Presentation on theme: "Forensic Drug Testing Part 1: Screening"— Presentation transcript:
1Forensic Drug Testing Part 1: Screening Roger L. Bertholf, Ph.D.Associate Professor of PathologyChief of Clinical Chemistry & Toxicology
2What is forensic drug testing? MDs order drug tests to evaluate the medical condition of a patientMedical drug testing, orClinical ToxicologyEmployers order drug tests to determine whether someone uses illegal drugsDrug testing for legal purposes, orForensic Drug Testing
3Medical vs. forensic drug testing Patient consent not requiredIdentity of specimen is presumedScreening result is sufficient for medical decisionResults are used for medical evaluationSubject must consent to be testedIdentity of specimen must be provedOnly confirmed results can be considered positiveResults are used for legal action
4Illegal Drug Use in the U.S. (1998 Household Survey) 13.6 million Americans use illicit drugs25 million in 19798.3% of youths age use marijuana14.2% in 19791.8 million Americans use cocaine5.7 million in 1985
7History of workplace drug testing 1960s – 1970s: The Department of Defense begins testing military personnel for illegal drug use.1986: President Reagan establishes the “Federal Drug-Free Workplace”.1988: Mandatory Guidelines for Federal Workplace Drug Testing Programs is published in the Federal Register.
8The “NIDA” programNIDA (now SAMHSA) requirements for drug testing were drafted by Research Triangle InstituteThe RTI established the National Laboratory Certification Program (NLCP)Drug testing for federal agencies (DOT, NRC, etc.) must be performed in a NLCP-certified laboratory
9Florida Drug-Free Workplace The Florida HRS (now AHCA) established a drug-free workplace program in 1990Specifications for the State of Florida program are similar to federal requirements, but there are notable differencesEmployees of Florida Drug-Free Workplace-compliant businesses must be tested in AHCA-licensed laboratories
10Comparison of NLCP Certified and AHCA Licensed Laboratories Florida Drug Free Workplace Program10 drugs + ethanolInspected every 6 monthsQuarterly proficienciesDirector must be board-certifiedFederal employees, federally-regulated jobs5 drugsInspected every 6 monthsQuarterly proficienciesDirector must be board-certified
11ScreeningSensitivity vs. specificity of analytical methods
12Performance characteristics of screening tests (80)(100)(50)(20)(15)(12)(10)1 - Sensitivity(5)Receiver Operator Characteristic(2)(1)Specificity
13Screening Procedure is designed to eliminate all negatives Positive screens are presumptiveNegative screens can be reviewed and released by a Scientific Review OfficerPositive screens are submitted for confirmatory testing
14Challenge question . . .We regularly use immunochemical methods for quantifying therapeutic drugs, but consider them “screening” methods for drugs of abuse.Why?
15Introduction to Homogeneous Immunoassay What is the distinguishing feature of homogeneous immunoassays?They do not require separation of bound and free ligandsDo homogeneous methods have any advantage(s) over heterogeneous methods?YesWhat are they?SpeedAdaptability
17Homogeneous immunoassays Virtually all homogeneous immunoassays are one-siteVirtually all homogeneous immunoassays are competitiveVirtually all homogeneous immunoassays are designed for small antigensTherapeutic/abused drugsSteroid/peptide hormones
18Typical design of a homogeneous immunoassay No signalSignal
19Enzyme-multiplied immunoassay technique (EMIT™) Developed by Syva Corporation (Palo Alto, CA) in 1970s--now owned by Behring DiagnosticsOffered an alternative to RIA or HPLC for measuring therapeutic drugsSparked the widespread use of TDMAdaptable to virtually any chemistry analyzerHas both quantitative (TDM) and qualitative (DAU) applications; forensic drug testing is the most common use of the EMIT methods
21EMIT™ signal/concentration curve Signal (enzyme activity)Antigen concentrationFunctional concentration range
22Fluorescence polarization immunoassay (FPIA) Developed by Abbott Diagnostics, about the same time as the EMIT was developed by SyvaRoche marketed FPIA methods for the Cobas FARA analyzer, but not have a significant impact on the marketLike the EMIT, the first applications were for therapeutic drugsCurrently the most widely used method for TDMRequires an Abbott instrument
23Molecular electronic energy transitions SingletAVRTripletICFPsecsecE0
25Fluorescence polarization Fluoresceininout( sec)Orientation of polarized radiation is maintained!
26Fluorescence polarization But. . .OHCRotational frequency 1010 sec-1inout( sec)Orientation of polarized radiation is NOT maintained!
27Fluorescence polarization immunoassay Polarization maintainedSlow rotationRapid rotationPolarization lost
28FPIA signal/concentration curve Signal (I/I)Antigen concentrationFunctional concentration range
29Cloned enzyme donor immunoassay (CEDIA™) Developed by Microgenics in 1980s (purchased by BMC, then divested by Roche)Both TDM and DAU applications are availableAdaptable to any chemistry analyzerCurrently trails EMIT and FPIA applications in market penetration
30Cloned enzyme donor Spontaneous Monomer (inactive) Active tetramer AcceptorMonomer(inactive)-Galactosidase
32CEDIA™ signal/concentration curve Signal (enzyme activity)Antigen concentrationFunctional concentration range
33Screening thresholds Why do we need screening thresholds? To ensure that results in all participating laboratories agreeWho determines the thresholds?The agency sponsoring the drug testing program (e.g., SAMHSA, State of Florida, or individual employer)
38Amphetamine stereochemistry Pharmacological preparations of amphetamine can be racemic d,l mixtures (Benzedrine) or pure d-amphetamine (Dexedrine)Most immunoassays are calibrated with d,l-amphetamine
39Methamphetamine stereochemistry d-Methamphetamine is 10 times more potent than the l isomerl-Desoxyephedrine is used in some non-prescription nasal decongestants
47SummaryScreening is the first step of a two-step process in forensic drug testingScreening methods are designed to eliminate negative specimensPositive screens are presumptiveSeveral homogeneous immunoassays have been developed for drug screening