title: Discrimination of glycoproteins via two-color laser-induced fluorescence detection coupled with postcolumn derivatization in capillary electrophoresis.
abstract: Here, we report a novel method consisting of capillary electrophoretic separation followed by two-color LIF detection with postcolumn derivatization. The method can be used to discriminate glycoproteins in a protein mixture containing both glycosylated and unglycosylated proteins. The detector permitted simultaneous measurements of two electropherograms obtained by 450 nm (diode laser) and 532 nm (Nd:YAG laser) lasers excited native proteins following postcolumn derivatization with naphthalene-2,3-dicarboxaldehyde and concanavalin A (Con A) labeled with tetramethylrhodamine (rhodamine-labeled Con A), respectively. So, a protein can be assigned as glycosylated if it shows a peak at the same migration time in both electropherograms. According to the proposed principle, in a single run we discriminated a glycosylated protein (thyroglobulin) from an unglycosylated protein (albumin) in the presence of rhodamine-labeled Con A. Because the methodology permits the simultaneous detection of native proteins and their complexes with a fluorescently labeled probe, it should have broad applicability to binding assays.
authors.affiliation: Department of Chemistry, Graduate School of Natural Science and Technology, Okayama University, Okayama, Japan. kaneta@okayama-u.ac.jp
authors.forename: Ayumi
authors.initials: A
authors.lastname: Tabara
authors.forename: Takashi
authors.initials: T
authors.lastname: Kaneta
mesh_terms: D003208:Concanavalin A / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D007834:Lasers; D009281:Naphthalenes / Q000737:chemistry; D012235:Rhodamines / Q000737:chemistry; D012709:Serum Albumin / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods*
chemical_list: D006023:Glycoproteins; D009281:Naphthalenes; D012235:Rhodamines; D012709:Serum Albumin; D003208:Concanavalin A; C005358:tetramethylrhodamine; C027717:2,3-naphthalenedicarboxaldehyde
keywords: Capillary electrophoresis; Concanavalin A; Glycoprotein; Postcolumn derivatization; Two-color laser-induced fluorescence