Solid-phase permethylation for glycomic analysis.
Methods Mol Biol
; 534: 53-64, 2009.
Article
em En
| MEDLINE
| ID: mdl-19277536
ABSTRACT
This chapter discusses in detail a miniaturized version of the widely used permethylation technique which permits quantitative derivatization of oligosaccharides derived from minute quantities of glycoprotein. The approach involves packing of sodium hydroxide powder or beads in a microcolumn format, including spin columns, fused silica capillaries (500 microm i.d.) and plastic tubes (1 mm i.d.). The derivatization proceeds effectively in less than a minute time scale and it is applicable to glycans derived from femtomole quantities of glycoproteins. Prior to mass spectrometry (MS), methyl iodide is added to analytes suspended in dimethyl sulfoxide solution containing traces of water. The reaction mixture is then immediately infused through the microreactor. The packed sodium hydroxide powder or beads inside the microcolumns minimize oxidative degradation and peeling reactions which are otherwise commonly associated with the conventional permethylation technique. In addition, this solid-phase permethylation approach eliminates the need for excessive sample clean-up. As demonstrated below, picomole amounts of various types of glycans derived from model glycoproteins as well as real samples, including linear and branched, sialylated and neutral glycans were shown to become rapidly and efficiently permethylated through this approach.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Glicoproteínas
/
Microextração em Fase Sólida
/
Glicômica
Limite:
Animals
/
Humans
Idioma:
En
Revista:
Methods Mol Biol
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2009
Tipo de documento:
Article
País de afiliação:
Estados Unidos
País de publicação:
EEUU
/
ESTADOS UNIDOS
/
ESTADOS UNIDOS DA AMERICA
/
EUA
/
UNITED STATES
/
UNITED STATES OF AMERICA
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US
/
USA