Exonuclease III-assisted nucleic acid amplification fluorescence immunoassay for the ultrasensitive detection of chloramphenicol in milk

Herein, an integrated approach based on immunoassay and exonuclease III-assisted nucleic acid amplification (EANA), which is defined as exonuclease III-assisted nucleic acid amplification fluorescence immunoassay (EA-NAFIA) was developed for the detection of chloramphenicol (CAP). Au nanoparticles (AuNPs) modified with antibody and DNA were used as bifunctional probe, in which antibody captured the target, and DNA initiated the signal amplification through EANA. A detection linear range from 0.01 to 5 ng/L was obtained under optimized conditions, and the detection limit was as low as 0.001 ng/L. There was no cross reaction with streptomycin, kanamycin, gentamicin, thiamphenicol. This assay was successfully applied to detect CAP in skim milk, with acceptable recoveries. This kind of integrated strategy demonstrated the excellent hybrid performance of indirect immunoassay with ultrasensitive sensitivity.