(20 days)
The LZI Fentanyl III Enzyme Immunoassay is intended for the qualitative determination of fentanyl in human urine at the cutoff value of 1 ng/mL when calibrated against fentanyl. The assay is designed for prescription use with a number of automated clinical chemistry analyzers.
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 Fentanyl III Enzyme Immunoassay is a homogeneous enzyme immunoassay with ready-to-use liquid reagents. The assay is based on competition between the drug in the sample and the 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 fentanyl standard and referred to as fentanyl-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 a drug in the sample, fentanyl-labeled G6PDH conjugate is bound to the antibody, and the enzyme activity is inhibited. On the other hand, when the free drug is present in the sample, the antibody would bind to the free drug; the unbound fentanyl-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 Fentanyl III Enzyme Immunoassay is a kit comprised of two reagents, R1 and R2, which are bottled separately but sold together within the kit.
The R1 solution contains mouse monoclonal anti-fentanyl 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 fentanyl in buffer with sodium azide (0.09%) as a preservative.
The provided FDA 510(k) Clearance Letter for the LZI Fentanyl III Enzyme Immunoassay details the device's acceptance criteria (in terms of performance characteristics) and the study results that demonstrate the device meets these criteria.
Here's a breakdown of the requested information:
1. Acceptance Criteria and Reported Device Performance
The acceptance criteria are implicitly defined by the satisfactory performance demonstrated in the various studies, aiming to prove substantial equivalence to the predicate device. The performance characteristics sections show the device meets the expected qualitative and quantitative accuracy, precision, and specificity.
| Acceptance Criteria Category | Specific Criteria (Implicitly Derived from Study Design/Outcomes) | Reported Device Performance |
|---|---|---|
| Intended Use Equivalence | Device is substantially equivalent to the predicate (LZI Fentanyl II Enzyme Immunoassay) in terms of intended use, method principle, device components, and clinical performance. | The submission states, "The LZI Fentanyl III Enzyme Immunoassay is substantially equivalent to the LZI Fentanyl II Enzyme Immunoassay (K201938) manufactured by LZI in terms of intended use, method principle, device components, and clinical performance." The changes are clearly identified (target analyte from norfentanyl to fentanyl, updated cutoff, and assay application parameters), and subsequent studies address the impact of these changes. |
| Qualitative Accuracy | High concordance with LC/MS confirmation for true positives and true negatives, especially at and around the cutoff concentration. | Method Comparison - Clinical Samples (1 ng/mL Cutoff): |
- Negative by LC/MS analysis: 35 negatives, 0 positives by EIA.
- ** 50% above cutoff) by LC/MS:** 0 negatives, 61 positives by EIA.
Overall, 35/35 true negatives, 83/83 true positives (excluding near-cutoff positives and positives above cutoff). The remaining 32 discrepant samples (20 + 12) were positive by EIA but negative by LC/MS for fentanyl, indicating cross-reactivity with norfentanyl as a contributing factor. |
| Precision | Consistent and reliable qualitative results (positive/negative) across multiple runs and days, especially around the cutoff concentration. | Precision: 1 ng/mL Cutoff (N=88 total replicates over 22 days):
- 0 ng/mL, 0.25 ng/mL, 0.5 ng/mL, 0.75 ng/mL: 88/88 Negative (100% agreement).
- 1 ng/mL (Cutoff): 4 Neg / 84 Pos (95.5% Positive, 4.5% Negative - demonstrating near-cutoff variability, which is expected for qualitative assays at the cutoff).
- 1.25 ng/mL, 1.5 ng/mL, 1.75 ng/mL, 2 ng/mL: 88/88 Positive (100% agreement). |
| Cross-Reactivity (Specificity) | Minimal or no false positives from structurally unrelated compounds. Acceptable cross-reactivity with fentanyl metabolites and structurally related compounds as defined by the assay's intended specificity. | Fentanyl and Metabolites: Fentanyl (100%), Norfentanyl (40%).
Structurally Related Compounds: Varying levels of cross-reactivity (e.g., Acetyl fentanyl 25%, Acryl fentanyl 100%, Thiofentanyl 200%). Many substances showed "ND" (Not Detected) at high concentrations (e.g., 100,000 ng/mL).
Structurally Unrelated Pharmacological Compounds: No interference (all Neg/Neg/Pos results for 0 ng/mL Fentanyl, -50% Cutoff, +50% Cutoff respectively) at 100,000 ng/mL for almost all listed compounds, except Dextromethorphan, which showed interference at 20,000 ng/mL (Neg/Pos/Pos) but was acceptable at 15,000 ng/mL (Neg/Neg/Pos). This demonstrates good specificity against common drugs. |
| Interference | Test performance (positive/negative results) should not be significantly affected by common endogenous substances or preservatives found in urine, or by varying specific gravity and pH within physiological ranges. | Endogenous and Preservative Compound Interference: No interference observed for most compounds (e.g., Acetone, Ascorbic acid, Bilirubin, Glucose, Hemoglobin) at high concentrations (e.g., 100,000 mg/dL), showing Neg/Neg/Pos results for 0 ng/mL, -50% Cutoff, +50% Cutoff respectively. Boric acid showed interference at 1,000 mg/dL (Neg/Neg/Neg), indicating a potential issue if present at high concentrations. This is a known interference for certain urine assays.
Specific Gravity Interference: No interference observed across the range of 1.000 to 1.030 (all Neg/Neg/Pos).
pH Interference: No major interference observed between pH 3 to pH 11 (all Neg/Neg/Pos). |
2. Sample Size and Data Provenance
- Test Set Sample Size:
- Precision Study: 88 replicates for each concentration level (0 ng/mL to 2 ng/mL fentanyl).
- Method Comparison (Clinical Samples): 150 unaltered clinical samples.
- Cross-Reactivity: Duplicates for each compound and concentration tested.
- Interference (Endogenous/Preservative): Duplicates for each compound and concentration tested.
- Specific Gravity/pH Interference: Duplicates for each specific gravity/pH level and fentanyl concentration.
- Data Provenance:
- The clinical samples for the Method Comparison study were obtained by LZI and "through collaboration with various clinical laboratories across the United States and Canada." This indicates a prospective and/or retrospective collection of real-world clinical samples from diverse geographical regions. The nature (retrospective/prospective) of the collection for these specific 150 samples isn't explicitly stated but "clinical samples" usually implies samples collected from patients.
3. Number of Experts and Qualifications for Ground Truth
- The document does not specify the number of experts used to establish ground truth.
- Qualifications of Experts: Not explicitly stated. However, for LC/MS confirmation, it implicitly relies on the expertise of the laboratory personnel performing and interpreting the LC/MS (Liquid Chromatography-Mass Spectrometry) analysis, which is a gold standard for chemical identification and quantification. These would typically be trained analytical chemists or lab technicians with experience in mass spectrometry.
4. Adjudication Method for the Test Set
- The document does not describe an adjudication method for the test set in the typical sense of multiple human readers or reviewers resolving discrepancies.
- For the Method Comparison study, the "ground truth" was established independently by LC/MS. Discrepancies between the EIA result and the LC/MS result were reported and attributed to norfentanyl cross-reactivity. This is a comparison against a reference method, not a subjective adjudication by experts.
5. Multi Reader Multi Case (MRMC) Comparative Effectiveness Study
- No, an MRMC comparative effectiveness study was not done. This type of study is more common for diagnostic imaging AI algorithms where human interpretation is a primary component and AI aims to augment or replace it. This document describes an in vitro diagnostic (IVD) assay, an enzyme immunoassay, which is an automated lab test. Its performance is evaluated against a reference standard (LC/MS), not against human reader performance.
6. Standalone Performance
- Yes, standalone performance was done. The entire submission details the performance of the LZI Fentanyl III Enzyme Immunoassay as a standalone device. The qualitative accuracy, precision, cross-reactivity, and interference studies all evaluate the algorithm/device's performance independently in generating a preliminary analytical result from a urine sample. It does not involve human-in-the-loop performance; rather, it provides a result that a clinician then interprets in conjunction with other clinical data.
7. Type of Ground Truth Used
- The primary type of ground truth used for the quantitative confirmation of fentanyl concentration in the clinical samples (Method Comparison study) was LC/MS (Liquid Chromatography-Mass Spectrometry) analysis. This is considered a highly specific and sensitive reference method for drug quantification.
8. Sample Size for the Training Set
- The document does not explicitly state the sample size for the training set. The described studies are verification and validation activities for a modified assay (Special 510(k) for an existing predicate device, LZI Fentanyl II). For IVD assays, "training" often refers to the internal development and optimization of the assay performed by the manufacturer, which precedes the formal V&V studies presented for regulatory submission. The reported studies are the test set performance.
9. How the Ground Truth for the Training Set Was Established
- Since the training set size is not explicitly stated, the method for establishing ground truth for it is also not explicitly described. However, it is highly probable that internal development and optimization of the assay would have utilized similar rigorous analytical methods, likely LC/MS or other established analytical techniques, to calibrate and refine the assay's performance against known concentrations of fentanyl and its metabolites. This would be part of the manufacturer's design control and development process.
§ 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).