Development of a rapid detection method for enrofloxacin in food

Develop the ic-ELISA rapid detection method of Enrofloxacin (ENR). Corresponding antibodies are obtained by animal immunity to identify their titer and specificity. The optimal coating time was obtained by indirect competition ELISA, and the antigen coating time, suitable coating concentration, primary antibody dilution factor, blocking solution blocking time, primary antibody reaction time and secondary antibody reaction time were optimized, and the specificity and accuracy of the method were evaluated. The ic-ELISA rapid detection method of ENR, IC₅₀ was 9.13 ng/mL, and the linear detection range (IC₂₀-IC₈₀) was 4.16-20.03 ng/mL. The LOD limit is 2.11 ng/mL. The cross-reactivity rate of 9 fluoroquinolones was above 10%, and the average recovery rate was above 80%. The reason why the heterologous coating is more sensitive may be due to the fact that the piperazine group of ofloxacin is one less carbon atom than enrofloxacin, and ofloxacin is connected to the main ring by N and O hybridization, while enrofloxacin is connected to the main ring through a ternary ring, these two reasons may cause the charge density of extracyclic oxygen at the ofloxacin binding site to be higher than that of enrofloxacin, and the binding ability to antibodies is stronger. Therefore, when heterologous coating, the competitive inhibition rate against enrofloxacin is higher and the effect is better. The conclusion obtained through this experiment is that the detection method has strong broad spectrum and good sensitivity, and can quickly detect the total amount of enrofloxacin and its seven common fluoroquinolones in fish and eggs.