Fibulin-1 purification from human plasma using affinity chromatography on Factor H-Sepharose

• A method has been established to purify human Fibulin-1 from plasma.
• The last step is affinity chromatography on Factor H-Sepharose.
• The purification can be integrated with published methods enabling the purification of 30 proteins from human plasma.
• Fibulin-1 binding to Factor H, fibronectin and fibrinogen is augmented appreciably by calcium ion.
• At least two Fibulins have a binding capacity for Factor H.

A method is reported to purify Fibulin-1 from human plasma resulting in a 36% recovery. The steps involve removal of the cryoglobulin and the vitamin K dependent proteins followed by polyethylene glycol and ammonium sulfate precipitations, DEAE-Sephadex column chromatography and finally Factor H-Sepharose affinity purification. The procedure is designed to be integrated into an overall scheme for the isolation of over 30 plasma proteins from a single batch of human plasma. Results from mass spectroscopy, SDS-PAGE, and Western blotting indicate that human plasma Fibulin-1 is a single chain of the largest isotype. Functional binding assays demonstrated calcium ion dependent interaction of Fibulin-1 for fibrinogen, fibronectin, and Factor H. The procedure described is the first to our knowledge that enables a large scale purification of Fibulin-1 from human plasma.