Detection of ustilaginoidins in rice samples by immunoassay based on monoclonal antibodies prepared from hemiustilaginoidin-derived haptens

Weixuan Wang; Xiaoxiang Fu; Yihao Li; Mingpeng Jing; Yonglin Yang; Dan Xu; Daowan Lai; Mingan Wang; Baomin Wang; Ligang Zhou

doi:10.1016/j.heliyon.2023.e23036

Detection of ustilaginoidins in rice samples by immunoassay based on monoclonal antibodies prepared from hemiustilaginoidin-derived haptens

Heliyon. 2023 Nov 29;9(12):e23036. doi: 10.1016/j.heliyon.2023.e23036. eCollection 2023 Dec.

Authors

Weixuan Wang¹, Xiaoxiang Fu^{1

2}, Yihao Li³, Mingpeng Jing¹, Yonglin Yang¹, Dan Xu¹, Daowan Lai¹, Mingan Wang³, Baomin Wang⁴, Ligang Zhou¹

Affiliations

¹ Department of Plant Pathology, College of Plant Protection, China Agricultural University, Beijing 100193, China.
² Laboratory for Phytochemistry and Plant-Derived Pesticides, College of Agriculture, Jiangxi Agricultural University, Nanchang 330045, China.
³ Innovation Center of Pesticide Research, Department of Applied Chemistry, College of Science, China Agricultural University, Beijing 100193, China.
⁴ College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China.

Abstract

Ustilaginoidins are a class of bis-naphtho-γ-pyrone mycotoxins to threaten humans, animals and environment. Ustilaginoidins are produced by Villosiclava virens, the rice false smut pathogen. To prepare antibodies for quantitatively analyzing ustilaginoidins in rice samples, hemiustilaginoidins D and F from the laccase gene deficiency mutant of V. virens respectively reacted with diazonium 4-aminobenzoic acid to obtain haptens with a carboxyl group, which further reacted with bovine serum albumin or ovalbumin to get their complete antigens. Two monoclonal antibodies (mAbs) designated as 4A12C6 and 5F4F6 were developed by immunization. The relationships between mAb sensitivity and 20 ustilaginoidins were described. 4A12C6 was chosen for further analysis as it could recognize main ustilaginoidins and was more sensitive than 5F4F6. The achieved indirect competitive enzyme-linked immunosorbent assay (icELISA) based on 4A12C6 had a half maximal inhibitory concentration (IC₅₀) of 0.76 ng/mL and working range of 0.2-2.8 ng/mL to ustilaginoidin A. The results of ustilaginoidins-contaminated rice samples by icELISA detection were consistent with those determined by HPLC‒DAD detection. Therefore, we developed a new strategy to get haptens from the biosynthetic precursors with half structures of ustilaginoidins. The achieved icELISA was demonstrated as a convenient method to monitor ustilaginoidin content in rice samples, and showed that the contents of total ustilaginoidins from the rice cultivars with low resistance to rice false smut were more than those of high resistance cultivars.

Keywords: Haptens; Hemiustilaginoidin; Immunoassay; Monoclonal antibodies; Rice samples; Ustilaginoidins.