dual-label chemiluminescence strategy for multiplexed immunoassay of 20 fluoroquinolones, 15 β-lactams, 15 sulfonamides, and cap in milk

In this paper, a novel dual-label time-resolved chemiluminescent multiplexed immunoassay (dltrc-mia) based on the distinction of the kinetic characteristics of horseradish peroxidase (hrp) and alkaline phosphatase (alp) with approximate estimation approach for simultaneous determination of 20 fluoroquinolones (fqs), 15 β-lactams, 15 sulfonamides (sas), and chloramphenicol (cap) in milk was developed. the strategy integrated a single-chain variable fragment–alkaline phosphatase fusion protein (scfv-alp), a recombinant penicillin-binding protein (pbp) 2×*, a monoclonal antibody (mab), and a polyclonal antibody (pab) in one immunoassay and in a single well together to fulfill the simultaneous detection of 51 low-molecular weight contaminants (20 fqs, 15 β-lactams, 15 sas, and cap). the limits of detection for fqs, β-lactams, sas, and cap range from 0.29 μg l−1 for ciprofloxacin (cip) to 81.6 μg l−1 for trovafloxacin (tro), 0.27 μg l−1 for ceftiofur (cef) to 44.1 μg l−1 for cephalexin (cel), 0.089 μg l−1 for sulfadimethoxine (sdm) to 2.7 μg l−1 for sulfadiazine (sdz), and 0.028 μg l−1 for cap, respectively. the results demonstrated that the detection limits of dltrc-mia meet the requirement of detection levels for 51 drug residues in milk, suitable for high-throughput screening of low-molecular weight contaminants.