Development of a Monoclonal Antibody-Based icELISA for the Detection of Ustiloxin B in Rice False Smut Balls and Rice Grains.
Rice false smut is an emerging and economically-important rice disease caused by infection by the fungal pathogen Villosiclava virens. Ustiloxin B is an antimitotic cyclopeptide mycotoxin isolated from the rice false smut balls that formed in the pathogen-infected rice spikelets. A monoclonal antibody (mAb) designated as mAb 1B5A10 was generated with ustiloxin B-ovalbumin conjugate. A highly-sensitive and specific indirect competitive enzyme-linked immunosorbent assay (icELISA) was then developed. The median inhibitory concentration (IC50) of the icELISA was 18.0 ng/mL for the detection of ustiloxin B; the limit of detection was 0.6 ng/mL, and the calibration range was from 2.5 to 107.4 ng/mL. The LOD/LOQ values of the developed ELISA used for the determination of ustiloxin B in rice false smut balls and rice grains were 12/50 μg/g and 30/125 ng/g, respectively. The mAb 1B5A10 cross-reacted with ustiloxin A at 13.9% relative to ustiloxin B. Average recoveries of ustiloxin B ranged from 91.3% to 105.1% for rice false smut balls at spiking levels of 0.2 to 3.2 mg/g and from 92.6% to 103.5% for rice grains at spiking levels of 100 to 5000 ng/g. Comparison of ustiloxin B content in rice false smut balls and rice grains detected by both icELISA and high performance liquid chromatography (HPLC) demonstrated that the developed icELISA can be employed as an effective and accurate method for the detection of ustiloxin B in rice false smut balls, as well as rice food and feed samples.
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. xiaoxiaofu@cau.edu.cn.
Xiaoxiang
X
Fu
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. wangali526@163.com.
Ali
A
Wang
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. wangxiaohan99@126.com.
Xiaohan
X
Wang
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. fengkelin.123321aa@foxmail.com.
Fengke
F
Lin
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. hls0305@136.com.
Lishan
L
He
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. dwlai@cau.edu.cn.
Daowan
D
Lai
Institute of Agro-Products Processing Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China. liuyang01@caas.cn.
Yang
Y
Liu
Department of Molecular Biosciences and Bioengineering, University of Hawaii at Manoa, Honolulu, HI 96822, USA. qingl@hawaii.edu.
Qing X
QX
Li
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. lgzhou@cau.edu.cn.
Ligang
L
Zhou
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China. wbaomin@263.net.
Baoming
B
Wang
D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006999:Hypocreales / Q000378:metabolism; D020128:Inhibitory Concentration 50; D057230:Limit of Detection; D012275:Oryza / Q000737:chemistry / Q000382:microbiology*; D010456:Peptides, Cyclic / Q000032:analysis*; D010935:Plant Diseases / Q000382:microbiology
D000911:Antibodies, Monoclonal; D010456:Peptides, Cyclic; C089077:ustiloxin B; C087237:ustiloxin A
ELISA; Villosiclava virens; mycotoxin; phytotoxin; rice false smut; ustiloxin B