DRI Benzodiazepine Assay

{0} 1 # 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY ONLY TEMPLATE A. 510(k) Number: k173963 B. Purpose for Submission: Modification of a previously cleared device to replace the anti-benzodiazepine polyclonal goat antibodies with polyclonal sheep antibodies. C. Measurand: Benzodiazepines D. Type of Test: Qualitative and semi-quantitative homogeneous immunoassay E. Applicant: Microgenics Corporation F. Proprietary and Established Names: DRI Benzodiazepine Assay G. Regulatory Information: | Regulation section | Classification | Product Code | Panel | | --- | --- | --- | --- | | 21CFR 862.3170 | Class II | JXM | Toxicology (91) | H. Intended Use: 1. Intended use(s): Refer to Indications for Use below 2. Indication(s) for use: The DRI Benzodiazepine Assay is a homogeneous enzyme immunoassay intended for the qualitative and/or semi-quantitative determination of the presence of benzodiazepines and their metabolites in human urine at a cutoff concentration of 200 ng/mL. The assay is {1} intended to be used in laboratories and provides a simple and rapid analytical screening procedure to detect benzodiazepines in human urine. The assay is designed for use with a number of clinical chemistry analyzers. This assay is calibrated against Oxazepam. This product is intended to be used by trained professionals only. The semi-quantitative mode is for the purpose of enabling laboratories to determine an appropriate dilution of the specimen for confirmation by a confirmatory method such as Liquid Chromatography/tandem mass spectrometry (LC-MS/MS) or permitting laboratories to establish quality control procedures. The assay provides only a preliminary analytical test result. A more specific alternative chemical method must be used to obtain a confirmed analytical result. Gas chromatography/ mass spectrometry (GC/MS) or Liquid chromatography/tandem mass spectrometry (LC-MS/MS) is the preferred confirmatory method. Clinical and professional judgment should be applied to any drug of abuse test result, particularly when preliminary results are used. For In Vitro Diagnostic Use Only. 3. Special conditions for use statement(s): For prescription use only. 4. Special instrument requirements: Performance data was obtained using the Beckman AU680 clinical chemistry analyzer. I. Device Description: The assay consists of reagents (A and E): Reagent A: Contains sheep polyclonal anti-benzodiazepine antibodies, glucose-6-phosphate (G6P) and nicotinamide adenine dinucleotide (NAD) in Tris buffer with sodium azide as a preservative. Reagent E: Contains benzodiazepine derivative labeled with glucose-6-phosphate dehydrogenase (G6PDH) in Tris buffer with sodium azide as a preservative. J. Substantial Equivalence Information: 1. Predicate device name(s): DRI Benzodiazepine Assay 2. Predicate 510(k) number(s): k930529 {2} 3. Comparison with predicate: | Similarities | | | | --- | --- | --- | | Item | Candidate Device k173963 | Predicate Device k930529 | | Intended Use | The qualitative and/or semi-quantitative determination of the presence of benzodiazepines and their metabolites in human urine | Same | | Measured Analyte | Benzodiazepine and its metabolites | Same | | Test Matrix | Urine | Same | | Calibrator | Oxazepam | Same | | Cutoff Levels | 200 ng/mL | Same | | Methodology | Homogeneous enzyme immunoassay | Same | | Storage | 2–8°C until expiration | Same | | Differences | | | | --- | --- | --- | | Item | Candidate Device k173963 | Predicate Device k930529 | | Antibody | Polyclonal sheep antibody | Polyclonal goat antibody | K. Standard/Guidance Document Referenced (if applicable): - CLSI EP05-A3, Evaluation of Precision Performance of Quantitative Measurement Methods; Third Edition, 2014. - CLSI EP07-A2: Interference Testing in Clinical Chemistry, Approved Guideline – Second Edition, 2005. - CLSI EP09-A3; Method Comparison and Bias Estimation Using Patient Samples; Approved Guideline – Third Edition, 2013. - CLSI EP06-A: Evaluation of the Linearity of Quantitative Measurement Procedures, a Statistical Approach; Approved Guideline, 2003. - CLSI EP25-A: Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline, 2009. L. Test Principle: The DRI Benzodiazepine Assay is a homogeneous enzyme immunoassay with liquid ready-to-use reagents. The assay uses a specific antibody which can detect benzodiazepines and their metabolites in urine. The assay is based on the competition of an enzyme glucose-6-phosphate dehydrogenase (G6PDH) labeled drug and the drug from the urine sample for a {3} fixed amount of specific antibody binding sites. In the absence of free drug from the sample, the enzyme-labeled drug is bound by the specific antibody and the enzyme activity is inhibited. This phenomenon creates a relationship between drug concentration in urine and the enzyme activity. The enzyme G6PDH activity is determined spectrophotometrically at $340~\mathrm{nm}$ by measuring its ability to convert nicotinamide adenine dinucleotide (NAD) to NADH. # M. Performance Characteristics (if/when applicable): # 1. Analytical performance: # a. Precision/Reproducibility: Precision was evaluated using CLSI Guideline EP05-A3 as a guideline, at one site with one analyzer, two operators, and two lots each of reagents, calibrators and controls. Testing was carried out for 20 days with two runs per day, at least two hours apart and two replicates per run in both Qualitative and Semi-quantitative modes, giving a total of 80 determinants $(n = 80)$ . Drug-free negative urine was spiked with oxazepam to final concentrations of $-100\%$ , $-75\%$ , $-50\%$ , $-25\%$ , below cutoff and $+25\%$ , $+50\%$ , $+75\%$ and $+100\%$ , above cutoff, and the concentrations were confirmed by LC-MS/MS. Results are summarized below: Qualitative Mode Lot 1: | % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | Measured Conc. (ng/mL) | # of determinants | # Negative / # Positive | | --- | --- | --- | --- | --- | | -100 | 0 | N/A | 80 | 80/0 | | -75 | 50 | 56 | 80 | 80/0 | | -50 | 100 | 102 | 80 | 80/0 | | -25 | 150 | 161.5 | 80 | 80/0 | | Cut-off | 200 | 214 | 80 | 16/64 | | +25 | 250 | 255.5 | 80 | 0/80 | | +50 | 300 | 299 | 80 | 0/80 | | +75 | 350 | 348 | 80 | 0/80 | | +100 | 400 | 403 | 80 | 0/80 | Qualitative Mode Lot 2: | % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | Measured Conc. (ng/mL) | # of determinants | # Negative / # Positive | | --- | --- | --- | --- | --- | | -100 | 0 | N/A | 80 | 80/0 | | -75 | 50 | 56 | 80 | 80/0 | | -50 | 100 | 102 | 80 | 80/0 | | -25 | 150 | 161.5 | 80 | 80/0 | | Cut-off | 200 | 214 | 80 | 70/10 | {4} | % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | Measured Conc. (ng/mL) | # of determinants | # Negative / # Positive | | --- | --- | --- | --- | --- | | +25 | 250 | 255.5 | 80 | 0/80 | | +50 | 300 | 299 | 80 | 0/80 | | +75 | 350 | 348 | 80 | 0/80 | | +100 | 400 | 403 | 80 | 0/80 | Semi-Quantitative Mode Lot 1: | % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | Measured Conc. (ng/mL) | # of determinants | # Negative / # Positive | | --- | --- | --- | --- | --- | | -100 | 0 | N/A | 80 | 80/0 | | -75 | 50 | 56 | 80 | 80/0 | | -50 | 100 | 102 | 80 | 80/0 | | -25 | 150 | 161.5 | 80 | 80/0 | | Cut-off | 200 | 214 | 80 | 27/53 | | +25 | 250 | 255.5 | 80 | 0/80 | | +50 | 300 | 299 | 80 | 0/80 | | +75 | 350 | 348 | 80 | 0/80 | | +100 | 400 | 403 | 80 | 0/80 | Semi-Quantitative Mode Lot 2: | % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | Measured Conc. (ng/mL) | # of determinants | # Negative / # Positive | | --- | --- | --- | --- | --- | | -100 | 0 | N/A | 80 | 80/0 | | -75 | 50 | 56 | 80 | 80/0 | | -50 | 100 | 102 | 80 | 80/0 | | -25 | 150 | 161.5 | 80 | 80/0 | | Cut-off | 200 | 214 | 80 | 79/1 | | +25 | 250 | 255.5 | 80 | 0/80 | | +50 | 300 | 299 | 80 | 0/80 | | +75 | 350 | 348 | 80 | 0/80 | | +100 | 400 | 403 | 80 | 0/80 | b. Linearity/assay reportable range: A recovery study was performed using CLSI EP06-A guidelines. Samples were prepared by spiking a drug free urine pool with a high concentration of oxazepam and generating serial dilutions to achieve concentrations ranging from 100 ng/mL to 1000 ng/mL and were analyzed in replicates of 5 in semi-quantitative mode on one Beckman AU680 analyzer. The results are shown below. {5} | Target concentration (ng/mL) | Observed concentration (ng/mL) | % Recovery | | --- | --- | --- | | 0 | -1.0 | N/A | | 100 | 104.5 | 104.5 | | 200 | 196.2 | 98.1 | | 300 | 314.7 | 104.9 | | 400 | 455.3 | 113.8 | | 500 | 565.2 | 113.0 | | 600 | 661.0 | 110.2 | | 700 | 764.7 | 109.2 | | 800 | 872.1 | 109.0 | | 900 | 937.3 | 104.1 | | 1000 | 1024.9 | 102.5 | c. Traceability, Stability, Expected values (controls, calibrators, or methods): Traceability: The primary calibrators are traceable to the oxazepam drug purchased from a commercial source which is established at 98% purity. The concentration of the primary calibrator stocks is confirmed by LC-MS/MS from three independent laboratories. d. Detection limit: Not applicable. e. Analytical specificity: Cross-Reactivity Of Benzodiazepine Compounds and Metabolites The cross reactivity of benzodiazepine compounds and their metabolites was evaluated by adding known amounts of each compound to drug-free negative urine. The specificity (cross-reactivity) study was performed using one lot of reagents, calibrators and controls in both qualitative and semi-quantitative modes. Percent cross-reactivity was calculated as (Cut-off concentration / Lowest concentration of cross reactant that gives a positive result) x 100. Results are summarized below: | Structurally related and unrelated compounds | Lowest concentration producing a positive result (ng/mL) | Cross-reactivity (%) | | --- | --- | --- | | α-Hydroxyalprazolam | 110 | 182 | | α-Hydroxytriazolam | 140 | 143 | | Alprazolam | 110 | 182 | | 7-Aminoclonazepam | 2,500 | 8 | | 7-Aminoflunitrazepam | 300 | 67 | {6} | Structurally related and unrelated compounds | Lowest concentration producing a positive result (ng/mL) | Cross-reactivity (%) | | --- | --- | --- | | 7-Aminonitrazepam | 300 | 67 | | Bromazepam | 170 | 118 | | Chlordiazepoxide | 700 | 29 | | Clobazam | 150 | 133 | | Clonazepam | 210 | 95 | | Clorazepate | 135 | 148 | | Delorazepam | 150 | 133 | | Demoxepam | 220 | 91 | | Desalkylflurazepam | 130 | 154 | | Diazepam | 110 | 182 | | Estazolam | 100 | 200 | | Flunitrazepam | 120 | 167 | | Flurazepam | 150 | 133 | | 2-Hydroxyethylflurazepam | 120 | 167 | | Lorazepam | 700 | 29 | | Lorazepam glucuronide | 50,000 | <0.4 | | Lormetazepam | 275 | 73 | | Medazepam | 325 | 62 | | Midazolam | 180 | 111 | | Nitrazepam | 130 | 154 | | Norchlordiazepoxide | 800 | 25 | | Nordiazepam | 110 | 182 | | Oxaprozin | 125,000 | 0.16 | | Oxazepam | 200 | 100 | | Oxazepam glucuronide | 50,000 | 0.4 | | Prazepam | 200 | 100 | | Temazepam | 160 | 125 | | Temazepam glucuronide | 50,000 | <0.4 | | Triazolam | 130 | 154 | ## Interference Testing of Structurally Unrelated Compounds Interference from structurally unrelated compounds was evaluated by spiking these compounds into urine samples containing near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of oxazepam. The compounds listed in the table below did not cause any positive or negative interference at the concentrations shown: {7} | Structurally Unrelated Compounds | Tested Concentration (ng/mL) | | --- | --- | | 6-Acetyl morphine | 100,000 | | 10,11 Dihydrocarbamazepine | 100,000 | | 11-nor-Δ^{9}-THC-COOH | 100,000 | | Acetaminophen | 1,000,000 | | Acetylsalicylic acid | 1,000,000 | | Amitriptyline | 100,000 | | Amoxicillin | 100,000 | | Amphetamine | 100,000 | | Amisulpride | 100,000 | | Benzotropine Mesylate | 100,000 | | Benzoylecgonine | 100,000 | | Brompheniramine | 100,000 | | Buprenorphine | 100,000 | | Caffeine | 100,000 | | Captopril | 100,000 | | Chlorpromazine | 100,000 | | Chloroquine | 100,000 | | Cimetidine | 100,000 | | Clomipramine | 100,000 | | Codeine | 100,000 | | Desipramine | 100,000 | | Dextromethorphan | 100,000 | | Digoxin | 100,000 | | Dihydrocodeine | 100,000 | | Diphenhydramine | 500,000 | | Doxepine HCl | 100,000 | | EDDP | 100,000 | | EMDP | 25,000 | | Enalapril | 100,000 | | Fentanyl | 100,000 | | Fluoxetine | 500,000 | | Fluophenazine | 100,000 | | Haloperidol | 100,000 | | Heroin | 100,000 | | Hydrocodone | 100,000 | | Hydromorphone | 100,000 | | Hydroxychloroquine | 100,000 | | Hydroxyzine | 100,000 | | Ibuprofen | 100,000 | | Imipramine | 100,000 | 8 {8} | Structurally Unrelated Compounds | Tested Concentration (ng/mL) | | --- | --- | | LAAM | 100,000 | | Levorphanol | 100,000 | | Levothyroxine | 100,000 | | Maprotiline | 100,000 | | Meperidine | 100,000 | | Methadone | 100,000 | | Methamphentamine | 100,000 | | Morphine | 100,000 | | Morphine-3β-D-glucuronide | 100,000 | | Morphine-6β-D-glucuronide | 100,000 | | Nalbuphine | 100,000 | | Nalorphine | 100,000 | | Naloxone | 100,000 | | Naltrexone | 100,000 | | Naproxen | 100,000 | | Nifedipine | 100,000 | | Norcodeine | 100,000 | | Norhydrocodone | 100,000 | | Norfluoxetine | 500,000 | | Noroxycodone | 100,000 | | Noroxymorphone | 100,000 | | Norpropoxyphene | 100,000 | | Norsertraline | 62,500 | | Nortryptiline | 100,000 | | Oxycodone | 100,000 | | Oxymorphone | 100,000 | | Paroxetine | 100,000 | | Perphenazine | 100,000 | | Phencyclidine | 100,000 | | Phenobarbital | 100,000 | | Procyclidine | 100,000 | | Propoxyphene | 100,000 | | Protriptyline | 100,000 | | Ranitidine | 100,000 | | Secobarbital | 100,000 | | Sertraline | 62,500 | | Sulpiride | 100,000 | | Tapentadol | 100,000 | | Thioridazine | 100,000 | | Tramadol | 100,000 | | Triprolidine | 100,000 | 9 {9} 10 | Structurally Unrelated Compounds | Tested Concentration (ng/mL) | | --- | --- | | Verapamil | 100,000 | | Zaleplon | 100,000 | | Zolpidem | 100,000 | | Zopiclone | 100,000 | ## Interference Testing of Endogenous Compounds Potential interference from endogenous compounds was evaluated by spiking these compounds into urine samples containing near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of oxazepam. The compounds or conditions listed in the table below did not cause any positive or negative interference, either in the qualitative or semi-quantitative modes, at the concentrations shown in the table below: | Compounds | Tested Conc. (mg/dL) | | --- | --- | | Acetone | 500 | | Ascorbic Acid | 150 | | Creatinine | 400 | | Ethanol | 1000 | | Galactose | 5 | | Glucose | 1,000 | | Hemoglobin | 150 | | Human Serum Albumin (HSA) | 200 | | Oxalic acid | 50 | | Riboflavin | 3 | | Sodium Chloride | 1,000 | | Urea | 1,000 | ## Interference Testing of Specific Gravity and pH: Interference from specific gravity and pH was evaluated by adjusting the specific gravity and pH of samples with near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of oxazepam. The following specific gravity or pH levels did not cause any positive or negative interference: Specific gravity of 1.004, 1.005, 1.007, 1.010, 1.011, 1.013, 1.019, 1.023, 1.025, 1.029. pH of 3, 4, 5, 6, 7, 8, 9, 10, and 11. {10} f. Assay cut-off: Characterization of how the device performs analytically around the claimed cutoff concentration is described in the precision section, M.1.a. above. 2. Comparison studies: A method comparison study was performed in accordance with CLSI Guideline EP09-A3. One hundred and six patient urine samples were analyzed by the DRI Benzodiazepine Assay in both qualitative and semi-quantitative modes and the results were compared to LC-MS/MS. The results were the same for the qualitative and semi-quantitative modes and are summarized below. Candidate Device Results vs. LC/MS-MS Values | Candidate Device Results | Negative by LC-MS/MS | < 50% of Cutoff concentration by LC-MS/MS (< 100ng/mL) | Near Cutoff Negative (Between 50% below the cutoff and the cutoff concentration as determined by LC-MS/MS) (100 - 199 ng/mL) | Near Cutoff Positive (Between the cutoff and 50% above the cutoff concentration as determined by LC-MS/MS) (200 - 300 ng/mL) | High Positives (Greater than 50% above cutoff concentration (>300 ng/mL) | | --- | --- | --- | --- | --- | --- | | Positive | 0 | 1* | 3* | 5 | 45 | | Negative | 48 | 2 | 2 | 0 | 0 | *Discordant samples | Sample ID | Qualitative | Semi-Quantitative | LC-MS/MS | | | | --- | --- | --- | --- | --- | --- | | | Negative/Positive | Negative/Positive | Oxazepam (ng/mL) | 7-Amino-clonazepam (ng/mL) | α-Hydroxy-alprazolam (ng/mL) | | CA160606-045 | Positive | Positive | 86.20 | 3154.6 | NA | | CA160926-057 | Positive | Positive | 175.08 | 410.7 | 13.5 | | CA170605-001 | Positive | Positive | 151.52 | 560.4 | 1.43 | | CA160908-003 | Positive | Positive | 192.87 | NA | 96.3 | {11} b. Matrix comparison: Not applicable. Urine is the only claimed matrix for the candidate device. 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 Parts 801 and 809, as applicable. O. Conclusion: The submitted information in this premarket notification is complete and supports a substantial equivalence decision. 12