Related Experiment Video
Updated: May 15, 2026
Using Reference Reagents to Confirm Robustness of Cytokine Release Assays for the Prediction of Monoclonal Antibody Safety
Published on: September 15, 2023
Cytokine measurements and possible interference from heterophilic antibodies--problems and solutions experienced with
Else Marie Bartels1, Søren Ribel-Madsen
1The Parker Institute, Department of Rheumatology, Copenhagen University Hospitals Bispebjerg and Frederiksberg, Denmark. emb@frh.regionh.dk
Insights
Reliable cytokine measurements using immunoassays require understanding and mitigating antibody interference. Methods like PEG precipitation can reduce interference from rheumatoid factor and other antibodies, ensuring accurate diagnostic data.
Area of Science:
- Immunology
- Biochemistry
- Medical Diagnostics
Background:
- Cytokines are crucial for understanding immune responses in health and disease.
- Accurate cytokine measurement is vital for diagnostics, relying on immunometric methods like ELISA and MIA.
- Various antibodies in biological fluids can interfere with immunoassay accuracy.
Purpose of the Study:
- To highlight potential pitfalls in cytokine measurement using immunoassays.
- To provide strategies for optimizing cytokine measurements with ELISA and MIA.
- To address interference from specific antibodies like heterophilic, human anti-animal, and rheumatoid factor (RF).
Main Methods:
- Focus on enzyme-linked immunosorbent assay (ELISA) and multiplex immunoassays (MIA).
- Discuss interference from heterophilic antibodies, human anti-animal antibodies, and rheumatoid factor (RF).
- Evaluate PEG precipitation as an affordable method to reduce RF interference, alongside other blocking agents.
Main Results:
- Immunometric techniques are particularly susceptible to interference from heterophilic and human anti-animal antibodies.
- Rheumatoid factor (RF), especially RF IgM, significantly interferes with immunometric cytokine measurements.
- PEG precipitation offers a cost-effective solution to mitigate RF interference.
Conclusions:
- Understanding and addressing antibody interference is essential for reliable cytokine data.
- Assay development and application must consider potential interference from RF, heterophilic, and human anti-animal antibodies.
- Implementing interference-reduction strategies ensures the validity of cytokine measurements in diagnostics.
Abstract:
Cytokines are important in the understanding of the immune process in health and disease and are valuable indicators in diagnostics. Measurements of cytokines are based on immunometric methods, and it is important to understand possible pitfalls in these methods to produce reliable cytokine data. This paper focuses on obtaining optimal measurements when applying enzyme-linked immunosorbent assay (ELISA) or multiplex immunoassays (MIA). Cytokines are measured in serum or plasma, as well as in various other body fluids, all containing a series of antibodies and the possibility of interference from these. Some antibodies, such as heterophilic and human anti-animal antibodies, are able to interfere with all immunoassays, but the immunometric techniques are most prone to serious interference from this source. Another type, rheumatoid factor (RF) is a composite of different autoimmune antibodies which can be present in both blood and synovial fluid. RF is present in some arthritic diseases as well as in some other medical conditions. When present, especially RF IgM is known to interfere with the immunometric measurements. A possible and affordable solution to diminish this interference is PEG precipitation, but other efficient, but more expensive, methods, such as precipitation using Protein L or commercially available blocking agents, are also available. Interference of RF is at present not tested in all cytokine assays, but degree of interference from RF, human anti-animal and heterophilic antibodies, as well as from other possible disease-specific antibodies, must always be considered when developing and applying new assays for cytokine measurements.
