Department of Chemical Engineering, Imperial College London, SW7 2AZ, London, UK.
Analyst. 2013 Jul 21;138(14):4040-7. doi: 10.1039/c3an00327b. Epub 2013 Mar 21.
The label-free, non-destructive chemical analysis offered by FTIR spectroscopic imaging is a very attractive and potentially powerful tool for studies of live biological cells. FTIR imaging of live cells is a challenging task, due to the fact that cells are cultured in an aqueous environment. While the synchrotron facility has proven to be a valuable tool for FTIR microspectroscopic studies of single live cells, we have demonstrated that high quality infrared spectra of single live cells using an ordinary Globar source can also be obtained by adding a pair of lenses to a common transmission liquid cell. The lenses, when placed on the transmission cell window, form pseudo hemispheres which removes the refraction of light and hence improve the imaging and spectral quality of the obtained data. This study demonstrates that infrared spectra of single live cells can be obtained without the focus shifting effect at different wavenumbers, caused by the chromatic aberration. Spectra of the single cells have confirmed that the measured spectral region remains in focus across the whole range, while spectra of the single cells measured without the lenses have shown some erroneous features as a result of the shift of focus. It has also been demonstrated that the addition of lenses can be applied to the imaging of cells in microfabricated devices. We have shown that it was not possible to obtain a focused image of an isolated cell in a droplet of DPBS in oil unless the lenses are applied. The use of the approach described herein allows for well focused images of single cells in DPBS droplets to be obtained.
傅里叶变换红外光谱成像提供的无标记、非破坏性化学分析是研究活生物细胞的一种非常有吸引力和潜在强大的工具。由于细胞是在水相环境中培养的，因此对活细胞进行傅里叶变换红外成像技术是一项具有挑战性的任务。虽然同步加速器设施已被证明是用于单活细胞傅里叶变换显微镜研究的有价值工具，但我们已经证明，通过在普通透射液池上添加一对透镜，也可以用普通的 Globar 光源获得单活细胞的高质量红外光谱。当透镜放置在透射池窗口上时，形成的伪半球可以消除光线的折射，从而改善获得的数据的成像和光谱质量。这项研究表明，在不同的波数处，由于色差引起的聚焦偏移效应，也可以获得单活细胞的红外光谱。单细胞的光谱证实，在所测量的整个范围内，测量的光谱区域仍保持聚焦，而没有透镜的单细胞光谱则由于焦点的偏移显示出一些错误的特征。还证明了可以将透镜的添加应用于微制造设备中细胞的成像。我们已经表明，除非使用透镜，否则无法在油中的 DPBS 液滴中获得单个细胞的聚焦图像。使用本文所描述的方法，可以获得 DPBS 液滴中单细胞的良好聚焦图像。
