Single cell measurement of micro-viscosity by ratio imaging of fluorescence of styrylpyridinium probe.
Single cell measurement of micro-viscosity by ratio imaging of fluorescence of styrylpyridinium probe.
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DOI：
10.1016/j.bios.2004.06.045
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发表时间：
2005-03
影响因子：
12.6
通讯作者：
B. Wandelt;P. Cywiński;G. D. Darling;B. Stranix
中科院分区：
文献类型：
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作者：
B. Wandelt;P. Cywiński;G. D. Darling;B. Stranix
在水溶液中，含有苯乙烯基吡啶鎓基团的化合物显示出双重荧光，其中在469或360 nm处的激发各自产生约600 nm的发射带。两条带的荧光强度比值（R = I469/I360）对局部粘度敏感。发现DMASP的N-羧甲基丁酯能够不可逆地加载到活细胞中;推测通过涉及细胞脂肪酶的水解，其被转化为膜不可渗透的荧光羧酸盐。使用荧光成像显微镜和荧光分光光度计以双激发单发射模式产生来自单个细胞的比率R的图。在校准探针在水/甘油溶液中的比例后，获得大鼠胚胎胸主动脉平滑肌的单个细胞的细胞内粘度。细胞内粘度在细胞内有差异，所得值18- 7 cP符合其他实验室报道的所有值。探针的荧光发射（500- 650 nm）处于非常有利的区域，适合与可见荧光显微镜一起使用，不会受到细胞或组织自发荧光的干扰。结果呈现出检测和跟踪荧光强度比率的微小变化的能力，并且显然是微粘度的微小变化。
In aqueous solution, compounds containing the styrylpyridinium group showed dual fluorescence, in which excitation at either 469 or 360nm each produced an emission band around 600nm. The ratio of fluorescence intensities of the two bands (R = I469/I360) was sensitive to local viscosity. The N-carboxymethyl butyl ester of DMASP was found to be able to irreversibly load into a living cell; presumably by hydrolysis involving cellular lipases it was transformed to a membrane-impermeable fluorescent carboxylate. A map of the ratio, R, from a single cell was generated using fluorescence imaging microscopy with a spectrofluorimeter in dual-excitation single-emission mode. After calibrating the ratio for the probe in water/glycerol solutions, the intracellular viscosities were obtained for a single cell of smooth muscle of a rat embryonic thoracic aorta. The intracellular viscosity is differentiated inside the cell and the obtained values 18–7cP obey all the values reported by other laboratories. Fluorescence emission of the probe (500–650nm) is in a very favourable region for its use with visible fluorescence microscopy, without interferences from cell or tissue auto-fluorescence. The results present ability to detect and follow small changes in the ratio of fluorescence intensities, and apparently of the micro-viscosity.