Matrix effects immunoassay interference

The analytical response generated by an immunoassay is caused by the interaction of the analyte with the antibody. Although immunoassays have greater specificity than many other analytical procedures, they are also subject to significant interference problems. Interference is defined as any alteration in the assay signal different from the signal produced by the assay under standard conditions. Specific (cross-reactivity) and nonspecific (matrix) interferences may be major sources of immunoassay error and should be controlled to the greatest extent possible. Because of their different impacts on analyses, different approaches to minimize matrix effects and antibody cross-reactivity will be discussed separately. 

Evaluation, characterization, and testing of a particular analytical method is necessary to ensure the intended use of the method is met. In general, this process requires the determination of intra-and interlaboratory studies for precision and bias, method detection limits, matrix effects, interferences, limits of reliable measurements and ruggedness of the method. Before the EPA commits time and resources for an in-depth evaluation study, the developer must meet certain developmental criteria or justify why they were not met. The developer must also clearly define all necessary reagents as well as the underlying basis of the immunoassay. 

It can be concluded that immunoassays can serve very well in the surveillance and operational monitoring mandated within the WFD. Nevertheless, the possibility of interference from cross-reacting compounds and matrix effects must be taken into account. 

Matrix effects can be a problem for immunoassays, especially for a method without any prior sample clean-up. It can be caused by either nonspecific or specific interferences from the sample matrix and reagents. Possible matrix effects can... 

Matrix effects are particularly troublesome in immunoassay methods. Possible matrix interference and non-specific binding must be evaluated and documented in a number of different ways during the method vahdation ... 

Immunoassays are subject to some effects and interferences unique to immunoassay and some common to all chemical assays (Wu 2000). Establishment of an assay should include some assessment of antibody interference, signal interference, and matrix and hook effects. Interference may be caused by cross-reactivity or by an endogenous metabolite (e.g., in steroid hormone immunoassays) or a xenobi-otic (or metabolite) with a similar structure this problem occurs more frequently... 

Since FPIAs are conducted as homogeneous immunoassays, they are susceptible to effects from endogenous fluorophores and from intersample variations. Such problems and others due to the sample matrix are largely avoided by sample dilutions of several hundredfold. Low-affinity, nonspecific binding of tracers to sample proteins, when present in sufficiently high concentrations, can result in a falsely elevated polarization signal. Interference from sample proteins can be eliminated when warranted, by proteolytic hydrolysis with pepsin.(46)... 

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