伦敦白胶和茶：一种既快又安全的用于透射电子显微镜的细菌样品制备方法（英文）

LR-White resin and tea: a quick and safe bacterial cells preparation method for transmission electron microscopy

摘要:

【目的】检测乌龙茶提取物是否可作为电子染色剂取代醋酸双氧铀用于细菌细胞染色，使其能在透射电子显微镜下进行观察。【方法】利用伦敦白胶对细菌样品(大肠杆菌和金黄色葡萄球菌)进行胶块的制备，再在复染铅与不复染铅这两种情况下对超薄切片样品进行3种不同染色剂的电子染色，之后在透射电子显微镜下观察比较其不同之处。这3种不同的染色剂分别是醋酸双氧铀、0.05%乌龙茶提取物以及0.1%乌龙茶提取物。首先将带有超薄切片样品的铜网悬浮于不同的待比较染液中10?15 min，若需进一步用柠檬酸铅复染，则将经3次蒸馏水冲洗过后的铜网再次悬浮于柠檬酸铅染液中8?10 min。【结果】复染铅的情况下，在透射电子显微镜下无论是大肠杆菌还是金黄色葡萄球菌，利用3种电子染色剂进行染色的结果均非常相似。【结论】实验结果表明，在观察细菌结构中，乌龙茶提取物可以替代醋酸双氧铀进行透射电子显微镜样品的电子染色。

Abstract:

[Objective] Oolong tea extract (OTE) was tested for its potential as an electron staining reagent to substitute for uranyl acetate (UA) in electron microscopy of bacterial cells. [Methods] Three electron microscopic staining methods (standard, 0.05% OTE, and 0.1% OTE) were compared for electron microscopic observation of ultrathin sections with or without lead citrate (Pb). First floating the sections on the compared staining solutions for 10?15 min, then if need to post-stained with lead citrate, after rinsing with deionized water three times, the sections were re-floated on lead citrate for 8?10 min. Electron microscopy was performed on sections from LR-White-embedded cells of a Gram-negative bacterium, Escherichia coli, and a Gram-positive bacterium, Staphylococcus aureus. [Results] Both concentrations of the OTE preparations showed the staining results were similar to that of standard method with UA and Pb in E. coli and S. aureus cells. [Conclusion] This study demonstrated that OTE could potentially be used as an alternative to UA in electron microscopy staining for certain structures.