Journal article
Affinity purification of secreted alkaline phosphatase produced by baculovirus expression vector system
Applied biochemistry and biotechnology, Vol.90(2), pp.125-136
02/2001
DOI: 10.1385/ABAB:90:2:125
PMID: 11297388
Abstract
Human secreted alkaline phosphatase (SEAP) was produced in a stablytransformed Spodoptera frugiperda Sf-9 insect cell line (Sfb4GalT) following infection with a recombinant Autographa californica multiple nuclear polyhedrovirus containing the SEAP gene under control of the polyhedrin promoter. An affinity chromatographic column prepared by linking 4-aminobenzylphosphonic acid to histidyl-expoxy-Sepharose was used to isolate SEAP from the cell supernatant following removal of cells and virus and 10-fold concentration through ultrafiltration. We found that the binding of SEAP on the affinity matrix follows the Langmuir isotherm model. In addition, either recycling SEAP sample through the column for 24 h or loading high SEAP concentrations resulted in a high-purity product. Some nonspecific binding of protein on the matrix occurred when low concentrations of SEAP sample were loaded. Finally, we found that SEAP binding occurs rapidly, i.e., within 30 min of adding the SEAP sample to the affinity matrix.
Details
- Title: Subtitle
- Affinity purification of secreted alkaline phosphatase produced by baculovirus expression vector system
- Creators
- Fuming Zhang - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IAMichael Wolff - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IADavid Williams - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IAKatie Busch - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IASybil Lang - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IADavid Murhammer - Department of Chemical and Biochemical Engineering University of Iowa 52242 Iowa City IARobert Linhardt - Department of Chemistry and Division of Medicinal and Natural Products Chemistry University of Iowa 52242 Iowa City IA
- Resource Type
- Journal article
- Publication Details
- Applied biochemistry and biotechnology, Vol.90(2), pp.125-136
- Publisher
- Humana Press; Totowa
- DOI
- 10.1385/ABAB:90:2:125
- PMID
- 11297388
- ISSN
- 0273-2289
- eISSN
- 1559-0291
- Language
- English
- Date published
- 02/2001
- Academic Unit
- Chemical and Biochemical Engineering
- Record Identifier
- 9984003401402771
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