Purification of anionic fluorescent probes through precise fraction collection with a two‐point detection system using multiple‐stacking preparative capillary transient isotachophoresis
| Autor: | Shingo Saito, Miyauchi Saori, Tomoko Haraga, Hiroto Tsujimura, Takuya Kamimura, Masami Shibukawa |
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| Rok vydání: | 2020 |
| Předmět: |
Anions
Chromatography Isotachophoresis Aptamer Ligand binding assay 010401 analytical chemistry Clinical Biochemistry Stacking Electrophoresis Capillary 02 engineering and technology Fractionation Aptamers Nucleotide 021001 nanoscience & nanotechnology 01 natural sciences Biochemistry Fluorescence 0104 chemical sciences Analytical Chemistry chemistry.chemical_compound chemistry Reagent Fluorescein 0210 nano-technology Fluorescent Dyes |
| Zdroj: | ELECTROPHORESIS. 41:1152-1159 |
| ISSN: | 1522-2683 0173-0835 |
| DOI: | 10.1002/elps.201900399 |
| Popis: | A novel combination of CE-based separation techniques was used for the precise fractionation of ionic compounds from impurities. The combination of on-capillary concentration and separation using transient isotachophoresis, with multiple injections and a two-point detection system provided higher efficiency, and accuracy at a microliter-scale injection volume, than when CE was individually used for purification. In this paper, we present successful applications of the CE fractionation techniques for the purification of fluorescein, fluorescein-4-isothiocyanate, two fluorescent metal ion probes, and a fluorescein-modified DNA aptamer. The purity of the isolated fluorescent probes ranged from 95 to 99%. Such high purity could not be achieved using chromatographic purification techniques. With relatively low dilution factors of 6-9, the purified probe solutions were practical for use as purified stock solutions. In addition, the fluorescein-modified DNA aptamer purified by our method was successfully used in a thrombin binding assay. The method developed was useful for the purification of anionic fluorescent reagents to be of ultratrace analytical grade for use with CE-LIF. |
| Databáze: | OpenAIRE |
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