Post-capillary reaction detection in capillary electrophoresis based on the streptavidin-biotin interaction. Optimization and application to single cell analysis

Autor:	A, Feltus, N G, Hentz, S, Daunert
Rok vydání:	2001
Předmět:	Spectrometry Fluorescence Sea Urchins Oocytes Animals Biotin Electrophoresis Capillary Streptavidin Hydrogen-Ion Concentration Cells Cultured Fluorescein-5-isothiocyanate Rats
Zdroj:	Journal of chromatography. A. 918(2)
ISSN:	0021-9673
Popis:	A class-selective post-capillary reaction detection method for capillary electrophoresis is described in which a streptavidin-fluorescein isothiocyanate (streptavidin-FITC) conjugate is used to detect biotin moieties. The selective binding of biotin moieties to the streptavidin-FITC conjugate causes an enhancement of fluorescence proportional to the concentration of biotin present. After capillary electrophoresis the separated analytes react with streptavidin-FITC in a coaxial reactor and are then detected either by a benchtop spectrofluorometer (2.5 microM detection limit) or by an epi-fluorescence microscope (1 x 10(-7) M detection limit). The method is used to examine biotinylated species in a crude mammalian cell lysate which was found to contain 83+/-3 fmol in 3600 cell volumes. In addition, it is used to examine the uptake of biotin by individual sea urchin oocytes. The results indicate that, in the oocytes, biocytin is the prevalent form of biotin and its concentration varies widely between cells (mean=2+/-2 microM).
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=pmid________::1ba69cf08b25347820fe892322ad246b https://pubmed.ncbi.nlm.nih.gov/11407585 Zobrazit plný text záznamu Full Text from ScienceDirect