Optimization of condition for conjugation of enrofloxacin to enzymes in chemiluminescence enzyme immunoassay

• Carbodiimide condensation was a proper method for conjugation.
• Dialysis time was the most important factor to purify conjugate.
• Dilution buffer had little effect on the application of conjugates in CLEIA.
• Conjugates could be stable at 4 °C with no additive up to 30 days.

In this study, in order to find out a proper method for conjugation of enrofloxacin to label enzymes, two methods were compared and carbodiimide condensation was proved to be better. The results showed that the binding ratio of enrofloxacin and alkaline phosphatase (ALP) was 8:1 and that of enrofloxacin and horseradish peroxidase (HRP) was 5:1. This indicated that conjugate synthesized by carbodiimide condensation was fit for chemiluminescence enzyme immunoassay (CLEIA). Furthermore, data revealed that dialysis time was an important parameter for conjugation and 6 days was best. Buffer to dilute conjugate had little effect on CLEIA. The storage condition for conjugates was also studied and it was shown that the conjugate was stable at 4 °C with no additive up to 30 days. These data were valuable for establishing CLEIA to quantify enrofloxacin.

Figure optionsDownload as PowerPoint slide