High performance gel filtration using monodisperse highly cross-linked agarose as a one-step system for mucin purification.
High performance gel filtration on monodisperse cross-linked agarose (Superose 6) has been assessed as a system for purification of mucus glycoproteins. Comparison with the conventional two-step purification of mucus glycoprotein by Sepharose CL4B gel filtration followed by caesium chloride density gradient centrifugation shows that purification by high performance gel filtration is at least as thorough, yielding mucin that is free from non-mucin glycoproteins as defined by buoyant density, mobility on sodium dodecyl sulphate-polyacrylamide gel electrophoresis and absence of Concanavalin A binding (mannose-containing) material. This technique allows mucus glycoprotein to be purified from lyophilized crude mucin in 120 min compared with approximately 72 h using the conventional techniques. This makes the comparative study of mucus glycoprotein changes in disease states much more feasible.
Parker
N
N
University Department of Medicine, Royal Liverpool University Hospital, UK.
Finnie
I A
IA
Raouf
A H
AH
Ryder
S D
SD
Campbell
B J
BJ
Tsai
H H
HH
Iddon
D
D
Milton
J D
JD
Rhodes
J M
JM
D002586:Cesium / Q000737:chemistry; D002712:Chlorides; D002850:Chromatography, Gel / Q000379:methods*; D003432:Cross-Linking Reagents; D004247:DNA / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008055:Lipids / Q000032:analysis; D009077:Mucins / Q000737:chemistry / Q000302:isolation & purification*; D011480:Protease Inhibitors / Q000737:chemistry; D015203:Reproducibility of Results; D012685:Sepharose / Q000737:chemistry*; D013849:Thimerosal / Q000737:chemistry
D002712:Chlorides; D003432:Cross-Linking Reagents; D008055:Lipids; D009077:Mucins; D011480:Protease Inhibitors; D002586:Cesium; D013849:Thimerosal; D004247:DNA; D012685:Sepharose; C028019:cesium chloride