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Immunoassay blockers are used in diagnostic assays to reduce non-specific binding and other interference that can lead to false-positive results. Specifically, antibody interference from human anti-mouse antibodies (HAMA), rheumatoid factor (RH), and heterophilic antibodies (HA) are a major concern in both paired monoclonal sandwich assays and competitive assays. In IgM detection assays, IgG antibodies which are present in 10-15x higher concentrations than IgM can also reduce assay sensitivity due to their sheer abundance. Specialized blockers are required to reduce antibody interference from HAMA, HA, RF and IgG to ensure an assay's accuracy and improve its sensitivity.
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TRU Block™ – Targeting all Heterophile Interferences
IgG Blockers for IgM Detection Assays
Competition between IgG and IgM antibodies for antigen binding sites can significantly reduce assay sensitivity in IgM detection assays. In addition, IgG can form immune complexes with Rheumatoid Factor (RF) and compete with specific IgM for substrate binding sites. Removal of IgG and RF-IgM can be accomplished by pre-treating the patient specimen with goat anti-human IgG or IgM diluent.
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FAQs: Immunoassay Interference Blockers
Immunoassays used for human in vitro diagnostics (IVD) typically use animal-derived antibodies to recognize specific disease markers. Some patients have antibodies in their blood that can react with animal antibodies in the immunoassay and cause a false result. Although the frequency of these interferences is low, false-positive results have a significant negative impact on the quality and competitiveness of a diagnostic assay as well as on the lives of those individuals who have been falsely diagnosed.
Human anti-mouse antibodies (HAMA) is the common antibody interference which causes both positive and negative interferences in two-site mouse monoclonal antibody-based assays.
No. HAMA only represents one type of heterophilic antibody (HA) interference – others include HA to animals such as goat (HAGA), sheep (HASA), and rabbit (HARA) which can cause false results when antibodies originating from these animals are used in immunoassays. In addition to HA there is another class of interference called Rheumatoid factor (RF), which is an autoantibody that reacts with the patient’s own immunoglobulin (Ig) and can cross react with animal Ig, similar to HA/HAMA
interference.
Rheumatoid factor is an autoantibody (usually IgM) directed against the Fc portion of IgG. It is common in people with rheumatoid factor, occurring in about half of all people with the disease, but about 1 in 20 people without rheumatoid arthritis can also have high levels and is more common in elderly patients.
RF, similar to heterophilic antibodies, has the potential to cause falsely elevated test results by cross-linking the assay antibodies, even in the absence of analyte, most often via binding to the Fc-part of assay antibodies. RF can react against different species of IgG, including human and rabbit.
Our recommendations are: a) use the species of the blocker the same as the host of the capture or detection antibody; 2) use at a concentration of Mouse or Goat IgG equivalent to ten times the concentration of the MAb/PAb being used in the assay. (e.g. if 5ug/mL of Ab/conjugate, add 50ug/mL of animal IgG)
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Have questions about a product? Want to learn more about Meridian’s molecular or immunoassay reagent portfolio? We want to hear from you!