS. All sample aliquots were blind-labeled and randomized by the person who prepared samples and did not take part in the sample testing. For each concentration, tests were performed two runs per day for 25 days by three different operators for

{19}------------------------------------------------

each format of devices. Different set of operators tested each format. The results obtained are summarized in the following tables:

	-100%Cut-off	-75%Cut-off	-50%Cut-off	-25%Cut-off	Cut-off	+25%Cut-off	+50%Cut-off	+75%Cut-off	+100%Cut-off
Drug
Lot 1	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 2	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 3	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-

Nortriptyline Dip Card Format

Nortriptyline Cup Format

	-100%Cut-off	-75%Cut-off	-50%Cut-off	-25%Cut-off	Cut-off	+25%Cut-off	+50%Cut-off	+75%Cut-off	+100%Cut-off
Drug
Lot 4	50-/0+	50-/0+	50-/0+	50-/0+	2-/48+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 5	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-
Lot 6	50-/0+	50-/0+	50-/0+	50-/0+	3-/47+	50+/0-	50+/0-	50+/0-	50+/0-

b. Linearity

Not applicable.

c. Stability

d. Cut-off

A total of 150 samples equally distributed at concentrations of -50% cut-off; cutoff; +25% cut-off; +50% cut-off were tested using three different lots of each device by three different operators. Results were all positive at and above +25% cut-off and all negative at and below -25% cut-off for Nortriptyline. The following cut-off value for the test devices have been verified.

Test	Calibrator	Cut-off (ng/mL)
First Sign® Drug of Abuse NortriptylineTest	Nortriptyline	1000

e. Interference

{20}------------------------------------------------

Compounds that showed no interference at a concentration of 100μg/mL are summarized in the following tables. There were no differences observed for different formats.

4-Acetamidophenol	Erythromycin	Oxycodone
Acetophenetidin	β-Estradiol	Oxymetazoline
N-Acetylprocainamide	Estrone-3-sulfate	Papaverine
Acetylsalicylic acid	Ethyl-p-aminobenzoate	Penicillin-G
Aminopyrine	Fenoprofen	Pentazocine hydrochloride
Amobarbital	Furosemide	Pentobarbital
Amoxicillin	Gentisic acid	Perphenazine
Ampicillin	Hemoglobin	Phencyclidine
L-ascorbic acid	Hydralazine	Phenelzine
DL-Amphetamine sulfate	Hydrochlorothiazide	Phenobarbital
Apomorphine	Hydrocodone	Phentermine
Aspartame	Hydrocortisone	β-Phenylethylamine
Atropine	O-Hydroxyhippuric acid	Trans-2-phenylcyclopropylamine hydrochloride
Benzilic acid	p-Hydroxyamphetamine	L-Phenylephrine
Benzoic acid	p-Hydroxy- methamphetamine	Phenylpropanolamine
Benzoylecgonine	3-Hydroxytyramine	Prednisolone
Benzphetamine	Ibuprofen	Prednisone
Bilirubin	Iproniazid	Procaine
(±) - Brompheniramine	(±) - Isoproterenol	DL-Propranolol
Caffeine	Isoxsuprine	D-Propoxyphene
Cannabidiol	Ketamine	D-Pseudoephedrine
Cannabinol	Ketoprofen	Quinacrine
Chloralhydrate	Labetalol	Quinidine
Chloramphenicol	Loperamide	Quinine
Chlorothiazide	MDE	Ranitidine
(±) Chlorpheniramine	Meperidine	Salicylic acid
Chlorpromazine	Meprobamate	Secobarbital
Chloroquine	Methadone	Serotonin
Cholesterol	(L)Methamphetamine	Sulfamethazine
Clonidine	Methoxyphenamine	Sulindac
Cocaethylene	(±)-3,4-Methylenedioxyamphetamine hydrochloride	Tetracycline
Cocaine hydrochloride	(+)3,4-Methylenedioxymethamphetamine hydrochloride	Tetrahydrocortisone 3-(β-D-glucuronide)
Codeine	Morphine 3-β-Dglucuronide	Tetrahydrozoline
Cortisone	Morphine sulfate	Thiamine
(-) Cotinine	Nalidixic acid	Thioridazine
Creatinine	Naloxone	DL-Tyrosine
Deoxycorticosterone	Naltrexone	Tolbutamide
Dextromethorphan	Naproxen	Triamterene
Diclofenac	Niacinamide	Trifluoperazine
Diflunisal	Nifedipine	Trimethoprim
Digoxin	Norcodeine	Tryptamine
Diphenhydramine	Norethindrone	DL-Tryptophan
Doxylamine	D-Norpropoxyphene	Tyramine
Ecgonine	Noscapine	Uric acid
hydrochloride
Ecgonine methylester	Oxalic acid	Verapamil
Ephedrine	Oxazepam	Zomepirac
(L) - Epinephrine	Oxolinic acid

{21}------------------------------------------------

f. Specificity

Drug	Concentration (ng/ml)	% Cross-Reactivity
Nortriptyline	1000	100%
Amitriptyline	1500	67%
Clomipramine	15000	6.7%
Desipramine	1000	100%
Doxepine	2000	50%
Imipramine	600	167%
Nordoxepin	1000	100%
Promazine	24000	4%
Trimipramine	4000	25%
Cyclobenzaprine	1500	67%
Maprotiline	Negative at 100000	Not Detected
Promethazine	Negative at 100000	Not Detected
Norclomipramine	Negative at 100000	Not Detected

g. Effect of Urine Specific Gravity and Urine pH

{22}------------------------------------------------

h. Comparison Studies
The method comparison studies for the First Sign® Drug of Abuse Nortriptyline Test was performed in-house with three different laboratory assistants for each format of the device. Operators ran 80 (40 negative and 40 positive) unaltered clinical. The samples were blind labeled and compared to GC/MS results. The results are presented in the tables below:

DipCard format		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% and cut-off)	Near CutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan+50%)
Viewer A	Positive	0	0	1	14	26
	Negative	10	10	19	0	0
Viewer B	Positive	0	0	1	13	26
	Negative	10	10	19	1	0
Viewer C	Positive	0	0	1	13	26
	Negative	10	10	19	1	0

Discordant Results

Viewer	Sample Number	GC/MS (ng/mL)Result	DipCard FormatViewer Results
Viewer A	2014122464	863	Positive
Viewer B	2014122434	1069	Negative
Viewer B	2014122358	851	Positive
Viewer C	2014122445	1125	Negative
Viewer C	2014122607	879	Positive

Cup format		Negative	LowNegativeby GC/MS(less than -50%)	Near CutoffNegative byGC/MS(Between -50% and cut-off)	Near CutoffPositive byGC/MS(Betweenthe cut-offand +50%)	HighPositive byGC/MS(greaterthan+50%)
Viewer A	Positive	0	0	1	13	26
	Negative	10	10	19	1	0
Viewer B	Positive	0	0	1	14	26
	Negative	10	10	19	0	0

{23}------------------------------------------------

Viewer C	Positive	0	0	0	13	26
	Negative	10	10	20	1	0

Discordant Results

Viewer	Sample Number	GC/MS (ng/mL) Result	Cup Format Viewer Results
Viewer A	2014122358	851	Positive
Viewer A	2014122809	1084	Negative
Viewer B	2014122620	870	Positive
Viewer C	2014122390	1135	Negative

i. Lay-user study

A lay user study was performed at three intended user sites with 280 lay persons testing the Nortriptyline devices. They had diverse educational and professional backgrounds and ranged in age from 21 to > 50 years. Urine samples were prepared at the following concentrations; negative, +/-75%, +/-50%, +/-25% of the cutoff by spiking drugs into drug free-pooled urine specimens. The concentrations of the samples were confirmed by GC/MS. Each sample was aliquoted into individual containers and blind-labeled. Each participant was provided with the package insert, 1 blind labeled sample and a device. The results are summarized below.

% of Cutoff	Number of samples	Nortriptyline Concentration by GC/MS (ng/mL)	Lay person results		The percentage of correct results (%)
-100% Cutoff	20	0	0	20	100%
-75% Cutoff	20	261	0	20	100%
-50% Cutoff	20	495	0	20	100%
-25% Cutoff	20	720	1	19	95%
+25% Cutoff	20	1180	20	0	100%
+50% Cutoff	20	1485	20	0	100%
+75% Cutoff	20	1687	20	0	100%

Comparison between GC/MS and Lay Person Results (Nortriptyline DipCard)

Comparison between GC/MS and Lay Person Results (Nortriptyline Cup)

% of Cutoff	Numberofsamples	NortriptylineConcentration by GC/MS(ng/mL)	Lay person results		Thepercentage ofcorrect results(%)
			No. ofPositive	No. ofNegative
-100% Cutoff	20	0	0	20	100%
-75% Cutoff	20	261	0	20	100%
-50% Cutoff	20	495	0	20	100%
-25% Cutoff	20	720	1	19	95%
+25% Cutoff	20	1180	19	1	95%
+50% Cutoff	20	1485	20	0	100%

{24}------------------------------------------------

+75% Cutoff	20	1687	20	0	100%
-------------	----	------	----	---	------

j. Clinical Studies
Not applicable.

11. Conclusion

Based on the test principle and acceptable performance characteristics including precision, cut-off, interference, specificity and method comparison of the devices, it's concluded that the First Sign® Drug of Abuse MDMA Test and First Sign® Drug of Abuse EDDP Test and First Sign® Drug of Abuse Nortriptyline Test are substantially equivalent to the predicate.

Regulation Number and Section

§ 862.3620 Methadone test system.

(a)
Identification. A methadone test system is a device intended to measure methadone, an addictive narcotic pain-relieving drug, in serum and urine. Measurements obtained by this device are used in the diagnosis and treatment of methadone use or overdose and to determine compliance with regulations in methadone maintenance treatment.(b)
Classification. Class II (special controls). A methadone 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).