Rhone-poulenc Rorer Pharmaceuticals Inc.

Thomas L Darling

Michael E Hrinda

Jonathan J Mitschelen

Lida Y Akhnana

Recovery of the 52/48 kDa tryptic fragment of vWF or peptide subfragments thereof produced in the form of inclusion bodies from recombinant host cells is carried out by providing a washed recombinant host cell suspension, adding a detergent and subjecting the cells to mechanical disruption. A second detergent is added, followed by another mechanical disruption and centrifugation to a pellet. The pellet is resuspended in a buffer and subjected to another mechanical disruption. Inclusion bodies are washed, resuspended and then recovered. In addition, endotoxins and DNA are removed from inclusion bodies containing the 52/48 kDa tryptic fragment of vWF or peptide subfragments thereof by mechanically disrupting the inclusion bodies in an aqueous buffer containing a detergent. A washed pellet is formed from these mechanically disrupted inclusion bodies and dissolved in a denaturant. Alkylation is then performed and the suspension is subjected to column chromatography, where the alkylated fragment or subfragment is recovered by elution.

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Lafayette Hill, PA, United States of America

Thomas L Darling