Anti-fading media for live cell GFP imaging.
Anti-fading media for live cell GFP imaging.
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DOI：
10.1371/journal.pone.0053004
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Lukyanov KA
中科院分区：
文献类型：
--
作者：
Bogdanov AM;Kudryavtseva EI;Lukyanov KA
光稳定性是荧光显微镜用染料最重要的特性之一。最近，我们证明了成像介质中存在的维生素极大地加速了增强型绿色荧光蛋白(EGFP)和许多其他绿色荧光蛋白和可光激活蛋白的光漂白。在这里，我们逐一测试了常用介质(如Dulbecco‘s Modify Eagle Medium，DMEM)中的所有维生素，发现只有核黄素和吡哆醛两种维生素降低了EGFP的光稳定性。因此，不含核黄素和吡哆醛的DMEM可以作为成像介质，以最小的生化干扰为代价确保GFP的高光稳定性。然后，我们测试了一些抗氧化剂，发现一种植物类黄酮芦丁在活细胞显微镜下极大地提高了EGFP的光稳定性。在完全DMEM中，芦丁提高了EGFP的光稳定性，达到了缺乏维生素的DMEM的水平。此外，将芦丁添加到缺乏维生素的DMEM中，能够进一步抑制绿色荧光蛋白的光漂白。潜在地，新的介质配方可以广泛用于表达GFP的细胞的荧光显微镜，并在各种成像应用中模拟多细胞生物体，其中光稳定性是一个挑战。
Photostability is one of the most important characteristic of a dye for fluorescence microscopy. Recently we demonstrated that vitamins present in imaging media dramatically accelerate photobleaching of Enhanced Green Fluorescent Protein (EGFP) and many other green fluorescent and photoactivatable proteins. Here we tested all vitamins of commonly used media (such as Dulbecco's Modified Eagle Medium, DMEM) one-by-one and found that only two vitamins, riboflavin and pyridoxal, decrease photostability of EGFP. Thus, DMEM without riboflavin and pyridoxal can be used as an imaging medium, which ensures high photostability of GFPs at the expense of minimal biochemical disturbance. Then, we tested some antioxidants and found that a plant flavonoid rutin greatly enhances photostability of EGFP during live cell microscopy. In complete DMEM, rutin increased EGFP photostability up to the level of vitamin-depleted DMEM. Moreover, being added to vitamin-depleted DMEM, rutin was able to further suppress EGFP photobleaching. Potentially, new medium formulations can be widely used for fluorescence microscopy of GFP-expressing cells and model multicellular organisms in a variety of imaging applications, where photostability represents a challenge.
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影响因子：
6.1
作者：
Huvaere, Kevin;Sinnaeve, Bart;Skibsted, Leif H.
通讯作者：
Skibsted, Leif H.
影响因子：
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作者：
MIKHAILOV, AV;GUNDERSEN, GG
通讯作者：
GUNDERSEN, GG
影响因子：
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
Lukyanov, Konstantin A.
影响因子：
6.7
作者：
Bhandary, Bidur;Piao, Cheng Shi;Chae, Han-Jung
通讯作者：
Chae, Han-Jung