蛋白质的非定点荧光标记——推荐一个化学生物学实验
Non-Site-Specific Fluorescent Labeling of Proteins as a Chemical Biology Experiment
洪歆怡 1薛太玲 1徐洲 1谢恩蓉 1吴明凯 1王青青 1吴丽娜1
作者信息
- 1. 厦门大学化学化工学院,化学国家级实验教学示范中心(厦门大学),福建 厦门 361005
- 折叠
摘要
本论文介绍了一项化学生物学实验,旨在利用异硫氰酸荧光素的异硫氰基与蛋白质氨基共价结合的原理,实现对体外蛋白(以牛血清白蛋白为例)和体内蛋白(以大肠杆菌总蛋白为例)的非定点荧光标记.实验采用凝胶过滤法作为后续纯化手段,并运用紫外-可见分光光度计、流式细胞术以及荧光显微镜对标记结果进行全面表征.该实验涵盖了多种基础和前沿实验技能,与科研工作中常见的抗体荧光标记技术密切相关,难度适中.通过参与这一实验,学生不仅能够深入理解化学生物学基础知识,提高综合实践能力,还能够拓宽科研视野并培养科研思维.
Abstract
This paper presents a chemical biology experiment designed to exploit the principle of covalent binding between fluorescein isothiocyanate and protein amines.This process is aimed at achieving non-site-specific fluorescent labeling of both extracellular proteins,exemplified by bovine serum albumin,and intracellular proteins,using Escherichia coli total protein as a case study.Gel filtration chromatography is employed as a subsequent purification method,with comprehensive characterization of the labeling results performed using UV-Visible spectrophotometry,flow cytometry,and fluorescence microscopy.This experiment encompasses a range of foundational and cutting-edge laboratory skills.It is closely aligned with the commonly employed antibody fluorescence labeling techniques in scientific research,presenting a moderate level of complexity.Participation in this experiment allows students to gain a profound understanding of fundamental concepts in chemical biology,enhance their comprehensive practical skills,broaden their research perspectives and nurture scientific thinking.
关键词
非定点荧光标记/蛋白质/异硫氰酸荧光素/化学生物学实验Key words
Non-site-specific fluorescent labeling/Protein/Fluorescein isothiocyanate/Chemical biology experiment引用本文复制引用
出版年
2024
国家科技期刊平台
NSTL
万方数据