In this study, we used stimulated Raman spectroscopy (SRS) microscopy to collect Raman signatures from live Saccharomyces cerevisiae cells in the spectral range 2804−3060 cm⁻¹ and 830−2000 cm⁻¹ with a spectral resolution of 8 cm⁻¹. To effect this, we tuned the pump beam to several distinct wavelengths and thus acquired a series of chemical maps in order to reconstruct SRS spectra based on the intensity of the pixels, an approach also referred as hyperspectral SRS (hsSRS). One of the advantages of hsSRS over spontaneous Raman is that it is not overtly plagued by fluorescence and so fluorescent samples like yeast can be analysed. We show however that Raman signatures acquired by this approach may be subject to spectral artefacts that manifest as drops in intensity of Raman signal due to the movement of lipid droplets (LDs) within the yeast cells. To overcome this issue, yeast cells were chemically fixed with 4% formaldehyde and no artefacts were observed in the Raman signatures acquired from ‘stationary’ samples. Our findings indicate that caution must be applied when analysing SRS signatures obtained through hsSRS from mobile LDs and/or any other moving target within a system, whether biological or not.

在这项研究中，我们使用受激拉曼光谱 (SRS) 显微镜从活的酿酒酵母细胞中收集光谱范围 2804-3060 cm ^-1和 830-2000 cm ^{-1 的}拉曼特征，光谱分辨率为 8 cm ^-1. 为了实现这一点，我们将泵浦光束调整到几个不同的波长，从而获得了一系列化学图，以便根据像素强度重建 SRS 光谱，这种方法也称为高光谱 SRS (hsSRS)。hsSRS 相对于自发拉曼的优势之一是它不受荧光的明显困扰，因此可以分析酵母等荧光样品。然而，我们表明，通过这种方法获得的拉曼特征可能会受到光谱伪影的影响，这些伪影表现为由于酵母细胞内脂滴 (LD) 的运动而导致的拉曼信号强度下降。为了克服这个问题，酵母细胞用 4% 的甲醛化学固定，并且在从“固定”样品中获得的拉曼特征中没有观察到任何人工制品。