Institute for Science and Technology in Medicine, Guy Hilton Research Centre, Keele University, Thornburrow Drive, Stoke on Trent, UK ST4 7QB.
Analyst. 2009 Jun;134(6):1176-81. doi: 10.1039/b821112d. Epub 2009 Mar 11.
Single-cell studies have important implications in biomedicine. An accurate investigation of biochemical behaviour and status requires a biomolecular probe such as vibrational microscopy. Amongst other approaches, synchrotron infrared microspectroscopy is an appropriate analytical tool for single-cell investigation. However, it is important to understand the precise origin of spectral differences as they are directly related to the cell biochemistry. Beside biomolecular changes, physical properties can interfere in the resulting information, and the two effects need separating. Both cells and nuclei induce Mie scattering effects due to their equivalent size with the probe wavelength. This results in a large modification of the spectra, and its precise contribution has to be determined in order to extract the true spectral information. On this basis, we carried out this study in order to evaluate the exact contribution of cell nuclei to Mie scattering. To this purpose, we isolated whole cancer cell nuclei and obtained, for the first time, their FTIR spectra with good signal to noise ratio. The synchrotron-based FTIR (S-FTIR) spectra of nuclei showed changes in lipids, proteins, and DNA absorptions when compared to spectra of whole lung cancer cells. Importantly, we estimated the Mie scattering properties of single cells and single nuclei spectra and were consequently able to separate optical and chemical properties of single cells and nuclei. This is the first study which sheds new light on the identification of the precise spectral biomarkers of a whole cell and those of the cell nucleus.
单细胞研究在生物医学中有重要意义。准确研究生物化学行为和状态需要生物分子探针，如振动显微镜。在其他方法中，同步辐射红外微光谱学是单细胞研究的一种合适的分析工具。然而，了解光谱差异的确切来源非常重要，因为它们与细胞生物化学直接相关。除了生物分子变化外，物理性质也会干扰最终的信息，需要将这两种效应分开。由于细胞和细胞核的尺寸与探针波长相当，因此都会引起米氏散射效应。这会导致光谱发生很大的变化，因此必须确定其精确贡献，以提取真实的光谱信息。基于此，我们进行了这项研究，以评估细胞核对米氏散射的确切贡献。为此，我们分离了完整的癌细胞核，并首次获得了具有良好信噪比的 FTIR 光谱。与全肺癌细胞的光谱相比，基于同步加速器的 FTIR（S-FTIR）光谱显示细胞核中脂质、蛋白质和 DNA 吸收的变化。重要的是，我们估计了单细胞和单细胞核光谱的米氏散射特性，从而能够分离单细胞和细胞核的光学和化学性质。这是第一项阐明整个细胞和细胞核的精确光谱生物标志物的研究。
