Development of monoclonal antibody-based sensitive ELISA for the determination of Cry1Ie protein in transgenic plant

doi:10.1007/s00216-016-9938-5

	Development of monoclonal antibody-based sensitive ELISA for the determination of Cry1Ie protein in transgenic plant
	Zhang, Yuwen 1,2; Zhang, Wei 2; Liu, Yan 2; Wang, Jianhua 2; Wang, Guoying 1; Liu, Yunjun 1
刊名	ANALYTICAL AND BIOANALYTICAL CHEMISTRY
	2016
卷号	408 期号:28 页码:8231-8239
关键词	Bacillus thuringiensis Cry1Ie Sandwich ELISA Transgenic plant
ISSN号	1618-2642
DOI	10.1007/s00216-016-9938-5
通讯作者	Zhang, Yuwen
英文摘要	Cry1Ie is a kind of Bacillus thuringiensis (Bt) toxin protein which has a different action model than the Cry1Ab and Cry1Ac protein. The transgenic maize expressing Cry1Ie might be commercially used in the near future and it is urgent to develop a method to detect Cry1Ie protein in transgenic plants and their products. To develop an ELISA method, Cry1Ie protein was expressed in Escherichia coli strain Transetta DE3, purified with the Ni-NTA spin columns, and then validated by sequencing. Bioassay results showed that the purified Cry1Ie protein was highly toxic to the Asian corn borer. The polyclonal antibody (pAb) and the specific monoclonal antibody (mAb) 1G(4)2D(6) were generated from rabbit and mice which were immunized with Cry1Ie protein, respectively. Western blotting of crude Cry1Ie protein extracts was established by employing mAb 1G(4)2D(6), whereas the mAb 1G(4)2D(6) negligibly recognized other Bt proteins. Sandwich ELISA against Cry1Ie protein was established by coating with pAb and detecting with mAb 1G(4)2D(6). The limit of detection (LOD), the limit of quantification (LOQ), and the quantification range of the assay in different matrices of maize plant were determined as 0.27-0.51, 0.29-0.78, and 0.45-15.71 ng/mL, respectively. Recoveries of Cry1Ie protein spiked in different maize tissues ranged from 75.1 to 99.5 %. The established sandwich ELISA was verified using transgenic maize overexpressing Cry1Ie. The results in this study suggested that the established ELISA method is effective for detecting Cry1Ie protein in transgenic plants.
学科主题	Chemistry, Analytical
语种	英语
出版者	SPRINGER HEIDELBERG
WOS记录号	WOS:000386693000034
内容类型	期刊论文
源URL	[http://111.203.20.206/handle/2HMLN22E/105299]
专题	作物科学研究所_分子生物学系
作者单位	1.Chinese Acad Agr Sci, Inst Crop Sci, South St 12, Beijing 100081, Peoples R China 2.China Agr Univ, Coll Agr & Biotechnol, Yuanmingyuan West Rd 2, Beijing 100193, Peoples R China
推荐引用方式 GB/T 7714	Zhang, Yuwen,Zhang, Wei,Liu, Yan,et al. Development of monoclonal antibody-based sensitive ELISA for the determination of Cry1Ie protein in transgenic plant[J]. ANALYTICAL AND BIOANALYTICAL CHEMISTRY,2016,408(28):8231-8239.
APA	Zhang, Yuwen,Zhang, Wei,Liu, Yan,Wang, Jianhua,Wang, Guoying,&Liu, Yunjun.(2016).Development of monoclonal antibody-based sensitive ELISA for the determination of Cry1Ie protein in transgenic plant.ANALYTICAL AND BIOANALYTICAL CHEMISTRY,408(28),8231-8239.
MLA	Zhang, Yuwen,et al."Development of monoclonal antibody-based sensitive ELISA for the determination of Cry1Ie protein in transgenic plant".ANALYTICAL AND BIOANALYTICAL CHEMISTRY 408.28(2016):8231-8239.