Here we present the realization of a novel fluorescence detection method based on the electromigration of fluorescent molecules within a nanocapillary combined with the laser excitation through a platinum (Pt)-coated nanocapillary. By using the Pt nanocapillary assisted focusing of a laser beam, we completely remove the background scattering on the tip of the electrophoretic nanocapillary. In this excitation geometry, we demonstrate a 1000-fold sensitivity enhancement (1.0 nM to 1.0 pM) compared to the detection in microcapillaries with epifluorescence illumination and fluorescence spectrophotometry. Due to a significant electroosmotic flow, we observe a decelerating migration of DNA molecules close to the tip of the electrophoretic nanocapillary. The reduced DNA translocation velocity causes a two-step stacking process of molecules in the tip of the nanocapillary and can be used as a way to locally concentrate molecules. The sensitivity of our method is further improved by a continuous electrokinetic injection of DNA molecules followed by sample zone stacking on the tip of the nanocapillary. Concentrations ranging from 0.1 pM to 1.0 fM can be directly observed on the orifice of the electrophoretic nanocapillary. This is a 1000-fold improvement compared to traditional capillary electrophoresis with laser-induced fluorescence.

中文翻译：

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正交的尖端到尖端纳米毛细管对准允许轻松检测飞摩尔浓度中的荧光发射体

在这里，我们介绍了一种新型的荧光检测方法，该方法基于纳米毛细管内荧光分子的电迁移并结合了通过铂（Pt）涂层的纳米毛细管的激光激发。通过使用Pt纳米毛细管辅助激光束聚焦，我们可以完全消除电泳纳米毛细管尖端上的背景散射。在这种激发几何结构中，与通过落射荧光照明和荧光分光光度法在微毛细管中进行检测相比，我们证明了灵敏度提高了1000倍（1.0 nM至1.0 pM）。由于大量的电渗流，我们观察到了接近电泳纳米毛细管尖端的DNA分子的减速迁移。降低的DNA易位速度导致纳米毛细管尖端中分子的两步堆叠过程，可以用作局部浓缩分子的方法。通过连续电动注入DNA分子，然后将样品区域堆积在纳米毛细管的尖端，可以进一步提高我们方法的灵敏度。可以直接在电泳纳米毛细管的孔上观察到浓度范围为0.1 pM至1.0 fM。与使用激光诱导的荧光的传统毛细管电泳相比，这是1000倍的改进。可以直接在电泳纳米毛细管的孔上观察到浓度范围为0.1 pM至1.0 fM。与使用激光诱导的荧光的传统毛细管电泳相比，这是1000倍的改进。可以直接在电泳纳米毛细管的孔上观察到浓度范围为0.1 pM至1.0 fM。与使用激光诱导的荧光的传统毛细管电泳相比，这是1000倍的改进。