Real-time measurements of dissolved oxygen inside live cells by organically modified silicate fluorescent nanosensors
Real-time measurements of dissolved oxygen inside live cells by organically modified silicate fluorescent nanosensors
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DOI：
10.1021/ac035493f
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发表时间：
2004-05-01
影响因子：
7.4
通讯作者：
Philbert, MA
中科院分区：
文献类型：
--
作者：
Koo, YEL;Cao, YF;Philbert, MA
光学 PEBBLE（通过生物局部嵌入封装的探针）纳米传感器已开发用于使用有机改性硅酸盐（ormosil）纳米颗粒作为基质来检测溶解氧。 Ormosil纳米颗粒是通过基于溶胶-凝胶的方法制备的，该方法包括以苯基三甲氧基硅烷作为前体形成核颗粒，然后以甲基三甲氧基硅烷作为前体形成涂层。所得颗粒的平均直径为120nm。这些传感器采用氧敏感铂卟啉染料作为指示剂，并采用氧不敏感染料作为比率强度测量的参考。使用两对指示剂染料和参比染料，分别是八乙基卟啉铂(II)和3,3'-二十八烷基氧羰花青高氯酸盐，以及八乙基卟啉铂(II)酮和八乙基卟啉。该传感器具有出色的灵敏度，总体淬灭响应率为 97%，并且在整个溶解氧浓度范围 (0-43 ppm) 上 Stern-Volmer 图 (r(2) = 0.999) 具有出色的线性度。在大鼠 C6 神经胶质瘤细胞中，用基因枪注射 PEBBLE，监测体外细胞内由于细胞呼吸而产生的溶解氧变化。对于九组不同的细胞，观察到显着变化，1 小时后荧光比率增加高达 500%，这对应于溶解氧浓度减少 90%。这些结果清楚地表明了 PEBBLE 传感器细胞内研究的传递方法的有效性，以及实现细胞内溶解氧浓度实时测量所需的高灵敏度。
Optical PEBBLE (probes encapsulated by biologically localized embedding) nanosensors have been developed for dissolved oxygen using organically modified silicate (ormosil) nanoparticles as a matrix. The ormosil nanoparticles are prepared via a sol-gel-based process, which includes the formation of core particles with phenyltrimethoxysilane as a precursor followed by the formation of a coating layer with methyltrimethoxysilane as a precursor. The average diameter of the resultant particles is 120 nm. These sensors incorporate the oxygen-sensitive platinum porphyrin dye as an indicator and an oxygen-insensitive dye as a reference for ratiometric intensity measurement. Two pairs of indicator dye and reference dye, respectively, platinum(II) octaethylporphine and 3,3'-dioctadecyloxacarbocyanine perchlorate, and platinum(II) octaethylporphine ketone and octaethylporphine, were used. The sensors have excellent sensitivity with an overall quenching response of 97%, as well as excellent linearity of the Stern-Volmer plot (r(2) = 0.999) over the whole range of dissolved oxygen concentrations (0-43 ppm). In vitro intracellular changes of dissolved oxygen due to cell respiration were monitored, with gene gun injected PEBBLEs, in rat C6 glioma cells. A significant change was observed with a fluorescence ratio increase of up to 500% after 1 h, for nine different sets of cells, which corresponds to a 90% reduction in terms of dissolved oxygen concentration. These results clearly show the validity of the delivery method for intracellular studies of PEBBLE sensors, as well as the high sensitivity, which is needed to achieve real-time measurements of intracellular dissolved oxygen concentration.