干细胞拉曼光谱收集条件选择及间充质干细胞与胚胎干细胞拉曼光谱的比较

Optimization of Raman spectra acquisition conditions and its application in a comparison study on mesenchymal stem cells and embryonic stem cells

背景:拉曼光谱在检测细胞状态方面相比于传统技术具有快速、非介入、非标记等特点,因此在生物医学领域中的应用正日益受到关注,但是该技术应用于干细胞检测的条件还有待进一步优化。目的:探索激光波长与光栅刻度密度对于干细胞拉曼光谱分辨率与收集时间等参数的影响,明确可以用于干细胞分析的拉曼光谱收集条件。方法:首先比较532,638,785 nm 3个波长激光及600,1 200,1 800 gr/mm 3种光栅刻度密度条件对人间充质干细胞与人胚胎干细胞拉曼光谱的影响;之后比较这2个条件不同组合得到的拉曼光谱的光谱分辨率与收集时间,选择出最佳的组合条件;最后使用选出的最佳条件结合主成分分析方法比较胚胎干细胞与间充质干细胞拉曼光谱的差异以验证优化条件的适用性。结果与结论:(1)激光波长与光栅刻度密度均会不同程度影响干细胞拉曼光谱的峰强,进而影响特征性拉曼峰之间的比值;(2)785 nm波长激光与1 200 gr/mm光栅刻度密度条件组合是最适合收集条件;(3)对比分析胚胎干细胞与间充质干细胞的拉曼光谱图,提示胚胎干细胞具有更高的核酸含量,而间充质干细胞在一些蛋白和脂类含量上更为丰富。

BACKGROUND： Raman spectrum, compared with conventional detection technologies, is a rapid, non-invasive and label-free optical method. Its application has become an issue of concern in biomedical research. However, further studies are warranted to optimize the acquisition condition of Raman spectra from different stem cells. OBJECTIVE： To explore the effect of the wavelength of laser and the groove frequency of gratings to obtain the optimized parameter combination for Raman spectrum collection in human stem cells. METHODS： Using human mesenchymal stem cells as samples, the effects of different laser wavelengths （532, 38,785 nm） and different grating groove frequency （600, 1200, 1800 gr/mm） on Raman spectra were compared respectively. Then the different combinations of the wavelength and groove frequency were used and compared in terms of the spectra resolution and acquisition time, and the best acquisition condition was selected and applied in a comparison study on the Raman spectra from human mesenchymal stem cells and human embryonic stem cells. RESULTS AND CONCLUSION： The wavelengths of lasers and groove frequencies of gratings showed compound impacts on both the spikes at different wavenumbers and the ratio between spikes; the combination of 785 nm and 1200 gr/mm was confirmed to be the best spectrum features for human mesenchymal stem cells. The comparison of Raman spectra from human mesenchymal stem cells and human embryonic stem cells implies that the embryonic stem cells contain higher nucleic acids than the mesenchymal stem cells, while the mesenchymal stem cells appear to contain more proteins and lipids.