An Improved Label-Free Indirect Competitive SPR Immunosensor and Its Comparison with Conventional ELISA for Ractopamine Detection in Swine Urine.

Sai Wang,Jiahui Liu,Shuai Zhao,Shan Zhang,Xiao Wei,Yiyang Dong

doi:10.3390/s17030604

Sai Wang, Jiahui Liu + Show 4 more

Open Access

https://doi.org/10.3390/s17030604

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Abstract

Ractopamine (RCT) is banned for use in animals in many countries, and it is urgent to develop efficient methods for specific and sensitive RCT detection. A label-free indirect competitive surface plasmon resonance (SPR) immunosensor was first developed with a primary antibody herein and then improved by a secondary antibody for the detection of RCT residue in swine urine. Meanwhile, a pre-incubation process of RCT and the primary antibody was performed to further improve the sensitivity. With all the key parameters optimized, the improved immunosenor can attain a linear range of 0.3–32 ng/mL and a limit of detection (LOD) of 0.09 ng/mL for RCT detection with high specificity. Furthermore, the improved label-free SPR immunosenor was compared thoroughly with a conventional enzyme-linked immunosorbent assay (ELISA). The SPR immunosensor showed advantages over the ELISA in terms of LOD, reagent consumption, analysis time, experiment automation, and so on. The SPR immunosensor can be used as potential method for real-time monitoring and screening of RCT residue in swine urine or other samples. In addition, the design using antibody pairs for biosensor development can be further referred to for other small molecule detection.

Highlights

Ractopamine (RCT), a typical agonist belonging to the β-agonist family, was originally used as veterinary medicine for respiratory diseases and bronchodilators [1,2]; some unscrupulous manufacturers ignore the promulgated laws and regulations [3], and apply excess RCT to promote animal growth
RCT residue detection was traditionally performed with high-performance liquid chromatography (HPLC), liquid chromatography/mass spectrometry/mass spectrometry (LC–MS/MS), and immunoassays such as the enzyme-linked immunosorbent assay (ELISA) [4,5,6,7,8,9]
We focus on the improvement of the limit of detection (LOD) and linear range of the surface plasmon resonance (SPR) immunosensor for RCT detection in swine urine samples, and we developed an ELISA immunosensor for validation and comparison

Summary

Introduction

Ractopamine (RCT), a typical agonist belonging to the β-agonist family, was originally used as veterinary medicine for respiratory diseases and bronchodilators [1,2]; some unscrupulous manufacturers ignore the promulgated laws and regulations [3], and apply excess RCT to promote animal growth. RCT residue detection was traditionally performed with high-performance liquid chromatography (HPLC), liquid chromatography/mass spectrometry/mass spectrometry (LC–MS/MS), and immunoassays such as the enzyme-linked immunosorbent assay (ELISA) [4,5,6,7,8,9]. LC–MS/MS and HPLC obtain a limit of detection (LOD) of about 2 μg/kg [4,5,6], but require a complicated sample extraction process, a long time and a high cost. ELISA or fluorescence-based immunoassays for β-agonist detection allow LODs of less than 2 μg/kg with good specificity [7,8,9], but they take a long time due to incubation steps, require multiple washing steps

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