Ciobanu Madalina, Taylor Dale E, Wilburn Jeremy P, Cliffel David E
Department of Chemistry, Vanderbilt University, Nashville, Tennessee 37235, USA.
Anal Chem. 2008 Apr 15;80(8):2717-27. doi: 10.1021/ac7021184. Epub 2008 Mar 18.
We have developed glucose and lactate ultramicroelectrode (UME) biosensors based on glucose oxidase and lactate oxidase (with enzymes immobilized onto Pt UMEs by either electropolymerization or casting) for scanning electrochemical microscopy (SECM) and have determined their sensitivity to glucose and lactate, respectively. The results of our evaluations reveal different advantages for sensors constructed by each method: improved sensitivity and shorter manufacturing time for hand-casting, and increased reproducibility for electropolymerization. We have acquired amperometric approach curves (ACs) for each type of manufactured biosensor UME, and these ACs can be used as a means of positioning the UME above a substrate at a known distance. We have used the glucose biosensor UMEs to record profiles of glucose uptake above individual fibroblasts. Likewise, we have employed the lactate biosensor UMEs for recording the lactate production above single cancer cells with the SECM. We also show that oxygen respiration profiles for single cancer cells do not mimic cell topography, but are rather more convoluted, with a higher respiration activity observed at the points where the cell touches the Petri dish. These UME biosensors, along with the application of others already described in the literature, could prove to be powerful tools for mapping metabolic analytes, such as glucose, lactate, and oxygen, in single cancer cells.
我们基于葡萄糖氧化酶和乳酸氧化酶开发了葡萄糖和乳酸超微电极（UME）生物传感器（通过电聚合或浇铸将酶固定在铂超微电极上），用于扫描电化学显微镜（SECM），并分别测定了它们对葡萄糖和乳酸的灵敏度。我们的评估结果揭示了每种方法构建的传感器的不同优势：手工浇铸的传感器灵敏度提高且制造时间缩短，电聚合的传感器重现性提高。我们为每种制造的生物传感器超微电极获取了安培法逼近曲线（ACs），这些ACs可用于将超微电极以已知距离定位在底物上方。我们使用葡萄糖生物传感器超微电极记录单个成纤维细胞上方的葡萄糖摄取情况。同样，我们使用乳酸生物传感器超微电极通过SECM记录单个癌细胞上方的乳酸产生情况。我们还表明，单个癌细胞的氧呼吸曲线并不模仿细胞形貌，而是更加复杂，在细胞接触培养皿的点处观察到更高的呼吸活性。这些超微电极生物传感器，连同文献中已描述的其他传感器的应用，可能被证明是用于绘制单个癌细胞中葡萄糖、乳酸和氧气等代谢分析物图谱的强大工具。
