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CHEE450: Engineering Biology. IEX Chromatography. Presented by: Nikki Apostolakis Helen So Tiffany Yu. Chromatography Introduction.
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CHEE450: Engineering Biology IEX Chromatography Presented by: Nikki Apostolakis Helen So Tiffany Yu
Chromatography Introduction The term “chromatography” applies to a wide range of separation techniques that are based on the differential interaction of molecules between a moving phase and a stationary phase. Anion Exchanger Cation Exchanger vs. Column chromatography is a commonly used technique for protein purification. Different column chromatography methods exploit different properties of the protein; Ion exchange chromatography separates molecules based on charge.
Ion Exchange Principles • There are four main steps to IEX Chromatography: • Equilibration • Load and Wash • Elution • Regeneration Resins are the medium for separation and represent the stationary phase. The surface of these particles provide sites for selective binding. Separation is based on reversible adsorption of ions onto resin
Ion Exchange Principles Stationary Equilibration Load Wash Elution Regeneration
Ion Exchange Purpose • Intermediate separation and purification of MABs from bulk of impurities • Inlet stream Composition: • Acetic Acid • Impurities • MAB • Polysorbate80 • WFI
Design Considerations • COLUMN DESIGN • Flow rates • Aspect ratio (H, d) • Column material • Space constraints • COLUMN INTERNALS • Packing material • Buffer pH’s • Buffer composition
Chosen Design Chromaflow Columns • • Easily validated • • Cost-effective, safe packing/unpacking • • Appropriatefor most chromatography • techniques • Suitable for large scale GMP production
Chosen Design Chromaflow Columns Resin Type: Sepharose High Performance Resin Volume: 207.88 L Column Type: GE Chromaflow Column Dimensions: Inner diameter: 1800 mm Height: 100-300mm Pressure Rating: 3 bar
IEX Column Costing BPG BioProcess Columns
Alternatives CONFIGURATION Can make batch process continuous by running columns in parallel SEPARATION Change resin to have impurities bind and allow MAB to pass Use a different basis for separation (e.g. molecular size) Downstream Processing
THE END THANKS FOR LISTENING. ANY QUESTIONS?