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Principles of Immunoassay

The Immunoassay is a method that relies on antibody/antigen reaction. As the name implies, Immunoassay; Immuno from the word immune or immunity  the assay to analyzed. The instruments that use this method include Syva EMIT, Abbott ADX/TDX, Roche Cobas Mira, just to name a few. The IA is non-specific, which means if a person is taking Diazepam (Valium), a member of drug class of Benzodiazepines, the Immunoassay drug screen result would report it as positive for Benzodiazepines, and not positive for Diazepam. To determine specifically what member of the Benzodiazepine Drug Class, a confirmation usually by GC/MS is required

As mentioned earlier, the Immunoassay method involves the interaction between an antibody and an antigen. An antigen can be a drug such as Cocaine. The antibody is the protein that is specific for the drug Cocaine. The antibody binds specifically to the antigen's binding site. In other words, there must be an exact match (binding of the antibody to the antigen) for an antibody/antigen reaction to take place Figure 1. Think of a lock and key concept. If the key fits the lock, then the lock opens.

Figure 1 Antibody - Antigen reaction diagram referring to one of the main defenses against the infection

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      Antigen (drug)    +     Antibody                           

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Antibody binding with the  Antigen binding Therefore, a positive drug test test


Immunoassay False Positive Result

Since the IA method involves the antibody/antigen reaction, a false positive result can occur when multiple drugs or substances interact to form an antigen binding site similar to the antigen binding site of a different drug. For example, if a person is taking Ibuprofen and Aspirin simultaneously, after undergoing metabolisms can possibly produce an antigen binding site similar to the antigen binding site of Marijuana. Therefore, a false positive result for Marijuana is obtained assuming that the person has never used this drug.

Immunoassay False Negative Result

A false negative result is due in large part to the presence of adulterant that renders the antibody's ability to recognize and subsequently bind with the antigen (drug). Therefore, antibody/antigen reaction is inhibited. In simple terms, it is a lot like placing a blanket over the antigen so that the antibody is unable to bind with the antigen. " No binding, No reaction, No drugs". See Figure 2.

Figure 2. Effects of presence of adulterants
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Antigen (Drug)      +      Antibody 
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with adulterant 

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Therefore, No binding,  No Reaction, No drugs!!!

Limit of Sensitivity - see Cut-Off Limits

13. Negative Result - A result that means there is absolutely no drugs present in the sample.

14. Non-Detect Result t - A reported result that is below the limit of sensitivity for a particular drug.




Copyright 1999 [Toxicology Associates, Inc.]. All rights reserved.
Revised: January 13, 2010