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Table. 3.

Table. 3.

Troubleshooting the most frequent types of immunoassay interference

Interference type Error type Identification Troubleshooting
Method-specific interference
Biotin Competitive assay: ↑
Sandwich assay: ↓
Patient’s file
Adsorption if validated
Withdraw biotin if possible, and obtain a new sample
Test with an alternative method
Anti-streptavidin antibody Competitive assay: ↑
Sandwich assay: ↓
Adsorption if validated Test with an alternative method
Signal-specific antibody interference (anti- ruthenium, anti-alkaline phosphatase) Competitive assay: ↑
Sandwich assay: ↓
Alternative method comparison Alternative method needed
Fluorescence signal ↑ or ↓ Fluorescein injection Obtain a new sample after washout
Alkaline phosphatase label ↑ or ↓ Recombinant alkaline phosphatase therapy Obtain a new sample after washout, if possible
Common interference
Hook effect (prolactin, HCG, Tg, calcitonin) Progressive dilutions Two coherent successive dilutions
Cross-reaction Competitive assay: ↑ Sandwich assay: ↑ or ↓ (depending on specificity characteristics) Patient’s file, therapeutic information Appropriate interpretation
LC-MS/MS control (steroid assays)
Macro analyte (prolactin, TSH, FSH, LH) PEG precipitation
Recovery test
Gel filtration chromatography
Appropriate interpretation and percent recovery post-PEG
Anti-insulin Ab (insulin assay) ↑ or normal Assay anti-insulin Ab
Anti-Tg Ab (TG assay) Assay anti-Tg Control TG with LC-MS/MS, if available
Burosumab therapy (intact FGF23 assay) Patient’s file, therapeutic information C-terminal FGF23 assay, if available
Heterophile Ab, HAAA, RF (all assays) Mostly ↑
↓ possible
Dilution test
HBT pretreatment
PEG pretreatment
RF assay
RF adsorbent
Adding non-immune Ig
Patients’ file: looking for immunotherapy, auto-immune context
Alternative method needed
Gammopathy (all assays)
↓ possible
Dilution test
HBT pretreatment
PEG pretreatment
Protein electrophoresis
Alternative method needed
FDH, thyroid hormone autoantibody (FT4, FT3 assays) Specific assay Notify in patient’s file, alternative methods needed

Interferences are either method-specific or common to all immunoassays (although errors of different magnitudes are recorded among the different methods).

Abbreviations: Ab, antibody; HAAA, human anti-animal antibody; FDH, familial dysalbuminemic hyperthyroxinemia; FGF, fibroblast growth factor; FSH, follicle stimulating hormone; FT4, free thyroxine; FT3, free triiodothyronine; HBT, heterophile blocking tube; LC-MS/MS, liquid chromatography coupled to tandem mass spectrometry; LH, luteotropin hormone; PEG, polyethylene glycol; RF, rheumatoid factor; TSH, thyroid stimulating hormone; Tg, thyroglobulin.

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