Development of a Direct Competitive Enzyme-Linked Immunosorbent Assay for the Detection of Fluoroquinolone Residues in Shrimp

Abstract

Rabbit polyclonal antibodies were generated against norfloxacin, purified, and used as the basis of a direct competitive enzyme-linked immunosorbent assay for the screening of fluoroquinolones in shrimp. The developed method used a simple ethanol/acetic acid solvent extraction, which resulted in a 1.0-ng-norfloxacin/g limit of detection (based on the analysis of known negative and fortified shrimp samples). Norfloxacin extraction efficiencies were evaluated at two fortification concentrations and were greater than 70%, with an intra-assay variation less than 30%. The assay displayed greater than 10% cross reactivity against enro-, cipro-, sara-, and difloxacin. Incurred and known negative shrimp samples were analyzed and compared to the results obtained from an independent liquid chromatography tandem mass spectrometric method. All three instances in which fluoroquinolones were present at concentrations near or above the assay limit of detection (1.0 to 17 ng/g) were identified as positive by the newly developed assay, demonstrating the usefulness of this assay as a screening tool.

Synthesis of novel haptens against ciprofloxacin and production of generic monoclonal antibodies for immunoscreening of fluoroquinolones in meat

“…As described above, the anti‐SAR antibody showed lower CRs to other fluoroquinolones than did the anti‐NOR antibodies . In our opinion, this was because the molecular differences between SAR and other fluoroquinolones (except DIF) were larger than those between NOR and other fluoroquinolones.…”

“…The anti-NOR antibody showed high CRs to ENR (133%), NOR (100%), PEF (92%), CIP (50%) and DIF (13%), but showed low CRs to other analogues (2-9%). Tittlemier et al 22 synthesised the hapten and produced the antibody of NOR as the previous report 21 and the resultant antibody showed CRs of higher than 10% to ENR, CIP, SAR and DIF.…”

Biomimetic norfloxacin sensors made of molecularly-imprinted materials for potentiometric transduction

“…The same comments may apply to the electrophoretic-based procedures reported in literature [16]. Other methods are based on immunoassays [17]; although they provide specific responses, the overall procedure is long and too expensive for routine analytical measurements.…”

Performance of a New Microbial Test for Quinolone Residues in Muscle

“…Enzyme immunoassay kits for the quantitative analysis of quinolones in several matrices have been commercialised by different manufacturers. Generally, these kits have lower limits of detection than microbial methods but cross reactivities between the different quinolones detected are not optimal in most cases (Tittlemier et al 2008;Scortichini et al 2009). Antibodies used in ELISA tests for detection of quinolones have shown cross-reactivity values lower than 10% against several evaluated quinolones.…”

“…Analytical methods described in the literature for the analysis of quinolones in foods include instrumental techniques (Hernández-Arteseros et al 1998;Rose et al 1998;Roybal et al 2002), rapid immunological techniques (Tittlemier et al 2008;Scortichini et al 2009) and microbial methods (Currie et al 1998;Pikkemaat et al 2007;Fuselier et al 2000;Calderón et al 1996;Myllyniemi et al 2001;Cantwell and O'keeffe 2006;Gaudin et al 2009). …”