Mechanism Of Interference

In the competition type immunoassays, interference from antibody can be explained from the following mechanisms.

1. If the assay uses labeled analyte (or its analog), the interfering antibody may interact with the label causing reduced signal and false-positive results. The same phenomenon may occur even if the assay antibody is labeled and the competing analyte (or its analogs) on the "capture reagent" binds to the interfering antibody thus reducing signal.

2. However, if only the analyte (when the assay uses an analog of the analyte in the reagent) interacts with the interfering endogenous antibody, then analyte concentration in the assay reaction decreases, causing false-negative results

3. Heterophilic antibody may interact with the assay antibody, reducing its effective concentration (via steric hindrance). Such interaction decreases assay signal generating a false-positive result.

The competition immunoassays, which are commonly used in analyzing small (TDM/DAU) molecules, are in general less affected by heterophilic antibody. In contrast, two-site immunometric assays (sandwich type assays) which are used for analytes with relatively higher molecular weights than drug molecules (cardiac troponin I, hCG, and so on) suffer more from interference of heterophilic antibodies. In these type of assays, the heterophilic antibody commonly bridges between the two assay antibodies ("capture" and "label") used, creating false sandwich complexes and false positive results.

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