Low-cost customizable microscale toolkit for rapid screening and purification of therapeutic proteins
Autor: | Erick Gutierrez, David Burgenson, Sevda Deldari, Govind Rao, Chandrasekhar Gurramkonda, Abhay Andar, Douglas D. Frey, Mustafa Al-Adhami, Leah Tolosa, Yordan Kostov |
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Rok vydání: | 2018 |
Předmět: |
0106 biological sciences
0301 basic medicine Materials science Multiphysics Microfluidics Bioengineering CHO Cells 01 natural sciences Applied Microbiology and Biotechnology High-performance liquid chromatography Chromatography Affinity 03 medical and health sciences Cricetulus 010608 biotechnology Protein purification Granulocyte Colony-Stimulating Factor Animals Microscale chemistry Chromatography High Pressure Liquid Chromatography Equipment Design Biological materials 030104 developmental biology Biopharmaceutical Reagent Algorithms Biotechnology |
Zdroj: | Biotechnology and bioengineering. 116(4) |
ISSN: | 1097-0290 |
Popis: | Biopharmaceutical separations require tremendous amounts of optimization to achieve acceptable product purity. Typically, large volumes of reagents and biological materials are needed for testing different parameters, thus adding to the expense of biopharmaceutical process development. This study demonstrates a versatile and customizable microscale column (µCol) for biopharmaceutical separations using immobilized metal affinity chromatography (IMAC) as an example application to identify key parameters. µCols have excellent precision, efficiency, and reproducibility, can accommodate any affinity, ion-exchange or size-exclusion-based resin and are compatible with any high-performance liquid chromatography (HPLC) system. µCols reduce reagent amounts, provide comparable purification performance and high-throughput, and are easy to automate compared with current conventional resin columns. We provide a detailed description of the fabrication methods, resin packing methods, and µCol validation experiments using a conventional HPLC system. Finite element modeling using COMSOL Multiphysics was used to validate the experimental performance of the µCols. In this study, µCols were used for improving the purification achieved for granulocyte colony stimulating factor (G-CSF) expressed using a cell-free CHO in vitro translation (IVT) system and were compared to a conventional 1 ml IMAC column. Experimental data revealed comparable purity with a 10-fold reduction in the amount of buffer, resin, and purification time for the μCols compared with conventional columns for similar protein yields. |
Databáze: | OpenAIRE |
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