CAJ | 학술논문

[目的]建立一种基于荧光显微成像技术的潜在肝毒性物质体外快速筛查方法.[方法]采用荧光染料TMRM和Hoechst 33342分别标记HepG2细胞的线粒体和细胞核,在细胞荧光显微成像平台上分别采集两种荧光信号,考察双色荧光染色法的稳定性、线性动态范围,同时选用肝毒性和无肝毒性药物验证方法的可靠性,并将考察结果与经典的MTT法相比较.[结果]两种荧光的荧光强度值在平均荧光强度值的3个SD范围内波动,具有较好的稳定性;荧光强度值与细胞接种数量之间呈现良好的线性相关,R2值分别为0.9817和0.9852;双色荧光染色法考察肝毒性和无肝毒性药物的结果与MTT法基本一致.[结论]基于HepG2细胞模型的双色荧光染色法稳定、可靠,可用于潜在肝毒性药物的体外筛查.
[목적]건립일충기우형광현미성상기술적잠재간독성물질체외쾌속사사방법.[방법]채용형광염료TMRM화Hoechst 33342분별표기HepG2세포적선립체화세포핵,재세포형광현미성상평태상분별채집량충형광신호,고찰쌍색형광염색법적은정성、선성동태범위,동시선용간독성화무간독성약물험증방법적가고성,병장고찰결과여경전적MTT법상비교.[결과]량충형광적형광강도치재평균형광강도치적3개SD범위내파동,구유교호적은정성;형광강도치여세포접충수량지간정현량호적선성상관,R2치분별위0.9817화0.9852;쌍색형광염색법고찰간독성화무간독성약물적결과여MTT법기본일치.[결론]기우HepG2세포모형적쌍색형광염색법은정、가고,가용우잠재간독성약물적체외사사.
Objective]To establish a novel fluorescent imaging-based assay for rapid screening potential hepatotoxic compounds in vitro. [Method] The signals of fluorescent dyes TMRM and Hoechst 33342, which were used to label mitochondrion and nucleus of cel s, were col ected by platform of cel fluorescent microscopy imaging in our study. The stability and linearity of two-color fluorescence assay(TCF assay) were tested. And then, the reliability of TCF assay was validated with hepatotoxic compounds and nonhepatotoxic compounds, which was compared with the results of 3-(4,5-dimethylthiazol-2-yl) -2,5-diphenyl tetrazolium bromide(MTT) assay. [Result] The fluorescence intensity of these two fluorescent dyes fluctuated within 3 folds of standard deviation(SD) of the mean fluorescence intensity, indicating that TCF assay was stable. Meanwhile, TCF assay had a good linearity, their R2 was 0.9817 and 0.9852 for TMRM and Hoechst 33342, respectively. Furthermore, the result of reliability validation for TCF assay was in concordance with MTT assay. [Conclusion] The new proposed TCF assay was stable and reliable, and it could be used for screening potential hepatotoxic compounds.