Sensitive immunoassays based on a monoclonal antibody for detection of marbofloxacin in milk

Abstract

Marbofloxacin (MBF) is a key class of synthetic fluoroquinolone antibiotic that is commonly used as a veterinary drug. However, excess residue of MBF in animal-derived food samples, such as milk, is harmful to consumers. In this study, 4 mAb against MBF, namely, M4E3, M7A6, M3C7, and M5C6, were produced. Indirect competitive (ic) ELISA revealed that the M4E3 exhibited the highest sensitivity with a half-maximal inhibitory concentration (IC 50 ) of 0.07 ng/mL and a limit of detection of 0.01 ng/mL for detection of MBF.The results of the cross-reactivity experiment revealed that the M4E3 could detect lomefloxacin, ofloxacin, fleroxacin, pefloxacin, danofloxacin, and enrofloxacin sensitively with IC 50 of 0.02, 0.07, 0.18, 0.27, 0.30, and 0.32 ng/mL, respectively. Meanwhile, a cross-reactivity experiment showed that the M4E3 had a crossover rate of more than 20% with these fluoroquinolone analogs. A weak crossover rate below 1.11% was observed with 14 other fluoroquinolone analogs. The recovery rate of MBF in milk ranged from 72.28% to 129.19%, which showed that the developed indirect competitive ELISA was effective in detecting MBF in milk. Furthermore, a visual colloidal gold-based immunochromatographic assay was developed for detecting MBF with a cut-off value of 1 ng/mL in both phosphate-buffered saline and a milk sample by using this mAb. Given this sensitive mAb, both indirect competitive ELISA and colloidal gold-based immunochromatographic assay could be effective screening tools for detection of MBF in milk.

Europium Fluorescent Nanoparticles-Based Multiplex Lateral Flow Immunoassay for Simultaneous Detection of Three Antibiotic Families Residue

“…Under optimised conditions, single-EuNPs-FIA could simultaneously detect 13 types of SAs, seven types of FQs and four types of TCs respectively, determining its potential as a competitive tool for primary mass screening in comparison to other analytical methods. However, Europium has been applied to the detection of streptomycin in milk, however, the detection was limited by a single target ( Luo et al, 2020 ). The increase in the number of controlled compounds in medical diagnostics and ecological assays makes a single detection system inadequate for the detection needs ( Supplementary Table S3 ).…”

Quantum dot bead-based competitive immunochromatographic assay for enterotoxin aureus A detection in pasteurized milk

“…Herein, the anti-SEA mAb is prepared by injecting the SEA protein in BALB/c mice, and a QB-based ICA (QB-ICA) with competitive format has been developed for the rapid and quantitative detection of SEA in pasteurized milk, in which highly luminescent QB are synthesized by encapsulating numerous CdSe/ZnS quantum dots (QD) into the polymer matrix by using the microemulsion technique (Luo et al, 2020). The QB-ICA is prepared by labeling anti-SEA mAb on QB, whereas the SEA protein is sprayed on the nitrocellulose (NC) membrane as T line (Shao et al, 2018).…”

Fluorescent strip sensor for rapid and ultrasensitive determination of fluoroquinolones in fish and milk

“…Nowadays, a variety of analytical techniques for analysis of FQs in animal-derived foods have been developed based on microbiological assay, 12,13 chromatographic techniques, 10,14 electrochemical sensors, 11,15,16 chemiluminescent methods, 17 and immunoassays. [18][19][20][21] Among them, immunochromatography is a powerful strategy for the development of rapid detection methods due to its obvious characteristics of economic efficiency, sensitivity and simplicity. [22][23][24] Immunochromatographic assay (ICA) can realize qualitative and quantitative analysis of one or a group of compounds in complex sample matrices with simple pretreatment.…”