Supported molecular matrix electrophoresis: a new tool for characterization of glycoproteins.
A new concept of separation technology, supported molecular matrix electrophoresis (SMME), is described. In SMME, analytes migrate in a molecular matrix supported by backbone materials. Here we introduce a novel strategy for the separation and characterization of mucins using SMME. Mucin, a highly tumor-associated glycoprotein, has great potential as clinical biomarker for diagnosis of various malignant tumors. However, due to their large size, polymeric nature, and heterogeneous glycosylation, analysis of mucins has been left behind by modern techniques. For mucin analysis, we employed a poly(vinylidene difluoride) (PVDF) membrane and poly(vinyl alcohol) (PVA) as the backbone material and the matrix molecule, respectively. Combining SMME with mass spectrometry and capillary electrophoresis, we demonstrate that a crude porcine stomach mucin consists of a neutral and a sulfated mucin and is contaminated by chondroitin sulfate-containing proteoglycan and hyaluronic acid. Furthermore, to demonstrate the feasibility of the strategy for biomarker discovery, we analyzed mucins in human pancreatic juice, which is an important source for clinical biomarkers of pancreatic tumors. This work revealed the presence of three types of mucin with distinct glycan profiles in human pancreatic juice.
Matsuno
Yu-ki
YK
Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology, Open Space Laboratory C-2, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8568, Japan.
Saito
Takuro
T
Gotoh
Mitsukazu
M
Narimatsu
Hisashi
H
Kameyama
Akihiko
A
D019075:Electrophoresis, Capillary / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis*; D006801:Humans; D008567:Membranes, Artificial; D009077:Mucins / Q000032:analysis; D010189:Pancreatic Juice / Q000737:chemistry; D011145:Polyvinyls / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D005465:Fluorocarbon Polymers
D006023:Glycoproteins; D008567:Membranes, Artificial; D009077:Mucins; D011145:Polyvinyls; C024865:polyvinylidene fluoride; D005465:Fluorocarbon Polymers