How do we confirm heterophile antibody (heterophile ab) interference in thyroid function tests (TFT)?

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Confirming Heterophile Antibody Interference in Thyroid Function Tests

To confirm heterophile antibody interference in thyroid function tests (TFT), use heterophile antibody blocking tubes (HBT) for sample pretreatment and compare results across multiple assay platforms, as this is the most reliable approach to identify this interference.

Understanding Heterophile Antibody Interference

  • Heterophile antibodies can cause falsely elevated or decreased results in immunoassays, with falsely elevated values being more common in most thyroid function test platforms 1
  • These interferences occur when antibodies in the patient's sample cross-react with assay reagents, leading to thyroid function test results that are inconsistent with the patient's clinical presentation 2
  • The prevalence of heterophile antibody interference is estimated to be approximately 1.5-3% in commonly used automated thyroid assays 3

When to Suspect Heterophile Antibody Interference

Suspect heterophile antibody interference when:

  • There is a discrepancy between clinical presentation and thyroid function test results 2
  • Results from different thyroid hormone measurements (TSH, free T4, free T3) are inconsistent with each other 4
  • Thyroid function test results do not change appropriately in response to treatment or physiological stimulation 5
  • There are unexplained fluctuations in thyroid function test results over time 3

Confirmation Methods for Heterophile Antibody Interference

1. Sample Pretreatment with Heterophile Blocking Agents

  • Use heterophile antibody blocking tubes (HBT) to pretreat samples before repeating the assay 3
  • A significant decrease in measured values (>3 standard deviations or >56.91% change) after HBT treatment strongly suggests heterophile antibody interference 3
  • This is considered the gold standard approach for confirming heterophile antibody interference 1

2. Testing on Multiple Assay Platforms

  • Analyze the sample using different immunoassay platforms from various manufacturers (e.g., Siemens, Abbott, Roche, Beckman) 1
  • Significant discrepancies between platforms (e.g., normal results on one platform but abnormal on another) suggest interference 1
  • In one reported case, TSH results varied dramatically across platforms: 5.52 μIU/ml (Siemens), 0.54 μIU/ml (Abbott), 0.12 μIU/ml (Roche), and <0.015 μIU/ml (Beckman) 1

3. Dilution Testing

  • Perform serial dilutions of the sample and test for linearity 5
  • Non-linear results with serial dilutions suggest the presence of interfering antibodies 5
  • This method can be used as a complementary approach to confirm interference 3

4. Alternative Methodology

  • For thyroglobulin testing, mass spectrometry (MS) methods can be used to confirm interference, as they are not affected by heterophile antibodies 5
  • If available, compare immunoassay results with those obtained by alternative methodologies not based on antibody binding 5

Practical Approach to Confirm Heterophile Antibody Interference

  1. First step: When interference is suspected, repeat the test using heterophile antibody blocking tubes 3
  2. Second step: If available, test the sample on multiple assay platforms from different manufacturers 1
  3. Third step: Perform serial dilutions to check for linearity of results 5
  4. Fourth step: Consider alternative methodologies (e.g., mass spectrometry) if available 5

Common Pitfalls to Avoid

  • Failing to consider heterophile antibody interference when thyroid function test results don't match the clinical picture 2
  • Initiating unnecessary treatment based on falsely abnormal results without confirming their accuracy 3
  • Confusing heterophile antibody interference with other causes of abnormal thyroid function tests, such as euthyroid sick syndrome 6
  • Not distinguishing between heterophile antibody interference and anti-thyroid antibody interference (e.g., anti-Tg antibodies), which are different phenomena 3

Where to Get Testing Done

  • Most reference laboratories and large hospital laboratories can perform heterophile antibody blocking tube testing 3
  • For complex cases, specialized endocrine reference laboratories may offer more comprehensive interference testing panels 5
  • Consult with your laboratory director or endocrinologist for guidance on where to send samples for specialized testing 2

References

1
Research

Heterophilic antibody interference with TSH measurement on different immunoassay platforms.

Clinica chimica acta; international journal of clinical chemistry, 2021

2
Research

The Complex Web of Interferences With Thyroid Function Tests.

Endocrine practice : official journal of the American College of Endocrinology and the American Association of Clinical Endocrinologists, 2025

3
Research

Phantoms in the assay tube: heterophile antibody interferences in serum thyroglobulin assays.

The Journal of clinical endocrinology and metabolism, 2003

5
Research

False Positives in Thyroglobulin Determinations Due to the Presence of Heterophile Antibodies: An Underrecognized and Consequential Clinical Problem.

Endocrine practice : official journal of the American College of Endocrinology and the American Association of Clinical Endocrinologists, 2021

6
Guideline

Diagnostic Approach for Hypothyroidism

Praxis Medical Insights: Practical Summaries of Clinical Guidelines, 2025

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Professional Medical Disclaimer

This information is intended for healthcare professionals. Any medical decision-making should rely on clinical judgment and independently verified information. The content provided herein does not replace professional discretion and should be considered supplementary to established clinical guidelines. Healthcare providers should verify all information against primary literature and current practice standards before application in patient care. Dr.Oracle assumes no liability for clinical decisions based on this content.

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