(29 days)
The LZI Tramadol Enzyme Immunoassay is intended for the qualitative and semi-quantitative determination of tramadol in human urine at the cutoff value of 100 ng/mL when calibrated against tramadol. The assay is designed for prescription use with a number of automated clinical chemistry analyzers.
The semi-quantitative mode is for purposes of (1) enabling laboratories to determine an appropriate dilution of the specimen for confirmation by a confirmatory method such as gas or liquid chromatography/mass spectrometry (GC/MS or LC/MS) or (2) permitting laboratories to establish quality control procedures.
The assay provides only a preliminary analytical result. A more specific alternative chemical method (e.g., gas or liquid chromatography and mass spectrometry) must be used in order to obtain a confirmed analytical result. Clinical consideration and professional judgment should be exercised with any drug of abuse test result, particularly when the preliminary test result is positive.
The LZI Tramadol Enzyme Immunoassay is a homogeneous enzyme immunoassay with readyto-use liquid reagents. The assay is based on competition between drug in the sample and drug labeled with the enzyme glucose-6-phosphate dehydrogenase (G6PDH) for a fixed amount of antibody in the reagent. The drug-labeled G6PDH conjugate is traceable to a commercially available tramadol standard and referred to as tramadol-labeled G6PDH conjugate. Enzyme activity decreases upon binding to the antibody, and the drug concentration in the sample is measured in terms of enzyme activity. In the absence of drug in the sample, tramadol-labeled G6PDH conjugate is bound to antibody, and the enzyme activity is inhibited. On the other hand, when free drug is present in the sample, antibody would bind to free drug; the unbound tramadol-labeled G6PDH then exhibits its maximal enzyme activity. Active enzyme converts nicotinamide adenine dinucleotide (NAD) to NADH, resulting in an absorbance change that can be measured spectrophotometrically at 340 nm.
The LZI Tramadol Enzyme Immunoassay is a kit comprised of two reagents, an R1 and R2 which are bottled separately but sold together within the kit. The LZI Tramadol Enzyme Immunoassay is traceable to a commercially available tramadol standard.
The Ri solution contains mouse monoclonal anti-tramadol antibody, glucose-6-phosphate (G6P) nicotinamide adenine dinucleotide (NAD), stabilizers, and sodium azide (0.09 %) as a preservative. The R2 solution contains glucose-6-phosphate dehydrogenase (G6PDH) labeled with tramadol in buffer with sodium azide (0.09 %) as a preservative.
The provided text describes the performance characteristics of the LZI Tramadol Enzyme Immunoassay. Here's a breakdown of the requested information:
1. Table of Acceptance Criteria and Reported Device Performance
The document does not explicitly list "acceptance criteria" for precision, recovery, or method comparison in a consolidated table with specific numerical targets like "sensitivity > X%" or "specificity > Y%". Instead, it presents the raw performance data and concludes that the results are acceptable or show no significant interference. However, using the precision study's implicit goal (correct qualitative classification for samples ± 25% of cutoff) and the method comparison's agreement, and the recovery study's target, we can construct the following:
| Acceptance Criterion (Implicit) | Reported Device Performance | Study |
|---|---|---|
| Precision: Samples at 0%, 25%, 50%, 75% of cutoff should be Negative. Samples at 125%, 150%, 175%, 200% of cutoff should be Positive. (For 100% cutoff, some variance is expected due to inherent assay variability near the cutoff). | 100% Qualitative Agreement for 0, 25, 50, 75, 125, 150, 175, 200 ng/mL samples over 22 days/88 replicates. Samples at 100 ng/mL appropriately showed mixed results. | Precision (Qualitative) |
| Analytical Recovery: Determined average percent recovery for spiked samples to be between 85% and 115% of target concentration. | Recovery ranged between 89% - 116%. | Analytical Recovery |
| Method Comparison (Qualitative Accuracy): High percentage agreement with LC/MS for clinical samples. (Implicitly close to 100% after addressing near-cutoff discrepancies). | Overall % Agreement: 97.7% (Semi-Quantitative) and 95.3% (Qualitative) | Method Comparison - Clinical Samples |
| Cross-reactivity: Structurally unrelated compounds should not cause false positives or interfere with results at ±25% of cutoff concentrations when present at high concentrations (e.g., 100,000 ng/mL). | Most tested compounds showed no cross-reactivity and did not interfere with negative or positive trampadol samples. | Cross-reactivity & Structurally Unrelated Compounds |
| Endogenous/Preservative Interference: Common endogenous and preservative compounds should not cause false positives or interfere with results at ±25% of cutoff concentrations when present at high concentrations (e.g., 1000 mg/dL). | Boric Acid interfered at 1% w/v but no other significant interference observed. | Endogenous and Preservative Compound Interference |
| Specific Gravity Interference: No interference observed across a range of specific gravity (1.000 to 1.030) for samples at 0 ng/mL, 75 ng/mL, and 125 ng/mL tramadol. | No interference observed. | Specific Gravity Interference |
| pH Interference: No major interference observed between pH 3 to pH 11 for samples at 0 ng/mL, 75 ng/mL, and 125 ng/mL tramadol. | No major interference observed. | pH Interference |
2. Sample Size Used for the Test Set and Data Provenance
- Precision Test Set: Not clinical samples. Tramadol samples were prepared by spiking a tramadol standard into a pool of negative human urine.
- For qualitative and semi-quantitative modes: 8 concentrations (0, 25, 50, 75, 100, 125, 150, 175, 200 ng/mL). Each concentration was tested in duplicate, two runs a day for 22 days, totaling 88 replicates per concentration.
- Analytical Recovery: Not clinical samples. A drug-free urine pool spiked with tramadol at 400 ng/mL was serially diluted. Each dilution was run in 10 replicates.
- Method Comparison - Clinical Samples: Eighty-six (86) unaltered clinical samples.
- Provenance: Samples were collected by Lin-Zhi International, Inc. (LZI) and the University of California at San Francisco (UCSF, San Francisco). This indicates a retrospective collection of clinical samples from a US-based origin.
- Cross-reactivity, Endogenous/Preservative Interference, Specific Gravity, pH Interference: Not clinical samples. Prepared by spiking substances into pooled negative human urine. All samples were tested in duplicates (except for Analytical Recovery, which was 10 replicates).
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications
N/A. This device is an in-vitro diagnostic assay for tramadol in urine. The ground truth for the clinical samples was established using a "more specific alternative chemical method," specifically LC/MS (Liquid Chromatography/Mass Spectrometry), which is a gold standard analytical technique for drug quantification, not human expert interpretation. For the spiked samples (precision, recovery, interference studies), the ground truth was the known concentration of the spiked substance, not expert consensus.
4. Adjudication Method for the Test Set
N/A. The ground truth for clinical samples was established by LC/MS analysis, which is an objective chemical measurement. There was no human expert adjudication of results, as there would be in an image analysis or diagnostic interpretation study.
5. If a Multi Reader Multi Case (MRMC) Comparative Effectiveness Study was done
No, an MRMC comparative effectiveness study was not done. This device is an automated enzyme immunoassay, not an AI-assisted diagnostic tool that involves human reader interpretation. Therefore, there is no "human readers improve with AI vs without AI assistance" effect size to report. The study focuses on the analytical performance of the immunoassay itself in comparison to a confirmatory method (LC/MS).
6. If a Standalone (i.e. algorithm only without human-in-the-loop performance) was done
Yes, the studies presented are standalone (algorithm only) performance evaluations of the LZI Tramadol Enzyme Immunoassay. The assay is automated on a clinical chemistry analyzer, and its results are compared directly to known concentrations or LC/MS results without human interpretative input as part of its primary performance.
7. The Type of Ground Truth Used
- For Clinical Samples (Method Comparison): LC/MS (Liquid Chromatography/Mass Spectrometry) was used as the confirmatory gold standard for tramadol concentrations.
- For Spiked Samples (Precision, Analytical Recovery, Cross-reactivity, Interference Studies): The ground truth was the known concentration of tramadol or interfering substance spiked into negative human urine.
8. The Sample Size for the Training Set
N/A. This is an immunoassay, not a machine learning model. It does not have a "training set" in the context of AI/ML. The assay's performance is based on its chemical and enzymatic reactions, calibrated using known standards.
9. How the Ground Truth for the Training Set was Established
N/A. As mentioned above, there is no "training set" for this immunoassay device. The assay uses calibrators (0, 50, 100, 225, and 400 ng/mL tramadol) to establish its calibration curve, which determines the relationship between enzyme activity and tramadol concentration. These calibrators represent known, established concentrations.
§ 862.3650 Opiate test system.
(a)
Identification. An opiate test system is a device intended to measure any of the addictive narcotic pain-relieving opiate drugs in blood, serum, urine, gastric contents, and saliva. An opiate is any natural or synthetic drug that has morphine-like pharmocological actions. The opiates include drugs such as morphine, morphine glucoronide, heroin, codeine, nalorphine, and meperedine. Measurements obtained by this device are used in the diagnosis and treatment of opiate use or overdose and in monitoring the levels of opiate administration to ensure appropriate therapy.(b)
Classification. Class II (special controls). An opiate 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).