Using Fluorescence Polarization Immunoassay for the Determination of Flunixin in Milk

A fluorescence polarization immunoassay method is developed to detect residual amounts of the nonsteroidal anti-inflammatory drug flunixin, commonly used for prevention and treatment in veterinary medicine and animal husbandry. The assay required only 15 min, including sample preparation, and demonstrated a linear detection range of 20–5000 ng/mL with a limit of detection of 2 ng/mL. The immunoreagents exhibited high selectivity for flunixin, with cross-reactivity coefficients for structurally related compounds remaining below 0.01%. the method for detecting flunixin in milk was validated using a spiking (standard addition) approach and it was confirmed that the sample preparation procedure reliably detected the analyte within the linear range. These findings demonstrate that the method, which employs polyclonal antibodies, enables the rapid, sensitive, and selective detection of flunixin in milk.