School of Engineering, Nanotechnology, University of Ulster at Jordanstown, Newtownabbey, Co. Antrim, Northern Ireland, UK.
J Mater Sci Mater Med. 2010 Aug;21(8):2317-24. doi: 10.1007/s10856-009-3965-0. Epub 2009 Dec 18.
Raman spectroscopy has been used to determine the chemical composition of materials for over 70 years. Recent spectacular advances in laser and CCD camera technology creating instruments with higher sensitivity and lower cost have initiated a strong resurgence in the technique, ranging from fundamental research to process control methodology. One such area of increased potential is in tissue engineering and regenerative medicine (TERM), where autologous cell culture, stem cell biology and growth of human cells on biomaterial scaffolds are of high importance. Traditional techniques for the in vitro analysis of biochemical cell processes involves cell techniques such as fixation, lysis or the use of radioactive or chemical labels which are time consuming and can involve the perpetuation of artefacts. Several studies have already shown the potential of Raman spectroscopy to provide useful information on key biochemical markers within cells, however, many of these studies have utilised micro- or confocal Raman to do this, which are not suited to the rapid and non-invasive monitoring of cells. For this study a versatile fit-for-purpose Raman spectrometer was used, employing a macro-sampling optical platform (laser spot size 100 mum at focus on the sample) to discriminate between different TERM relevant cell types and viable and non-viable cells. The results clearly show that the technique is capable of obtaining Raman spectra from live cells in a non-destructive, rapid and non-invasive manner, however, in these experiments it was not possible to discriminate between different cell lines. Despite this, notable differences were observed in the spectra obtained from viable and non-viable cells, showing significant changes in the spectral profiles of protein, DNA/RNA and lipid cell constituents after cell death. It is evident that the method employed here shows significant potential for further utilisation in TERM, providing data directly from live cells that fits within a quality assurance framework and provides the opportunity to analyse cells in a non-destructive manner.
拉曼光谱技术已经被用于确定材料的化学组成超过 70 年。近年来，激光和 CCD 相机技术的惊人进步，创造了具有更高灵敏度和更低成本的仪器，引发了该技术的强烈复兴，从基础研究到过程控制方法学。一个增加潜力的领域是在组织工程和再生医学（TERM）中，其中自体细胞培养、干细胞生物学和人类细胞在生物材料支架上的生长具有重要意义。用于生化细胞过程的体外分析的传统技术涉及细胞技术，例如固定、裂解或使用放射性或化学标记，这些技术既耗时又可能导致伪像的延续。已经有几项研究表明拉曼光谱在提供细胞内关键生化标志物的有用信息方面具有潜力，然而，这些研究中的许多都利用了微拉曼或共聚焦拉曼来做到这一点，而这并不适合快速和非侵入性的细胞监测。在这项研究中，使用了一种多功能的、适用于目的的拉曼光谱仪，采用宏观采样光学平台（激光在样品焦点上的光斑尺寸为 100 微米）来区分不同的 TERM 相关细胞类型和有活力和无活力的细胞。结果清楚地表明，该技术能够以非破坏性、快速和非侵入性的方式从活细胞中获得拉曼光谱，然而，在这些实验中，无法区分不同的细胞系。尽管如此，在从有活力和无活力的细胞中获得的光谱中观察到了明显的差异，显示出细胞死亡后蛋白质、DNA/RNA 和脂质细胞成分的光谱轮廓发生了显著变化。显然，这里采用的方法在 TERM 中具有进一步利用的巨大潜力，直接从活细胞中提供符合质量保证框架的数据，并提供以非破坏性方式分析细胞的机会。
