Preparation of Monoclonal Antibody against Deoxynivalenol and Development of Immunoassays

Abstract

Fusarium toxins are the largest group of mycotoxins, which contain more than 140 known secondary metabolites of fungi. Deoxynivalenol (DON) is one of the most important compounds of this class due to its high toxicity and its potential to harm mankind and animals and a widespread contaminant of agricultural commodities, such as wheat, corn, barley, oats, bread, and biscuits. Herein, a hybridoma cell 8G2 secreting mAb against DON was produced by fusing the splenocytes with a tumor cell line Sp2/0. The obtained mAb had a high affinity (2.39 × 109 L/mol) to DON. An indirect competitive Enzyme-Linked Immunosorbent Assay (ic-ELISA) showed that the linear range for DON detection was 3.125–25 μg/mL, and the minimum inhibitory concentration (IC50) was 18.125 μg/mL with a limit of detection (LOD) of 7.875 μg/mL. A colloidal gold nanoparticle (AuNP) with 20 nm in diameter was synthesized for on-site detection of DON within 10 min with vLOD of 20 μg/mL. To improve the limit of detection, the gold nanoflower (AuNF) with a larger size (75 nm) was used to develop the AuNF-based strip with vLOD of 6.67 μg/mL. Compared to the vLOD of a convectional AuNP-based strip, the AuNF-based strip was three times lower. Herein, three immunoassay methods (ic-ELISA and AuNP/AuNF-based strips) were successfully developed, and these methods could be applied for the DON detection in agricultural products.

Improvement of the sensitivity of lateral flow systems for detecting mycotoxins: Up‐to‐date strategies and future perspectives

“…Xirui et al ( 2021) synthesized three gold nanobeads (GNBs) in sizes of 94, 129, and 237 nm for LFAs of FB 1 in maize samples, and GNBs showed the enhanced colorimetric signal intensity compared with 20-40 nm AuNPs (Figure 4A). Similarly, Mokhtar et al (2022) used AuNPs with a diameter of 20 nm and gold nanoflowers (AuNF) at 75 nm as detection probes in LFAs developed based on the DON monoclonal antibody, and the results showed that the visual detection limit of 6.67 μg/mL in test strips developed based on AuNF was three times higher than that of AuNPs. Silver nucleation is another common strategy used to enlarge the extinction coefficient of AuNPs by increasing their particle size (Yeh et al, 2009).…”

“…They are used in various fields for labeling, delivery of biomolecules, and sensing (Sperling et al., 2008). Since their first reported application in LFAs (Shyu et al., 2002), AuNPs have become the most widely used nanomaterials in LFA‐based devices due to their ease of production, stability, and compatibility with different biomolecules and their versatility (Elsir et al., 2022; Xirui et al., 2021). When excited with specific wavelengths, the conduction electrons of the NPs’ surface oscillate collectively, resulting in strong absorption and scattering of light (Liz‐Marzan, 2004), referred to as localized surface plasmon resonance, and this property of AuNPs is mainly harnessed for the development of sensitive colorimetric LFAs.…”

Deoxynivalenol: Emerging Toxic Mechanisms and Control Strategies, Current and Future Perspectives

“…Moreover, ELISA is a feasible technique for low-level detection and quantification of mycotoxins. Indirect competitive ELISA and immunoassay methods based on colloidal gold nanoparticles and gold nanoknife have been successfully developed and can be used to detect DON in agricultural products . A portable and reusable evanescent wave optofluidic immunosensor has been used to measure DON in situ based on the principle of an indirect competitive immunoassay …”

Comparison of Synthetic Methods and Identification of Several Artificial Antigens of Deoxynivalenol

“…In recent years, immunological detection technology has been widely used for residue analysis of small-molecule pollutants such as mycotoxins, pesticides, antibiotics, etc. [ 19 ]. The core reagent and challenge for establishing immunological detection technology is the preparation of mAbs, and the basis of high-quality core mAb reagents is the preparation of high-quality artificial antigens.…”