Magneto Immunofluorescence Assay for Quinolone Detection in Bovine Milk Silvina V. Kergaravat 1,2 & Orlando G. Nagel 3 & Rafael L. Althaus 3 & Silvia R. Hernández 1 Received: 1 November 2019 /Accepted: 22 March 2020 # The Author(s) 2020 Abstract Quinolone is a family of widely prescript antibiotics in veterinary medicine. Their excessive use can be causative of quinolone residues in foodstuffs of animal origin. Therefore, a magneto immunofluorescence assay in microwell plate has been developed and validated to screen quinolone in bovine milk. The assay is based on an indirect competitive method where the quinolone in the milk sample competed with the quinolone immobilized on magnetic beads for the anti-quinolone antibody. Then, an anti-IgG antibody–labeled peroxidase and their respective substrates were added and the fluorescent signal was indirectly related with the quinolone concentration. Recoveries of quinolones from 82 to 102% were obtained in skimmed, whole, and raw milk samples in concordance with the results obtained by a reference method. Seven quinolones were evaluated quantitatively by the method, obtaining limits of detection of 10 μgL -1 for ciprofloxacin and marbofloxacin, 13 μgL -1 for enrofloxacin and danofloxacin, 22 μgL -1 for norfloxacin, 29 μgL -1 for sarafloxacin, and 30 μgL -1 for ofloxacin. For the screening detection of quinolones, ciprofloxacin was used as model with decision limit (CCα) and detection capability (CCβ) of 103 and 106 μgL -1 , respectively. High cross-reactivity (> 100%) was obtained to all assayed quinolones while negligible cross-reactivity (< 0.1%) to other antibiotics (β-lactams, aminoglycosides, macrolides, tetracyclines, and sulfamides) was observed. The validation provided evidence that the method is suitable to be applied in routine analysis for the detection of quinolones in bovine milk. Keywords Quinolone . Magnetic bead . Immunofluorescence assay . Bovine milk . Screening method Introduction Antibiotics are widely used for disease prevention and treat- ment as well as growth promotion in animal husbandry (Van Boeckel et al. 2015). Unfortunately, the use of antibiotics, especially if not used according to veterinary’ s prescription, can result in drug residues in animal-origin food such as milk. The quinolones (QNs), together with β-lactams, tetracycline, and sulfonamides, are the most often used antibiotics in dairy cattle management (Song et al. 2015). The presence of antibiotic residues in milk also gives rise to public health concerns due to the development of drug resistance in intesti- nal bacteria populations (Bilandžić et al. 2011). To ensure food safety, different control agencies have established maxi- mum residue limits (MRLs) for foodstuffs of animal origin, including bovine milk (Codex Alimentarius 2010 ; Commission Regulation 2010). Specifically, for quinolones, the European Union has established MRLs of 30 μgL -1 for danofloxacin (DAN), 75 μgL -1 for marbofloxacin (MAR), and 100 μgL -1 for both enrofloxacin (ENR) and ciprofloxa- cin (CIP) (Commission Regulation 2010). Currently, the available methods to determine QNs in milk are mainly dependent on the complex high-performance liq- uid chromatography (HPLC) with diode-array detection (Herrera-Herrera et al. 2013) or coupled mass spectrometry (Zhang et al. 2015; Petrie et al. 2016), capillary electrophore- sis (Zhou et al. 2008; Xu et al. 2015) and microbiological method (Linage et al. 2007; Nagel et al. 2013; Tumini et al. 2017). However, these methods generally require highly qual- ified operators and expensive equipment, and, in general, are time consuming, therefore, delay a result to solve the prob- lematic situation. * Silvina V. Kergaravat skergaravat@fbcb.unl.edu.ar 1 Laboratorio de Sensores y Biosensores, Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, 3000 Santa Fe, Argentina 2 Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), CCT Santa Fe, 3000 Santa Fe, Argentina 3 Cátedra de Biofísica, Departamento de Ciencias Básicas, Facultad de Ciencias Veterinarias, Universidad Nacional del Litoral, 3080 Esperanza, Argentina Food Analytical Methods https://doi.org/10.1007/s12161-020-01749-9