[Purification and separation of mannan-binding lectin (MBL) and MBL-associated serine proteases complex from human plasma].
OBJECTIVE
To separate mannan-binding lectin (MBL) and MBL-associated serine proteases (MASPs) from human plasma.


METHODS
A two-step affinity chromatography on underivatized sepharose 4B was employed for purification of MBL-MASP complex, followed by gel filtration on a Sephacryl S-300 column for separation of MBL and MASPs from the complex. The purification procedures were performed at 4 degrees Celsius with the addition of two proteolytic inhibitors, phenyl methylsulfonyl fluoride and 1,10-phenanthroline during affinity chromatography but not in the gel filtration buffer.


RESULTS
Preparations of highly purified MBL and proenzyme MASPs were obtained. The purified MBL was shown by SDS-PAGE and Western blotting to be a functional multimer composed of 28,000 and 32,000 peptide chains, with high bioactivity as demonstrated by ligand-binding assay and yeast agglutination experiment.


CONCLUSIONS
A simple and convenient procedure is established successfully for the purification of MBL and the proenzyme MASPs.
Chen
Yue
Y
Department of Immunology, Southern Medical University, Guangzhou 510515, China. chenyue@fimmu.com
Zhang
Li-yun
LY
Chen
Zheng-liang
ZL
D002846:Chromatography, Affinity; D006801:Humans; D037601:Mannose-Binding Lectin / Q000302:isolation & purification*; D050606:Mannose-Binding Protein-Associated Serine Proteases / Q000302:isolation & purification*; D010949:Plasma / Q000737:chemistry*
D037601:Mannose-Binding Lectin; D050606:Mannose-Binding Protein-Associated Serine Proteases