A Sensitive Monoclonal-Antibody-Based ELISA for Forchlorfenuron Residue Analysis in Food Samples

Abstract

In this study, forchlorfenuron (CPPU) was coupled with succinic anhydride to yield a CPPU hapten (CPPU-COOH), and a high-affinity monoclonal antibody (mAb) that can specifically recognize CPPU was produced. Using this mAb as a recognition reagent, a sensitive indirect competitive enzyme-linked immunosorbent assay (icELISA) for CPPU was optimized, which exhibits an IC50 of 1.04 ng/mL, a limit of detection of 0.16 ng/mL, and a linear range of 0.31–3.43 ng/mL for CPPU. Cross-reactivity percentages with six analogues were all below 6%. The average recovery rates for cucumber and orange samples were from 85.23% to 119.14%. The analysis results of this icELISA showed good consistency with those from liquid chromatography mass spectrometry. These results suggest that the proposed icELISA provides a sensitive, specific, and reliable strategy for CPPU detection in food samples.

A Magnetic-Bead-Based Immunoassay with a Newly Developed Monoclonal Antibody for Rapid and Highly Sensitive Detection of Forchlorfenuron

“…As listed in Table 1 , anti-CPPU mAb 14G1 showed extremely inconspicuous CRs with analogues in the MB-based assay and icELISA systems. When compared with previous studies [ 14 , 16 ], the ELISA-based antibodies exhibited obvious cross-reaction with thidiazuron. However, in the present study, an MB-based assay and icELISA-based mAb 14G1 had negligible cross-reaction with thidiazuron and other listed compounds, which exhibited better selectivity and could effectively guarantee the specific detection of CPPU.…”

“…Additionally, we have provided a detailed comparison of the analytical performance of these two developed methods with those of the previously documented immunoassay methods for CPPU detection, as shown in Table S2 . The comparison results showed that the total analysis time of the developed MB-based assay was much less than other microplate-based enzyme immunoassays for CPPU [ 13 , 14 , 15 , 16 ], and they were even comparable to strip-based immunochromatography, which can complete the assay in 30 min [ 15 ]. Moreover, the IC 50 and LOD values of the MB-based assay for CPPU were far lower than the reported immunoassays.…”

“…Alternatively, antibody-mediated immunoassays, such as enzyme-linked immunosorbent assay (ELISA) and nanomaterial-based strips, have been widely developed for food hazardous determination with advantages of simplicity, rapidity, cost effectiveness, and high throughput [ 12 ]. Up until now, there have only been a few immunoassay reports on CPPU, differing in their assay performance [ 13 , 14 , 15 , 16 ]. Antibodies serve as core reagents in immunoassays, which largely determine the specificity and sensitivity of the resulting immunodetection technique.…”

“…Therefore, another mAb, P6#42, was employed to establish a LFIA for the rapid detection of CPPU, which can complete the detection in 30 min, but the detection sensitivity (IC 50 of 286 ng/L) was significantly decreased compared with that of dcELISA (IC 50 of 50 ng/L). Recently, a new mAb against CPPU has been produced and reported to be used to develop an indirect competitive ELISA (icELISA) [ 16 ]. However, the established icELISA exhibited an IC 50 value of 1.04 ng/mL, and the procedure needs as long as 80 min for incubation steps, which showed less superiority over the reported dcELISA and LFIA.…”

Chemiluminescence method based on the KIO4−K2CO3−Mn2+ reaction for rapid and sensitive determination of forchlorfenuron in dried fruit

“…To date, various analysis methods of CPPU have been reported, such as high-performance liquid chromatography (HPLC) [7], surfaceenhanced Raman spectroscopy (SERS) [8], enzyme-linked immunosorbent assay (ELISA) [9], and electrochemistry (EC) [10]. These methods not only have the advantages of high sensitivity and specificity, but also have disadvantages such as long time, complex operation and high costs [11].…”