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

ABSTRACT

BACKGROUND: 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. RESULTS: 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. CONCLUSION: 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.