Department of Micro and Nanotechnology, Technical University of Denmark, Kongens Lyngby, Denmark.
Electrophoresis. 2010 Nov;31(22):3696-702. doi: 10.1002/elps.201000322. Epub 2010 Oct 22.
Porous liquid crystalline lipid-based nanoparticles are shown here to enable protein analysis in microchip electroseparation by reducing sample adsorption. Additionally, higher stability and reproducibility of the separations were observed. The method was tested by separating green fluorescent protein (GFP) in hot embossed cyclic olefin polymer microchips with integrated fiber grooves for LIF detection. The sample adsorption was indirectly quantified by measuring the height, width and asymmetry of the separation peaks for various concentrations of nanoparticles in the sample and background electrolyte. Without nanoparticles, electropherograms displayed typical signs of extensive adsorption to the channel walls, with low, broad tailing peaks. Higher, narrower more symmetric peaks were generated when 0.5-10% nanoparticles were added, showing a dramatic reduction of sample adsorption. The current through the separation channel decreased with nanoparticle concentration, reducing to half its value when the nanoparticle concentration was increased from 0.5 to 4%. Addition of nanoparticles enabled separations that were otherwise hindered by extensive adsorption, e.g. separation of GFP mutants differing by only one amino acid. It was also observed that increasing the nanoparticle concentration increased the number of impurities that could be resolved in a GFP sample. This indicates that the adsorption is further reduced, and/or that the nanoparticles provide an interacting pseudostationary phase for electrochromatography.
多孔液晶脂基纳米粒子可减少样品吸附，从而实现微芯片电分离中的蛋白质分析。此外，还观察到分离的稳定性和重现性更高。该方法通过在带有集成光纤槽的热压环烯烃聚合物微芯片中分离绿色荧光蛋白 (GFP) 进行了测试，用于 LIF 检测。通过测量不同浓度纳米粒子在样品和背景电解质中的样品和背景电解质中的分离峰的高度、宽度和不对称性，间接定量了样品吸附。没有纳米粒子时，电泳图谱显示出与通道壁广泛吸附的典型迹象，峰低而宽，拖尾严重。当添加 0.5-10%的纳米粒子时，会产生更高、更窄、更对称的峰，表明样品吸附明显减少。分离通道中的电流随纳米粒子浓度的增加而降低，当纳米粒子浓度从 0.5 增加到 4%时，电流降低到其值的一半。添加纳米粒子可以实现否则因广泛吸附而受阻的分离，例如仅相差一个氨基酸的 GFP 突变体的分离。还观察到增加纳米粒子浓度会增加 GFP 样品中可分辨的杂质数量。这表明吸附进一步减少，和/或纳米粒子为电色谱提供了相互作用的拟稳态相。
