Zhang Yun, Phillips Gregory J, Yeung Edward S
Ames Laboratory-USDOE and Department of Chemistry, Iowa State University, Ames, Iowa 50011, USA.
Anal Chem. 2007 Jul 15;79(14):5373-81. doi: 10.1021/ac070477u. Epub 2007 Jun 6.
The small size of bacteria makes it difficult to study the biochemistry inside single cells. The amount of material inside is limited; therefore, an ultrasensitive method is required to interrogate single cells. Using a sensitive ICCD detector to record chemiluminescence (CL) from an optimized firefly luciferase-ATP bioluminescence reaction system, we report for the first time real-time imaging of lysis and leakage of single bacterium with 10-s temporal resolution. Movies are generated to visualize how the cell membrane was damaged by phage lysis, antibiotics attack, or dehydration, as well as the wall repair and cell recovery processes. The results show single-cell variations that are not obtainable from bulk measurements, confirming that CL microscopy of luciferase-expressing bacteria is a powerful tool for studying the fundamental biology of cells.
细菌体积微小，这使得研究单个细胞内部的生物化学过程变得困难。细胞内的物质数量有限，因此需要一种超灵敏的方法来检测单个细胞。我们使用灵敏的ICCD探测器记录优化后的萤火虫荧光素酶-ATP生物发光反应系统产生的化学发光（CL），首次以10秒的时间分辨率对单个细菌的裂解和泄漏进行实时成像。生成的影像展示了细胞膜是如何被噬菌体裂解、抗生素攻击或脱水破坏的，以及细胞壁修复和细胞恢复过程。结果显示了批量测量无法获得的单细胞差异，证实了对表达荧光素酶的细菌进行CL显微镜检查是研究细胞基础生物学的有力工具。
