1708058008190c503spV3.

MDN2
ONLINE DAT Methadone II
Order information
Analyzer(s) on which cobas c pack(s) can be used
08058008190 ONLINE DAT Methadone II (850 tests) System‑ID 2088 001 cobas c 303, cobas c 503
03304671190 Preciset DAT Plus I calibrator CAL 5 Code 20435
07978766190 Serum DAT Control Low (ACQ Partner Channel*)
07978740190 Serum DAT Control High (ACQ Partner Channel*)
08063494190 NaCl Diluent 9 % (123 mL) System‑ID 2906 001
*Roche does not hold the product registration for Partner Channels. The legal manufacturer indicated on the kit is solely responsible for all of the design,
legal, and regulatory aspects of the product.

English Reagents - working solutions

System information R1 Conjugated methadone derivative; buffer; bovine serum albumin;
MDQ3S: ACN 20883 (Serum/plasma): for qualitative assay, 300 ng/mL 0.09 % sodium azide
Intended use R2 Microparticles attached to methadone antibody (mouse
The ONLINE DAT II assay for methadone is an in vitro diagnostic test for monoclonal); buffer; bovine serum albumin; 0.09 % sodium azide
the qualitative detection of methadone in human serum and plasma on
Roche/Hitachi cobas c systems at a cutoff concentration of 300 ng/mL. R1 is in position B and R2 is in position C.
The assay provides only a preliminary analytical test result. A more Precautions and warnings
specific alternate chemical method must be used in order to obtain a For in vitro diagnostic use.
confirmed analytical result. Gas chromatography/mass spectrometry Exercise the normal precautions required for handling all laboratory
(GC‑MS) or liquid chromatography coupled with tandem mass reagents.
spectrometry (LC‑MS/MS) is the preferred confirmatory method.1 Disposal of all waste material should be in accordance with local guidelines.
Clinical consideration and professional judgment should be applied to Safety data sheet available for professional user on request.
any drug of abuse test result, particularly when preliminary positive
results are used. Reagent handling
Ready for use
Summary
Methadone is a synthetic diphenylpropylamine used for detoxification and Carefully invert reagent container several times prior to use to ensure that
temporary maintenance of narcotic addiction, as well as treatment of acute the reagent components are mixed.
and chronic pain. Methadone has many of the pharmacologic properties of Storage and stability
morphine, and its analgesic potency is similar. Unlike morphine, repeated
administration causes marked sedative effects due to drug accumulation in Shelf life at 2‑8 °C: See expiration date on
the body. Methadone withdrawal syndrome is qualitatively similar to cobas c pack label
morphine, yet it differs in that it develops more slowly, is less intense, and is
more prolonged.2 For these reasons, methadone is used in the On‑board in use and refrigerated on the 26 weeks
management of narcotic dependence, hopefully eliminating the need for analyzer:
illicit opiate drugs. Overdoses of methadone are characterized by stupor, Do not freeze.
respiratory depression, cold and clammy skin, hypotension, coma, and
circulatory collapse.3 Specimen collection and preparation
Methadone is given intramuscularly for analgesic purposes and orally for For specimen collection and preparation only use suitable tubes or
methadone maintenance therapy. Following ingestion, the drug is well collection containers.
absorbed from the gastrointestinal tract and is widely distributed to the liver, Only the specimens listed below were tested and found acceptable.
lung, kidney, spleen, blood, and urine. The fact that methadone is highly Serum: Serum tubes with and without separating gel.
bound to tissue protein may explain its cumulative effects.4 Methadone is Plasma: K2‑ or K3‑EDTA, lithium heparin.
metabolized largely by mono- and di‑N‑demethylation. Spontaneous
cyclization of the resulting unstable compounds forms the major Stability: 5 days capped at 15‑25 °C
metabolites, 2‑ethylidene‑1,5‑dimethyl‑3,3‑diphenylpyrrolidine (EDDP) and
2‑ethyl‑5‑methyl‑3,3‑diphenylpyrroline (EMDP). Both are hydrolyzed to 14 days capped at 2‑8 °C
some extent, with subsequent glucuronidation.5,6 In maintenance patients, 6 months capped at ‑20 °C
excretion of unchanged methadone can account for 5‑50 % of the dose.
Urinary pH affects the percentage of unchanged drug excreted, as does The sample types listed were tested with a selection of sample collection
urinary volume, dose, and individual metabolism.7,8 tubes that were commercially available at the time of testing, i.e. not all
available tubes of all manufacturers were tested. Sample collection systems
Test principle from various manufacturers may contain differing materials which could
The assay is based on the kinetic interaction of microparticles in a solution affect the test results in some cases. When processing samples in primary
(KIMS)9,10 as measured by changes in light transmission. In the absence of tubes (sample collection systems), follow the instructions of the tube
sample drug, soluble drug conjugates bind to antibody-bound manufacturer.
microparticles, causing the formation of particle aggregates. As the Centrifuge samples containing precipitates before performing the assay.
aggregation reaction proceeds in the absence of sample drug, the
absorbance increases. See the limitations and interferences section for details about possible
sample interferences.
When a serum sample contains the drug in question, this drug competes
with the drug derivative conjugate for microparticle‑bound antibody. Specimens can be repeatedly frozen and thawed up to 3 times.
Antibody bound to sample drug is no longer available to promote particle Invert thawed specimens several times prior to testing.
aggregation, and subsequent particle lattice formation is inhibited. The CAUTION: Specimen dilutions should only be used to interpret results of
presence of sample drug diminishes the increasing absorbance in Calc.? and Samp.? alarms, or when estimating concentration in preparation
proportion to the concentration of drug in the sample. Sample drug content for GC‑MS. Dilution results are not intended for patient values. Dilution
is determined relative to the value obtained for a known cutoff concentration procedures, when used, should be validated.
of drug.11
Materials provided
See “Reagents – working solutions” section for reagents.

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MDN2
ONLINE DAT Methadone II

Materials required (but not provided) Values obtained should fall within the defined limits. Each laboratory should
See “Order information” section establish corrective measures to be taken if values fall outside the defined
limits.
General laboratory equipment
Follow the applicable government regulations and local guidelines for
Assay quality control.
For optimum performance of the assay follow the directions given in this
document for the analyzer concerned. Refer to the appropriate operator’s Results
manual for analyzer‑specific assay instructions. The cutoff calibrator is used as a reference in distinguishing between
preliminary positive and negative samples. Samples producing a positive or
The performance of applications not validated by Roche is not warranted “0” absorbance value are considered preliminary positive. Preliminary
and must be defined by the user. positive samples are flagged with >Test. Samples producing a negative
Application for serum and plasma absorbance value are considered negative. Negative samples are preceded
by a minus sign.
Test definition Preliminary positive results should be confirmed by another method.
Qualitative Limitations - interference
Reporting time 10 min Criterion: No cross‑over at initial values of samples of 150 ng/mL and
450 ng/mL (control levels).
Wavelength (sub/main) – /546 nm
See the "Specific performance data" section of this document for
Reagent pipetting information on substances tested with this assay. There is the possibility
R1 59 µL that other substances and/or factors may interfere with the test and cause
erroneous results (e.g., technical or procedural errors).
R2 26 µL A preliminary positive result with this assay indicates the presence of
Sample volumes Sample methadone and/or its metabolites in serum. It does not measure the level of
intoxication.
300 ng/mL cutoff
Icterus:12 No significant interference up to an I index of 60 for conjugated
Normal 2.3 µL and unconjugated bilirubin (approximate conjugated and unconjugated
bilirubin concentration: 1026 µmol/L or 60 mg/dL).
Decreased 2.3 µL
Hemolysis:12 No significant interference up to an H index of 1000
Increased 2.3 µL (approximate hemoglobin concentration: 622 µmol/L or 1000 mg/dL).
For further information about the assay test definitions refer to the Lipemia (native lipaemic samples):12 No significant interference up to an
application parameters setting screen of the corresponding analyzer and L index of 100. There is poor correlation between the L index (corresponds
assay. to turbidity) and triglycerides concentration.
Calibration Rheumatoid factors: No significant interference from rheumatoid factors up
to a concentration of 1200 IU/mL.
Calibrators Qualitative application Immunoglobulins: No significant interference from immunoglobulins up to a
300 ng/mL cutoff assay concentration of 16 g/L (simulated by human immunoglobulin A), up to a
concentration of 70 g/L (simulated by human immunoglobulin G) and up to
S1: Preciset DAT Plus I calibrator ‑ CAL 5, a concentration of 10 g/L (simulated by human immunoglobulin M).
2000 ng/mL with automatic pre‑dilution Albumin: No significant interference from human serum albumin up to a
The drug concentration of the calibrator has been concentration of 70 g/L.
verified by GC‑MS. As with any assay employing mouse antibodies, the possibility exists for
interference by human anti‑mouse antibodies (HAMA) in the sample, which
Calibration K factor For the qualitative application a K factor of -1000 is could cause falsely lowered results.
predefined in the application settings.
For diagnostic purposes, the results should always be assessed in
Calibration mode Qualitative application conjunction with the patient’s medical history, clinical examination and other
findings.
Linear
ACTION REQUIRED
Calibration frequency Full calibration Special Wash Programming: The use of special wash steps is mandatory
- after reagent lot change when certain test combinations are run together on cobas c systems. All
- every 84 days special wash programming necessary for avoiding carry‑over is available
- as required following quality control procedures via the cobas link. The latest version of the carry‑over evasion list can be
found with the NaOHD/SMS/SCCS Method Sheet for information. For
For the cutoff calibrator a value of "0" is encoded in the e‑barcode in order further instructions refer to the operator’s manual.
to ensure flagging of positive samples with >Test and negative absorbance
values for negative samples. Expected values
Calibration interval may be extended based on acceptable verification of Qualitative assay
calibration by the laboratory. Results of this assay distinguish preliminary positive (≥ 300 ng/mL) from
Traceability: This method has been standardized against a primary negative samples only. The amount of drug detected in a preliminary
reference method (GC‑MS). positive sample cannot be estimated.

Quality control Specific performance data

For quality control, use control materials as listed in the “Order information” Representative performance data on the analyzers are given below. These
section. In addition, other suitable control material can be used. data represent the performance of the analytical procedure itself.
Drug concentrations of the high and low controls have been verified by Results obtained in individual laboratories may differ due to heterogenous
LC‑MS/MS. sample materials, aging of analyzer components and mixture of reagents
running on the analyzer.
The control intervals and limits should be adapted to each laboratory’s
individual requirements. It is recommended to perform quality control Precision
always after lot calibration and subsequently at least every 26 weeks. Precision was determined using human samples and controls in
accordance with the CLSI (Clinical and Laboratory Standards Institute)
EP05‑A3 requirements with repeatability (n = 84) and intermediate precision

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ONLINE DAT Methadone II

(2 aliquots per run, 2 runs per day, 21 days). Results for repeatability and Samples were tested and the following results were obtained on a cobas c
intermediate precision were obtained on the cobas c 503 analyzer. 501 analyzer.
Qualitative precision - 300 ng/mL Compound Comp. conc. Neg. Pos.
Cutoff (300) Number Correct Confidence level mg/L level level
tested results Acetaminophen 200 neg pos
Serum -75 % 84 84 > 95 % negative reading Acetylcysteine 1660 neg pos
ACQ‑L 84 84 > 95 % negative reading Acetylsalicylic acid 1000 neg pos
Cutoff serum 84 n.a.* n.a.* Amitriptyline 1.00 neg pos
ACQ‑H 84 84 > 95 % negative reading Ampicillin‑Na 1000 neg pos
Serum +75 % 84 84 > 95 % negative reading Ascorbic acid 300 neg pos
*n.a. = not applicable Caffeine 59.8 neg pos
Accuracy Cefoxitin 2500 neg pos
103 serum samples, screened negative for methadone on a cobas c 501 Cyclosporine 5.00 neg pos
analyzer were evaluated with the Methadone II assay on a cobas c 503
analyzer. 100 % of these normal serum samples were negative with the d‑Amphetamine 1.36 neg pos
Methadone II assay on a cobas c 503 analyzer. 51 serum samples
screened positive for methadone relative to the 300 ng/mL cutoff on a Diazepam 5.13 neg pos
cobas c 501 analyzer were evaluated with the Methadone II assay on a Doxycycline 50.0 neg pos
cobas c 503 analyzer. At the 300 ng/mL cutoff, 100 % of the samples were
positive on both the cobas c 501 analyzer and the cobas c 503 analyzer. d‑Pseudoephedrine 9.98 neg pos
Erythromycin 59.9 neg pos
Methadone II correlation (cutoff = 300 ng/mL)
Fenoprofen 195 neg pos
cobas c 501 analyzer
Furosemide 59.9 neg pos
+ -
Gentisic acid 18.0 neg pos
cobas c 503 analyzer + 51 0
Heparin 5000 U/L neg pos
- 0 103
Hydrochlorothiazide 6.02 neg pos
52 serum samples, screened negative for methadone on a cobas c 501
analyzer were evaluated with the Methadone II assay on a cobas c 303 l‑Amphetamine 1.00 neg pos
analyzer. 100 % of these normal serum samples were negative with the Ibuprofen 500 neg pos
Methadone II assay on a cobas c 303 analyzer. 52 serum samples
screened positive for methadone relative to the 300 ng/mL cutoff on a Imipramine 0.70 neg pos
cobas c 501 analyzer were evaluated with the Methadone II assay on a Ketamine 10.0 neg pos
cobas c 303 analyzer. At the 300 ng/mL cutoff, 100 % of the samples were
positive on both the cobas c 501 analyzer and the cobas c 303 analyzer. Levodopa 20.0 neg pos
Methadone II correlation (cutoff = 300 ng/mL) Lidocaine 12.0 neg pos
cobas c 501 analyzer Methyldopa + 1.5 H2O 20.0 neg pos
+ - Metronidazole 200 neg pos
cobas c 303 analyzer + 52 0 Morphine 0.50 neg pos
- 0 52 Naproxen 499 neg pos
Phenylbutazone 400 neg pos
Analytical specificity
The specificity of this assay for structurally similar compounds was Procaine 39.9 neg pos
determined by generating inhibition curves for each of the compounds listed Promethazine 1.20 neg pos
and determining the approximate quantity of each compound that is
equivalent in assay reactivity to a 300 ng/mL assay cutoff. Caution should Quinidine 12.0 neg pos
be taken when interpreting results of patient samples containing structurally Quinine 48.0 neg pos
related compounds having greater than 0.5 % cross‑reactivity. The
following results were obtained on a cobas c 501 analyzer. Rifampicin 60.0 neg pos
Compound ng/mL Approximate % Tetracycline 15.1 neg pos
Equivalent to cross‑reactivity Theophylline 100 neg pos
300 ng/mL Trifluoperazine 1.00 neg pos
methadone
In very rare cases, gammopathy, in particular type IgM (Waldenström’s
Chlorpromazine 70752 0.42 macroglobulinemia), may cause unreliable results.13
Metabolite Lu AA34443 6341 4.73 References
Methadone 308 97.3 1 Karch SB, ed. Drug Abuse Handbook. Boca Raton, FL: CRC Press
LLC 1998.
Vortioxetine 8344 3.60
2 Council Reports: Treatment of morphine-type dependence by
Drug interference withdrawal methods. JAMA 1972;219(12):1611-1615.
Interfering substances were added to serum containing methadone at 3 Smialek JE, Monforte JR, Aronow R, et al. Methadone deaths in
‑50 % and +50 % of the cutoff level at the concentration listed below. children: A continuing problem JAMA 1977;238(23):2516-2517.

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MDN2
ONLINE DAT Methadone II

4 Garriott JC, Sterner WQ, Mason MF. Toxicologic findings in six

fatalities involving methadone. Clin Toxicol 1973;6:163-173.
5 Sullivan HR, Due SL, McMahon RE. The identification of three new
metabolites of methadone in man and in the rat. J Am Chem Soc
1972;94(11):4050-4051.
6 Baselt RC, Bickelt MH. Biliary excretion of methadone by the rat:
identification of a para-hydroxylated major metabolite. Biochem Pharm
1973;22:3117-3120.
7 Baselt RC, Casarett LJ. Urinary excretion of methadone in man. Clin
Pharmacol Ther 1972 Jan-Feb;13(1):64-70.
8 Bellward GD, Warren PM, Howald W, et al. Methadone maintenance:
Effect of urinary pH on renal clearance in chronic high and low doses.
Clin Pharmacol Ther 1977;22(1):92-99.
9 Armbruster DA, Schwarzhoff RH, Pierce BL, et al. Method comparison
of EMIT II and ONLINE with RIA for drug screening. J Forensic Sci
1993;38:1326-1341.
10 Armbruster DA, Schwarzhoff RH, Hubster EC, et al. Enzyme
immunoassay, kinetic microparticle immunoassay, radioimmunoassay,
and fluorescence polarization immunoassay compared for drugs-of-
abuse screening. Clin Chem 1993;39:2137-2146.
11 Bates M, Brandle J, Casaretto E, et al. An Abuscreen immunoassay for
opiates in urine on the COBAS MIRA automated analyzer. Amer Acad
Forensic Sci. Abstract 1991;37(6):1000.
12 Glick MR, Ryder KW, Jackson SA. Graphical Comparisons of
Interferences in Clinical Chemistry Instrumentation.
Clin Chem 1986;32:470-475.
13 Bakker AJ, Mücke M. Gammopathy interference in clinical chemistry
assays: mechanisms, detection and prevention.
Clin Chem Lab Med 2007;45(9):1240-1243.
A point (period/stop) is always used in this Method Sheet as the decimal
separator to mark the border between the integral and the fractional parts of
a decimal numeral. Separators for thousands are not used.
Symbols
Roche Diagnostics uses the following symbols and signs in addition to
those listed in the ISO 15223‑1 standard (for USA: see dialog.roche.com for
definition of symbols used):
Contents of kit
Volume for reconstitution
GTIN Global Trade Item Number

ABUSCREEN, COBAS, COBAS C, ONLINE DAT and PRECISET are trademarks of Roche.
All other product names and trademarks are the property of their respective owners.
Additions, deletions or changes are indicated by a change bar in the margin.
© 2021, Roche Diagnostics

Roche Diagnostics GmbH, Sandhofer Strasse 116, D-68305 Mannheim

www.roche.com

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