| Autor: |
M, Schuste, E, Wasserbauer, A, Neubauer, A, Jungbauer |
| Rok vydání: |
2001 |
| Předmět: |
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| Zdroj: |
Bioseparation. 9(5) |
| ISSN: |
0923-179X |
| Popis: |
Immuno-affinity chromatography exploiting the Ca2+ dependent interaction of the anti-Flag antibody and Flag-tagged proteins has been investigated. The antibody has been immobilized on porous glass beads (Prosep) containing gigapores and on a monolith, the polymethacrylate based Convective Interactive Media (CIM) column at a ligand density of 2 mg/g and 10 mg/ml respectively. The performance of the columns was assessed by applying clarified yeast culture supernatant containing overexpressed Flag-human serum albumin. Dynamic binding capacity and purity was checked at various flow rates ranging from 100 cm/h to 800 cm/h. 95% purity could be obtained. Anti Flag-CIM columns showed a higher unspecific adsorption, requiring a longer wash cycle to obtain the same purity compared to the Prosep column. Anti Flag-CIM columns showed a flow independent performance, which is explained by its monolithic structure. A decreasing dynamic binding capacity with flow was observed with anti-Flag-Prosep columns. Both columns are suited to purify milligrams of protein out of a yeast culture supernatant within a few minutes. We considered them as promising candidates for high throughput screening, where fast purification is a necessity. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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