In capillary labeling and online electrophoretic separation of N-glycans from glycoproteins.
Bin YangThanh Duc MaiNguyet Thuy TranMyriam TavernaPublished in: Journal of separation science (2022)
In this study, we present a new approach for in-capillary fluorescent labeling of N-glycans prior to their analysis with CE coupled with laser-induced fluorescent detection. This integrated approach allows using a CE capillary as a microreactor to perform several steps required for labeling glycans with 8-aminopyrene-1,3,6 trisulfonic acid and at the same time as a separation channel for CE of fluorescently labeled glycans. This could be achieved through careful optimization of all different steps, including sequential injections of fluorescent dye and glycan plugs, mixing by transverse diffusion of laminar flow profiles, incubation in a thermostatic zone, and finally separation and detection with CE. Such a complex sample treatment protocol for glycan labeling that is feasible thus far only in batchwise mode can now be converted into an automated and integrated protocol. Our approach was applied successfully to analyze fluorescently labeled N-linked oligosaccharides released from human immunoglobulin G and rituximab, a monoclonal antibody used for cancer treatment. We demonstrated the superiority of this in-capillary approach over the conventional in-tube protocol, with fourfold less reagent consumption and full automation without remarkable degradation of the glycan separation profile obtained by capillary electrophoresis.
Keyphrases
- cell surface
- capillary electrophoresis
- label free
- liquid chromatography
- monoclonal antibody
- quantum dots
- mass spectrometry
- randomized controlled trial
- energy transfer
- living cells
- loop mediated isothermal amplification
- endothelial cells
- pet imaging
- diffuse large b cell lymphoma
- health information
- ultrasound guided
- hodgkin lymphoma
- combination therapy
- smoking cessation
- platelet rich plasma