The application of capillary electrophoresis for assisting whole-cell aptamers selection by characterizing complete ssDNA distribution

• CE applied to characterize complete ssDNA sequence distribution in whole-cell SELEX process.
• CE applied for whole-cell target and ssDNA interaction evaluation.
• Binding comparison of three cell lines with a ssDNA sequence and a ssDNA library

Whole-cell SELEX faces more difficulties than SELEX against purified molecules target. In this work, we demonstrate the application of capillary electrophoresis for assisting whole-cell aptamers selection by characterizing complete ssDNA distribution. We chose three cancer cell lines U251, Hela and PC3 as target, FAM labeled Sgc8c (a 41mer aptamer) and FAM labeled 41mer random ssDNA library as ssDNA model. CE conditions of running buffer and capillary length and inner diameter as well as UV and LIF detection were optimized. The distribution percentage of Sgc8c and ssDNA library against U251, Hela and PC3 was demonstrated, the relative peak area of their complex is 8.94%, 1.05% and 0.44% for Sgc8c and 9.03%, 1.04% and 0.12% for ssDNA library respectively. Under the chosen experimental conditions, binding ability comparison of three cell lines was U251 > Hela > PC3, which was validated by laser confocol microscope. For each cell, distribution percentage of ssDNA library was compared with that of Sgc8c. Finally, whole-cell complex of U251–Sgc8c was confirmed by increase incubation time and fraction CE analysis.