Preparation of anti-Pefloxacin Antibody and Development of an Indirect Competitive Enzyme-Linked Immunosorbent Assay for Detection of Pefloxacin Residue in Chicken Liver

Abstract

Pefloxacin has been increasingly used in veterinary medicine to treat microbial infections. To avoid using a labor-intensive instrumental method to detect the residue of pefloxacin in food, a simple and convenient indirect competitive enzyme-linked immunosorbent assay method has been developed in this study. The antibody generated from immunogen cationized bovine serum albumin-pefloxacin showed high sensitivity toward pefloxacin with an IC50 value of 6.7 ppb in buffer and was suitable for a screening assay to detect the residue of pefloxacin in food products. The antibody has been assessed using rapid enzyme immunoassays to exploit its specificity. The antibody prepared shows cross-reactivity with a few other (fluoro)quinolones including fleroxacin (116%), enrofloxacin (88%), and ofloxacin (10%). The assay measured drug residue in chicken liver spiked with pefloxacin with an interassay coefficient of variation of 13.6% or less and an intra-assay coefficient of variation of 10.9% or less. The average recovery rates at 0.5, 5, 10, 50, and 100 ppb were in the range of 86-106% for interassay and in the range of 87-103% for intra-assay, respectively.

Preparation of high-affinity rabbit monoclonal antibodies for ciprofloxacin and development of an indirect competitive ELISA for residues in milk

“…The immunogen CPFX-BSA and the coating antigen CPFX-OVA were both prepared by carbodiimide active ester method as per a previous paper (Lu et al, 2006). A total of 3.86 mg of CPFX, 20.64 mg of EDC, and 5.76 mg of NHS were added to 0.23 ml DMF in order.…”

“…Therefore, more and more organizations have issued maximum residue levels (MRLs) and withdrawal criterion for FQs. According to different food resources and drugs, the MRLs of FQs are set in a range of 30-1500 μg/kg (Lu et al, 2006;Zhao et al, 2007).…”

“…Numerous studies have been reported on developing ELISA methods for the detection of FQs, including CPFX (Mellgren and Sternesjö, 1998), enrofloxacin (Mellgren and Sternesjö, 1998;Watanabe et al, 2002), norfloxacin (Watanabe et al, 2002;Huet et al, 2006), pefloxacin (Mellgren and Sternesjö, 1998;Huet et al, 2006), and sarafloxacin (Lu et al, 2006). Moreover, Bucknall et al (2003) reported results concerning a polyclonal antibody against CPFX with a limit of detection (LOD) of 4 ng/ml (Bucknall et al, 2003).…”

Development of an electrochemical immunoassay for detection of gatifloxacin in swine urine

“…The cross-reactivity studies were carried out with the electrochemical immunoassay by adding various competitors in concentrations from 0.01 to 1000 ng/ml (Lu et al, 2006). The results confirmed that the anti-GAT antibody was highly specific and showed little cross-reactivity with lomefloxacin, ciprofloxacin, or ofloxacin (Table 1).…”

An indirect competitive enzyme-linked immunosorbent assay for determination of norfloxacin in waters using a specific polyclonal antibody

“…Enzyme-linked immunosorbent assay (ELISA) has been extensively used for screening of veterinary drug residues including fluoroquinolones in food and animal tissues (e.g., muscle, kidney, liver, serum, milk, and egg) due to its sensitivity, specificity, rapidity, and simplicity [20][21][22]. Nevertheless, using ELISA for determination of fluoroquinolones and other pharmaceuticals in water samples has still been rarely reported [23].…”