期刊
MICROSCOPY AND MICROANALYSIS
卷 27, 期 5, 页码 1265-1272出版社
OXFORD UNIV PRESS
DOI: 10.1017/S1431927621012289
关键词
microorganisms; transmission electron microscopy; ultra-low lead staining; ultrastructure
资金
- National Key R&D Program of China [2020YFC1808000]
- National Natural Science Foundation of China [31770548]
- Program for Student Innovation Through Research and Training [S20190010, 201910307090P, 202013YX18]
The ULLS technique offers a novel approach for preparing ultra-thin sections for TEM analysis, simplifying the sample preparation process and providing acceptable toxicity levels for most microbial cells. It holds promise for playing a significant role in studying the ultrastructure of microbial cells.
The staining procedure is critical for investigating intra- and extra-cellular ultrastructure of microorganisms by transmission electron microscopy (TEM). Here, we propose a new ultra-low lead staining (ULLS) technique for preparing the ultrathin sections for TEM analysis. Sections of Enterobacter sp. (bacteria), Aspergillus niger (filamentous fungi), Rhodotorula mucilaginosa (fungi), and Chlamydomonas reinhardtii (microalgae) were tested. Compared with the sections prepared by the typical double-staining technique, ULLS-based sections showed evident advantages: (i) the staining process only required the addition of Pb (NO3)(2); (ii) the Pb level during incubation was set as low as 1 mg/L, which had negligible toxicity to most microbial cells; (iii) the Pb cations were added during microbial culture, which avoided complicated sample preparation as in typical double staining. Taking C. reinhardtii as an example, the ULLS technique allowed fine investigation of microbial ultrastructure, e.g., starch granule, mitochondrion, Golgi apparatus, vacuole, and vesicle. Meanwhile, the physiological processes of the cells such as cell lysis and exocytosis were successfully captured, with relatively high contrast. This study hence shows a bright future on preparation of the high-quality ultrathin sections of microbial cells by the ULLS technique.
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