Thursday, September 13, 2007

False Negatives & False Positives

Qualitative Results
A qualitative drug test is one that provides a dichotomous result, that is, it indicates whether a sample is positive or negative for a specified drug. However, there are four possible results of a qualitative drug test.

A true-positive result occurs when the test correctly identifies the presence of a drug in the sample taken.
A false-positive result is one where the test incorrectly detects the presence of a drug where in fact no drug is present.
A true-negative result occurs when the test correctly confirms the absence of a drug.
A false-negative result is one where the test fails to detect the presence of a drug when it is in fact present.

Interpreting a Positive Test Result

A positive result indicates that the specific drug (or class of drug) is present at or above the designated cut-off level. Typically, the cut-off concentration is set to the lowest concentration the drug can be reliably detected following consumption. It considers environmental and analytical variability caused by such factors as passive contamination/ingestion, technological limits, etc.

False-positives resulting from qualitative screening
A false-positive result can occur when a benign substance in the biological sample mimics the chemical effect of the targeted substance on the test. The test indicates a positive result even though the targeted drug was absent. Such results have reportedly occurred after ingestion of antihistamines, certain anti- inflammatory drugs, cold and flu medications, and poppy seeds (Selavka, 1991). The false-positive rate for particular testing methods is discussed in the relevant chapters below. Although levels are generally low, it does highlight the necessity of appropriate confirmatory testing with parent / metabolite quantification to identify and safeguard against this.

Interpreting a Negative Test Result
In the majority of cases a negative result indicates that the parent drug (typcially the active ingredient) and / or its metabolites are absent in the biological sample. It does not mean that the person has not used the substance in the days or weeks prior to testing. The amount of drug present in the sample at the time of sample collection, and thus whether a positive result is obtained, is determined by a number of factors which include: the cut-off level used; the testing schedule employed; the biological sample analyzed; when the drug was ingested; the amount of drug ingested; the form in which it was ingested; and physical and pharmacological characteristics of the user.

When an initial screen result in negative and (1.) the individual ingests a drug and the concentration of the drug in the sample is at our above the cut-off, or (2.) the individual ingests a drug and the concentration of of the sample is below the the cut-off due to sample adulteration or substitution, the result referred to as a “false-negative”.
Relative to urinalysis, there are a number of actions an individual can take, to increase the likelihood of a false-negative result. An individual can adulterate the specimen via dilution by drinking excessive amounts of water (in vivo adulteration), or by adding water or chemicals that will affect the test (in vitro adulteration) (Coleman & Baselt, 1997). Hair testing may be susceptible to excessive washing (Rohrich, Zorntlein, Potsch, et al., 2000), bleaching (Yegles, Marson & Wennig, 2000) and other cosmetic hair treatment (Skopp, Potsch & Moeller, 1997). There are no currently proven methods to adulterate or substitute oral fluid.

Quantitative Results
Quantitative drug testing involves the determination of the specific concentrations of a parent drug and/or its metabolite(s) in a sample, typically via GC/MS and LC/MS/MS. In addition to confirmatory testing of qualitative screening results, quantitative results quantitative results using blood or oral fluid / saliva specimens can provide additional information regarding the quantity and frequency of drug use (Cone, 1997). Blood and/or oral fluid / saliva may also be useful when establishing impairment levels for certain drug classes. Urine and hair specimens are generally considered effective for historical use only. With knowledge of the drug’s pharmacokinetic parameters, including its half-life, an estimate of the frequency of new drug use can be obtained using quantitative analysis (Cone, 1997, Huestis and Cone 1998).

The Physiology of Urine Production
Urine is produced continuously by the kidneys and may be considered an ultrafiltrate of blood. During urine production the kidneys reabsorb essential substances. Excess water and waste products, such as urea, organic substances and inorganic substances, are eliminated from the body. Parent drugs (typically the active ingredient) are often present in urine in very low concentrations or not detected at all. Therefore distinguishing between codeine, heroin and morphine use, for example, can be difficult. Furthermore, inter-subject variations in urine drug concentrations, even after similar dosing, is high.
Absorption into urine is usually slow when a drug is orally administered and excretion may be delayed for several hours (approximately 6-9 hours) .Generally, a urine specimen will contain the highest concentration of parent drug and metabolite at this time period. As drug elimination usually occurs at an exponential rate, for most illicit drugs a dose will be eliminated almost completely within 48 hours. A number of factors influence detection times including the quantity of drug administered, parent drug and metabolite half- life, cut-off level used, and a number of physiological factors. It is also noted that for many of drugs, frequent, multiple dosing over extended periods of time can cause the drug to accumulate in the body resulting in significantly extended detection times.

The Physiology of Oral Fluid
Salivary gland is a term used to include any tissue that normally discharges a secretary product into the oral cavity. Thus, oral fluid refers to the mixture of fluid in the oral cavity. Saliva is a complex aqueous fluid (99% water) containing electrolytes (principally sodium,potassium, chloride and bicarbonate), proteins (mostly enzymes, including amylase) and muncin (Kidwell, Holland & Athanaselis, 1998). The mucin gives oral fluid its sticky character. Saliva also contains cell and food debris and oral microorganisms. The composition and production of oral fluid is determined by the relative contribution of the different glands, which in turn is dependant on a variety of factors including nutritional and emotional state, sex, age, season of the year, time of day, and a variety of diseases and pharmacological agents (Höld, 1996; United Nations, 1998)
The three major salivary glands are: (1) the parotid, at the top of the mouth, (2) the submandibular, at the base of the tongue, and (3) the sublingual, at the sides of the oral cavity. The parotid gland, responsible for about 25% of the saliva produced, excretes saliva derived primarily from blood plasma (serous fluid); the submandibular and sublingual glands excrete both serous fluid and mucin and contribute approximately 71% and 4% respectively (Kidwell, Holland & Athanaselis, 1998). The volume of saliva produced by an adult ranges from 500 to 1500 ml per day. Unstimulated saliva has a pH range between 5.6 and 7. Stimulation increases the pH to a maximum of 8 (Kidwell, Holland & Athanaselis, 1998).

A thin layer of epithelial cells separates the salivary ducts from the systemic blood circulation (capillaries). The lipid membrane of these cells determines which molecules may be transferred from blood plasma into oral fluid. Three routes have been identified that may transport a drug across the lipid membrane; these include active transport (secretion), passive diffusion through the membrane across a concentration gradient, and diffusion through pores in the membrane (ultrafiltration) (Höld, de Boer, Zuidema & Maes, 1996,United Nations, 1998). Some molecules with a low molecular mass (i.e. ethanol) may diffuse through the water-filled pores in the membrane. Other small molecules are primarily transported through secretion. For larger molecules (most drugs of abuse), passive diffusion across a concentration gradient is thought to be the major factor in transport (Höld, de Boer, Zuidema & Maes, 1996; Huestis & Cone, 1998). Equilibrium occurs between plasma and saliva. In plasma a large proportion of a drug is bound to proteins. Drug concentrations in oral vary with the free fraction of drug in plasma, and therefore mimic concentrations found in blood (Cone, 1993).
Interpretation of Drug Concentrations in Saliva
Saliva has been shown to be a suitable matrix for the detection of drugs of abuse, specifically cocaine and benzoylecgonine (e.g. Cone, 1993; Schramm, Craig, Smith, et al., 1993), heroin, 6-MAM and morphine (e.g. Goldberger, Darwin, Grant, et al., 1993), codeine (in Huestis & Cone, 1998b), methadone (e.g. Wolff, 1991) and amphetamines (Cone, 1993). Cannabis use is somewhat more difficult to detect in saliva though it has been shown to be possible (e.g. Menkes, Howard, Spears, et al., 1991). Saliva drug concentrations generally correlate well with the free fraction of drug in blood (Cone, 1993; Kidwell, Holland & Athanaselis, 1998).
Saliva can be used to provide both qualitative and quantitative information on the drug status of an individual undergoing testing for all drugs of abuse reviewed (Cone, 1993).

Much research into saliva testing has examined its utility as an alternative test matrix to blood and urine.

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