Most therapeutic drug monitoring (TDM) and drugs of abuse (DAU) testing are performed in clinical laboratories by using automated systems. There are two main technologies used in such determination. The most common one is immunoassay, which has been described in detail in Chapter 3. Immunoassay has the benefits of high sensitivity and specificity and thus has become the major technique in TDM and DAU assays. The second method used in TDM/DAU is enzymatic and/or chemistry methods. In these assays, the specificity of an enzyme is used to generate analyte-specific signals (in direct or indirect reaction), and the concentration is calculated based on generated signal. There are four major sources of "endogenous" interference that derive from specimen condition, especially in serum or plasma (Table 1).

Table 1

Specimen Appearance and Causative Interferents

Specimen appearance

Causative Interferent


Greenish, icteric

Red, hemolyzed


Nothing specific. Hyper-proteinemia may make the specimen viscous. There may be protein fibers in stored plasma or serum samples

Bilirubin; Reference Range (NR): < 1 mg/dL; High levels up to 20 mg/dL may be found in jaundiced patients

Hemoglobin; NR: < 1 mg/dL; Higher concentrations are found in vivo (hemolytic diseases) or in vitro (preanalytic causes)

Lipids (chylomicrons and VLDL)

Protein (includes paraproteins)

Different conjugates of bilirubin; interference caused by photometric or fluorimetric properties of bilirubin. Bilirubin may participate in side reactions interfering with the assay. Additionally, bilirubin may interfere in free drug assays by changing the distribution of drug between free versus complexed compartments

Hemoglobin has photometric, fluorimetric, and chemiluminescent properties which may interfere in immunoassays using such signals

Lipids interfere mainly by light scattering and absorption of lipoprotein micelles Both hypo- and hyper-proteinemia may cause assay interference

VLDL, very-low-density lipoproteins.

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