Abstract：
Elemental analysis at the single-cell level is an emerging technique in the field of inductively coupled plasma mass spectrometry (ICP-MS). In comparison to the analysis of cell suspensions by fast time-resolved analysis, single-cell sampling by laser ablation (LA) allows the discriminatory analysis of single cells according to their size and morphology. In this study, we evaluated the changes in elemental contents in a single cell through differentiation of rat adrenal pheochromocytoma into neuron-like cells by LA-ICP-MS. The contents of seven essential minerals were increased about 2-3 times after the differentiation. In addition, we evaluated the degree of differentiation at the single-cell level by means of imaging cytometry after immunofluorescence staining of microtubule-associated protein 2 (Map2), a neuron-specific protein. The fluorescence intensities of Alexa Fluor 488-conjugated secondary antibody against the anti-Map2 primary antibody showed large variations among the cells after the onset of differentiation. Although the average fluorescence intensity was increased through the differentiation, there were still less-matured neuron-like cells that exhibited a lower fluorescence intensity after 5 days of differentiation. Since a positive correlation between the fluorescence intensity and the cell size in area was found, we separately measured the elemental contents in the less-matured smaller cells and well-matured larger cells by LA-ICP-MS. The larger cells had higher elemental contents than the smaller cells, indicating that the essential minerals are highly required at a later stage of differentiation.
摘要：
单细胞水平的元素分析是电感耦合等离子体质谱（ICP-MS）领域的新兴技术。与通过快速时间分辨分析进行细胞悬浮液分析相比，通过激光消融（LA）进行单细胞采样，可以根据其大小和形态对单细胞进行区别分析。在这项研究中,我们通过LA-ICP-MS将大鼠肾上腺嗜铬细胞瘤分化为神经元样细胞，评估了单个细胞中元素含量的变化。分化后7种必需矿物质的含量增加了约2-3倍。此外,我们在微管相关蛋白2（Map2）免疫荧光染色后，通过成像细胞计数评估了单细胞水平的分化程度，神经元特异性蛋白.在分化开始后,针对抗Map2一级抗体的AlexaFluor488缀合的二级抗体的荧光强度在细胞中显示出很大的变化。虽然平均荧光强度通过分化增加，分化5天后,仍有较不成熟的神经元样细胞表现出较低的荧光强度.由于发现荧光强度与细胞面积之间存在正相关，我们通过LA-ICP-MS分别测量了较不成熟的较小细胞和成熟的较大细胞中的元素含量。较大的细胞比较小的细胞有更高的元素含量，表明在分化的后期阶段高度需要必需矿物质。
