Determination of quinoxaline antibiotics in fish feed by enzyme-linked immunosorbent assay using a monoclonal antibody

Abstract

Olaquindox (OLA), mequindox (MEQ), and quincetone (QCT) are widely used synthetic antibiotics of the quinoxaline-1,4-dioxide family. However, no studies have focused on the detection of sum of OLA, MEQ, and QCT by mAb-based enzyme-linked immunosorbent assay (ELISA). In this study, a specific mAb 2F3 against OLA, MEQ, and QCT was successfully prepared. Furthermore, using the mAb 2F3, an indirect competitive ELISA (icELISA) was developed using MQCA (quinoxaline marker) coupled to OVA as the heterologous coating antigen. Under optimized assay conditions, the IC 50 values were 1.03, 1.54, and 1.73 ng ml À1 for OLA, MEQ, and QCT, respectively. The recoveries ranged from 82.1% to 96.3%, and no cross-reactivity with other compounds was detected except for carbadox (0.9%). The developed icELISA was rapid and reliable for the determination of sum of OLA, MEQ, and QCT in fish feed.

Using molecular descriptors for assisted screening of heterologous competitive antigens to improve the sensitivity of ELISA for detection of enrofloxacin in raw milk

“…An icELISA was performed to evaluate the affinity of the antibodies to the analyte in serum or superna-tants of hybridoma cells as described previously (Peng et al, 2015). The icELISA procedures were as follows: 100 μL/well of ENR-OVA solution (0.1 μg/mL, diluted in 0.01 M CB) was added to the 96-well plates and incubated at 37°C for 2 h. The plates were washed 3 times with 0.01 M PBS-Tween and blocked with 100 μL/well of 0.01 M CB containing 0.2% (wt/vol) gelatin at 37°C for 2 h. After washing, 50 μL of ENR (0, 0.05, 0.1, 0.5, 1, 5, and 10 ng/mL) in PBS and 50 μL of 0.1 μg/mL mAb in antibody dilution solution were added and incubated at 37°C for 30 min.…”

Development of a highly sensitive ELISA and immunochromatographic strip to detect pentachlorophenol

“…The assay was based on the icELISA format (Liu et al, 2015;Peng et al, 2015). Polystyrene 96-well microtiter plates were coated with 100 μl/well hapten-OVA (0.03 μg/ml in CBS) overnight at 4°C.…”

Sandwich ELISA and immunochromatographic strip of Kunitz trypsin inhibitor using sensitive monoclonal antibodies

“…KTI, previously dissolved in physiological saline, was emulsified (1 mg/ml, 100 µg) in FCA for the first immunization of each mouse. Booster immunizations were performed every 21 d, and the immunogen dose was adjusted to 50 µg KTI emulsified in FIA (Guan et al, 2015;Peng et al, 2015). After 7-10 d of the third immunization, sera were collected from the tail of each mouse, and the titer was detected using indirect ELISA (Kong, Song, et al 2015;Kong, Liu, Xing, & Xu, 2015).…”