摘要
为实现T-2毒素的可视化毒性筛查,利用T-2毒素毒性通路中AP-1的关键靶点应答元件TRE与红色荧光蛋白(mCherry)的启动子构建pcDNA3.1-TRE-mCherry荧光质粒,建立荧光HEK293细胞传感筛查模型;同时,为验证该细胞传感模型的适用性,对实际样品中残留的T-2毒素和T-2毒素的主要代谢产物HT-2毒素进行了毒性监测。结果表明,T-2毒素刺激模式细胞8 h时,细胞内的红色荧光强度达到最高值并趋于稳定。当T-2毒素质量浓度在1~25 ng/mL时,其与荧光强度呈线性相关,线性方程为y=1.14938x+64.72,R^(2)=0.969。根据剂量曲线得出T-2毒素的EC50为16.27 ng/mL,检测限为0.691 ng/mL。该细胞传感模型用于样品加标实验,平均加标回收率为86.13%~126.20%,对HT-2毒素进行毒性筛查得到EC50为27.65 ng/mL。
To realize the visual toxicity screening of T-2 toxin,the fluorescent plasmid pcDNA 3.1-TRE-mCherry was constructed using the key AP-1 target response element TRE and red fluorescent protein(mCherry)in the toxicity pathway of T-2 toxin and the sensing screening model of fluorescent HEK293 cells was established.To verify the applicability of the cell sensing model,the toxicity detection of T-2 toxin residue in the actual samples and HT-2 toxin which was the main metabolite of T-2 toxin was conducted.The results showed that the intracellular red fluorescence intensity reached the maximum value and tended to be stable when the T-2 toxin stimulated the model cells for 8 h.When the concentration of T-2 toxin was in the range of 1~25 ng/mL,it was linearly correlated with fluorescence intensity,and the linear equation was y=1.14938x+64.72,R^(2)=0.969.According to the dose curve,the EC50 of T-2 toxin was 16.27 ng/mL,and the detection limit was 0.691 ng/mL.The cell sensing model was used in the standard addition test.The average recovery of standard addition was 86.13%~126.20%,and the EC50 of HT-2 toxin was 27.65 ng/mL after toxicity screening.
作者
孙嘉笛
高璐
张银志
孙秀兰
SUN Jiadi;GAO Lu;ZHANG Yinzhi;SUN Xiulan(School of Food Science and Technology,Jiangnan University,Wuxi 214122,China)
出处
《食品与生物技术学报》
CAS
CSCD
北大核心
2021年第12期35-43,共9页
Journal of Food Science and Biotechnology
基金
国家“十三五”重点研发计划项目(2021YFA0910200)
国家自然科学基金项目(31772069)
江苏省自然科学基金青年基金项目(BK20190584)。
关键词
T-2毒素
毒性筛查
细胞传感
荧光
T-2 toxin
toxicity screening
cell sensing
fluorescence
