CEDIA Benzodiazepine Assay
Device Facts
| Record ID | K190968 |
|---|---|
| Device Name | CEDIA Benzodiazepine Assay |
| Applicant | Microgenics Corporation |
| Product Code | JXM · Clinical Toxicology |
| Decision Date | Dec 9, 2019 |
| Decision | SESE |
| Submission Type | Traditional |
| Regulation | 21 CFR 862.3170 |
| Device Class | Class 2 |
Indications for Use
The CEDIA™ Benzodiazepine Assay is a homogeneous enzyme immunoassay intended for the qualitative and/or semiquantitative determination of benzodiazepines in human urine at a cutoff concentration of 200 ng/mL. 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 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 consideration 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.
Device Story
CEDIA™ Benzodiazepine Assay is a homogeneous enzyme immunoassay for human urine; utilizes recombinant DNA technology with bacterial enzyme β-galactosidase split into inactive Enzyme Acceptor (EA) and Enzyme Donor (ED) fragments. In presence of benzodiazepines, fragments re-associate to form active enzyme, cleaving substrate to produce spectrophotometrically measurable color change. Requires addition of β-glucuronidase to hydrolyze glucuronidated metabolites. Operated by trained professionals on clinical analyzers (e.g., Beckman Coulter AU680). Provides preliminary qualitative/semi-quantitative results; semi-quantitative mode assists in determining specimen dilution for confirmatory testing (LC-MS/MS or GC/MS). Clinical decision-making requires professional judgment and confirmatory testing. Benefits include rapid preliminary screening for benzodiazepine presence in urine samples.
Clinical Evidence
Bench testing only. Performance evaluated on Beckman Coulter AU680. Precision (n=80) showed high concordance at cutoff. Linearity demonstrated up to 800 ng/mL. Method comparison against LC-MS/MS (n=128) showed 97% concordance in qualitative mode and 96% in semi-quantitative mode. Specificity and interference studies confirmed no significant cross-reactivity with structurally unrelated compounds or physiological substances (pH, specific gravity, common drugs).
Technological Characteristics
Homogeneous enzyme immunoassay; uses recombinant microbial β-galactosidase fragments (EA/ED). Reagents: sheep polyclonal anti-benzodiazepine antibody, enzyme-donor conjugate, β-glucuronidase. Spectrophotometric detection. Automated platform (e.g., Beckman AU680). Calibrators traceable to Oxazepam. Complies with CLSI guidelines EP05-A3, EP06-A, EP07-A2, and EP25-A.
Indications for Use
Indicated for qualitative and/or semiquantitative determination of benzodiazepines in human urine at a 200 ng/mL cutoff. For use by trained professionals in clinical laboratory settings to provide preliminary analytical results requiring confirmation.
Regulatory Classification
Identification
A benzodiazepine test system is a device intended to measure any of the benzodiazepine compounds, sedative and hypnotic drugs, in blood, plasma, and urine. The benzodiazepine compounds include chlordiazepoxide, diazepam, oxazepam, chlorzepate, flurazepam, and nitrazepam. Measurements obtained by this device are used in the diagnosis and treatment of benzodiazepine use or overdose and in monitoring levels of benzodiazepines to ensure appropriate therapy.
Special Controls
*Classification.* Class II (special controls). A benzodiazepine test system is not exempt if it is intended for any use other than employment or insurance testing or is intended for Federal drug testing programs. The device is exempt from the premarket notification procedures in subpart E of part 807 of this chapter subject to the limitations in § 862.9, provided the test system is intended for employment and insurance testing and includes a statement in the labeling that the device is intended solely for use in employment and insurance testing, and does not include devices intended for Federal drug testing programs (*e.g.,* programs run by the Substance Abuse and Mental Health Services Administration (SAMHSA), the Department of Transportation (DOT), and the U.S. military).
Predicate Devices
- CEDIA™ DAU Benzodiazepine Assay (k962734)
Submission Summary (Full Text)
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Food and Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993-0002
www.fda.gov
# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY
ASSAY ONLY
## I Background Information:
A 510(k) Number
k190968
B Applicant
Microgenics Corporation
C Proprietary and Established Names
CEDIA™ Benzodiazepine Assay
D Regulatory Information
| Product Code(s) | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| JXM | Class II | 21 CFR 862.3170 - Benzodiazepine Test System | TX - Clinical Toxicology |
## II Submission/Device Overview:
A Purpose for Submission:
New device
B Measurand:
Benzodiazepines
C Type of Test:
Qualitative and semi-quantitative homogeneous immunoassay
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## III Intended Use/Indications for Use:
### A Intended Use(s):
See Indications for Use below.
### B Indication(s) for Use:
The CEDIA™ Benzodiazepine Assay is a homogeneous enzyme immunoassay intended for the qualitative and/or semiquantitative determination of benzodiazepines in human urine at a cutoff concentration of 200 ng/mL.
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 consideration 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.
### C Special Conditions for Use Statement(s):
Rx - For Prescription Use Only
### D Special Instrument Requirements:
Performance data was obtained using the Beckman AU680 clinical chemistry analyzer.
## IV Device/System Characteristics:
### A Device Description:
The assay consists of two lyophilized and two liquid reagents:
- EA Reconstitution Buffer and ED Reconstitution Buffer: (lyophilized)
- EA Reagent and ED Reagent (liquid ready to-use)
- β-Glucuronidase reagent
The components include sheep polyclonal anti-benzodiazepine antibody, recombinant microbial "enzyme donor" – benzodiazepine conjugate, "enzyme acceptor", chlorophenol red β-D-galactopyranoside, stabilizers and preservatives. All specimens must be tested with β-glucuronidase enzyme. Add β-glucuronidase enzyme to the reconstituted EA solution before using the assay. This enzyme will hydrolyze the glucuronidated metabolites of benzodiazepines in the samples, thereby enabling the detection of benzodiazepine glucuronides.
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B Principle of Operation:
CEDIA™ technology uses recombinant DNA technology to produce a unique homogeneous enzyme immunoassay system. The assay is based on the bacterial enzyme β-galactosidase, which has been genetically engineered into two inactive fragments, Enzyme acceptor (EA) and Enzyme Donor (ED). These fragments spontaneously re-associate to form fully active enzyme that, in the assay format, cleaves a substrate. This generates a color change that can be measured spectrophotometrically.
V Substantial Equivalence Information:
A Predicate Device Name(s):
CEDIA™ DAU Benzodiazepine Assay
B Predicate 510(k) Number(s):
k962734
C Comparison with Predicate(s):
| Device & Predicate Device(s): | k190968 | k962734 |
| --- | --- | --- |
| Device Trade Name | CEDIA™ Benzodiazepine Assay | CEDIA™ DAU Benzodiazepine Assay |
| General Device Characteristic Similarities | | |
| Intended Use/Indications For Use | Intended for the qualitative and semiquantitative assay of benzodiazepines in human urine | Same |
| Measured Analyte | Benzodiazepine and its metabolites | Same |
| Antibody | Polyclonal sheep antibody | Same |
| Test Matrix | Urine | Same |
| Methodology | Homogeneous enzyme immunoassay | Same |
| General Device Characteristic Differences | CEDIA™ Benzodiazepine Assay | CEDIA™ DAU Benzodiazepine Assay |
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| Calibrator | Oxazepam | Nitrazepam |
| --- | --- | --- |
| Cutoff Levels | 200 ng/mL High Sensitivity | 200 ng/mL or 300 ng/mL |
VI Standards/Guidance Documents Referenced:
CLSI EP05-A3 – Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline – Third Edition.
CLSI EP06-A – Evaluation of the Linearity of Quantitative Measurement Procedures: A Statistical Approach; Approved Guideline.
CLSI EP07-A2 – Interference Testing In Clinical Chemistry; Approved Guideline – Second Edition
CLSI EP25-A – Evaluation of Stability of In Vitro Diagnostic Reagents; Approved Guideline
VII Performance Characteristics (if/when applicable):
A Analytical Performance:
1. Precision/Reproducibility:
The precision study was performed using CLSI Guideline EP05-A3 as a guideline. 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 spikes were confirmed by LC-MS/MS.
Qualitative Mode for 200 ng/ml cutoff:
| % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | N | # Negative / # Positive |
| --- | --- | --- | --- |
| -100 | 0 | 80 | 80/0 |
| -75 | 50 | 80 | 80/0 |
| -50 | 100 | 80 | 80/0 |
| -25 | 150 | 80 | 80/0 |
| Cut-off | 200 | 80 | 6/74 |
| +25 | 250 | 80 | 0/80 |
| +50 | 300 | 80 | 0/80 |
| +75 | 350 | 80 | 0/80 |
| +100 | 400 | 80 | 0/80 |
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Semi-Quantitative Mode for 200 ng/ml cutoff:
| % of Cutoff (200 ng/mL) | Target concentration (ng/mL) | N | # Negative / # Positive |
| --- | --- | --- | --- |
| -100 | 0 | 80 | 80/0 |
| -75 | 50 | 80 | 80/0 |
| -50 | 100 | 80 | 80/0 |
| -25 | 150 | 80 | 79/1 |
| Cut-off | 200 | 80 | 1/79 |
| +25 | 250 | 80 | 0/80 |
| +50 | 300 | 80 | 0/80 |
| +75 | 350 | 80 | 0/80 |
| +100 | 400 | 80 | 0/80 |
Linearity:
A linearity study was performed using CLSI EP06-A guidelines. To demonstrate the dilution linearity for purposes of sample dilution and quality control up to 800 ng/mL assay range, drug free urine was spiked to 900 ng/mL level calibrator using Oxazepam and diluted with drug free urine to generate 8 intermediate levels. Each sample was run in replicates of five in semi-quantitative mode and the average was used to determine percent recovery compared to the expected target value. The average percent recovery is summarized in the table below.
| Expected Concentration (ng/mL) | Observed concentration (ng/mL) | Average Recovery (%) | Range of Recovery (%) |
| --- | --- | --- | --- |
| 0 | -1.00 | N/A | 95.2 – 107.8 |
| 100 | 115.6107.8 | 115.60107.8 | |
| 200 | 198.0205.8 | 99.00102.9 | |
| 300 | 304.4289.4 | 101.4796.5 | |
| 400 | 448.6412.4 | 112.15103.1 | |
| 500 | 517.2 | 116.16103.4 | |
| 600 | 595.0 | 109.5099.2 | |
| 700 | 732.6666.2 | 104.6695.2 | |
| 800 | 766.2 | 118.8095.8 | |
| 900 | 907.6 | 131.16100.8 | |
2. Analytical Specificity/Interference:
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
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off concentration / lowest concentration of cross reactant that gives a positive result) x 100. Results are summarized below:
Cross Reactivity of Benzodiazepines and Metabolites- High Sensitivity 200 ng/mL Cutoff
| Benzodiazepine and metabolites | Lowest concentration producing a positive result (ng/mL) | Cross-reactivity (%) |
| --- | --- | --- |
| α-Hydroxyalprazolam | 110 | 182 |
| α-Hydroxytriazolam | 140 | 143 |
| Alprazolam | 100 | 200 |
| 7-Aminoclonazepam | 800 | 25 |
| 7-Aminoflunitrazepam | 225 | 89 |
| 7-Aminonitrazepam | 500 | 40 |
| Bromazepam | 300 | 67 |
| Chlordiazepoxide | 2000 | 10 |
| Clobazam | 450 | 44 |
| Clonazepam | 350 | 57 |
| Clorazepate | 100 | 200 |
| Delorazepam | 100 | 200 |
| Demoxepam | 1500 | 13 |
| Desalkylflurazepam (Norfludiazepam) | 110 | 182 |
| Diazepam | 80 | 250 |
| Estazolam | 115 | 174 |
| Flunitrazepam | 125 | 160 |
| Flurazepam | 70 | 286 |
| Lorazepam | 250 | 80 |
| Lorazepam glucuronide | 400 | 50 |
| Lormetazepam | 175 | 114 |
| Medazepam | 200 | 100 |
| Nitrazepam | 290 | 69 |
| Nordiazepam (Desmethyldiazepam) | 70 | 286 |
| Oxazepam | 200 | 100 |
| Oxazepam glucuronide | 350 | 57 |
| Prazepam | 140 | 143 |
| Temazepam | 130 | 154 |
| Temazepam glucuronide | 250 | 80 |
| Triazolam | 90 | 222 |
## Interference Testing of Structurally Unrelated Compounds
Interference from structurally unrelated compounds was evaluated by adding known amounts of each compound to urine samples containing near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of Oxazepam. Testing was performed in both
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qualitative and semiquantitative modes. The compounds listed in the table below did not cause any positive or negative interference at the concentrations shown:
High Sensitivity 200 ng/mL cutoff
| Structurally Unrelated Compounds | Tested Concentration (ng/mL) |
| --- | --- |
| 6-Acetyl Morphine | 100000 |
| 10,11 Dihydrocarbamazepine | 100000 |
| 11-nor-Δ9-THC-COOH | 100000 |
| Acetaminophen | 100000 |
| Acetylsalicylic Acid | 100000 |
| Amitriptyline | 75000 |
| Amoxicillin | 100000 |
| Amphetamine | 100000 |
| Benzoylecgonine | 100000 |
| Brompheniramine | 100000 |
| Buprenorphine | 100000 |
| Caffeine | 100000 |
| Captopril | 100000 |
| Cimetidine | 100000 |
| Codeine | 100000 |
| Desipramine | 100000 |
| Dextromethorphan | 100000 |
| Digoxin | 100000 |
| Diphenhydramine | 5000030000 |
| EDDP | 100000 |
| EMDP | 150003000 |
| Fentanyl | 100000 |
| Fluoxetine | 75000 |
| Fluphenazine | 75000 |
| Haloperidol | 100000 |
| Heroin | 100000 |
| Hydrocodone | 100000 |
| Hydromorphone | 100000 |
| Ibuprofen | 100000 |
| Levorphanol | 100000 |
| Levothyroxine | 100000 |
| Meperidine | 100000 |
| Methadone | 75000 |
| Methamphetamine | 100000 |
| Morphine | 100000 |
| Morhpine-3β-D-glucuronide | 100000 |
| Morhpine-6β-D-glucuronide | 100000 |
| Nalbuphine | 100000 |
| Nalorphine | 100000 |
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| Structurally Unrelated Compounds | Tested Concentration (ng/mL) |
| --- | --- |
| Naloxone | 100000 |
| Naltrexone | 100000 |
| Naproxen | 100000 |
| Nifedipine | 100000 |
| Oxaprozin | 5000 |
| Oxycodone | 100000 |
| Oxymorphone | 100000 |
| Perphenazine | 5000030000 |
| Phencyclidine | 10000090000 |
| Phenobarbital | 100000 |
| Procyclidine | 100000 |
| Propoxyphene | 100000 |
| Ranitidine | 100000 |
| Secobarbital | 100000 |
| Sertraline | 150007000 |
| Sulpiride | 100000 |
| Tapentadol | 100000 |
| Thioridazine | 100000 |
| Tramadol | 100000 |
| Triprolidine | 5000040000 |
| Verapamil | 100000 |
| Zolpidem | 5000040000 |
| Enalapril | 100000 |
| Salicyluric Acid | 100000 |
| Tolmetin | 100000 |
## Interference testing of endogenous and exogenous compounds
Potential interference from endogenous and exogenous compounds on recovery of Oxazepam was evaluated by adding known amounts of each compound into urine samples containing near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of Oxazepam for the 200 ng/mL cutoff. The compounds 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:
200 ng/mL Cutoffs
| Compounds | Tested Conc. (mg/dL) |
| --- | --- |
| Ascorbic Acid | 150 |
| Caffeine | 5 |
| Creatinine | 400 |
| Ethanol | 1000 |
| Galactose | 5 |
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| Compounds | Tested Conc. (mg/dL) |
| --- | --- |
| Glucose | 1000 |
| Hemoglobin | 150 |
| Human Serum Albumin | 200 |
| Ibuprofen | 10 |
| Oxalic acid | 50 |
| Riboflavin | 3 |
| Sodium Chloride | 1000 |
| Urea | 1000 |
Interference Testing of Specific Gravity and pH:
Drug free urine samples with specific gravity ranging in value from 1.002 to 1.029 were split and spiked with Oxazepam to final concentrations of 150 ng/mL and 250 ng/mL for 200 ng/mL cutoff. Samples were evaluated in qualitative and semi-quantitative modes. The following specific gravity did not cause any positive or negative interference: 1.002, 1.004, 1.005, 1.007, 1.010, 1.012, 1.014, 1.019, 1.023, 1.025 and 1.029.
Interference from pH was evaluated by adjusting the pH of urine samples containing near cutoff negative (150 ng/mL) and near cutoff positive (250 ng/mL) concentrations of Oxazepam for the 200 ng/mL cutoff. The following pH values did not cause any positive or negative interference: 3, 4, 5, 6, 7, 8, 9, 10 and 11.
3. Assay Reportable Range:
Not applicable.
4. 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.
5. Detection Limit:
Not applicable.
6. Assay Cut-Off:
Characterization of how the device performs analytically around the claimed cutoff concentration is described in the precision section, VII.A.1. above.
B Comparison Studies:
1. Method Comparison:
The method comparison study was performed in accordance with CLSI Guideline EP09-A3.
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One hundred and twenty-eight (128) samples were treated with β-glucuronidase reagent prior to analysis by the CEDIA™ Benzodiazepine Assay in both qualitative and semi-quantitative modes. The results were compared to LC-MS/MS where samples were also treated with β-glucuronidase.
The qualitative and semi-quantitative results are summarized in the tables below.
## Qualitative Mode Accuracy Study with LC-MS/MS as Reference Method High Sensitivity 200 ng/mL Cutoff
| Candidate Device Results | < 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 | 4*b | 13 | 55 |
| Negative | 54 | 2 | 0 | 0 |
Agreement among Positives: 68/68 = 100%
Agreement among Negative: 56/60 = 93%
## Semi-Quantitative Mode Accuracy Study with LC-MS/MS as Reference Method – High Sensitivity 200 ng/mL Cutoff
| Candidate Device Results | < 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 | 4*b | 12 | 55 |
| Negative | 54 | 2 | 1*b | 0 |
Agreement among Positives: 67/68 = 99%
Agreement among Negative: 56/60 = 93%
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*b Discordant sample results for high sensitivity 200 ng/mL cutoff
| Sample ID | EIA | | LC-MS/MS |
| --- | --- | --- | --- |
| | Qualitative | Semi-Quantitative | Total Benzodiazepine Parent Only (ng/mL) |
| CA160606-045*1 | Positive | Positive | 111 |
| CA170605-001*1 | Positive | Positive | 171 |
| CA160926-057*1 | Positive | Positive | 199 |
| CA180820-014*2 | Positive | Positive | 197 |
| CA170531-075*3 | Positive | Negative | 230 |
*1 These samples are discordant due to the presence of parent benzodiazepine and also benzodiazepine metabolites as follows: CA160606-045 contains 7-aminoclonazepam at 3155 ng/ml. CA170605-001 contains 7- aminoclonazepam at 560 ng/mL. CA160926-057 contains 7-aminoclonazepam at 411 ng/mL and 13 ng/mL of $\alpha$ hydroxyprazolam.
*2 Sample CA180820-014 is borderline negative by LC-MS/MS at 197 ng/ml compared to the 200 ng/ml cut-off.
*3 Sample CA170531-075 is borderline positive by LC-MS/MS at 230 ng/ml compared to the 200 ng/ml cut-off.
2. Matrix Comparison:
Not applicable. Urine is the only claimed matrix for the candidate device.
# C Clinical Studies:
1. Clinical Sensitivity:
Not applicable.
2. Clinical Specificity:
Not applicable.
3. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):
Not applicable.
# D Clinical Cut-Off:
Not applicable.
# Expected Values/Reference Range:
Not applicable.
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VIII Proposed Labeling:
The labeling supports the finding of substantial equivalence for this device.
IX Conclusion:
The submitted information in this premarket notification is complete and supports a substantial equivalence decision.
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