Impact of fixation on in vitro cell culture lines monitored with Raman spectroscopy
Impact of fixation on in vitro cell culture lines monitored with Raman spectroscopy
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DOI：
10.1039/b822408k
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发表时间：
2009-01-01
期刊：
影响因子：
4.2
通讯作者：
Deckert, Volker
中科院分区：
文献类型：
--
作者：
Mariani, Melissa M.;Lampen, Peter;Deckert, Volker
拉曼光谱提供了有关分析物组成的丰富的化学信息，是生物学研究的有力工具。由于能够研究特定的细胞成分或对整个细胞进行成像，因此必须采用兼容的样品保存方法以收集准确的光谱。不幸的是，许多常用的样品保存方法的效果尚未与培养的细胞进行探讨。在这项研究中，两种不同表型的人类细胞系被用来研究样品保存方法的影响。将细胞直接培养在石英基质上，并福尔马林固定、干燥或风干。结果表明，应用于细胞培养物的拉曼分析方法显著影响所得光谱数据的质量和再现性。福尔马林固定并没有被发现是普遍有效的，作为一种常用的固定剂的预期。这主要是由于细胞系之间样品保存的不一致性和信号强度的损失。样品风干被认为是在很大程度上不一致的光谱再现性。我们的研究表明，样品干燥显示出良好的光谱重现性，并导致在一个良好的信噪比。与空气干燥相比，活化和灭活细胞中的脂质和蛋白质含量得以保持，并提供了一种更可控的方法，表明干燥速度对样品保存很重要
Raman spectroscopy provides chemical-rich information about the composition of analytes and is a powerful tool for biological studies. With the ability to investigate specific cellular components or image whole cells, compatible methods of sample preservation must be implemented for accurate spectra to be collected. Unfortunately, the effects of many commonly used sample preservation methods have not been explored with cultured cells. In this study, two human cell lineages of varying phenotypes were used to investigate the effects of sample preservation methods. Cells were cultured directly onto quartz substrates and either formalin-fixed, desiccated or air dried. The results indicate that the methodology applied to cell cultures for Raman analysis significantly influences the quality and reproducibility of the resulting spectral data. Formalin fixation was not found to be as universally efficient as anticipated for a commonly used fixative. This was due largely to the inconsistency in sample preservation between cell lines and loss of signal intensity. Sample air-drying was found to be largely inconsistent in terms of spectral reproducibility. Our study shows that sample desiccation displayed good spectral reproducibility and resulted in a good signal-to-noise ratio. Lipid and protein content in both activated and inactivated cells were maintained and provided a more controlled method compared with air-drying, revealing that the speed of drying is important for sample preservation