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High-capacity composite adsorbents for nucleic acids
Peter Tiainen1, M Rokebul Anower, Per-Olof Larsson
1Division of Pure and Applied Biochemistry, Department of Chemistry, Lund University, P.O. Box 124, SE-221 00 Lund, Sweden.
Journal of Chromatography. A
|July 2, 2011
Summary
A novel composite material combining Cytopore™ and reticulated vitreous carbon (RVC) offers enhanced performance for column chromatography. This strong, high-capacity material efficiently binds DNA and RNA, enabling effective plasmid isolation.
Area of Science:
- Biomaterials Science
- Chromatography
- Cell Culture Technology
Background:
- Cytopore™ is a compressible, macroporous cellulose-based anion-exchange material for cell culture.
- Reticulated vitreous carbon (RVC) is a robust, non-compressible scaffold with high voidage.
- Combining these materials can leverage their complementary properties for advanced applications.
Purpose of the Study:
- To develop and characterize a composite material for column chromatography by integrating Cytopore™ with RVC.
- To evaluate the composite's mechanical stability, flow properties, and binding capacity.
- To demonstrate the composite's utility in isolating plasmid DNA.
Main Methods:
- Fabrication of cylindrical Cytopore™-RVC composites for chromatography columns.
- Assessment of bed compaction resistance and flow rates.
- Chromatographic runs with tracers to determine Height Equivalent to a Theoretical Plate (HETP).
- Determination of dynamic binding capacities for DNA and RNA.
- Isolation of pUC 18-type plasmids from alkaline lysate using the composite column.
- Confocal microscopy to visualize plasmid binding within the composite matrix.
Main Results:
- The Cytopore™-RVC composite exhibited excellent mechanical stability, withstanding very high flow rates (>25 column volumes/min) without bed compaction.
- Low HETP values (0.3 mm) indicated efficient pore flow.
- High dynamic binding capacities (10% breakthrough) of 240-370 mg/g dry weight Cytopore were observed for DNA and RNA.
- Successful isolation of pUC 18-type plasmids in good yield.
- Confocal microscopy revealed plasmid binding throughout the Cytopore™ matrix, explaining the high capacities.
Conclusions:
- The Cytopore™-RVC composite effectively combines the high binding capacity of Cytopore™ with the mechanical strength of RVC.
- This composite material is suitable for high-flow-rate column chromatography with superior binding performance.
- The strategy of combining weak, high-capacity materials with strong scaffolds is applicable to other chromatographic systems.
