Published: Journal of Analytical Toxicology, Volume 23, Number 4, July/August 1999, pp.299-300.

Disopyramide Cross-Reactivity in a Commercial Immunoassay Reagent for Methadone

The specificity of an immunoassay reagent is dependent on the quality of its antibody and the affinity of the antibody to form a matched complex with a complementary antigen. The challenge for immunoassay manufacturers is to design antibodies that have a high degree of specificity, which ultimately restricts the number of antigens able to bind with a given antibody. The advent of monoclonal antibodies has greatly reduced but not eliminated the potential for unintended antigens to cross-react with antibodies. In fact, the literature is replete with such examples (1–3).

Recently, our laboratory encountered a donor urine specimen submitted for drugs-of-abuse testing that exhibited an immunoassay response that was positive for methadone via the Microgenics CEDIA DAU methadone assay on the Boehringer Mannheim Hitachi 717. Extraction of the specimen by a previously described method (4) and subsequent analysis via gas chromatography–mass spectrometry (GC–MS) failed to confirm the presence of methadone. Testing of the specimen by thin-layer chromatography (TLC) and a full-scan analysis of the GC–MS extract produced results suggestive of disopyramide.

Disopyramide is an antiarrhythmic drug used clinically to decrease membrane responsiveness and to prolong action potentials in myocardium. Disopyramide is rapidly absorbed from the gastrointestinal tract. The kidney excretes most of the absorbed dose (50%) as unchanged drug and approximately 30% as nordisopyramide (5). A typical therapeutic dose of disopyramide is 400 to 800 mg daily. Given its pharmacokinetics, relatively high concentrations of disopyramide can be expected in the urine even at therapeutic levels.

In order to investigate the potential for disopyramide cross-reactivity, certified-negative urine samples were spiked with varying concentrations of a methanolic solution of disopyramide. The spiked urine samples were submitted for TLC, immunoassay screening, and GC–MS analysis. Results generated for the spiked samples were consistent with the results obtained for the donor specimen (Table I).

Identifying the exact mechanism(s) for the false-positive methadone result was beyond the scope of our abbreviated investigation; however, review of the chemical structures of disopyramide and methadone (6) revealed a striking similarity between these two drugs, suggesting a possible structural basis for the observed cross-reactivity (Figure 1).

As with most immunoassay tests, the CEDIA DAU methadone assay is intended to provide only preliminary test results. An appropriate confirmatory technique should be employed when such preliminary results are positive.

Paul Moorman, Marcia McCoy,
Bonnie Hague, and Debbie Huge
South Bend Medical Foundation
530 N. Lafayette Blvd.
South Bend, Indiana 46601

References

1. P. Mura, P. Kintz, R. Robert, and Y. Papet. Buflomedil is a potent interfering substance in immunoassays of tricyclic antidepressants.
   J. Anal. Toxicol. 22: 254 (1998).
2. T. Matuch-Hite, P. Jones, and J. Moriarity. Interference of oxaprozin with benzodiazepines via enzyme immunoassay technique. J. Anal. Toxicol. 19: 130 (1995).
3. J. Lotz, G. Hafner, J. Rohrich, S. Zorntlein, T. Kern, and W. Prellwitz. False positive LSD drug screening induced by a mucolytic medication. Clin. Chem. 44: 1580–1581 (1998).
4. D. Fretthold, P. Jones, G. Sebrosky, and I. Sunshine. Testing for basic drugs in biological fluids by solvent extraction and dual capillary GC–NPD. J. Anal. Toxicol. 10: 10–14 (1986).
5. M.J. Ellenhorn and D.G. Barceloux. Medical Toxicology. Elsevier, New York, NY, 1988.
6. R.C. Baselt and R.H. Cravey. Disposition of Toxic Drugs and Chemicals in Man. Chemical Toxicology Institute, Foster City, CA, 1995.

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