An Ultrasensitive Immunoassay Strip for Simultaneously Detecting Cyproheptadinehydrochloride and Six Phenothiazinesin Feedstuffs Based on a Monoclonal Antibody

Abstract Background: An antihistamine cyproheptadine (CPH) and phenothiazines (PZs)sedative hypnoticshave a similar tricyclic structure, these drugs are often illegally added to food animal feedstuffs due to their price advantage and significant effect in promoting growth and improving meat quality. However, the abuse of these drugs may lead totheir residues in animal products, thereby causing harmful effects such as allergies and dermatological reactions on human health.To supervise the use of prohibited drugs and ensure food safety, it is necessary to establish a simple and effective screening method to detect CPH and PZs.In this research, an artificial antigen against cyproheptadine(CPH) was successfully synthesized by bromoacetic acid substitution method. An ultrasensitive and broad-specific monoclonal antibody (mAb) able to recognize CPH and six phenothiazines (PZs) was produced. Based on the gold-labeled mAb, an immunochromatographic strip (ICS) was established. Results :The 50% inhibition concentration (IC 50 ) of the produced mAb against CPH was identified as 0.036 ng mL -1 by ELISA, and the cross-reactivities for six PZs were from 6.33% to 63.16%. The visual detection limits(cut-off values) of the developed ICS ranged from 5 to 100ngg -1 in feedstuffs. Take a reading by strip reader, the IC 50 was from 0.570 to 7.750ng g -1 . In addition,a recovery experiment was carried out to verify the reliability of the ICS. The intra-assay recoveries were from 79.83% to 103.38% with the highest coefficient of variation (CV) of 11.31%. The inter-assay recoveries were from 79.00% to 96.63% with the highest CV of 12.66%. Conclusions : We have successfully produced a broad-spectrum monoclonal antibody and established an ICS for simultaneouslydetecting CPH and six PZs drugs.In brief, the proposed ICS was considered suitable for qualitatively and quantitatively monitoring CPH and PZs in feedstuffs.