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活细胞超分辨率成像非传统的动力学和核孔综合体的组合
Xianxin Ye1, Minzhu Guan2, Yaorong Guo2
1National Biomedical Imaging Center, State Key Laboratory of Membrane Biology, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, Institute of Molecular Medicine, College of Future Technology, Peking University, Beijing 100871, China.
Biophysics reports
|March 22, 2024
概括
超分辨率显微镜现在允许活细胞成像稀疏的蛋白质,如核毛孔复合体 (NPC) 中的蛋白质. 这种新的Halo-SiR和Sparse-SIM方法提高了观察动态NPC结构的可见性和对比度.
科学领域:
- 细胞生物学 细胞生物学
- 生物物理学的生物物理.
- 显微镜的使用方法
背景情况:
- 超分辨率显微镜 (SRM) 面临着在细胞结构中成像低丰度蛋白质的挑战.
- 核孔复合体 (NPC) 具有有限的蛋白质,阻碍了对活细胞的持续观察和SRM评估.
研究的目的:
- 开发一种方法,用于在活细胞结构内的稀疏蛋白质的超高分辨率扩展成像.
- 克服在核孔复合体中观察动态蛋白种群的局限性.
主要方法:
- 使用Halo-SiR光探针标记POM121蛋白质.
- 使用结构化照明显微镜与稀疏解卷 (Sparse-SIM) 进行成像.
- 将Halo-SiR性能与串联的mCherry标签进行比较.
主要成果:
- 与mCherry相比,Halo-SiR的光强度超过六倍,对比度提高四倍.
- 在200SR成像过程中观察到Halo-SiR的微不足道的光漂白.
- 确定了各种NPC结构 (环状,集群状) 并观察了它们的动态重塑.
结论:
- Halo-SiR与Sparse-SIM相结合,可以对活细胞中的动态NPC结构进行强大的,扩展的SR成像.
- 这种技术对于可视化低拷贝数量的蛋白质是有效的,进步了细胞生物学研究.
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