Purification of anionic fluorescent probes through precise fraction collection with a two-point detection system using multiple-stacking preparative capillary transient isotachophoresis.
Capillary transient isotachophoresis
Multiple stacking
Precise fraction collection
Thrombin binding assay
Two-point detection CE-UV
Journal
Electrophoresis
ISSN: 1522-2683
Titre abrégé: Electrophoresis
Pays: Germany
ID NLM: 8204476
Informations de publication
Date de publication:
07 2020
07 2020
Historique:
received:
14
10
2019
revised:
27
03
2020
accepted:
29
03
2020
pubmed:
8
4
2020
medline:
23
3
2021
entrez:
8
4
2020
Statut:
ppublish
Résumé
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.
Identifiants
pubmed: 32253765
doi: 10.1002/elps.201900399
doi:
Substances chimiques
Anions
0
Aptamers, Nucleotide
0
Fluorescent Dyes
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
1152-1159Subventions
Organisme : Japan Society for the Promotion of Science
Pays : International
Organisme : Japan Atomic Energy Agency
ID : Cooperative Research Scheme on the Nuclear Fuel Cy
Pays : International
Informations de copyright
© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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