The application of capillary electrophoresis for assisting whole-cell aptamers selection by characterizing complete ssDNA distribution
| Autor: | Jieying Yan, Feng Qu, Erning Chen, Beilei Lou, Xinying Zhao |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Aptamer DNA Single-Stranded 01 natural sciences Biochemistry Chemistry Techniques Analytical Analytical Chemistry HeLa 03 medical and health sciences Capillary electrophoresis Cell Line Tumor Humans Distribution (pharmacology) Gene Library Chromatography biology Chemistry SELEX Aptamer Technique 010401 analytical chemistry Organic Chemistry Electrophoresis Capillary Reproducibility of Results General Medicine Aptamers Nucleotide biology.organism_classification 0104 chemical sciences Binding ability 030104 developmental biology Capillary length Whole cell Systematic evolution of ligands by exponential enrichment HeLa Cells |
| Zdroj: | Journal of Chromatography A. 1437:203-209 |
| ISSN: | 0021-9673 |
| DOI: | 10.1016/j.chroma.2016.01.073 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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