Chaudhary Neha, Que Nguyen Thi Nguyet, Maguire Adrian, Wynne Claire, Meade Aidan D
School of Physics, Technological University Dublin, Kevin Street, Dublin 8, Ireland.
Anal Methods. 2021 Feb 28;13(8):1019-1032. doi: 10.1039/d0ay02040k. Epub 2021 Feb 4.
The exquisite sensitivity of Raman spectroscopy to the molecular composition of biological samples has been a particular strength in its development towards clinical applicates. Its strength in this regard also presents challenges in the development of its diagnostic capabilities owing to its sensitivity, not only to the sample biochemistry, but also the preparation methodology employed prior to analysis. Here we have examined and optimised several approaches to the preparation of peripheral blood mononuclear cells (PBMCs), or immune cell subtypes of whole blood, for Raman spectroscopic analysis. Two approaches to the elimination of haemoglobin contamination, and two approaches to the purification of the lymphocyte portion of whole blood were investigated. It was found that a peroxide treatment of PBMCs prior to spectroscopic analysis was required for elimination of haemoglobin, while a negative selection approach involving magnetically labelled monoclonal antibodies was preferred for purification of individual leucocyte subpopulations in comparison to the plastic adherence method using an ex vivo culture. Further spectral fitting analysis has identified spectral features of interest which may be useful in the identification of individual leucocytes spectrally and warrant further investigation.
拉曼光谱对生物样品分子组成的极高灵敏度一直是其向临床应用发展的一项特殊优势。由于其不仅对样品生物化学敏感，而且对分析前采用的制备方法敏感，其在这方面的优势也给其诊断能力的发展带来了挑战。在此，我们研究并优化了几种用于制备外周血单个核细胞（PBMC）或全血免疫细胞亚型以进行拉曼光谱分析的方法。研究了两种消除血红蛋白污染的方法以及两种纯化全血淋巴细胞部分的方法。结果发现，光谱分析前需要用过氧化物处理PBMC以消除血红蛋白，而与使用体外培养的塑料贴壁法相比，涉及磁性标记单克隆抗体的阴性选择方法更适合用于纯化单个白细胞亚群。进一步的光谱拟合分析确定了感兴趣的光谱特征，这些特征可能有助于从光谱上识别单个白细胞，值得进一步研究。
