Glucose HK Gen.3; ONLINE DAT Methadone II; cobas pro integrated solutions

{0} FDA U.S. FOOD &amp; DRUG ADMINISTRATION # 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY ASSAY AND INSTRUMENT ## I Background Information: A 510(k) Number K253490 B Applicant Roche Diagnostics C Proprietary and Established Names Glucose HK Gen.3; ONLINE DAT Methadone II; cobas pro integrated solutions D Regulatory Information | Product Code(s) | Classification | Regulation Section | Panel | | --- | --- | --- | --- | | CFR | II | 21 CFR 862.2160 - Glucose Test System | CH - Clinical Chemistry | | DJR | II | 21 CFR 862.3620- Methadone Test System | CH - Clinical Chemistry | | JJE | I | 21 CFR 862.2160 - Discrete photometric chemistry analyzer for clinical use | CH - Clinical Chemistry | ## II Submission/Device Overview: A Purpose for Submission: Addition of previously cleared assays to a modified instrument platform B Measurand: Glucose and Methadone C Type of Test: Glucose: Quantitative photometric Methadone: Kinetic interaction of microparticles in a solution Food and Drug Administration 10903 New Hampshire Avenue Silver Spring, MD 20993-0002 www.fda.gov {1} III Intended Use/Indications for Use: A Intended Use(s): See Indications for Use below. B Indication(s) for Use: cobas pro integrated solutions is an automated analyzer, intended for running qualitative, semiquantitative, and quantitative clinical chemistry and immunochemistry assays as well as ion-selective measurements. Glucose HK Gen.3 is an in vitro test for the quantitative determination of glucose in human serum, plasma, urine and CSF on cobas c systems. Glucose measurements are used in the diagnosis and treatment of carbohydrate metabolism disorders including diabetes mellitus, neonatal hypoglycemia, and idiopathic hypoglycemia, and pancreatic islet cell tumors. ONLINE DAT Methadone II is an in vitro diagnostic test for the qualitative and semiquantitative detection of methadone in human urine on cobas c systems at a cutoff concentration of 300 ng/mL. Semiquantitative test results may be obtained that permit laboratories to assess assay performance as part of a quality control program. Semiquantitative assays are intended to determine an appropriate dilution of the specimen for confirmation by a confirmatory method such as gas chromatography/mass spectrometry (GC-MS). Methadone II provides only a preliminary analytical test result. A more specific alternate chemical method must be used in order to obtain a confirmed analytical result. GC-MS is the preferred confirmatory method. Clinical consideration and professional judgment should be applied to any drug of abuse test result, particularly when preliminary positive results are used. C Special Conditions for Use Statement(s): Rx - For Prescription Use Only D Special Instrument Requirements: cobas pro integrated solutions IV Device/System Characteristics: A Device Description: cobas pro integrated solutions The cobas pro integrated solutions is a fully automated, random-access, software-controlled system intended for in vitro quantitative and qualitative analysis of analytes in body fluids. The quantitative analysis also includes semi-quantitative. The cobas c 703 is being added to the cobas pro integrated solutions. K253490- Page 2 of 21 {2} K253490- Page 3 of 21 # Glucose HK Gen. 3 The reagent working solutions include: - R1: MES buffer: 5.0 mmol/L, pH 6.0; Mg²⁺, 24 mmol/L; ATP, ≥ 4.5 mmol/L; NADP, ≥ 7.0 mmol/L; preservative - R3: HEPES buffer: 200 mmol/L, pH 8.0; Mg²⁺, 4 mmol/L; HK (yeast), ≥ 300 μkat/L; G-6-PDH (E. coli), ≥ 300 μkat/L; preservative # ONLINE DAT Methadone II The reagent working solutions include: - R1: Conjugated methadone derivative; buffer; bovine serum albumin; 0.09 % sodium azide - R2: Microparticles attached to methadone antibody (mouse monoclonal); buffer; bovine serum albumin; 0.09 % sodium azide # B Principle of Operation: ## Glucose HK Gen. 3: Glucose is phosphorylated by hexokinase (HK) in the presence of adenosine triphosphate (ATP) and magnesium ions to produce glucose-6-phosphate (G-6-P) and adenosine diphosphate (ADP). Glucose-6-phosphate dehydrogenase (G-6-PDH) specifically oxidizes G-6-P to 6-phosphogluconate with the concurrent reduction of nicotinamide adenine dinucleotide (NAD) to nicotinamide adenine dinucleotide reduced (NADH). One micromole of NADH is produced for each micromole of glucose consumed. The NADH produced absorbs light at 340 nm and can be detected spectrophotometrically as an increased absorbance. The rate of NADPH formation during the reaction is directly proportional to the glucose concentration. ## ONLINE DAT Methadone II: The Methadone assay is based on the kinetic interaction of microparticles in a solution (KIMS) as measured by changes in light transmission. In the absence of sample drug, soluble drug conjugates bind to antibody-bound microparticles, causing the formation of particle aggregates. As the aggregation reaction proceeds in the absence of sample drug, the absorbance increases. When a urine sample contains the drug in question, this drug competes with the drug derivative conjugate for microparticle-bound antibody. Antibody bound to sample drug is no longer available to promote particle aggregation, and subsequent particle lattice formation is inhibited. The presence of sample drug diminishes the increasing absorbance in proportion to the concentration of drug in the sample. Sample drug content is determined relative to the value obtained for a known cutoff concentration of drug. # C Instrument Description Information: 1. Instrument Name: cobas pro integrated solutions 2. Specimen Identification: {3} The specimen is in a tube with a barcode label. The system identifies the specimen by scanning the barcode. This is the primary use case for sample identification. The system also supports manual entry of sample IDs to support the needs of the customer. 3. Specimen Sampling and Handling: Specimen sampling and handling procedures are analyte specific and documented in the respective reagent method sheets. 4. Calibration: The software of the cobas pure integrated solution automatically recommends calibration for all tests requiring calibration. Calibration methods and procedures are analyte specific and documented in the respective reagent method sheets. 5. Quality Control: Quality control procedures are analyte specific and documented in the respective reagent method sheets. V Substantial Equivalence Information: A Predicate Device Name(s): Glucose HK Gen. 3, cobas pro integrated solutions ONLINE DAT Methadone II B Predicate 510(k) Number(s): K191899, K021505 C Comparison with Predicate(s): Comparison with Glucose HK Gen. 3 | Device & Predicate Device(s): | K253490 | K191899 | | --- | --- | --- | | Device Trade Name | Glucose HK Gen.3 | Glucose HK Gen.3 | | General Device Characteristic Similarities | | | | Intended Use/Indications for Use | In vitro test for the quantitative determination of glucose in human serum, plasma, urine, and CSF. | Same | | General Device Characteristic Differences | | | | Instrument Platform | cobas c 703 in cobas pro integrated solutions | cobas c 503 in cobas pro integrated solutions | K253490- Page 4 of 21 {4} Comparison with ONLINE DAT Methadone II | Device & Predicate Device(s): | K253490 | K021505 | | --- | --- | --- | | Device Trade Name | ONLINE DAT Methadone II | ONLINE DAT Methadone II | | General Device Characteristic Similarities | | | | Intended Use/Indications for Use | In vitro test for the qualitative and semiquantitative detection of methadone in human urine at a cutoff concentration of 300 ng/mL. | Same | | General Device Characteristic Differences | | | | Instrument Platform | cobas c 703 in cobas pro integrated solutions | cobas c 503 in cobas pro integrated solutions | Comparison of the cobas pro integrated solutions | Device & Predicate Device(s): | K253490 | K191899 | | --- | --- | --- | | Device Trade Name | cobas pro integrated solutions | cobas pro integrated solutions | | General Device Characteristic Similarities | | | | Intended Use/Indications for Use | Intended for clinical chemistry and immunochemistry assays as well as ion-selective measurements | Same | | General Device Characteristic Differences | | | | Instrument platforms | cobas c 703 in cobas pro integrated solutions | cobas c 503 in cobas pro integrated solutions | VI Standards/Guidance Documents Referenced: - Clinical and Laboratory Standards Institute (CLSI) EP05-A3. Evaluation of Precision of Quantitative Measurement Procedures; Approved Guideline – Third Edition. - CLSI EP06 2nd Edition. Evaluation of the Linearity of Quantitative Measurement Procedures. - CLSI EP17-A2. Evaluation of Detection Capability for Clinical Laboratory Measurement Procedures; Approved Guideline – Second Edition. - IEC 61010-1 Edition 3.1 2017-01 consolidated version. Safety requirements for electrical equipment for measurement, control, and laboratory use – Part 1: General Requirements (including: Corrigendum 1). K253490- Page 5 of 21 {5} VII Performance Characteristics (if/when applicable): A Analytical Performance: 1. Precision/Reproducibility: Repeatability (within-run precision) and intermediate precision (within-laboratory precision) were evaluated following the recommendations in the CLSI guideline EP05-A3. Glucose HK Gen.3: Repeatability and Intermediate Precision The precision of the Glucose HK Gen.3 assay was evaluated on one candidate device using three lots of reagent. The protocol consisted of testing two sample replicates per run, two runs per day for 21 days. Repeatability and within-lab precision were calculated. The samples were randomized in each run separately. Two serum-based controls and five levels of individual serum samples were used (Serum 1 was diluted, Serum 2 was native, and Serum 3-5 were spiked). The protocol was repeated for each application (serum, urine, CSF). For urine, individual samples were also used (Urine 1 was native and Urine 2-5 were spiked) while for CSF, pooled samples were used (CSF 1-2 were diluted, CSF 3 was native, and CSF 4-5 were spiked). The results for a representative reagent lot are summarized in the table below. Summary of precision results – Glucose HK Gen.3 | Sample | N | Mean (mg/dL) | Repeatability | | Mean (mg/dL) | Within-Laboratory Precision | | | --- | --- | --- | --- | --- | --- | --- | --- | | | | | SD (mg/dL) | CV (%) | | SD (mg/dL) | CV (%) | | Serum Control 1 | 84 | 104 | 0.400 | 0.4 | 104 | 0.782 | 0.8 | | Serum Control 2 | 84 | 254 | 1.112 | 0.4 | 254 | 1.910 | 0.8 | | Serum 1 | 84 | 4.0 | 0.095 | 2.4 | 4.0 | 0.104 | 2.6 | | Serum 2 | 84 | 68 | 0.371 | 0.5 | 68 | 0.494 | 0.7 | | Serum 3 | 84 | 112 | 0.567 | 0.5 | 112 | 1.072 | 1 | | Serum 4 | 84 | 389 | 1.484 | 0.4 | 389 | 1.874 | 0.5 | | Serum 5 | 84 | 740 | 3.837 | 0.5 | 740 | 5.044 | 0.7 | | Sample | N | Mean (mg/dL) | Repeatability | | Mean (mg/dL) | Within-Laboratory Precision | | | --- | --- | --- | --- | --- | --- | --- | --- | | | | | SD (mg/dL) | CV (%) | | SD (mg/dL) | CV (%) | | Urine Control 1 | 84 | 27 | 0.267 | 1 | 27 | 0.297 | 1.1 | | Urine Control 2 | 84 | 290 | 1.748 | 0.6 | 290 | 2.414 | 0.8 | | Urine 1 | 84 | 3.8 | 0.263 | 6.9 | 3.8 | 0.263 | 6.9 | | Urine 2 | 84 | 14 | 0.207 | 1.5 | 14 | 0.225 | 1.6 | | Urine 3 | 84 | 70 | 0.407 | 0.6 | 70 | 0.567 | 0.8 | K253490- Page 6 of 21 {6} | Sample | N | Mean (mg/dL) | Repeatability | | Mean (mg/dL) | Within-Laboratory Precision | | | --- | --- | --- | --- | --- | --- | --- | --- | | | | | SD (mg/dL) | CV (%) | | SD (mg/dL) | CV (%) | | Urine 4 | 84 | 375 | 1.802 | 0.5 | 375 | 2.216 | 0.6 | | Urine 5 | 84 | 730 | 3.315 | 0.5 | 730 | 4.090 | 0.6 | | Sample | N | Mean (mg/dL) | Repeatability | | Mean (mg/dL) | Within-Laboratory Precision | | | --- | --- | --- | --- | --- | --- | --- | --- | | | | | SD (mg/dL) | CV (%) | | SD (mg/dL) | CV (%) | | CSF Control 1 | 84 | 57 | 0.303 | 0.5 | 57 | 0.368 | 0.6 | | CSF Control 2 | 84 | 30 | 0.231 | 0.8 | 30 | 0.254 | 0.9 | | CSF 1 | 84 | 4.5 | 0.254 | 5.7 | 4.5 | 0.265 | 5.9 | | CSF 2 | 84 | 38 | 0.234 | 0.6 | 38 | 0.335 | 0.9 | | CSF 3 | 84 | 71 | 0.335 | 0.5 | 71 | 0.402 | 0.6 | | CSF 4 | 84 | 375 | 1.748 | 0.5 | 375 | 2.774 | 0.7 | | CSF 5 | 84 | 726 | 3.081 | 0.4 | 726 | 4.234 | 0.6 | ONLINE DAT Methadone II The precision of the ONLINE DAT Methadone II test was evaluated on one candidate device using three reagent lots using both the qualitative and the semi-quantitative mode. The protocol consisted of testing two sample replicates per run, two runs per day for 21 days. Seven human samples and four control samples were used. The samples with the concentrations provided in the tables below were prepared by spiking drug into negative urine. The analyte concentration was confirmed using a validated method. The results for a representative reagent lot are summarized in the tables below. Summary of Within-Lab Precision – Qualitative (300 ng/mL cutoff) | Sample | N | Negative | Positive | | --- | --- | --- | --- | | Sample -100% (zero) | 84 | 84 | 0 | | Sample -75% | 84 | 84 | 0 | | Sample -50% | 84 | 84 | 0 | | DAT1N | 84 | 84 | 0 | | DATCN | 84 | 84 | 0 | | Cutoff | 84 | 34 | 50 | | DAT1P | 84 | 0 | 84 | | DATCP | 84 | 0 | 84 | | Sample +50% | 84 | 0 | 84 | | Sample +75% | 84 | 0 | 84 | | Sample +100% | 84 | 0 | 84 | K253490- Page 7 of 21 {7} Summary of Precision Results – Semi-Quantitative (300 ng/mL cutoff) | Sample | N | Mean (ng/mL) | Repeatability | | Mean (ng/mL) | Within-Laboratory Precision | | | --- | --- | --- | --- | --- | --- | --- | --- | | | | | SD (ng/mL) | CV (%) | | SD (ng/mL) | CV (%) | | Sample -100% (zero) | 84 | 15.1 | 13.7 | 90.6 | 15.3 | 16.1 | 105.0 | | Sample -75% | 84 | 113 | 6.12 | 5.4 | 113 | 9.97 | 8.8 | | Sample -50% | 84 | 170 | 5.94 | 3.5 | 165 | 12.3 | 7.5 | | DAT1N | 84 | 238 | 6.82 | 2.9 | 242 | 10.9 | 4.5 | | DATCN | 84 | 239 | 7.69 | 3.2 | 239 | 11.4 | 4.8 | | Cutoff | 84 | 295 | 10.2 | 3.5 | 296 | 22.5 | 7.6 | | DAT1P | 84 | 407 | 13.2 | 3.2 | 407 | 18.4 | 4.5 | | DATCP | 84 | 393 | 12.1 | 3.1 | 401 | 18.2 | 4.5 | | Sample +50% | 84 | 489 | 23.2 | 4.7 | 505 | 31.5 | 6.2 | | Sample +75% | 84 | 579 | 20.3 | 3.5 | 579 | 30.6 | 5.3 | | Sample +100% | 84 | 652 | 28.7 | 4.4 | 635 | 36.7 | 5.8 | 2. Linearity: The linearity studies were conducted following the recommendations in CLSI guideline EP06-Ed2. Glucose HK Gen.3 Linearity of the Glucose HK Gen.3 assay was assessed on candidate device in one run using three reagent lots, measuring four replicates per sample. High analyte concentration of human serum, plasma (collected in K2EDTA tubes), urine and CSF samples were diluted to obtain nine levels spanning the measuring range. The results of the linear regression analysis for a representative reagent lot are summarized in the table below. | Sample | Tested Range (mg/dL) | Linear Regression Data | Claimed Linear Range (mg/dL) | | --- | --- | --- | --- | | Serum | 1.82-777 | y = 0.997x - 0, R² = 0.9998 | 2-750 | | Plasma | 0.26-786 | y = 1.001x - 0, R² = 0.9999 | 2-750 | | Urine | 1.18-760 | y = 1.009x -0, R² = 1.0000 | 2-750 | | CSF | 0.99-773 | y = 1.003x - 0, R² = 1.0000 | 2-750 | The results demonstrated linearity of the claimed measuring range for serum, plasma, urine, and CSF (2.0 to 750 mg/dL or 0.11 to 41.6 mmol/L). ONLINE DAT Methadone II Linearity of the semi-quantitative ONLINE DAT Methadone II test was assessed on the candidate device in one run using one reagent lot, measuring four replicates per sample. A dilution series was prepared from human urine sample pools (spiked with methadone and K253490- Page 8 of 21 {8} diluted with human urine) to obtain nine levels that span the claimed range. The results of the linearity/recovery study are summarized in the table below. | Expected Value (ng/mL) | Observed Value (ng/mL) | Absolute Deviation (ng/mL) | % Recovery | | --- | --- | --- | --- | | 27.1 | 37.0 | 9.81 | | | 54.1 | 60.8 | 6.41 | | | 113 | 113 | -0.203 | | | 226 | 225 | -1.41 | | | 451 | 451 | | 100.0% | | 677 | 688 | | 101.6% | | 902 | 997 | | 110.5% | | 1128 | 1098 | | 97.3% | | 1353 | 1325 | | 97.9% | | 1579 | 1521 | | 96.3% | | 1804 | 1816 | | 100.6% | | 2030 | 1958 | | 96.5% | | 2255 | 2255 | | 100.0% | ## Dilution Recovery Studies ### Glucose HK Gen. 3 A dilution study was performed and the data support the rerun function 1:2 dilution claim in the Method Sheet. Five samples each (serum, urine, and CSF) were prepared using native and spiked samples and measured with the automatic rerun function of the candidate device. ### ONLINE DAT Methadone II Patient values should not be generated with diluted samples. ## 3. Analytical Specificity/Interference: ### Endogenous Interference The purpose of this study was to evaluate endogenous substances for potential interference when using candidate device. ### Glucose HK Gen. 3 The effect on the quantitation of analyte in the presence of potentially interfering endogenous substances using the Glucose HK Gen.3 assay was determined on the candidate device using serum, urine, and CSF samples. Glucose levels of approximately $38\mathrm{mg/dL}$ and $220\mathrm{mg/dL}$ were tested. Significant interference was defined as follows: - Serum/Plasma: bias between the samples with and without interferent is $&gt; \pm 7\mathrm{mg/dL}$ at glucose concentrations $\leq 70\mathrm{mg/dL}$ and $&gt; \pm 10\%$ at glucose concentrations $&gt;70\mathrm{mg/dL}$ - Urine: bias between the samples with and without interferent is $&gt; \pm 2.0\mathrm{mg/dL}$ at glucose concentrations $\leq 19.8\mathrm{mg/dL}$ and $&gt; \pm 10\%$ at glucose concentrations $&gt;19.8\mathrm{mg/dL}$ K253490- Page 9 of 21 {9} - CSF: bias between the samples with and without interferent is $\geq 4\mathrm{mg / dL}$ at glucose concentrations $\leq 39.6\mathrm{mg / dL}$ and $&gt; \pm 10\%$ at glucose concentrations $&gt;39.6\mathrm{mg / dL}$ The summary of results is presented in the table below. | Potential Interferent | | Highest Concentration Tested Without Significant Interference | | --- | --- | --- | | Serum | Albumin | 62.3 g/L | | | Conjugated Bilirubin | 64 mg/dL | | | Unconjugated Bilirubin | 77 mg/dL | | | Hemolysis | 1213 mg/dL | | | IgG | 65.1 g/L | | | Lipemia | 1102 mg/dL | | Urine | Albumin | 2.70 g/L | | | Calcium | 12.6 mmol/L | | | Citrate | 12.0 mmol/L | | | Creatinine | 89.7 mmol/L | | | Hemolysis | 876 mg/dL | | | IgG | 1.10 g/L | | | Magnesium | 27.3 mmol/L | | | Oxalate | 2.70 mmol/L | | | Phosphate | 137 mmol/L | | | Urea | 1825 mmol/L | | | Uric Acid | 6.0 mmol/L | | | Urobilinogen | 0.25 mmol/L | | CSF | Conjugated Bilirubin | 66 mg/dL | | | Hemolysis | 1201 mg/dL | # ONLINE DAT Methadone II The effects of potentially interfering endogenous substances on the ONLINE DAT Methadone II test were determined on one candidate device, using two reagent lots, one sample per concentration level, two human samples, in one run, with three replicates per sample. Methadone levels of approximately $225\mathrm{ng / mL}$ and $375\mathrm{ng / mL}$ (corresponding to $\pm 25\%$ of the cutoff concentration) were tested. Significant interference was defined as crossover from the expected result for each control level. The summary of results is presented in the table below. | Potential Interferent | Concentration tested | Spiked Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (Negative Control -25% Cutoff,) | 375 ng/mL (Positive Control, +25% Cutoff) | | Acetone | 1000 mg/dL | Neg | Pos | | Ascorbic acid | 1500 mg/dL | Neg | Pos | | Albumin | 500 mg/dL | Neg | Pos | | Bilirubin | 25 mg/dL | Neg | Pos | | Calcium | 133 mg/dL | Neg | Pos | K253490- Page 10 of 21 {10} K253490- Page 11 of 21 | Potential Interferent | Concentration tested | Spiked Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (Negative Control -25% Cutoff,) | 375 ng/mL (Positive Control, +25% Cutoff) | | Creatinine | 500 mg/dL | Neg | Pos | | Ethanol | 1000 mg/dL | Neg | Pos | | Glucose | 2000 mg/dL | Neg | Pos | | Hemoglobin | 750 mg/dL | Neg | Pos | | IgG | 110 mg/dL | Neg | Pos | | Magnesium | 238 mg/dL | Neg | Pos | | NaCl | 5800 mg/dL | Neg | Pos | | Oxalate | 200 mg/dL | Neg | Pos | | Phosphate | 2028 mg/dL | Neg | Pos | | Sodium citrate, dihydrate | 323.51 mg/dL | Neg | Pos | | Urea | 6000 mg/dL | Neg | Pos | | Uric Acid | 100.86 mg/dL | Neg | Pos | | Urobilinogen | 14.82 mg/dL | Neg | Pos | | pH | 3.9 – 9.1 | Neg | Pos | | Specific Gravity | 1.001-1.035 g/mL | Neg | Pos | ## Exogenous Interference The purpose of these studies was to evaluate drugs for potential interference when using the Glucose HK Gen.3 assay and the ONLINE DAT Methadone II test on the candidate device. The acceptance criteria were recovery within $100 \pm 10\%$, except methadone where significant interference was defined as crossover from the expected result for each control level. ## Glucose HK Gen. 3 The effect on the quantitation of analyte in the presence of potentially interfering drugs using the Glucose HK Gen.3 assay was determined on the candidate device using serum, urine and CSF samples. Glucose levels of approximately $40\mathrm{mg/dL}$ and $220\mathrm{mg/dL}$ were tested. The summary of results is presented in the table below. | Potential Interferent | | Highest Concentration without Interference [mg/L] | | --- | --- | --- | | Serum | N-Acetylcysteine | 150 | | | Phenylbutazone | 321 | | | Acetylsalicylic acid | 30 | | | Ampicillin-Na | 75 | | | Ascorbic acid | 52.5 | | | Cefoxitin | 750 | | | Doxycyclin | 18 | | | Heparin | 3300 IU/L | | | Levodopa | 7.5 | | | Methyldopa | 22.5 | {11} K253490- Page 12 of 21 | Potential Interferent | | Highest Concentration without Interference [mg/L] | | --- | --- | --- | | | Metronidazole | 123 | | | Rifampicin | 48 | | | Acetaminophen | 156 | | | Cyclosporine | 1.8 | | | Ibuprofen | 219 | | | Theophylline | 60 | | Urine | Acetaminophen | 3000 | | | N-Acetylcysteine | 10 | | | Ascorbic acid | 4000 | | | Cefoxitin | 12000 | | | Gentamycine sulfate | 400 | | | Levodopa | 1000 | | | Methyldopa | 2000 | | | Ofloxacin | 900 | | | Ibuprofen | 4000 | | | Phenazopyridine | 300 | | | Salicyluric acid | 100 | | | Tetracycline | 100 | ## ONLINE DAT Methadone II The effects of potential drug interference on the ONLINE DAT Methadone II assay were determined on one candidate device using urine samples. Urine samples containing methadone at positive and negative levels (a final methadone concentration of 225 ng/mL and 375 ng/mL (±25% of the cutoff concentration)) were prepared. In case interference was detected, the interferent was diluted and tested until no more interference was observed. The maximum drug concentration which did not interfere is presented in the table below. | Potential Interferent | Drug Concentration (ng/mL) | Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (-25% Cutoff, Negative Control) | 375 ng/mL (+25% Cutoff, Positive Control) | | Acetaminophen | 3000000 | Neg | Pos | | Acetyl cysteine | 10000 | Neg | Pos | | Acetylsalicylic acid | 100000 | Neg | Pos | | Aminopyrine | 100000 | Neg | Pos | | Amitriptyline | 100000 | Neg | Pos | | Amobarbital | 100000 | Neg | Pos | | d-Amphetamine | 100000 | Neg | Pos | | l-Amphetamine | 100000 | Neg | Pos | | Ampicillin | 100000 | Neg | Pos | {12} K253490- Page 13 of 21 | Potential Interferent | Drug Concentration (ng/mL) | Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (-25% Cutoff, Negative Control) | 375 ng/mL (+25% Cutoff, Positive Control) | | Ascorbic acid | 4000000 | Neg | Pos | | Aspartame | 100000 | Neg | Pos | | Atropine | 100000 | Neg | Pos | | Atropine | 100000 | Neg | Pos | | Atropine | 100000 | Neg | Pos | | Benzocaine | 100000 | Neg | Pos | | Benzoylecgonine | 100000 | Neg | Pos | | Benzphetamine | 100000 | Neg | Pos | | Butabarbital | 100000 | Neg | Pos | | Caffeine | 100000 | Neg | Pos | | Calcium dobesilate | 100000 | Neg | Pos | | Calcium hypochlorite | 100000 | Neg | Pos | | Carbamazepine | 100000 | Neg | Pos | | Cefoxitin | 12000000 | Neg | Pos | | Chlordiazepoxide | 100000 | Neg | Pos | | Chloroquine | 100000 | Neg | Pos | | Chlorpheniramine | 100000 | Neg | Pos | | Cocaine | 100000 | Neg | Pos | | Codeine | 100000 | Neg | Pos | | Cotinine | 100000 | Neg | Pos | | Cyclobenzaprine | 100000 | Neg | Pos | | Cyproheptadine | 100000 | Neg | Pos | | Desipramine | 100000 | Neg | Pos | | Dextrometorphan | 100000 | Neg | Pos | | Dextropropoxyphene | 100000 | Neg | Pos | | Diazepam | 100000 | Neg | Pos | | Diphenhydramine | 100000 | Neg | Pos | | Diphenylhydantoin | 100000 | Neg | Pos | | Disopyramide | 100000 | Neg | Pos | | Disopyramide | 1000000 | Neg | Pos | | Dopamine | 100000 | Neg | Pos | | Doxepin | 100000 | Neg | Pos | | Doxylamine | 100000 | Neg | Pos | | d-Phenylpropanolamine | 100000 | Neg | Pos | {13} K253490- Page 14 of 21 | Potential Interferent | Drug Concentration (ng/mL) | Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (-25% Cutoff, Negative Control) | 375 ng/mL (+25% Cutoff, Positive Control) | | d-Pseudoephedrine | 100000 | Neg | Pos | | Ecgonine | 100000 | Neg | Pos | | Ecgonine methyl ester | 100000 | Neg | Pos | | EDDP | 100000 | Neg | Pos | | EMDP | 100000 | Neg | Pos | | d-Ephedrine | 100000 | Neg | Pos | | dl-Ephedrine | 100000 | Neg | Pos | | l-Ephedrine | 100000 | Neg | Pos | | Epinephrine | 100000 | Neg | Pos | | Erythromycin | 100000 | Neg | Pos | | Estriol | 100000 | Neg | Pos | | Fenoprofen | 100000 | Neg | Pos | | Fluoxetine | 100000 | Neg | Pos | | Furosemide | 100000 | Neg | Pos | | Gentamicin sulfate | 400000 | Neg | Pos | | Gentisic acid | 100000 | Neg | Pos | | Glutethimide | 100000 | Neg | Pos | | Guaiacol glycerol ether | 100000 | Neg | Pos | | Haloperidol | 100000 | Neg | Pos | | Hydrochlorothiazide | 100000 | Neg | Pos | | Ibuprofen | 4000000 | Neg | Pos | | Imipramine | 100000 | Neg | Pos | | Isoproterenol | 100000 | Neg | Pos | | Ketamine | 100000 | Neg | Pos | | Levodopa | 1000000 | Neg | Pos | | Lidocaine | 100000 | Neg | Pos | | l-Pseudoephedrine | 100000 | Neg | Pos | | LSD | 100000 | Neg | Pos | | Maprotiline | 100000 | Neg | Pos | | MDA | 100000 | Neg | Pos | | MDMA | 100000 | Neg | Pos | | Melanin | 100000 | Neg | Pos | | Meperidine | 100000 | Neg | Pos | | d-Methamphetamine | 100000 | Neg | Pos | {14} K253490- Page 15 of 21 | Potential Interferent | Drug Concentration (ng/mL) | Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (-25% Cutoff, Negative Control) | 375 ng/mL (+25% Cutoff, Positive Control) | | l-Methamphetamine | 100000 | Neg | Pos | | Methaqualone | 100000 | Neg | Pos | | Methyldopa | 2000000 | Neg | Pos | | Methylphenidate | 100000 | Neg | Pos | | Mianserin | 100000 | Neg | Pos | | Morphine | 100000 | Neg | Pos | | Naloxone | 100000 | Neg | Pos | | Naltrexone | 100000 | Neg | Pos | | Naproxen | 100000 | Neg | Pos | | Niacinamide | 100000 | Neg | Pos | | Nicotine | 100000 | Neg | Pos | | Nordiazepam | 100000 | Neg | Pos | | Nordoxepin | 100000 | Neg | Pos | | Norethindrone | 100000 | Neg | Pos | | l-Norpseudoephedrine | 100000 | Neg | Pos | | Nortriptyline | 100000 | Neg | Pos | | Ofloxacin | 90000 | Neg | Pos | | Orphenadrine | 100000 | Neg | Pos | | Oxazepam | 100000 | Neg | Pos | | Penicillin G | 100000 | Neg | Pos | | Pentobarbital | 100000 | Neg | Pos | | Perphenazine | 100000 | Neg | Pos | | Phenazopyridine | 300000 | Neg | Pos | | Phencyclidine | 100000 | Neg | Pos | | Phenetylamine | 100000 | Neg | Pos | | Phenobarbital | 100000 | Neg | Pos | | Phenothiazine | 100000 | Neg | Pos | | Phentermine | 100000 | Neg | Pos | | Phenylbutazone | 100000 | Neg | Pos | | Phenylpropanolamine | 100000 | Neg | Pos | | Procaine | 100000 | Neg | Pos | | Protriptyline | 100000 | Neg | Pos | | Quetiapine-Carboxylic acid | 500000 | Neg | Pos | | Quetiapine-Fumarat | 750000 | Neg | Pos | {15} | Potential Interferent | Drug Concentration (ng/mL) | Methadone Concentration | | | --- | --- | --- | --- | | | | 225 ng/mL (-25% Cutoff, Negative Control) | 375 ng/mL (+25% Cutoff, Positive Control) | | Quetiapine-Sulfoxid | 500000 | Neg | Pos | | Quinidine | 100000 | Neg | Pos | | Quinine | 100000 | Neg | Pos | | Salicyluric acid | 6000000 | Neg | Pos | | Secobarbital | 100000 | Neg | Pos | | Sulindac | 100000 | Neg | Pos | | Tetracycline | 300000 | Neg | Pos | | Tetracycline | 300000 | Neg | Pos | | Tetracycline | 300000 | Neg | Pos | | Tetrahydrozoline | 100000 | Neg | Pos | | THC-9-carboxylic acid | 100000 | Neg | Pos | | Tramadol | 20000 | Neg | Pos | | Trifluoperazine | 100000 | Neg | Pos | | Tyramine | 100000 | Neg | Pos | | Verapamil | 100000 | Neg | Pos | ONLINE DAT Methadone II Cross Reactivity The specificity of the ONLINE DAT Methadone II assay for structurally similar compounds was determined by preparing concentration series for each of the compounds listed, using measured signals of the different potential cross reactant concentration to determine the approximate quantity of each compound that is equivalent in assay reactivity to the 300 ng/mL assay cutoff on one candidate device. Cross Reactivity was tested for the Semi-Quantitative application only. The percent cross-reactivity was calculated from the ratio of methadone concentration at cutoff level and the calculated cross reactant concentration, that yields the same signal. Structurally related pharmaceutical compounds: | Potential Interferent | Cross reactant units [ng/mL] equivalent to 300 ng/mL methadone | Cross-reactivity (%) | | --- | --- | --- | | Vortioxetine | 6531 | 4.59 | | LUAA34443 | 3914 | 7.66 | | Cyamemazine | 7282 | 4.12 | | Methotrimeprazine (Levomepromazine) | 8639 | 3.47 | | Chlorpromazine | 30891 | 0.97 | | Thiothixene | 77558 | 0.39 | | Promazine | 215852 | 0.14 | K253490- Page 16 of 21 {16} K253490- Page 17 of 21 | Potential Interferent | Cross reactant units [ng/mL] equivalent to 300 ng/mL methadone | Cross-reactivity (%) | | --- | --- | --- | | Clomipramine | 180534 | 0.17 | | Thioridazine | 206275 | 0.15 | | Chlorprothixene | 660667 | 0.045 | | Promethazine | 403915 | 0.074 | | Trimipramine | 572980 | 0.052 | ## 4. Detection Limit and Assay Reportable Range: The Limit of Blank (LoB), Limit of Detection (LoD) and Limit of Quantitation (LoQ) were determined in accordance with the CLSI EP17-A2 guideline. The LoB was determined as the 95th percentile of measurements of blank samples. For determination of the LoB for the Glucose HK Gen. 3 assay, one analyte-free sample was measured with three reagent lots in six runs, distributed over three days, 10 replicates per run, on one candidate device. In total, 60 determinations of analyte free samples were obtained. Data analysis was based on determination of the 95th percentile of the 60 measured values. In this design (n=60) the 95th percentile was the average of the 57th and 58th value. This process was repeated for each application (serum/plasma, urine, CSF). The LoD was determined as the lowest amount of analyte in a sample that can be detected with a 95% probability. For determination of the LoD for the Glucose HK Gen.3 assay, five samples with low analyte content were measured using three reagent lots with two-fold determination per run on one candidate device. Six runs distributed over three days on one instrument were performed. The LoD is defined as the concentration, at which there is a 95% probability that a sample contains analyte. LoD = LoB + 1.653 x SDtotal. This process was repeated for each application (serum/plasma, urine, CSF). The LoQ was determined as the lowest concentration of analyte which can be quantified with a maximum CV of 20% for the Glucose HK Gen.3 assay. Five samples with low analyte concentration were measured using three reagent lots on one candidate device. These samples were tested in one run per day over five days, five replicates per run for each LoQ sample (n=25 per sample). This process was repeated for each application (serum/plasma, urine, CSF). The results support the claimed LoB, LoD and LoQ for the Glucose HK Gen.3 assay which are unchanged from the predicate device (k191899) and are summarized in the table below: | Sample Type | Claimed LoB | Claimed LoD | Claimed LoQ | | --- | --- | --- | --- | | Serum/Plasma | 0.350 mg/dL (0.0194 mmol/L) | 0.486 mg/dL (0.0270 mmol/L) | 1.517 mg/dL (0.0842 mmol/L) | | Urine | 0.776 mg/dL (0.0431 mmol/L) | 1.160 mg/dL (0.0644 mmol/L) | 1.160 mg/dL (0.0644 mmol/L) | | CSF | 0.337 mg/dL (0.0187 mmol/L) | 0.506 mg/dL (0.0281 mmol/L) | 1.454 mg/dL (0.0807 mmol/L) | {17} 5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods): The traceability information for Glucose HK Gen. 3 assay was provided in K191899 and is unchanged. The traceability for ONLINE DAT Methadone II assay is unchanged from K220134. On-board Reagent Stability: Glucose HK Gen. 3 A reagent stability study confirmed that the Glucose HK Gen.3 reagent is stable for 26 weeks on-board the candidate device. ONLINE DAT Methadone II A reagent stability study confirmed that the ONLINE DAT Methadone II reagent is stable for 26 weeks on-board the candidate device. 6. Assay Cut-Off: Glucose HK Gen. 3 Not applicable. ONLINE DAT Methadone II Characterization of how the device performs analytically around the claimed cutoff concentration is described in the method comparison section VII.B.1. below. B Comparison Studies: 1. Method Comparison with Predicate Device: Glucose HK Gen. 3 Method comparison studies were performed for serum, urine and CSF using the Glucose HK Gen.3 assay on the candidate device versus the predicate device to assess the differences between the two test systems. Native human samples were tested in singlicate on each test system. Less than 10% of these samples were spiked or diluted to cover the measuring range. The results were analyzed using Passing-Bablok and linear regression analyses. The process was repeated for each application (serum, urine, CSF). A summary of results is presented in the table below. | Sample Type | N | Sample Concentration Range (mg/dL) | Passing-Bablok (Slope & Intercept) and Correlation (Kendall tau (t)) | Linear Regression (Slope & Intercept) and Correlation (Pearson (r)) | | --- | --- | --- | --- | --- | | Serum | 74 | 3.60 -724 | y = 0.995x - 0.0968 mg/dL t = 0.990 | y = 0.999x - 0.216 mg/dL r = 1.000 | K253490- Page 18 of 21 {18} | Sample Type | N | Sample Concentration Range (mg/dL) | Passing-Bablok (Slope & Intercept) and Correlation (Kendall tau (t)) | Linear Regression (Slope & Intercept) and Correlation (Pearson (r)) | | --- | --- | --- | --- | --- | | Urine | 67 | 2.04-730 | y = 0.998x + 0.209 mg/dL t = 0.950 | y = 1.001x + 0.214 mg/dL r = 1.000 | | CSF | 75 | 2.78- 730 | y = 0.987x - 0.234 mg/dL t = 0.984 | y = 0.989x - 0.259 mg/dL r = 1.000 | ## ONLINE DAT Methadone II Eighty seven (87) negative urine samples were evaluated with the ONLINE DAT Methadone II assay on the candidate device. 97.7 % of these normal urines were negative relative to the 300 ng/mL cutoff. Seventy nine (79) positive urine samples were evaluated with the ONLINE DAT Methadone II assay. 97.5 % of these samples were positive relative to the 300 ng/mL cutoff. Samples were evaluated using both the qualitative and the semi-quantitative modes and candidate device results were compared to GC-MS values. No differences were observed in the results, and results from the semi-quantitative mode are shown below. Semi-Quantitative Accuracy Study (cutoff = 300 ng/mL) | | Negative samples 0-150 | GC-MS values (ng/mL) | | | | | --- | --- | --- | --- | --- | --- | | | | Near cutoff | | Positive samples | | | | | 150-299 | 300-450 | >450 | | | Candidate Device | + | 0 | 2 | 15 | 62 | | | - | 59 | 26 | 2 | 0 | ## 2. Matrix Comparison: ### Glucose HK Gen. 3 To support the use of the Glucose HK Gen.3 assay on the candidate device with different sample matrices, a matrix comparison study was conducted by comparing values obtained from samples drawn into serum and plasma collection tubes. Some samples were spiked and/or diluted to cover the measuring range. The result of the serum sample was compared with the result of each plasma sample from the same donor. 43 serum/plasma pairs were tested for each type of anticoagulant in one run, in singlicate. The results were evaluated using Passing-Bablok regression analysis. A summary of results is presented in the table below. | Comparison | Slope | Intercept (mg/dL) | Correlation Coefficient | Concentration of Samples (mg/dL) | | --- | --- | --- | --- | --- | | Serum vs. Serum/gel separation tube | 1.007 | -0.432 | 1.000 | 3.17-737 | | Serum vs. K2-EDTA plasma | 1.003 | -0.0771 | 0.999 | 3.17-737 | | Serum vs. Li-Heparin plasma | 1.031 | -1.59 | 0.999 | 3.17-737 | K253490- Page 19 of 21 {19} | Comparison | Slope | Intercept (mg/dL) | Correlation Coefficient | Concentration of Samples (mg/dL) | | --- | --- | --- | --- | --- | | Serum vs. NaF/K-Oxalate plasma | 1.025 | -1.25 | 1.000 | 3.17-737 | | Serum vs. NaF/Na2-EDTA plasma | 1.037 | -1.78 | 0.999 | 3.17-737 | | Serum vs. KF/Na2-EDTA plasma | 1.028 | -0.587 | 0.999 | 3.17-737 | | Serum vs. NaF/Citrate/Na2-EDTA plasma | 1.005 | 1.87 | 0.999 | 2.88-730 | ## C Clinical Studies: 1. Clinical Sensitivity: Not applicable. 2. Clinical Specificity: Not applicable. 3. Clinical Cut-Off Not applicable. 4. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable): Not applicable. ## D Expected Values/Reference Range: Expected reference ranges for the assays are as follows: Glucose HK Gen.3 | Expected Values | | | | --- | --- | --- | | Sample Type | | Reference Range | | Serum, plasma | Adults | 74-106 mg/dL | | | 60-90 years | 82-115 mg/dL | | | > 90 years | 75-121 mg/dL | | | Children | 60-100 mg/dL | | | Neonates (1 day) | 40-60 mg/dL | | | Neonates (> 1 day) | 50-80 mg/dL | | Urine | 24-hour urine | < 2.78 mmol/24 h (<0.5 g/24 h) | | | Random urine | 1-15 mg/dL | | CSF | Children | 60-80 mg/dL | | | Adults | 40-70 mg/dL | K253490- Page 20 of 21 {20} Reference: Tietz NW, ed. Clinical Guide to Laboratory Tests, 4th ed. Philadelphia: WB Saunders Co 2006;444-451. ## ONLINE DAT Methadone II ### Qualitative assay Results of this assay distinguish preliminary positive (≥ 300 ng/mL) from negative samples only. The amount of drug detected in a preliminary positive sample cannot be estimated. ### Semiquantitative assay Results of this assay yield only approximate cumulative concentrations of the drug and its metabolites. ## E Other Supportive Instrument Performance Characteristics Data: 1. Carry-over studies were reviewed and found to be acceptable. 2. Electrical safety and electromagnetic compatibility (EMC) testing were performed, and the system was found to be acceptable. 3. Software and cybersecurity documentation was reviewed and found to be acceptable. ## VIII Proposed Labeling: The labeling supports or 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. K253490- Page 21 of 21