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首页> 外文期刊>The Analyst >Label-free molecular imaging of immunological synapses between dendritic and T cells by Raman micro-spectroscopy
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Label-free molecular imaging of immunological synapses between dendritic and T cells by Raman micro-spectroscopy

机译：树突状细胞和T细胞之间的免疫突触的无标记分子成像的拉曼光谱

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Confocal Raman micro-spectroscopy (CRMS) was used to measure spectral images of immunologicalnsynapse formation between dendritic and T cells without using molecular labels or other invasivenprocedures. The purpose-built inverted CRMS instrument integrated an environmental enclosure andna near-infrared laser to allow measurements on live cells maintained under physiological conditions.nThe integration of the wide-field fluorescence also enabled viability assays and direct comparisonnbetween Raman spectral images and gold-standard immuno-fluorescence images for specific molecules.nRaman spectral images of nucleus and proteins were built by fuzzy c-mean clustering method. ThenRaman images were found to be in good correspondence with the immuno-fluorescence images ofnDNA and actin. These results indicate that actin is a main contributor to the Raman spectrum of thencytoplasm of dendritic and T cells. While for control cells the Raman spectral images of proteinsnindicated a more homogeneous distribution of proteins in the cytoplasm of dendritic cells, theynindicated a higher accumulation of proteins at the immunological synapses when dendritic cells werenpre-treated with laminin. These conclusions were also supported by confocal immuno-fluorescencenimaging after cell fixation and labelling. This study demonstrates the potential of CRMS for label-freennon-invasive imaging of junctions between live cells. Therefore, this technique may become a usefulntool for studying cellular processes in live cells and where non-invasive molecular specific imaging isndesirable, such as cell–cell interactions.
机译：共焦拉曼显微光谱法（CRMS）用于测量树突细胞与T细胞之间免疫突触形成的光谱图像，而无需使用分子标记或其他侵入性程序。专门制造的倒置CRMS仪器集成了一个环境外壳和一个近红外激光，可以对在生理条件下维持的活细胞进行测量。通过模糊c均值聚类方法建立核和蛋白质的nRaman光谱图像。然后发现拉曼图像与nDNA和肌动蛋白的免疫荧光图像非常吻合。这些结果表明肌动蛋白是树突状和T细胞的胞质拉曼光谱的主要贡献者。对于对照细胞，蛋白质的拉曼光谱图像表明树突状细胞的细胞质中蛋白质分布更均匀，而当用层粘连蛋白预处理树突状细胞时，则表明免疫突触中蛋白质的积累更高。细胞固定和标记后共聚焦免疫荧光成像也支持了这些结论。这项研究证明了CRMS在活细胞之间连接的无标记无创成像中的潜力。因此，这项技术可能成为研究活细胞中细胞过程以及在不需要非侵入性分子特异性成像（例如细胞间相互作用）的情况下有用的工具。

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    《The Analyst》 |2010年第12期|p.3205-3212|共8页
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    Alina Bogumila Zoladek Ramneek Kaur Johal Samuel Garcia-Nieto Flavius Pascut Kevin M. Shakesheff Amir M. Ghaemmaghami and Ioan Notingher;

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    aSchool of Physics and Astronomy, University of Nottingham, Nottingham,UK. E-mail: ioan.notingher@nottingham.ac.ukbAllergy Research Group, School of Molecular Medical Sciences andRespiratory Biomedical Research Unit, University of Nottingham,Nottingham, UKcSchool of Pharmacy, University of Nottingham, Nottingham, UK† This article is part of a themed issue on Optical Diagnosis. This issueincludes work presented at SPEC 2010 Shedding Light on Disease:Optical Diagnosis for the New Millennium, which was held inManchester, UK June 26th–July 1st 2010.;

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