Development and Optimization of an Indirect Enzyme‐Linked Immunosorbent Assay for Thiamphenicol

Abstract

The specificity of an enzyme-linked immunosorbent assay (ELISA) was modified by changing the chemical structure of the haptenated coating antigen. A competitive indirect immunoassay was developed, evaluated, and validated to measure thiamphenicol (TAP) in eels. The working range of the assay was placed between 0.45 and 8.35 mg/L. Moreover, the influence of several physicochemical parameters, such as incubation time, ionic strength, detergent concentration, and pH, were studied. The specificity of this immunoassay was evaluated using 23 structurally related compounds. Finally, a good correlation was observed between the chromatographic and immunoassay techniques while analyzing five certified eel samples.

Development and Validation of a Sensitive Indirect Competitive Enzyme-Linked Immunosorbent Assay for the Screening of Florfenicol and Thiamphenicol in Edible Animal Tissue and Feed

“…As shown in Table 1, several haptens had been designed to obtain the special antibodies against TAP, FF, and/or FFA in previous studies. Hao et al (2006) and Guo et al (2015) selected TAP as the starting material to generate a hapten and then to prepare antigens. Wu et al (2008), Luo et al (2009b), andFodey et al (2013) selected FFA as their haptens.…”

“…Synthesis of Hapten FF-SA The hapten FF succinate (FF-SA) was synthesized by coupling FF with succinic anhydride (SA) based on the modified method of Hao et al (2006). Briefly, a solution of FF (358 mg) in dichloromethane (15 mL) was added to succinic anhydride (159 mg) at room temperature.…”

“…Recently, FF and TAP are even administered to fish by incorporation into feed for control of bacterial diseases (Hao et al 2006;Luo et al 2009a). Therefore, the need for an effective residue monitoring programme in place for FF and TAP is clear.…”

“…Hao et al (2006) developed an indirect ELISA to test TAP in eel using a polyclonal antibody (pAb) against TAP with IC 50 values 2.5 μg L −1 . Wu et al (Wu et al 2008) produced a pAb against FFA and developed an ic-ELISA to detect FFA with an IC 50 of 3.34 μg L −1 in muscle of swine and chicken.…”

Development of an ultrasensitive ic-ELISA and immunochromatographic strip assay for the simultaneous detection of florfenicol and thiamphenicol in eggs

“…The hapten FF-HS, which involved coupling FF with succinic anhydride, was synthesized in our laboratory following the method described by Hao, Guo, and Xu (2006). Specifically, FF (1.07 g, 3 mmol), succinic anhydride (0.6 g, 6 mmol) and pyridine (50 mL) were mixed and heated at 60°C for 4 h in a reflux condenser.…”