Eu³⁺ -labeled IgG-based time-resolved fluoroimmunoassay for highly sensitive detection of aflatoxin B₁ in feed

Aflatoxin B₁ (AFB₁ ) is a kind of toxic and carcinogenic mycotoxin. A time-resolved fluoroimmunoassay (TRFIA) was established for quantitative detection of AFB₁ in feed using Eu³⁺ -labeled IgG as tracer.

Monoclonal antibody (McAb) against AFB₁ (9B11-D7) was prepared through immunization and cell fusion and was identified as high affinity, specificity and sensibility by enzyme-linked immunosorbent assay (ELISA). The 50% inhibition value (IC₅₀ ) was 0.81 ng mL^-1 , the limit of detection (LOD) was 0.10 ng mL^-1 and detection range was 0.10-3.94 ng mL^-1 . Goat anti-mouse immunoglobulin G (IgG) was modified by Eu³⁺ -DATT, generating Eu³⁺ -labeled IgG. Under optimal assay conditions, TRFIA was shown to be highly sensitive and specific in detection of AFB₁ . The IC₅₀ and LOD were 94.73 pg mL^-1 and 3.55 pg mL^-1 , respectively, and detection range was 3.55-1.11 × 10³  pg mL^-1 . Cross-reactivity with AFM₁ , AFB₂ , AFG₁ and AFG₂ was 31.26%, 37.6%, 127.46% and 35.74%, respectively, but zero with other analogues. In determination of AFB₁ spiked in feed sample, TRFIA showed high accuracy and precision. The average recoveries ranged from 93.71% to 97.80%, and coefficient of variation was 1.25-3.73%. Good correlation between TRFIA and HPLC was demonstrated for determination of AFB₁ in feeds, confirming the reliability of the developed method.

The developed TRFIA exhibited good potential for employment in the ultrasensitive detection of AFB₁ in feed and could be used to determine total aflatoxins. © 2017 Society of Chemical Industry.