张鹏;彭涛;范安然;兰金芝;何志旭;舒莉萍;周艳华;
ZHANG Peng;PENG Tao;FAN Anran;LAN Jinzhi;HE Zhixu;SHU Liping;ZHOU Yanhua;National & Guizhou Joint Engineering Laboratory for Cell Engineering and Biomedicine Technique, Center for Tissue Engineering and Stem Cell Research, Guizhou Province Key Laboratory of Regenerative Medicine, Guizhou Medical University;Key Laboratory of Adult Stem Cell Translational Research,Chinese Academy of Medical Sciences;Department of Pediatrics,the Affiliated Hospital of Zunyi Medical University;

• 1:贵州医科大学细胞工程生物医药技术国家地方联合工程实验室组织工程与干细胞实验中心贵州省再生医学重点实验室
• 2:中国医学科学院成体干细胞转化研究重点实验室
• 3:遵义医科大学附属医院儿科学教研室

摘要(Abstract)：

目的:探讨Alexa Fluor 488 NHS Ester染料用于提高流式细胞周期检测结果的可行性。方法:以293细胞作为研究对象,采用两种方案进行细胞周期检测,每种方案设4组;方案一,按照细胞周期检测试剂盒的说明书对4组细胞进行染色;方案二,先对4组细胞用0.0、0.4、2.0及10.0 mg/L的Alexa Fluor 488 NH S Ester染料染色15 min,然后将4组细胞混合在一支试管中再用碘化丙啶(PI)进行染色;将两种染色方案获得的细胞上流式细胞仪检测,分析比较两种方案的4组细胞检测结果。结果:方案一染色的4组细胞,流式细胞仪检测结果不尽一致,个别组间由于细胞密度不同导致染色结果存在差异;方案二经不同浓度Alexa Fluor 488 NHS Ester染色的4组细胞虽然混合于一支试管,检测结果经Flowjo X软件分析时能明显分出4群细胞,对4群细胞分别进行细胞周期分析比较,结果一致。结论:采用不同浓度Alexa Fluor 488 NHS Ester染料预染细胞后混合在一起进行PI染色能够提高细胞周期检测结果的可信度,在神经退行性疾病研究中可节约病人的取样次数及数量。
Objective: To explore the feasibility of Alexa Fluor 488 NHS Ester dye for improving the results of flow cytometry cell cycle detection. Methods: The 293 cells were taken as the research object. Two schemes were used to detect cell cycle, and each scheme was divided into four groups.In the first scheme, cells in the four groups were stained according to the instructions of the cell cycle detection kit.In the second scheme, cells in the four groups were stained with 0.0, 0.4, 2.0 and 10.0 mg/L Alxa Fluor 488 NH S Ester dyes for 15 minutes, then mixed the cells in a test tube and stained with PI. Flow cytometry was used to detect the cell cycle, and the results were analyzed and compared between the two schemes. Results: The results of flow cytometry were inconsistent among groups in scheme 1 due to different cell densities. In scheme 2, the cells stained were firstly separated by Alexa Fluor 488 NHS Ester, and then further separated by PI, and the detection results of cell cycle were more consistent than those of scheme 1. Conclusion: Pre-stained cells with Alexa Fluor 488 NHS Ester dye can improve the reliability of cell cycle detection results, and also save the samples in the study of neurodegenerative disease.

关键词(KeyWords)： 细胞周期检测;流式细胞术;碘化丙啶;神经退行性疾病;Alexa Fluor 488 NHS Ester染料
cell cycle;flow cytometry;propidium iodide;neurodegenerative disease;Alexa Fluor 488 NHS Ester dye

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(31660316);; 贵州省应用基础研究计划重大专项[黔科合J重大字(2015)2003];; 贵州省科技支撑计划项目[黔科合支撑(2018)2757];; 贵州省科技计划项目[黔科合平台人才(2019)5406]

作者(Authors): 张鹏;彭涛;范安然;兰金芝;何志旭;舒莉萍;周艳华;
ZHANG Peng;PENG Tao;FAN Anran;LAN Jinzhi;HE Zhixu;SHU Liping;ZHOU Yanhua;National & Guizhou Joint Engineering Laboratory for Cell Engineering and Biomedicine Technique, Center for Tissue Engineering and Stem Cell Research, Guizhou Province Key Laboratory of Regenerative Medicine, Guizhou Medical University;Key Laboratory of Adult Stem Cell Translational Research,Chinese Academy of Medical Sciences;Department of Pediatrics,the Affiliated Hospital of Zunyi Medical University;

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