Immunoassays for drug screening in urine

Chances, challenges, and pitfalls
  • Harald SchützEmail author
  • Alexandre Paine
  • Freidoon Erdmann
  • Günter Weiler
  • Marcel A. Verhoff
Original Article


Immunoassays are presently used worldwide for the rapid screening of drugs. Despite the fact that they are a highly valuable tool for the testing of legal and illicit drugs, there is a real risk of false-positive and false-negative findings and many pitfalls must be taken into account when these tests are used in an uncritical manner and without valid confirmation procedures. In a long-standing successful cooperation with a large psychiatric hospital, we checked doubtful and nonplausible immunochemical findings in urine with gas chromatography-mass spectrometry (GC-MS) confirmation methods. The reported case histories demonstrate typical pitfalls, for example, influence of changing nutritional habits, use of sweeteners (cyclamate), uncritical creatinine correction, impaired elimination, or cross-reactivities of simultaneous therapeutic medication. In accordance with national and international associations, immunoassays are recommended as a useful tool for screening. However, confirmation analysis with conclusive methods (GC-MS or liquid chromatography-MS) is unavoidable for valid substance identification, discrimination between active and inactive metabolites, detection of congeners, and accurate determination of concentrations in body fluids.

Key words

Forensic toxicology toxicological screening immunoassays drugs illicit drugs pitfalls confirmation analysis 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Edwards R, ed. Immunoassay—Essential data. Chichester: Wiley, 1996.Google Scholar
  2. 2.
    Külpmann WR, ed. Klinisch-toxikologische Analytik. Weinheim: Wiley-VCH-Verlag, 2002.Google Scholar
  3. 3.
    Hand C, Baldwin D. Immunoassays. In: Moffat AC, MD, Widdop B, ed. Clarke's Analysis of Drugs and Poisons, 3rd ed., London, Chicago: Pharmaceutical Press, 2004, pp.Google Scholar
  4. 4.
    Law B, ed. Immunoassay—A Practical Guide. London: Taylor & Francis. 1996.Google Scholar
  5. 5.
    Schütz H. Drogenscreening mit Immunoassays. Pharmazie in unserer Zeit 1999;28:320–328.PubMedCrossRefGoogle Scholar
  6. 6.
    Wild D, ed. The Immunoassay Handbook. New York: Stockton Press, 1994.Google Scholar
  7. 7.
    Aderjan R, Schütz H, Käferstein H, Wilske J. Empfehlungen der Kommission Grenzwertfragen bei Arzneimitteln und Suchtstoffen zum Nachweis von Betäubungsmitteln im Blut: Immunologische Messungen von Substanzen im Blut reichen für den Nachweis einer Ordnungswidrigkeit im Sinne des §24 a StVG nicht aus. Blutalkohol 2003;40:337–342.Google Scholar
  8. 8.
    Al-Amri AM, El-Haj BM. Positive interference of the analgesic drug nefopam in the enzyme immunoassay (EMIT D.A.U.) for benzodiazepines. Bull Int Assoc Forens Toxicol 1999;29:4.Google Scholar
  9. 9.
    As van H, Stolk LML. Rifampicin cross-reacts with opiate immunoassay. J Anal Toxicol 1999;23:71.PubMedGoogle Scholar
  10. 10.
    Bogusz M, Maier RD, Hoenen H. Positive LSD immunoassay after sertraline intake. Bull Int Assoc Forensic Toxicol 1998;28:11, 12.Google Scholar
  11. 11.
    Brunk SD. False negative GC/MS assay for carboxy THC due to ibuprofen interference. Clin Chem 1988; 12:290, 291.Google Scholar
  12. 12.
    Krämer T, Bock K, Wenning R, Maurer HH. The antispasmodic mebeverine leads to positive amphetamine results with the fluorescence polarization immunoassay—studies on the metabolism and the toxicological detection in urine by GC/MS and FPIA. Mosbach: XI Symposium der GTFCh 1999: pp. 34–43.Google Scholar
  13. 13.
    Krämer T, Wennig R, Maurer HH. The antispasmodic mebeverine leads to positive amphetamine results by fluorescence polarization immunoassay (FPIA)—Studies on the toxicological analysis of urine by FPIA and GC-MS. J Anal Toxicol 2001;25:333–338.Google Scholar
  14. 14.
    Külpmann WR. Drug screening: Actual status, pitfalls and suggestions for improvement. J Lab Med 2004;28:317–325.Google Scholar
  15. 15.
    Larsen J, Fogerson R. Nonsteroidal anti-inflammatory drug interferences in TDx assays for abused drugs. Clin Chem 1988;34:987, 988.PubMedGoogle Scholar
  16. 16.
    Lewis JH. Interference of genfibrozil with Roche Testcup. J Anal Toxicol 1999;23:384.PubMedGoogle Scholar
  17. 17.
    Linder MW, Valdes R. Mechanism and elimination of aspirin induced interferences in EMIT II d.a.u. assays. Clin Chem 1994;40:1512–1516.PubMedGoogle Scholar
  18. 18.
    Lora-Tamayo C, Tena T. High concentrations of metronidazole in urine invalidates EMIT results. J Anal Toxicol 1991;15:159.Google Scholar
  19. 19.
    Lora-Tamayo C, Tena T, Rodriguez A. High concentration of ciprofloxacin in urine invalidates EMIT results. J Anal Toxicol 1996;20:334.PubMedGoogle Scholar
  20. 20.
    Lotz J, Hafner G, Röhrich J, Zörntlein S, Kern T, Prellwitz W. False positive LSD drug screening induced by a mucolytic medication. Clin Chem 1998;44:1580, 1581.PubMedGoogle Scholar
  21. 21.
    Luceri F, Godi F, Messeri G. Reducing false-negative tests in urinary drugs-of-abuse screening. J Anal Toxicol 1997;21:244,245.PubMedGoogle Scholar
  22. 22.
    Marson C, Schneider S, Meys F, Wennig R. Structural elucidation of an uncommon phenylethylamine analogue in urine responsible for discordant amphetamine immunoassay results. J Anal Toxicol 2000;24:17–21.PubMedGoogle Scholar
  23. 23.
    Matuch-Hite T, Jones P Jr, Moriarity J. Interference of oxaprozine with benzodiazepines via enzyme immunoassay technique. J Anal Toxicol 1995;19:130.PubMedGoogle Scholar
  24. 24.
    Meatherall R, Dai J. False-positive EMIT II opiates from ofloxacin. Ther Drug Monit 1997;19:98, 99.PubMedCrossRefGoogle Scholar
  25. 25.
    Moorman P, McCoy M, Hague B, Huge D. Disopyramide crossreactivity in a commercial immunoassay for methadone. J Anal Toxicol 1999;23:299, 300.PubMedGoogle Scholar
  26. 26.
    Mura P, Kintz P, Robert R, Papet Y. Buflomedil is a potent interfering substance in immunoassays of tricyclic antidepressants. J Anal Toxicol 1998;22:254.PubMedGoogle Scholar
  27. 27.
    Nishikawa T, Kamijo Y, Ohtani H, Fraser AD. Oxaprozin interference with urinary benzodiazepine immunoassay and noninterfrence with receptor assay. J Anal Toxicol 1999;23:125, 126.PubMedGoogle Scholar
  28. 28.
    Notarriani LJ, Belk D, Collins AJ. False positives and negatives in routine testing for drugs of abuse. Lancet 1995;345:1115.CrossRefGoogle Scholar
  29. 29.
    Poklis A, O'Neal C. Potential for false-positive results by the TRIAGETM panel of drugs-of-abuse immunoassay. J Anal toxicol 1996;20:209, 210.PubMedGoogle Scholar
  30. 30.
    Podkowik BI, Repka ML, Smith ML. Interference by ritodrine in GC/MS confirmation of delta-9-tetrahydrocannabinol-9-carboxylic acid in urine. Clin Chem 1991;37:1305, 1306.PubMedGoogle Scholar
  31. 31.
    Röhrich J, Zörntlein S, Lotz J, Becker J, Rittner C. False-positive LSD testing in urine samples from intensive care patients. J Anal Toxicol 1998;22:393–395.PubMedGoogle Scholar
  32. 32.
    Rollins DE, Jennison TA, Jones G. Investigation of interference by nonsteroidal anti-inflammatory drugs in urine tests for abused drugs. Clin Chem 1990;36:602–606.PubMedGoogle Scholar
  33. 33.
    Schneider S, Wennig R. Interference of diphenhydramine with the EMIT II immunoassay for propoxyphene. J Anal Toxicol 1999;23:637, 638.PubMedGoogle Scholar
  34. 34.
    Schütz H. Screening von Drogen und Arzneimitteln mit Immunoassays. Ein Leitfaden für die Praxis. Wiesbaden: Wiss. Verlagsabteilung Abbott GmbH, 1992.Google Scholar
  35. 35.
    Schütz H, Erdmann F, Magiera ES, Weiler G. Fehlerhafte Bestätigungsanalytik bei falsch-positiven Immunoassays und ihre Auswirkungen. Arch Kriminol 1998;35:93–96.Google Scholar
  36. 36.
    Schütz H, Erdmann F, Rochholz G, Weiler G. False Positive and False Negative Immunological Findings—A Permanent Risk of Analytical Pitfalls. Jpn J Forensic Toxicol 2000;18:14–20.Google Scholar
  37. 37.
    Schütz H, Erdmann F, Weiler G. Folgen unbestätigter Immunoassays mit miss verständlichem Befundbericht. Toxichem+Krimtech 1997;64:95.Google Scholar
  38. 38.
    Schütz H, Erdmann F, Verhoff MA, Weiler G. Pitfalls of toxicological analysis. Legal Medicine 2003;5:6–19.CrossRefGoogle Scholar
  39. 39.
    Schütz H, Rochholz G, Weiler G. Zur Problematik der falsch negativen Benzodiazepin-Immunoassays. Klin Lab 1992;38:150–152.Google Scholar
  40. 40.
    Schütz H, Verhoff MA, Erdmann F, Weiler G. Immunchemisches Screening: Manipulationen, Störungen und Fehlinterpretationen, dargestellt am Beispiel der Benzodiazepine. Arch Kriminol 2003;212:141–150.PubMedGoogle Scholar
  41. 41.
    Wagener RE, Linder MW, Valdes R. Decreased signal in EMIT assays of drug abuse in urine after ingestion of aspirin: potential of false-negative results. Clin Chem 1994;40:608–612.PubMedGoogle Scholar
  42. 42.
    Schütz H, Auch J, Erdmann F, Weiler G, Verhoff MA. Immunochemical drug screening—evidence for additional cross-reactivities. Arzneim-Forsch (Drug Res) (submitted for publication).Google Scholar
  43. 43.
    George S, Braithwaite RA, An investigation into the extent of possible dilution of specimens received for urinary drugs of abuse screening. Addiction 1995;90:967–970.PubMedCrossRefGoogle Scholar
  44. 44.
    Goldberger BA, Loewenthal B, Darwin WD, Cone EJ. Intrasubject variation of creatinine and specific gravity measurements in consecutive urine specimens of heroin users. Clin Chem 1995;41:116–117.PubMedGoogle Scholar
  45. 45.
    Lafolie P, Beck O, Blennow G, et al. Importance of creatinine analyses of urine when screening for abused drugs. Clin Chem 1991;37:1927–1931.PubMedGoogle Scholar
  46. 46.
    Needleman SB, Porvaznik M, Ander D. Creatinine analysis in single collection urine specimens. J Forens Sci 1992;37:1125–1133.Google Scholar
  47. 47.
    Pfleger K, Maurer HH, Weber A. Mass Spectral and GC Data of Drugs, Poisons, Pesticides, Pollutants and Their Metabolites (Parts 1–4), Weinheim, Wiley-VCH-Verlag, 2000.Google Scholar
  48. 48.
    Schütz H, Weiler G, Erdmann F, Magiera ES, Schade S. Risiken von Sparmass nahmen beim Drogen-Screening. Blutakohol 1998;35:139–144.Google Scholar
  49. 49.
    Baiker C, Serrano L, Lindner B. Hypochlorite adulteration of urine causing decreased concentration of delta-9-THC-COOH by GC/MS. J Anal Toxicol 1994;18:101–103.PubMedGoogle Scholar
  50. 50.
    Bronner W, Nymann P, von Minden D. Detectability of phen-cyclidine and 11-nor-delta-9-tetrahydrocannabinol-9-carboxylic acid in adulterated urine by radiommunoassay and fluorescence polarization immunoassay. J Anal Toxicol 1990;14: 368–371.PubMedGoogle Scholar
  51. 51.
    Cody JT. Specimen adulteration in drug urinalysis. Forens Sci Rev 1990;2:63–75.Google Scholar
  52. 52.
    Cody JT. Adulteration of urine specimens. In: Liu RH and BA Goldberger, ed. Handbook of Workplace Drug Testing. Washington DC: AACC Press 1995, pp.181–208.Google Scholar
  53. 53.
    Cody JT, Valtier S. Effects of stealth adulterant on immunoassay testing for drugs of abuse. J Anal Toxicol 2001;25:466–470.PubMedGoogle Scholar
  54. 54.
    Cody JT, Valtier S, Kuhlman J. Analysis of morphine and codeine in samples adulterated with stealthTM. J Anal Toxicol 2001;25:572–575.PubMedGoogle Scholar
  55. 55.
    Davis KH. Adulterants update paper presented at the annual meeting of the Society of Forensic Toxicologists, San Juan, Puerto Rico. 1999.Google Scholar
  56. 56.
    Edwards C, Fyfe MJ, Liu RH, Walia AS. Evaluation of common urine specimen adulteration indicators. J Anal Toxicol 1993;17:251–252.PubMedGoogle Scholar
  57. 57.
    ElSohly MA, Feng S, Kopycki WJ, et al. A procedure to overcome interferences caused by adulterant Klear in the GC/MS analysis of 11-nor-delta-9-THC-9-COOH. J Anal Toxicol 1997;21:240–242.PubMedGoogle Scholar
  58. 58.
    George S, Braithwaite RA. The effect of glutaraldehyde adulteration of urine specimens on Syva EMIT II drugs-of-abuse assays. J Anal. Toxicol 1996;20:195, 196.PubMedGoogle Scholar
  59. 59.
    Goldberger BA, Caplan YH. Effect of glutaraldehyde (Urin Aid) on detection of abused drugs in urine by immunoassay. Clin Chem 1994;40:1605, 1606.PubMedGoogle Scholar
  60. 60.
    Hagemann P, Siegrist M. Verfälschungsstoffe beim Drogennachweis. Lab Med 1990;14:114–120.Google Scholar
  61. 61.
    Liu RH. Important comsiderations in the interpretation of forensic drug test results. Forensic Sci Rev 1992;4:52–65.Google Scholar
  62. 62.
    Martz W. Untersuchungen zu propagierten Methoden der Urinverfälschung vor dem Drogentest. Toxichem+Krimtech 1997;64:88–94.Google Scholar
  63. 63.
    Mikkelsen SL, Ash KO. Adulterants causing false negatives in illicit drug testing. Clin Chem 1988;34:2333–2336.PubMedGoogle Scholar
  64. 64.
    Paul BD, Martin KK, Maguilo J Jr, Smith ML. Effects of pyridinium chlorochromate adulterant (urine luck) on testing for drugs of abuse and a method for quantitative detection of chromium (VI) in urine. J Anal Toxicol 2000;24:233–237.PubMedGoogle Scholar
  65. 65.
    Pearson SD, Ash KO, Urry FM. Mechanism of false-negative urine cannabinoid immunoassay screens by VisineTM eyedrops. Clin Chem 1989;35:636–638.PubMedGoogle Scholar
  66. 66.
    Sansom HL, Freser MD, Botelho C, Juntz DC, Foltz RL. Detection of urine specimens adulterated with UrinAid. Presented at the Society of Forensic Toxicologists anual meeting, Phoenix (AZ), 1993.Google Scholar
  67. 67.
    Schwarhoff R, Cody JT. He effects of adulterating agents on FPIA analysis of urine for drugs of abuse. J Anal Tox 1993;17: 14–17.Google Scholar
  68. 68.
    Singh J, Elberling JS, Hemphill DG, Holmstrom J. The measurements of nitrite in adulterated urine samples by high-performance ion chromatography. J Anal Toxicol 1999;23: 137–140.PubMedGoogle Scholar
  69. 69.
    Skopp G, Pötsch L, Becker J, Röhrich J, Mattern R. Zur prä-analytischen Phase chemisch-toxikologischer Untersuchungen. I. Immunchemisches Drogenscreening im Urin-Erkennbarkeit von Manipulationen und Strategien bei rechtsmedizinischer Fragestellung. Rechtsmedizin 1998;8:163–167.CrossRefGoogle Scholar
  70. 70.
    Smith BL, Wilson FW. Effect of adulterants on urine tests. Ther Drug Monit 1997;5:586.CrossRefGoogle Scholar
  71. 71.
    Tsai SC, ElSohly MA, Dubrovsky T, Twarowska B, Towt J, Salamone SJ. Determination of five abused drugs in nitrite-adulterated spectrometry. J Anal Toxicol 1998;22:474–480.PubMedGoogle Scholar
  72. 72.
    Tytgat J, Boven van M, Daenens P. Cannabinoid mimics in chocolate utilized as an argument in court. Int J Legal Med 2000;113:137–139.PubMedCrossRefGoogle Scholar
  73. 73.
    Urry FM, Komaromy-Hiller G, Staley B, et al. Nitrite adulteration of workplace urine drug-testing speciments. I. Sources and associated concentrations of intritein urine and distinction between natural sources and adulteration. J Anal Toxicol 1998;22:89–95.PubMedGoogle Scholar
  74. 74.
    Warner A. Interference of common household chemicals in immunoassay methods for drugs of abuse. Clin Chem 1989; 35:648–651.PubMedGoogle Scholar
  75. 75.
    Wu AH, Bristol B, Sexton K, Cassella-McLane G, Holtman V, Hill DW. Adulteration of urine by Urine Luck. Clin Chem 1999;45:1051–1057.PubMedGoogle Scholar
  76. 76.
    Wu AHB, Forte E, Casella G, Sun K, Hemphill G, Forey R, Schazenbach H. Cedia for screening drugs of abuse in urine adulterants. J Forens Sci 1995;40:614–618.Google Scholar
  77. 77.
    Wu A, Schmalz J, Bennett W. Identification of Urine Aid adulterated specimens by fluorimetric analysis. Clin Chem 1994; 40:845–846.PubMedGoogle Scholar

Copyright information

© Humana Press Inc 2006

Authors and Affiliations

  • Harald Schütz
    • 1
    Email author
  • Alexandre Paine
    • 1
  • Freidoon Erdmann
    • 1
  • Günter Weiler
    • 1
  • Marcel A. Verhoff
    • 1
  1. 1.Institute of Legal MedicineJustus-Liebig-UniversityGiessen

Personalised recommendations